Bone Substitutes and Membranes










913
35
Bone Substitutes and
Membranes
RALPH POWERS
S
ince acknowledging titanium as an inert substance capable of
binding to bone in humans, implant dentistry has ourished.
1,2
Dr. Per-Ingvar Brånemark is known as the “father of the mod-
ern dental implant” for taking a serendipitous nding in orthopedic
research and applying it to dentistry.
3
It took decades for him to
convince the medical and dental communities that titanium could
be integrated into living tissues, a process he called osseointegration.
Now dental implants are considered a standard of care and enjoy a
very high success rate. Having a reliable implant system is only part
of the equation. It is well understood that both bone volume and
bone quality are essential for implant placement and survival.
4
Most dental implants require some type of bone augmentation.
For many years autograft (considered the gold standard) was the
only material available as a bone void ller, and it still has a place in
many applications.
5
However, dramatic discoveries in biomaterials
and improvements in processing, preservation, and packaging have
made bone graft substitutes (BGS) available that are safe, eective,
accessible in sucient quantities, and suitable for almost all clinical
situations. In fact, the global BGS and dental membrane market is
expected to grow at a compound annual growth rate of 9.9% per year
through 2025.
6
Much of this increase is driven by the large expected
increase in dental implant and prosthetics work. e key factors driv-
ing the growth of this market include the growing geriatric popula-
tion and corresponding dental disorders, rising incidence of tooth
decay and edentulism, and the ability for more dental practitioners
to place implants and provide more complete treatment solutions.
It is up to the clinician to create an osseo-adaptive situation
when grafting, that is, having a host site prepared and a patient
sucient in health to allow for natural and predictable healing
after any type of augmentation procedure. e wise clinician will
be familiar with many types of BGS, as well as membranes, and
their properties. e clinician should develop a decision tree for
every clinical scenario based on a combination of empirical and
scientic support. It is hoped the following information will pro-
vide a basis for understanding the many bone graft and membrane
materials currently available.
Terminology for Bone Repair and
Regeneration
Bone remodeling is the ongoing process by which bone is resorbed
and replaced in a dynamic steady-state process that maintains
the health of bone. e process aects the entire skeleton all of
the time. Although bone may appear supercially as a static tis-
sue, it is actually very dynamic, undergoing constant remodeling
throughout the life of the vertebrate organism. Bone remodeling
is triggered by a need for calcium in the extracellular uid, but it
also occurs in response to mechanical stresses (microfracture) on
the bone tissue.
For the remodeling to occur, appropriate cell signaling occurs
to trigger osteoclasts to resorb the surface of the bone, followed by
deposition of bone by osteoblasts. Together the cells in any given
particular region of the bone surface that are responsible for bone
remodeling are known as the basic multicellular unit (BMU). e
action of osteoclasts and osteoblasts is synchronized: cells that
resorb and deposit bone, respectively.
To understand bone remodeling, you need to know about three
cell types found in bone:
• Osteoclastsarebone-resorbingcells (-clast means “to break”;
osteoclasts break down bone). ey are large, multinucleate
cells that form through the fusion of precursor cells. Unlike
osteoblasts, which are related to broblasts and other connec-
tive tissue cells, osteoclasts are descended from stem cells in the
bone marrow that also give rise to monocytes (macrophages).
Oneessentialfeatureistheingrowthofvasculartissue(neovas-
cularization); this is an essential feature of remodeling in that
the new vessels will carry cells and nutrients.
• Osteoblastsarebone-formingcells.eyareconnectivetissue
cells found at the surface of bone. ey can be stimulated to
proliferate and dierentiate as osteocytes. ey are recruited to
the area and form the lining of the newly created tunnel.
• Osteocytesarematurebonecells.Osteocytesmanufacturetype
I collagen and other substances that make up the bone extra-
cellular matrix. Osteocytes will be found enclosed in bone
7
(Fig. 35.1).
Fig. 35.2 includes multiple cells and shows the phases of bone
remodeling. e group of cells creating the tunnel through the
boneistheBMU(forming the cutting cone”).Osteoclastsare
followed by preosteoblasts that adhere to the new wall of the area
resorbed. As they mature to osteoblasts, they begin to secrete oste-
oid (immature bone matrix) in which they eventually are trapped
and where they live out the remainder of their existence as osteo-
cytes. is marks the start of reversal of the resorptive phase. e
nal product is the formation of Haversian canals (Fig. 35.3), or
osteons,” familiar to all as a normal observation in bone histology.

914
PART VII Soft and Hard Tissue Rehabilitation
In infants, bone “turnover” is rapid and may result in a 100%
new skeleton within 1 year; in adults, it is approximately 10%.
8
Statistics of adult bone remodeling are as follows:
• LifespanofBMUis6 to 9 months.
• SpeedofBMUis 25 μm/day.
• BonevolumereplacedbyasingleBMUis0.025 mm
3
.
• Lifespanofosteoclastsis2 weeks.
• Lifespanofosteoblastsis3 weeks.
• Intervalbetweensuccessiveremodelingeventsatthesameloca-
tion is 2 to 5 years.
• Rateofturnoverofwholeskeletonis10% per year.
e 10% per year approximation for the entire skeleton is
based on an average 4% turnover per year in cortical bone, which
represents roughly 75% of the entire skeleton, and an average
28% per year in trabecular (cancellous) bone, which represents
roughly 25% of the skeleton. As you can see, remodeling occurs
slowly and continuously.
Bone modeling adapts structure to loading and removes dam-
age as to maintain bone strength. It involves independent sites of
resorption and formation that change the size and shape of bones.
e stimulus is additional localized stress (as in orthodontic tooth
movement and weight training). is is described by Wol’s law,
which proposes that bone in a healthy person or animal will adapt
to the loads under which it is placed.
9
Modeling occurs at the cel-
lular level in a fashion similar to that described for remodeling.
Bone repair is a proliferative physiologic process in which the
body facilitates the repair of a bone fracture. Repair occurs in
response to trauma (fracture or overuse) and is the result of a com-
plicated cascade of events.
Bone regeneration is the regrowth of lost tissue. is requires
the use of surgical protocols that enable regeneration of the de-
cient sites, using the principles of osteogenesis, osteoinduction,
and osteoconduction (see more detailed denitions later in this
chapter).
• Guided bone regeneration (GBR) refersto alveolar ridge aug-
mentation or bone redevelopment (for implant placement or
to preserve the site for xed or removable bridgework); this
often requires the presence of a membrane to protect the
grafted area and restrict the entrance of unwanted cells. When
a graft is placed into a site (in, for example, a fresh extraction
socket), a competition occurs between soft tissue and bone-
forming cells to ll the surgical site. Soft tissue cells (epithe-
lial cells and broblasts) migrate at a very fast rate compared
with bone-formers. A properly chosen and placed membrane
Preosteoblasts
Osteoblasts
Lining cells
Osteocytes
Monocyte
Osteoid
New bone
New bone
Blood
vessel
Capillary
Osteoclast
MSC
MSC
Fig. . The Basics of Bone Remodeling. Involved are osteoclasts
(removing old or impaired bone), preosteoblast and osteoblasts (forming
new immature bone), and osteocytes trapped within the new bone matrix.
None of this can occur without new vasculature to bring in essential cells
and fluids. Mesenchymal stme cells (MSC) are the precursors to the bone
forming “blast” cells.
Tim
e
Cutting
cone
Closing
cone
Reversal
zone
Developing
resorption
cavity
Osteoclast
Fibroblast
Osteoblast
Quiescent
osteoblast
Blood
vessel
Resorption
cavity
Forming
osteon
Completed
osteon
Fig. . This is a more advanced look at the basic multicellular unit forming
the “cutting cone.” ( Adapted from: Roberts WE Garetto LP, Arbuckle GR et al:
What are the risk factors of osteoporosis? Assessing bone health, J Am Dent
Assoc 122:59-61, 1991. Source: https://basicmedicalkey.com/functional-
anatomy-of-the-musculoskeletal-system/)
Fig. . Osteons, or Haversian canals. Evidence that bone remodeling
occurred in the area.

915
CHAPTER 35 Bone Substitutes and Membranes
will reduce the competition. Below the membrane, regenera-
tion occurs. It involves the proliferation of new blood vessels
(angiogenesis) and the migration of bone-forming cells (osteo-
genesis). An initial blood clot will form, which is replaced by
brous bone. is material (called woven bone) is characterized
by a haphazard organization of collagen and is mechanically
weak.Laterthis will be transformedintoabetterorganized,
load-bearing “lamellar bone” (via normal bone remodeling).
10
• Guided tissue regeneration(GTR)involvesthesametechniques
used in GBR, but for redeveloping (regenerating) lost peri-
odontal tissues (cementum, periodontal ligament, alveolar
bone) to retain the natural dentition. Early research in this area
was instrumental in the development of the modern mem-
brane.
11
econcept of GBR was described rst in 1959 when cell-
occlusive membranes were employed for spinal fusions.
12
e
terms guided bone regeneration(GBR)andguided tissue regeneration
(GTR)oftenareusedsynonymouslyandratherinappropriately.
GTRdealswiththe regenerationofthesupportingperiodontal
apparatus, including cementum, periodontal ligament, and alveo-
larbone,whereasGBRreferstothepromotionofboneformation
alone.GBRandGTRarebasedonthesameprinciplesthatuse
barrier membranes for space maintenance over a defect, promot-
ing the ingrowth of osteogenic cells and preventing migration of
undesired cells from the overlying soft tissues into the wound.
Protection of a blood clot in the defect and exclusion of gingival
connective tissue and provision of a secluded space into which
osteogenic cell from the bone can migrate are essential for a suc-
cessful outcome. e sequence of bone healing is aected not only
by invasion of nonosteogenic tissue but more so by the defect size
and morphology.
Bone preservation indicates long-term stability of the alveolar
ridge. is is a general term for all of the previous terms in this
section.
Mechanisms of Bone Repair and
Regeneration
From the time of Hippocrates it has been known that bone has
considerable potential for regeneration and repair. Nicholas
Senn,
13
asurgeonatRushMedicalCollegeinChicago,described
the utility of antiseptic decalcied bone implants in the treatment
of osteomyelitis and certain bone deformities. Pierre Lacroix
14
proposed that there might be a hypothetical substance, osteo-
genin, that might initiate bone growth.
MarshallR.Uristprovidedthebiologicalbasisofbonemor-
phogenesis. Urist
15
made the key discovery that demineralized,
lyophilized segments of bone induced new bone formation when
implanted in muscle pouches in rabbits. is discovery was pub-
lished in 1965 in Science.
15
e term bone morphogenetic protein
(BMP) rst appeared in the scientic literature via the Journal of
Dental Research in 1971.
16
Bone morphogenetic (or morphoge-
neic) proteins are now referred to as BMPs for convenience.
Bone induction is a sequential multistep cascade. e key steps
in this cascade are chemotaxis, mitosis, and dierentiation.Chemo-
taxis is movement of a motile cell in a direction corresponding to
a gradient of increasing or decreasing concentration of a particu-
lar substance (such as a BMP). Mitosis is a type of cell division
that results in two daughter cells each having the same number
and kind of chromosomes as the parent nucleus. is is typical of
ordinary tissue growth. Dierentiation is the process by which a
cell becomes specialized to perform a specic function, as in the
case of a bone cell, a blood cell, or a neuron. ere are more than
250 general types of cells in the human body. For bone induction,
BMPs (uncovered by normal bone remodeling or exposed in the
matrix of a properly demineralized graft) signal for chemotaxis of
bone-forming cells to the bone void. e cells divide to increase
their number and mature to a more specialized form to produce
new immature bone material (osteoid). Over time this area is
remodeled to provide a better structure.
EarlystudiesbyHariReddiunraveledthesequenceofevents
involvedin bone matrix–induced bone morphogenesis. On the
basis of this work, it seemed likely that “morphogens” were present
in the bone matrix. A systematic study, using a battery of bioassays
for bone formation, was undertaken to isolate and purify putative
bone morphogenetic proteins.
17
It is well recognized that BMPs
can be found in properly prepared demineralized bone products
in the correct proportion to induce the sequential steps needed for
bone regeneration. To date, 20 BMPs have been identied.
18
Now
laboratory-produced recombinant human BMPs (rhBMPs) are
used in orthopedic applications (rhBMP-2 and rhBMP-7), such
as spinal fusions and nonunions. rhBMP-2 is U.S. Food and Drug
Administration (FDA) approved for some dental use.
In general, osteoinduction is the process by which osteogen-
esis is induced. It is a phenomenon regularly seen in any type of
bonehealingprocess.Osteoinductionimpliestherecruitmentof
immature cells and the stimulation of these cells to develop into
preosteoblasts. In a bone healing situation such as a fracture the
majorityofbonehealingisdependentonosteoinduction.Osteo-
conduction means that bone grows on a surface.
Currentlyosteogenesis can occur only with autografts. Examples
are the use of the rib, grafts from the chin, ascending ramus, ilium,
tibia, or outer table of the cranium, or from bone collected during
extractionorother dentalprocedure.Osteogenesisoccurswhen
vital osteoblasts originating from the bone graft material contrib-
ute to new bone growth along with bone growth generated via
osteoinduction and osteoconduction.
19
Osteoinduction has a rich research history and has been well
studied.Osteoinductivematerialswillrecruittheproperbonecells
toasite,andthesecellswillformbone.Osteoinductioncanpro-
duce bone where bone is not normally found (ectopic or hetero-
tropic sites). In fact, the early tests for a materials osteoinductive
potential” were placement of that material into the muscle pouch
of an animal. An example of a test animal is the “nude” mouse.
A nude mouse is a laboratory mouse from a strain with a genetic
mutation that causes a deteriorated or absent thymus, resulting in
an inhibited immune system because of a greatly reduced num-
ber of T cells. e phenotype (main outward appearance) of the
mouse is a lack of body hair, which gives it the “nude” nickname.
e nude mouse is valuable to research because it can receive
many dierent types of tissue grafts, as it mounts no rejection
response. erefore if new bone forms in the muscle pouch of a
nude mouse, it provides evidence of the potential for osteoinduc-
tivity. Now invitro tests have been developed to assess potential
osteoinductivity, although the invivo animal assay is considered
the gold standard.
20
Osteoconduction is the formation of bone on a surface. All
inert materials possess this characteristic. In bone regeneration,
healthy bone must be present adjacent to the site where the graft is
placed. Because bone will move from the healthy host site through
the grafting material placed in the bone void, this is commonly
referred to as “creeping substitution.Fig. 35.4 illustrates this phe-
nomenon with a cancellous-based product.

916
PART VII Soft and Hard Tissue Rehabilitation
It occurs similarly with a cortical material, but at a slower rate.
e rate of creeping substitution is based on the available space
for vascular ingrowth. Available space is based on particle spacing
(if a “powder” type material is used), as well as macroporosity and
microporosity of the material.
21
• Macroporosity(poresizegreaterthan100μm) is usually required
to facilitate the osteogenesis and angiogenesis. Interconnected
macropores are necessary to promote body uid circulation and
cell migration to the core of the implant. An example of macro-
porosity is the normal marrow space formed by trabeculation in
the cancellous portion of bone, or the interparticle spacing that
is created when particulate grafts are placed.
• Microporosity(poresizelessthan10μm) has importance, as
does the unique surface properties of microporous scaolds.
ese have considerable inuence on uid distribution and
protein adsorption. Moreover, capillary force generated by the
microporosity can improve the attachment of bone-related
cells on the scaolds surface and even make the cells achieve
penetration into the micropores smaller than them (Fig. 35.5).
Osteopromotion involves the enhancement of osteoinduction
without the material possessing osteoinductive properties. As an
example, enamel matrix derivative (xenograft based) has been
12
34
Fig. . “Creeping substitution.” This occurs with all materials to
some extent because all BGSs are osteoconductive. It requires the pres-
ence of healthy host bone in close proximity to the graft. Neovasculariza-
tion occurs in cells that will remove/repopulate the space. (Source: https://
pocketdentistry.com/basics-of-bone-grafting-and-graft-materials/ )
Granules
Macropore (>100µm)
Mesopore (10 - 100µm)
Micropore (<10µm)
Fig. . Different Types of Porosity. (Courtesy SigmaGraft Biomaterials, Fullerton, Calif.: http://sigmagraft.
com/inteross/ )

917
CHAPTER 35 Bone Substitutes and Membranes
shown to enhance the osteoinductive eect of demineralized
freeze-dried bone allograft (DFDBA) but will not stimulate new
bone growth alone.
22
Platelet-rich plasma and other substances
derived from the patients own blood are also examples.
Ideal Bone Graft Substitute
Reconstruction of bone defects or preparation of a site for
implant placement remains a challenge. On the one hand,
autografts harbor most features of “ideal” BGS; on the other
hand, they have a lot of insurmountable disadvantages (higher
level of surgical skill needed, insucient quantity, second-site
morbidity, increased operative time, etc.). An ideal bone graft
substitute should:
• bebiomechanicallystable;
• degradewithinanappropriatetimeframe;
• exhibitosteoconductive,osteogenic,andosteoinductiveprop-
erties; and
• provideafavorableenvironmentforinvadingbloodvesselsand
bone-forming cells.
Even though osteoconductivity of biomaterials for bone tissue
engineering (BTE) strategies can be directed by their composi-
tion, surface character, and internal structure, osteoinductive and
osteogenic features (discussed later) can be enhanced by the addi-
tion of osteopromotive materials.
23
Having the ideal substitute would address only a part of what
is needed for successful bone regeneration. As illustrated in Fig.
35.6, the bone healing triad is based on the complex process
involved in tissue repair, wherein the matrix/scaold (osteocon-
ductive materials), signaling proteins (osteoinductive, located
within the matrix), and tissue-forming cells (osteogenic, osteo-
clasts and osteoblasts) work in concert to form new tissue (bone)
in the healthy (osseo-adaptive) host over time.
Ideal Membrane Material
GBRisacommontechniqueinimplantdentistryforthetreat-
ment of bone defects. As discussed earlier, a critical component
oftheGBRprocedureistheuseofabarriermembrane.ese
materials are used to prevent the invasion of cells that are not
needed or would interfere with bone formation. e primary
goal is selective cell repopulation.
24
An ideal barrier membrane
should:
• Bebiologicallycompatible.ereshouldbenoinammation
or interaction between the barrier material and the host.
• Provide space maintenance. When desired, the membrane
should have the ability to prevent defect collapse.
• Stabilizethebloodclotthatformsaspartofnaturalhealing.
is will allow the regeneration process to progress and reduce
unwanted tissue integration into the defect.
• Provide cell occlusion. is is the primary function of the
membrane, but many membranes allow the passage of uid
that may assist in healing.
• Havesomedegreeofmechanicalstrength(basedonend-user
needs). Strength is needed, and in some cases shape memory is
desired.
• Resorbpredictably,pertheend-userrequirements.Fortunately,
many congurations with varying rates of resorption exist.
• Beeasytomodifyandmanipulate.
As you will see, there are many types of membranes to choose
from, most of which possess those properties. It is up to the clini-
cian to choose the barrier membrane that best provides for the
desired clinical outcome.
Classification of Bone Graft Substitutes and
Membranes
Transplant refers to the transfer of an organ from one body to
another, or from a certain section of the patient’s own body to
another area. is procedure is usually performed to replace a
damaged or missing organ. Tissues can be transferred from one
individual to another, and because they are generally placed to
encourage the body to heal itself, thus being incorporated into the
host, they are considered transplants.
Implants are medical devices intended to replace a missing body
part, support a damaged part, or enhance the body in some way.
Titanium dental implants are a good example. Some research-
ers consider allografts and xenografts (biological material) to be
implants because they are nonliving. It is acceptable to call them
either transplants or implants.
Autograft (or autotransplantation) is the transplantation of
functioning organs, tissues, or even particular proteins from one
part of the body to another in the same person. Examples in
implant dentistry are grafts from the ascending ramus, chin, or
iliac crest.
Allograft (or homograft) is the transplantation of cells, tissues,
or organs to a recipient from a genetically nonidentical donor
of the same species. It can also be called an allogeneic transplant.
Relatedtothisareisografts—agraftoftissuebetweentwoindi-
viduals who are genetically identical (i.e., monozygotic twins).
Demineralized freeze-dried bone and acellular dermis are exam-
ples used in implant dentistry.
Xenograft (or heterograft) is a tissue graft or organ transplant
from a donor of a dierent species from the recipient. Bovine
or porcine sourced materials (cancellous bone or collagen mem-
branes) are good examples. An interesting example of xenografts
is coral-derived materials. ese are considered xenografts (as
opposed to alloplasts) because of their organic nature.
Alloplast is an inorganic material used as a bone substitute or an
implant.Hydroxyapatite(HA)andtricalciumphosphate(TCP)
materials are examples.
Oversight
A 510(k) is a premarket submission made to the FDA to demon-
strate that a medical device is as safe and eective, that is, substan-
tiallyequivalent,asalegallymarketeddevice.Oncethedeviceis
determined to be substantially equivalent, it can then be marketed
Scaffolds
Adaptive
Time
Host
MoleculesCells
Bone
Fig. . The Bone Healing Triad. (Adapted from Murphy CM, O’Brien
FJ, Little DG, Schindeler A. Cell-scaffold interactions in the bone tissue
engineering triad. Eur Cell Mater. 2013;26:120-132.)

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91335Bone Substitutes and MembranesRALPH POWERSSince acknowledging titanium as an inert substance capable of binding to bone in humans, implant dentistry has ourished.1,2 Dr. Per-Ingvar Brånemark is known as the “father of the mod-ern dental implant” for taking a serendipitous nding in orthopedic research and applying it to dentistry.3 It took decades for him to convince the medical and dental communities that titanium could be integrated into living tissues, a process he called osseointegration. Now dental implants are considered a standard of care and enjoy a very high success rate. Having a reliable implant system is only part of the equation. It is well understood that both bone volume and bone quality are essential for implant placement and survival.4Most dental implants require some type of bone augmentation. For many years autograft (considered the gold standard) was the only material available as a bone void ller, and it still has a place in many applications.5 However, dramatic discoveries in biomaterials and improvements in processing, preservation, and packaging have made bone graft substitutes (BGS) available that are safe, eective, accessible in sucient quantities, and suitable for almost all clinical situations. In fact, the global BGS and dental membrane market is expected to grow at a compound annual growth rate of 9.9% per year through 2025.6 Much of this increase is driven by the large expected increase in dental implant and prosthetics work. e key factors driv-ing the growth of this market include the growing geriatric popula-tion and corresponding dental disorders, rising incidence of tooth decay and edentulism, and the ability for more dental practitioners to place implants and provide more complete treatment solutions.It is up to the clinician to create an osseo-adaptive situation when grafting, that is, having a host site prepared and a patient sucient in health to allow for natural and predictable healing after any type of augmentation procedure. e wise clinician will be familiar with many types of BGS, as well as membranes, and their properties. e clinician should develop a decision tree for every clinical scenario based on a combination of empirical and scientic support. It is hoped the following information will pro-vide a basis for understanding the many bone graft and membrane materials currently available.Terminology for Bone Repair and RegenerationBone remodeling is the ongoing process by which bone is resorbed and replaced in a dynamic steady-state process that maintains the health of bone. e process aects the entire skeleton all of the time. Although bone may appear supercially as a static tis-sue, it is actually very dynamic, undergoing constant remodeling throughout the life of the vertebrate organism. Bone remodeling is triggered by a need for calcium in the extracellular uid, but it also occurs in response to mechanical stresses (microfracture) on the bone tissue.For the remodeling to occur, appropriate cell signaling occurs to trigger osteoclasts to resorb the surface of the bone, followed by deposition of bone by osteoblasts. Together the cells in any given particular region of the bone surface that are responsible for bone remodeling are known as the basic multicellular unit (BMU). e action of osteoclasts and osteoblasts is synchronized: cells that resorb and deposit bone, respectively.To understand bone remodeling, you need to know about three cell types found in bone:• Osteoclastsarebone-resorbingcells (-clast means “to break”; osteoclasts break down bone). ey are large, multinucleate cells that form through the fusion of precursor cells. Unlike osteoblasts, which are related to broblasts and other connec-tive tissue cells, osteoclasts are descended from stem cells in the bone marrow that also give rise to monocytes (macrophages). Oneessentialfeatureistheingrowthofvasculartissue(neovas-cularization); this is an essential feature of remodeling in that the new vessels will carry cells and nutrients.• Osteoblastsarebone-formingcells.eyareconnectivetissuecells found at the surface of bone. ey can be stimulated to proliferate and dierentiate as osteocytes. ey are recruited to the area and form the lining of the newly created tunnel.• Osteocytesarematurebonecells.OsteocytesmanufacturetypeI collagen and other substances that make up the bone extra-cellular matrix. Osteocytes will be found enclosed in bone7 (Fig. 35.1).Fig. 35.2 includes multiple cells and shows the phases of bone remodeling. e group of cells creating the tunnel through the boneistheBMU(forming the “cutting cone”).Osteoclastsarefollowed by preosteoblasts that adhere to the new wall of the area resorbed. As they mature to osteoblasts, they begin to secrete oste-oid (immature bone matrix) in which they eventually are trapped and where they live out the remainder of their existence as osteo-cytes. is marks the start of reversal of the resorptive phase. e nal product is the formation of Haversian canals (Fig. 35.3), or “osteons,” familiar to all as a normal observation in bone histology. 914PART VII Soft and Hard Tissue RehabilitationIn infants, bone “turnover” is rapid and may result in a 100% new skeleton within 1 year; in adults, it is approximately 10%.8 Statistics of adult bone remodeling are as follows:• LifespanofBMUis∼6 to 9 months.• SpeedofBMUis∼ 25 μm/day.• BonevolumereplacedbyasingleBMUis∼0.025 mm3.• Lifespanofosteoclastsis∼2 weeks.• Lifespanofosteoblastsis∼3 weeks.• Intervalbetweensuccessiveremodelingeventsatthesameloca-tion is ∼2 to 5 years.• Rateofturnoverofwholeskeletonis∼10% per year.e 10% per year approximation for the entire skeleton is based on an average 4% turnover per year in cortical bone, which represents roughly 75% of the entire skeleton, and an average 28% per year in trabecular (cancellous) bone, which represents roughly 25% of the skeleton. As you can see, remodeling occurs slowly and continuously.Bone modeling adapts structure to loading and removes dam-age as to maintain bone strength. It involves independent sites of resorption and formation that change the size and shape of bones. e stimulus is additional localized stress (as in orthodontic tooth movement and weight training). is is described by Wol’s law, which proposes that bone in a healthy person or animal will adapt to the loads under which it is placed.9 Modeling occurs at the cel-lular level in a fashion similar to that described for remodeling.Bone repair is a proliferative physiologic process in which the body facilitates the repair of a bone fracture. Repair occurs inresponse to trauma (fracture or overuse) and is the result of a com-plicated cascade of events.Bone regeneration is the regrowth of lost tissue. is requires the use of surgical protocols that enable regeneration of the de-cient sites, using the principles of osteogenesis, osteoinduction, and osteoconduction (see more detailed denitions later in this chapter).•  Guided bone regeneration (GBR) refersto alveolar ridge aug-mentation or bone redevelopment (for implant placement or to preserve the site for xed or removable bridgework); this often requires the presence of a membrane to protect the grafted area and restrict the entrance of unwanted cells. When a graft is placed into a site (in, for example, a fresh extraction socket), a competition occurs between soft tissue and bone-forming cells to ll the surgical site. Soft tissue cells (epithe-lial cells and broblasts) migrate at a very fast rate compared with bone-formers. A properly chosen and placed membrane PreosteoblastsOsteoblastsLining cellsOsteocytesMonocyteOsteoidNew boneNew boneBloodvesselCapillaryOsteoclastMSCMSC• Fig. . The Basics of Bone Remodeling. Involved are osteoclasts (removing old or impaired bone), preosteoblast and osteoblasts (forming new immature bone), and osteocytes trapped within the new bone matrix. None of this can occur without new vasculature to bring in essential cells and fluids. Mesenchymal stme cells (MSC) are the precursors to the bone forming “blast” cells.TimeCuttingconeClosingconeReversalzoneDevelopingresorptioncavityOsteoclastFibroblastOsteoblastQuiescentosteoblastBloodvesselResorptioncavityFormingosteonCompletedosteon• Fig. . This is a more advanced look at the basic multicellular unit forming the “cutting cone.” ( Adapted from: Roberts WE Garetto LP, Arbuckle GR et al: What are the risk factors of osteoporosis? Assessing bone health, J Am Dent Assoc 122:59-61, 1991. Source: https://basicmedicalkey.com/functional- anatomy-of-the-musculoskeletal-system/)• Fig. . Osteons, or Haversian canals. Evidence that bone remodeling occurred in the area. 915CHAPTER 35 Bone Substitutes and Membraneswill reduce the competition. Below the membrane, regenera-tion occurs. It involves the proliferation of new blood vessels (angiogenesis) and the migration of bone-forming cells (osteo-genesis). An initial blood clot will form, which is replaced by brous bone. is material (called woven bone) is characterized by a haphazard organization of collagen and is mechanically weak.Laterthis will be transformedintoabetterorganized,load-bearing “lamellar bone” (via normal bone remodeling).10•  Guided tissue regeneration(GTR)involvesthesametechniquesused in GBR, but for redeveloping (regenerating) lost peri-odontal tissues (cementum, periodontal ligament, alveolar bone) to retain the natural dentition. Early research in this area was instrumental in the development of the modern mem-brane.11econcept of GBR was described rst in 1959 when cell-occlusive membranes were employed for spinal fusions.12 e terms guided bone regeneration(GBR)andguided tissue regeneration (GTR)oftenareusedsynonymouslyandratherinappropriately.GTRdealswiththe regenerationofthesupportingperiodontalapparatus, including cementum, periodontal ligament, and alveo-larbone,whereasGBRreferstothepromotionofboneformationalone.GBRandGTRarebasedonthesameprinciplesthatusebarrier membranes for space maintenance over a defect, promot-ing the ingrowth of osteogenic cells and preventing migration of undesired cells from the overlying soft tissues into the wound. Protection of a blood clot in the defect and exclusion of gingival connective tissue and provision of a secluded space into which osteogenic cell from the bone can migrate are essential for a suc-cessful outcome. e sequence of bone healing is aected not only by invasion of nonosteogenic tissue but more so by the defect size and morphology.Bone preservation indicates long-term stability of the alveolar ridge. is is a general term for all of the previous terms in this section. Mechanisms of Bone Repair and RegenerationFrom the time of Hippocrates it has been known that bone has considerable potential for regeneration and repair. Nicholas Senn,13asurgeonatRushMedicalCollegeinChicago,describedthe utility of antiseptic decalcied bone implants in the treatment of osteomyelitis and certain bone deformities. Pierre Lacroix14 proposed that there might be a hypothetical substance, osteo-genin, that might initiate bone growth.MarshallR.Uristprovidedthebiologicalbasisofbonemor-phogenesis. Urist15 made the key discovery that demineralized, lyophilized segments of bone induced new bone formation when implanted in muscle pouches in rabbits. is discovery was pub-lished in 1965 in Science.15 e term bone morphogenetic protein (BMP) rst appeared in the scientic literature via the Journal of Dental Research in 1971.16 Bone morphogenetic (or morphoge-neic) proteins are now referred to as BMPs for convenience.Bone induction is a sequential multistep cascade. e key steps in this cascade are chemotaxis, mitosis, and dierentiation.Chemo-taxis is movement of a motile cell in a direction corresponding to a gradient of increasing or decreasing concentration of a particu-lar substance (such as a BMP). Mitosis is a type of cell division that results in two daughter cells each having the same number and kind of chromosomes as the parent nucleus. is is typical of ordinary tissue growth. Dierentiation is the process by which a cell becomes specialized to perform a specic function, as in the case of a bone cell, a blood cell, or a neuron. ere are more than 250 general types of cells in the human body. For bone induction, BMPs (uncovered by normal bone remodeling or exposed in the matrix of a properly demineralized graft) signal for chemotaxis of bone-forming cells to the bone void. e cells divide to increase their number and mature to a more specialized form to produce new immature bone material (osteoid). Over time this area isremodeled to provide a better structure.EarlystudiesbyHariReddiunraveledthesequenceofeventsinvolvedin bone matrix–induced bone morphogenesis. On thebasis of this work, it seemed likely that “morphogens” were present in the bone matrix. A systematic study, using a battery of bioassays for bone formation, was undertaken to isolate and purify putative bone morphogenetic proteins.17 It is well recognized that BMPs can be found in properly prepared demineralized bone products in the correct proportion to induce the sequential steps needed for bone regeneration. To date, 20 BMPs have been identied.18 Now laboratory-produced recombinant human BMPs (rhBMPs) are used in orthopedic applications (rhBMP-2 and rhBMP-7), such as spinal fusions and nonunions. rhBMP-2 is U.S. Food and Drug Administration (FDA) approved for some dental use.In general, osteoinduction is the process by which osteogen-esis is induced. It is a phenomenon regularly seen in any type of bonehealingprocess.Osteoinductionimpliestherecruitmentofimmature cells and the stimulation of these cells to develop into preosteoblasts. In a bone healing situation such as a fracture the majorityofbonehealingisdependentonosteoinduction.Osteo-conduction means that bone grows on a surface.Currentlyosteogenesis can occur only with autografts. Examples are the use of the rib, grafts from the chin, ascending ramus, ilium, tibia, or outer table of the cranium, or from bone collected during extractionorother dentalprocedure.Osteogenesisoccurswhenvital osteoblasts originating from the bone graft material contrib-ute to new bone growth along with bone growth generated via osteoinduction and osteoconduction.19Osteoinduction has a rich research history and has been well studied.Osteoinductivematerialswillrecruittheproperbonecellstoasite,andthesecellswillformbone.Osteoinductioncanpro-duce bone where bone is not normally found (ectopic or hetero-tropic sites). In fact, the early tests for a material’s osteoinductive “potential” were placement of that material into the muscle pouch of an animal. An example of a test animal is the “nude” mouse. A nude mouse is a laboratory mouse from a strain with a genetic mutation that causes a deteriorated or absent thymus, resulting in an inhibited immune system because of a greatly reduced num-ber of T cells. e phenotype (main outward appearance) of the mouse is a lack of body hair, which gives it the “nude” nickname. e nude mouse is valuable to research because it can receive many dierent types of tissue grafts, as it mounts no rejection response. erefore if new bone forms in the muscle pouch of a nude mouse, it provides evidence of the potential for osteoinduc-tivity. Now invitro tests have been developed to assess potential osteoinductivity, although the invivo animal assay is considered the gold standard.20Osteoconduction is the formation of bone on a surface. All inert materials possess this characteristic. In bone regeneration, healthy bone must be present adjacent to the site where the graft is placed. Because bone will move from the healthy host site through the grafting material placed in the bone void, this is commonly referred to as “creeping substitution.” Fig. 35.4 illustrates this phe-nomenon with a cancellous-based product. 916PART VII Soft and Hard Tissue RehabilitationIt occurs similarly with a cortical material, but at a slower rate. e rate of creeping substitution is based on the available space for vascular ingrowth. Available space is based on particle spacing (if a “powder” type material is used), as well as macroporosity and microporosity of the material.21• Macroporosity(poresizegreaterthan100μm) is usually required to facilitate the osteogenesis and angiogenesis. Interconnected macropores are necessary to promote body uid circulation and cell migration to the core of the implant. An example of macro-porosity is the normal marrow space formed by trabeculation in the cancellous portion of bone, or the interparticle spacing that is created when particulate grafts are placed.• Microporosity(poresizelessthan10μm) has importance, as does the unique surface properties of microporous scaolds. ese have considerable inuence on uid distribution and protein adsorption. Moreover, capillary force generated by the microporosity can improve the attachment of bone-related cells on the scaolds surface and even make the cells achieve penetration into the micropores smaller than them (Fig. 35.5).Osteopromotion involves the enhancement of osteoinduction without the material possessing osteoinductive properties. As an example, enamel matrix derivative (xenograft based) has been 1234• Fig. . “Creeping substitution.” This occurs with all materials to some extent because all BGSs are osteoconductive. It requires the pres-ence of healthy host bone in close proximity to the graft. Neovasculariza-tion occurs in cells that will remove/repopulate the space. (Source: https://pocketdentistry.com/basics-of-bone-grafting-and-graft-materials/ )GranulesMacropore (>100µm)Mesopore (10 - 100µm)Micropore (<10µm)• Fig. . Different Types of Porosity. (Courtesy SigmaGraft Biomaterials, Fullerton, Calif.: http://sigmagraft.com/inteross/ ) 917CHAPTER 35 Bone Substitutes and Membranesshown to enhance the osteoinductive eect of demineralized freeze-dried bone allograft (DFDBA) but will not stimulate new bone growth alone.22 Platelet-rich plasma and other substances derived from the patient’s own blood are also examples. Ideal Bone Graft SubstituteReconstruction of bone defects or preparation of a site forimplant placement remains a challenge. On the one hand,autografts harbor most features of “ideal” BGS; on the other hand, they have a lot of insurmountable disadvantages (higher level of surgical skill needed, insucient quantity, second-site morbidity, increased operative time, etc.). An ideal bone graft substitute should:• bebiomechanicallystable;• degradewithinanappropriatetimeframe;• exhibitosteoconductive,osteogenic,andosteoinductiveprop-erties; and• provideafavorableenvironmentforinvadingbloodvesselsandbone-forming cells.Even though osteoconductivity of biomaterials for bone tissue engineering (BTE) strategies can be directed by their composi-tion, surface character, and internal structure, osteoinductive and osteogenic features (discussed later) can be enhanced by the addi-tion of osteopromotive materials.23Having the ideal substitute would address only a part of what is needed for successful bone regeneration. As illustrated in Fig. 35.6, the bone healing triad is based on the complex process involved in tissue repair, wherein the matrix/scaold (osteocon-ductive materials), signaling proteins (osteoinductive, located within the matrix), and tissue-forming cells (osteogenic, osteo-clasts and osteoblasts) work in concert to form new tissue (bone) in the healthy (osseo-adaptive) host over time. Ideal Membrane MaterialGBRisacommontechniqueinimplantdentistryforthetreat-ment of bone defects. As discussed earlier, a critical component oftheGBRprocedureistheuseofabarriermembrane.esematerials are used to prevent the invasion of cells that are not needed or would interfere with bone formation. e primary goal is selective cell repopulation.24 An ideal barrier membrane should:• Bebiologicallycompatible.ereshouldbenoinammationor interaction between the barrier material and the host.• Provide space maintenance. When desired, the membraneshould have the ability to prevent defect collapse.• Stabilizethebloodclotthatformsaspartofnaturalhealing.is will allow the regeneration process to progress and reduce unwanted tissue integration into the defect.• Provide cell occlusion. is is the primary function of themembrane, but many membranes allow the passage of uid that may assist in healing.• Havesomedegreeofmechanicalstrength(basedonend-userneeds). Strength is needed, and in some cases shape memory is desired.• Resorbpredictably,pertheend-userrequirements.Fortunately,many congurations with varying rates of resorption exist.• Beeasytomodifyandmanipulate.As you will see, there are many types of membranes to choose from, most of which possess those properties. It is up to the clini-cian to choose the barrier membrane that best provides for the desired clinical outcome. Classification of Bone Graft Substitutes and MembranesTransplant refers to the transfer of an organ from one body to another, or from a certain section of the patient’s own body to another area. is procedure is usually performed to replace a damaged or missing organ. Tissues can be transferred from one individual to another, and because they are generally placed to encourage the body to heal itself, thus being incorporated into the host, they are considered transplants.Implants are medical devices intended to replace a missing body part, support a damaged part, or enhance the body in some way. Titanium dental implants are a good example. Some research-ers consider allografts and xenografts (biological material) to be implants because they are nonliving. It is acceptable to call them either transplants or implants.Autograft (or autotransplantation) is the transplantation of functioning organs, tissues, or even particular proteins from one part of the body to another in the same person. Examples in implant dentistry are grafts from the ascending ramus, chin, or iliac crest.Allograft (or homograft) is the transplantation of cells, tissues, or organs to a recipient from a genetically nonidentical donor of the same species. It can also be called an allogeneic transplant. Relatedtothisareisografts—agraftoftissuebetweentwoindi-viduals who are genetically identical (i.e., monozygotic twins). Demineralized freeze-dried bone and acellular dermis are exam-ples used in implant dentistry.Xenograft (or heterograft) is a tissue graft or organ transplant from a donor of a dierent species from the recipient. Bovine or porcine sourced materials (cancellous bone or collagen mem-branes) are good examples. An interesting example of xenografts is coral-derived materials. ese are considered xenografts (as opposed to alloplasts) because of their organic nature.Alloplast is an inorganic material used as a bone substitute or an implant.Hydroxyapatite(HA)andtricalciumphosphate(TCP)materials are examples. OversightA 510(k) is a premarket submission made to the FDA to demon-strate that a medical device is as safe and eective, that is, substan-tiallyequivalent,asalegallymarketeddevice.Oncethedeviceisdetermined to be substantially equivalent, it can then be marketed ScaffoldsAdaptiveTimeHostMoleculesCellsBone• Fig. . The Bone Healing Triad. (Adapted from Murphy CM, O’Brien FJ, Little DG, Schindeler A. Cell-scaffold interactions in the bone tissue engineering triad. Eur Cell Mater. 2013;26:120-132.) 918PART VII Soft and Hard Tissue Rehabilitationin the United States. Two years after the Medical Device Amend-ments of 1976 were enacted, the FDA issued its nal draft of the medical device Good Manufacturing Practice (GMP) regulation, a series of requirements that prescribed the facilities, methods, and controls to be used in the manufacture, packaging, and storage of medical devices. All 510(k) products must be manufactured under GMP, and proof that they are is part of the process for obtaining approval. Xenograft and alloplast materials fall under the 501(k) requirements. Products going through the 510(k) pathway can have intended applications or claims.An investigational device exemption (IDE) allows the investiga-tional device to be used in a clinical study to collect safety and eectiveness data. Clinical studies are most often conducted tosupportapremarketapproval.Onlyasmallpercentageof510(k)srequire clinical data to support the application. Investigational use also includes clinical evaluation of certain modications or new intended uses of legally marketed devices. All clinical evaluations of investigational devices, unless exempt, must have an approved IDE before the study is initiated. It is rare that a product intended to be used as a BGS or membrane in dentistry requires an IDE and premarket approval. If the device does need clinical evaluation and has not been cleared for marketing, these are required:• an investigational plan approved by an institutional reviewboard• informedconsentfromallpatients• labelingstatingthatthedeviceisforinvestigationaluseonly• monitoringofthestudy• requiredrecordsandreportsGoodClinicalPracticereferstothe regulations and require-ments that must be complied with while conducting a clinical study. ese regulations apply to the manufacturers, sponsors, clinical investigators, institutional review boards, and the medical device.Products containing or consisting of human cells or tissues that are intended for implantation, transplantation, infusion, or trans-fer into a human recipient, and are considered minimally manipu-lated, are called human cellular and tissue-based products(HCT/Ps).HCT/Psmustalsobeintendedforhomologoususeandcan-not be combined with another article (except for water, crystal-loids, or a sterilizing, preserving, or storage agent). e product must have no systemic eect and cannot be dependent on the metabolic activity of living cells for the primary function.Allograft products are considered medical devices when the FDA determines that they have been more than minimally pro-cessed. To be considered beyond minimally processed, an original characteristic of a structural tissue must be altered, and that char-acteristic has to be relevant in that it has a potential eect on the utility of the tissue for reconstruction, repair, or replacement.ManufacturersofHCT/Ps(i.e.,tissueprocessors)arerequiredbytheFDAtocomplywithCurrentGoodTissuePractice.isincludes proper handling, processing, labeling, and recordkeeping procedures. Under these regulations, tissue banks must screen and test all donors for risk factors and clinical evidence of relevant communicable disease agents.With the rapid growth of all areas of tissue banking, there has been an increasing need for accountability and for measures that ensure that safe, quality tissues are available for clinical use. Qual-ity improvement can be aected through voluntary standards, and most tissue banks have incorporated the achievement of high standards into their goals. e American Association of Tissue Banks (AATB) has established comprehensive standards for donor screening, recovery and processing of musculoskeletal, cardiac, vascular, and skin tissues, and reproductive cells.25 In addition, the standards contain institutional requirements; descriptions of required functional components of a tissue bank; requirements for construction and management of records and development of procedures; requirements for informed consent, tissue labeling, storage, and release; expectations for handling adverse outcomes, investigations, and tissue recalls; requirements for establishment of a quality program; specications for equipment and facilities; and guidelines for tissue-dispensing services and tissue distribu-tion intermediaries. AATB’s Standards for Tissue Banking are con-sulted not only by tissue bankers but also by end-user healthcare facilities, other standard-setting organizations, and regulators worldwide. In 2018, more than 100 tissue banks in North Amer-ica held AATB accreditation. Best practice for checking a tissue bank’s accreditation status is to perform an accredited bank search on the AATB website.26eJointCommissionhasstandardsforstorageandissuanceof tissue for hospitals and ambulatory surgery centers. ese standards apply to bone, tendon, fascia, and cartilage, as well as cellular tissues of both human and animal (xenograft) origin. e standards address key functions, including the need to develop procedures for tissue acquisition and storage, recordkeeping and tracking, and follow-up of adverse events and suspected allograft-caused infections, which must be reported to the tis-sue bank from which the tissue was obtained. Similar to federal regulations and AATB Standards, the minimal record retention period is specied to be 10 years from the date of transplanta-tion, distribution, other disposition, or expiration, whichever is latest.FDA authority to create and “enforce regulations necessary to prevent the introduction, transmission, or spread of com-municable diseases between the States or from foreign countries into the States” under Section 361(a) of the U.S. Public Health ServiceAct(42USC264)appliestohumantissueintendedfortransplantation. Formal enforcement policy and regulations did notexistuntilDecember14,1993(codiedin21CFRParts16and1270),whenthe“InterimRule:HumanTissueIntendedforTransplantation,” which required donor screening, infectious dis-ease testing, and recordkeeping “to prevent transmission of infec-tious diseases through human tissue used in transplantation,” was adopted in response to reports of HIV transmission by human tissue and of potentially unsafe bone imported into the United States.27ese regulations were supplanted by a series of federal reg-ulations, published in stages, rst announced in the Proposed ApproachtotheRegulationofCellularandTissue-BasedProductsinMarch1997.Analrule,“HumanCells,Tissues,andCellularandTissue-BasedProducts:EstablishmentRegistrationandList-ing,” published in January 2001, required organizations that are engaged in tissue recovery, donor qualication, tissue processing, and/or tissue-related laboratory testing to register as a tissue estab-lishment with the FDA. e rule (21CFR Part 1271)becameeective for all tissue banks on March 29, 2004.A nal rule, “Eligibility Determination for Donors of Human Cells,Tissues,andCellularandTissue-BasedProducts,”publishedMay 25, 2004, set forth donor eligibility requirements, including health history screening and laboratory testing. Another nal rule, “CurrentGoodTissuePracticeforHumanCell,TissueandCel-lular and Tissue-Based Product Establishments; Inspection and Enforcement,” published on November 24, 2004, established ele-ments of good tissue practice, analogous to GMP for blood banks. Both rules became eective May 25, 2005.  919CHAPTER 35 Bone Substitutes and MembranesSterilitySterility will be discussed from the viewpoint of allografts because they are the most dicult to sterilize because of their fragile bio-logic nature and variety.28Tissue sterilization is dened as the killing or elimination of all microorganisms from allograft tissue, whereas disinfection refers to the removal of microbial contamination. e Association for the Advancement of Medical Instrumentation, a standard-setting organization for the medical instrumentation and technology industry,denessterilityassurancelevel(SAL)astheprobabil-ity that an individual device, dose, or unit is nonsterile (i.e., one or more viable microorganisms being present) after it has been exposed to a validated sterilization process. Although absolute sterility in theory would represent an absence of any pathogen, SALis generally applied only tothe levelof possible contami-nation with bacteria or parasites. In contrast with log reduction of viruses determined in assessments of virus reduction methods, SALisanabsolutedeterminedbytheabilityofthemethodtoeradicate or reduce microorganisms, the susceptibility of organ-isms that may be present to the sterilization method applied, and the maximal bioburden that could occur in the initial material. Forexample,aSALof10−6 means that there is less than a 1 in 1,000,000 chance of a viable microorganism remaining after the sterilization procedure. e FDA requires that medical devices be sterilizedusingamethodvalidatedtoachieveaSALof10−6. A medical device derived from or that includes a biological product componentmustalsomeet aSALof10−6 if it is to be labeled sterile.ASALof10−3, or a 1 in 1000 chance of a viable micro-organism being present, is a more achievable goal selected by some processors for aseptically processed tissues if the processor has been unable to validate their process to the more stringent SALof10−6 level, or if the tissues are unable to withstand the harshtreatmentneededtoachieveamorerestrictiveSALwithoutan impairment of tissue function. Such tissues may not then be labeled as sterile.e complex physical structures and density of musculoskel-etal tissues pose challenges for adequate penetration of antimi-crobial agents to eradicate microorganisms. Allografts will not tolerate methods usually applied to metal and plastic medical devices because such treatment would impair the mechanical and biologic properties necessary for clinical utility. As an alternative, sterilization of tissues has been accomplished by several methods, includingheat,chemicals,ethyleneoxidegas,supercriticalCO2, and gamma or electron beam irradiation. However, not all steril-ants have adequate tissue penetration. is is particularly the case for gases and liquids. e initial bioburden, which may be high in some tissues, must be considered. Some tissues are treated with antibiotics invitro before storage, but this treatment decontami-nates only the surface and may be eective against bacteria only.A variety of methods, including chemical treatments and irra-diation, has been used to reduce or eliminate pathogens in tissue intended for transplantation. e introduction of bone steriliza-tion by ethylene oxide gas simplied bone processing and facili-tated the widespread use of sterilized air-dried and lyophilized bone products. e eects of ethylene oxide treatment on the biomechanical and osteoinductive capacity of bone allografts have been questioned, although animal studies have yielded incon-sistent results. ese concerns, combined with those regarding the carcinogenic potential of ethylene oxide and its breakdown products, have largely led to abandonment of this method in the United States and the United Kingdom.First introduced in the 1960s, gamma irradiation of bone is still used widely, usually employing a cobalt-60 source. e gamma rays penetrate bone eectively and work by generating free radicals, which may have adverse eects on collagen and limit utility in soft tissues unless performed in a controlled dose fashion at ultra-low temperature. e minimal bactericidal level of gamma irradiation is 10 to 20 kGy (1 kGy = 100,000 rad). Uncontrolled human studies have shown irradiated, calcied, and demineralized bone grafts to be clinically eective. Numerous studies have shown that mineralized bone allografts irradiated at 25 to 30 kGy are also clinically eec-tive, with high success rates reported. In controlled studies the clini-cal eectiveness of bone allografts subjected to 25 kGy irradiation was comparable with that of nonirradiated bone grafts, although doses exceeding 25 kGy for cortical bone and 60 kGy for cancellous bone have been found to induce cross-linking of collagen and to impair mechanical function in a dose-dependent fashion. ere is invitro evidence that high irradiation reduces osteoclast activity and increases osteoblast apoptosis (programmed cell death), and that residual bacterial products induce inammatory bone resorption after macrophage inactivation. However, the clinical signicance of these ndings has not been established. Newer processes employ-ing radioprotectants have preserved bone allograft integrity when doses ≥25 kGy are applied, and controlled-dose methods permit successful irradiation at lower doses (see Proprietary Sterilization Methods to follow). Irradiated demineralized bone has potential osteoinductive activity and has been eective in nonstructural clini-cal applications.Concerns about pathogen transmission and the limitations ofirradiation, especially for soft tissues, have prompted improve-ments in sterilization methods and in the validation of these meth-ods. A number of proprietary chemical-based processing methods have been developed with aims of eectively penetrating tissues and reducing, killing, or inactivating microorganisms and viruses without unacceptable adverse eects on the tissue’s biomechanical properties. In addition, for use in transplantation, the agents must either be able to be eectively removed or be nontoxic. All methods in current use are applied only to tissue from donors who have met stringent criteria for medical history and behavioral risk assessment as well as negative results on infectious disease marker testing.Proprietary Sterilization Methodse Tutoplast process (Tutogen Medical, Gainesville, Fla.) was the rst process to sterilize and preserve tissue without aecting biological or mechanical properties. e process has been in use since the early 1970s for a variety of hard and soft tissues, includ-ing bone, fascia lata, pericardium, skin, amniotic membrane, and sclera. Initially lipids are removed in an ultrasonic acetone bath that also inactivates enveloped viruses and reduces prion activity. Bac-teria are destroyed using alternating hyperosmotic saline and puri-ed water baths that also wash out cellular debris. Soluble proteins, nonenveloped viruses, and bacterial spores are destroyed in mul-tiple hydrogen peroxide baths, and a 1N sodium hydroxide treat-ment further reduces prion infectivity by 6 logs. A nal acetone wash removes any residual prions and inactivates any remaining enveloped viruses. Vacuum extraction dehydrates the tissue before the grafts are shaped and then double-barrier packaged. Terminal sterilizationusinglow-dosegammairradiationyieldsaSALof10−6.e Allowash XG process (LifeNet Health [LNH], VirginiaBeach, Va.) employs six steps: (1) bioburden control, (2) biobur-den assessment, (3) minimization of contamination during pro-cessing, (4) rigorous cleaning, (5) disinfection steps, and (6) a nal 920PART VII Soft and Hard Tissue Rehabilitationstep of low-temperature, controlled-dose gamma irradiation. e processhasbeenvalidatedtoachieveaSALof10−6. Holtzclaw etal.29 in 2008 compared the Allowash XG and Tutoplast meth-ods, and found that each achieved medical-grade sterility with no eect on biological or biomechanical properties.eBioCleanseprocess(RegenerationTechnologies,Alachua,Fla.) uses low-temperature addition of chemical sterilants, such as hydrogen peroxide and isopropyl alcohol, which permeate the tissue’s inner matrix, followed by pressure variations intended to drivethesterilantsintoandoutofthetissue.RegenerationTech-nologies reports a SAL of 10−6 for soft tissues without adverse eects on the initial allograft mechanical properties.eClearantprocess(Clearant,LosAngeles,Calif.)isdesignedto avoid the negative eects of gamma irradiation through addi-tion of free radical scavengers, using pretreatment dimethyl sulf-oxide and propylene glycol as radioprotectants. Although the processsubjectstissueto50kGyradiationandachievesaSALof10−6 for bacteria, fungi, yeast, and spores, the tissue’s biomechani-cal properties are retained.e Musculoskeletal Transplant Foundation (Edison, N.J.) uses a series of chemicals, including nonionic detergents, hydro-gen peroxide, and alcohol, to treat cortical and cancellous bone grafts. For soft tissues an antibiotic mixture containing gentami-cin, amphotericin B, and Primaxin is added and then washed out to a nondetectable concentration. e Musculoskeletal Transplant FoundationclaimsaSALof10−3 for its products. Incoming tis-sues whose bioburden exceeds prescribed parameters are pretreated with low-dose gamma irradiation.NovaSterilis (Lansing, N.Y.) has developed a sterilizationtechnique that uses supercritical carbon dioxide at low tem-peratures and relatively low pressures, resulting in transient acidication, which is lethal to bacteria and viruses, with good penetration reported. However, this technique only recently became available for clinically available allografts, and data on clinical ecacy and retention of allograft mechanical proper-ties are limited. End-User ResponsibilitiesInformed Consent. e clinician is, as dened by the AATB and other organizations, the end-user. As such, the clinician’s responsibility is the safest and most ecacious treatment for his or her patient. is begins with a complete understanding of the characteristics and limitations of any material used in treatment. Second, the patient expectations and concerns must be assessed. is is part of the process of informed consent. It is important that proper informed consent is obtained.eDoctrineof InformedConsent30 is based on signicant history:•  Schloendor v. Society of New York Hospital in 1914: An opera-tion was performed against the patient’s wishes.•  Salgo v. Leland Stanford Jr. University Board of Trustees in 1957: e patient had not been informed of the risks involved with the surgery performed.•  Natanson v. Kline in 1960: is verdict established a standard that the risks as understood by a reasonable practitioner must be disclosed.•  Canterbury v. Spence in 1972: is verdict required practitio-ners to disclose the risks that a reasonable patient would want to know.• AMAPositionPaperon“informedconsent”in1981:isdoc-ument established the “best standards” concerning informed consent in medicine.Consentistoreectallapplicablelocal,state,andfederallawsand regulations, as well as internal policies and evolving best prac-tices. e informed consent discussion has several components31:• the nature of the proposed treatment, including necessity,prognosis, time element, and cost;• viablealternativestotheproposedtreatment,includingwhataspecialist might oer or the choice of no treatment; and• whataretheforeseeablerisks,includingthingslikelytooccurand risks of no treatment?When obtaining informed consent, the dental professional should:• Uselanguagethatiseasilyunderstandable.• Provide opportunities for patient questions, such as “Whatmore would you like to know?” or “What are your concerns?”• Assesspatientunderstandingbystating,“IfIhavenotexplainedthe proposed dentistry clearly or if you have diculty under-standing, please tell me so we can discuss anything you do not understand.”When gaining consent for the use of allografts, please use lan-guage that does not degrade the spirit of the gift of donation (i.e., use “deceased donor” instead of “cadaver” and “recovered” instead of “harvested”). Remember that all tissue is recovered “asepti-cally” (not “sterile” and not necessarily in an operating room). For all types of materials (allograft, xenograft, alloplast), proces-sors and manufacturers often provide patient education materials for use by the end-user. ese do not replace the informed con-sent discussion but exist to augment the eort. In addition, most companies that provide materials have a toll-free number or web-site for answering patient inquiries. Patients’ ability to recollect and comprehend treatment information plays a fundamental role in their decision making.32 Although patients in general report that they understand information given to them, they may have limited comprehension. Additional media may improve conven-tional informed consent processes in dentistry in a meaningful way. Proper Handling of Materials. e end-user is responsible for reviewing the “product insert” (also known by other names, e.g., “instructions for use,” “package insert”). is document provides valuable information on storage, indications, contraindications, tracking, among others. Some materials have been treated with chemicals or antibiotics, and this must be listed to avoid allergic reactions in their patients.Materials should be inspected on receipt. Are the materials the ones ordered? Is the packaging intact? Were the materials pro-tected from extremes in temperature during shipment? What is the expiration date? If the material appears to be compromised in any way, or is not what was expected, the distributor should be notied.Materials should be stored as indicated until time of use. It is imperative that certain material not be frozen or kept in areas with extreme heat (both conditions can destroy the char-acteristics of some materials). In addition, materials should be “logged” or tracked internally at time of receipt, when used on a patient, or when discarded or returned. All materials bear a unique identier, and it is the responsibility of the end-user to have a system for identifying when and where each mate-rial is used. In the case of allografts, federal law states that each allograft unit have a distinct identier, and that a system exists to track the graft from processor to consignee (and back). is makes it the responsibility of the end-user to track from his or her oce inventory to the patient, meaning that the unique identier can be associated with the patient’s unique identier (chart number, etc.).33 921CHAPTER 35 Bone Substitutes and MembranesTracking of xenograft and alloplast, although not always required, is a good practice. Take the example of a patient who experiences a localized reaction at a site where augmentation occurred; allograft bone was mixed with alloplast, and the site was covered with a xenograft membrane. Identication numbers for each material would be needed to determine, with the help of the individual processors, the root cause of the localized reaction. In this case, having the information at hand as part of the patient surgical notes would be a great advantage.In the unlikely chance that a recall occurs, it is much easier to determine patients receiving aected units if that information is in a central database or log. erefore in addition to information in the patient’s operative note, it is wise to have a central log where grafts received into inventory are logged (date and time), as well as theirnaldisposition(usedonapatient[andIDofthatpatient],returned to distributor, discarded, etc., noting date and time). Expiration Dates. Materials are not to be used past their expi-ration date. Disposition should be recorded in the oce log, and the material should be discarded in a proper manner. Material Safety Data Sheets. Material Safety Data Sheets (MSDS)arenotrequiredforBGSsandmembranes.eOccupa-tional Safety & Health Administration requires chemical manu-facturers and importers to develop an MSDS for each hazardous chemical produced or imported, and these must be provided to a distributor or end-user before or at the time of shipment. “Haz-ardous” chemicals are dened as any chemical that is a physical or health hazard. A “physical hazard” is a chemical where scientic evidence shows that it is combustible, a compressed gas, explosive, ammable, an oxidizer, or unstable (reactive). A “health hazard” is a chemical for which statistical evidence exists that shows acute or chronic health eects in exposed individuals. BGSs and mem-branes do not pose a physical or health hazard.As always, refer to the Instructions for Use document accom-panying any BGS or membrane. is will contain specic infor-mation on indications for use, contraindications, precautions, and preparation instructions. “Single-Patient Use”. All BGSs and membranes, if packaged individually, are designated for single patient use. is means that the material cannot be used for treatment of more than one patient. e AATB demands that “Single-Patient Use” appears on the label of every allograft produced by AATB-accredited tis-sue processors. Processors are working with the AATB to ensure that each graft is used for treatment of only one patient, and that each graft can be tracked (personal communication, Jon Boyd,Director of Certication and Online Learning, AATB,McLean,Va.).Althoughtheterminologymaydier(e.g.,“sin-gle use”), similar intent exists for xenografts and alloplasts (and thisinformationmayappearonapackageinsert).Considerthefollowing:• To reuse asingle-use device or material without consideringthe consequences could expose patients and sta to risks that outweigh the perceived benets.• A device or material designated as “single use” must not bereused. It should be used only on an individual patient during a single procedure and then discarded. It is not intended to be reprocessed and used again, even on the same patient.• ereuseofsingle-usedevicescanaecttheirsafety,perfor-mance, and eectiveness, exposing patients and sta to unnec-essary risk.• ereuseofsingle-usedeviceshaslegalimplications:Anyonewho reprocesses or reuses a device intended by the manufac-turer for use on a single occasion bears full responsibility for its safety and eectiveness. End-User Queries. When in doubt, the end-user should con-tact his or her product distributor for additional information. It is the responsibility of the distributor to nd answers, or to refer the end-user to a subject matter expert, which may include direct discussion with the manufacturer. In most cases the well-trained distributor representative or customer service representative can answer questions related to manufacture, clinical use, and safety. Allograft Source, Processing, and DistributionAllograft tissue banking has a rich history. e rst tissue bank was established by the U.S. Navy in 1949 by Dr. George Hyatt.34 Hyatt wasanorthopedicsurgeonattheNavalMedicalCenterinBethesda,Maryland. e navy program was the rst of its kind in the world and established many of the standards that are followed today (Fig. 35.7). During the 1950s, the identication of appropriate donor criteria for tissue donation, the development of procurement and processing methods, the establishment of a graft registry and documentation, and the clinical evaluation of a variety of tissues were pioneered at thisfacility.Cryopreservation,freeze-drying,irradiationsterilizationof tissue, and immunologic principles of tissue transplantation were developed during the 50 years of research and development by navy scientists.Organpreservation,cadavericbonemarrowrecovery,andimmunosuppressive protocols were also developed at the Navy Tis-sue Bank. e navy was also instrumental in the establishment of the National Marrow Donor Program and the AATB in the United States. Although the Navy Tissue Bank has ceased activity after 50 years of excellence, it should be recognized as the rst standard set-ter for the world community of tissue banks. e rst civilian tissue banks were formed by ex-navy surgeons who trained at Bethesda.At this time, all tissue banking in the United States is depen-dentonorganprocurementorganizations(OPOs).eOPOsarethe mechanism through which families can elect to donate not only their loved one’s lifesaving organs but also eyes, skin, heart valves, veins, arteries, bone, tendon, ligaments, and other “tissue” that can be used to improve health.• Fig. . Photographs from the first tissue bank, U.S. Navy at Bethesda Naval Hospital in Bethesda, Maryland. The picture on the left shows tech-nicians at work processing deceased donor material. Out of respect for the donor, processing was done in silence with technicians communicat-ing nonverbally. The sign on the door translates to “from death, life.” The right photograph shows a technician removing frozen grafts from a −80°C chest freezer. (From Strong DM. The US Navy Tissue Bank: 50 years on the cutting edge. Cell Tissue Bank. 2000;1:9-16.) 922PART VII Soft and Hard Tissue RehabilitationOPOsrepresentauniquecomponentofhealthcare.35 By federal law, they are the only organizations that can perform the lifesaving mission of recovering organs from deceased donorsfortransplantation.WhentheNationalOrganTrans-plant Act was signed into law in 1984, it created the national Organ Procurement and Transplantation Network (OPTN)formatchingdonororganstowaitingrecipients.TheOPTNboth standardized the process through which organs are donated and shared across the country, and created the system of federally designated OPOs throughout the United Statesanditsterritories.TheOPTNincludesallOPOsandtrans-plant centers, and is managed under contract by the United NetworkforOrganSharinglocatedin Richmond,Virginia.Therearecurrently58OPOsintheUnitedStates,andallarenonprofit entities.BecausethefocusoftheOPOisorgandonation,tissuerecov-ery and processing are managed by separate entities (tissue, eye, skin,heart valves)asdesignated by each OPO(through a con-tract).ItistheresponsibilityoftheOPOtomakesureevery family has the opportunity to fulll their loved one’s donation wishes.TissueprocessorsmayuseeithertheOPOorregionalrecov-ery programs for the recovery of tissue. All donor suitability assessment, recovery, transport, storage, processing, postproduc-tion testing, and distribution are the responsibility of the tissue processor.Donor suitability is standardized throughout the United States and consists of three parts: initial medical interview with next of kin, physical assessment by the recovery team, and laboratory test-ing for communicable disease. e medical history and history of present illness are critical, and ndings result in a high number of decisions not to proceed with tissue donation. When a poten-tial donor is approved for recovery, they are assigned a unique identier; this number will be associated with all tissue grafts subsequently produced. Tissue recovery must take place quickly: within 15 hours from time of asystole, or up to 24 hours from asystole if the deceased donor was refrigerated within 12 hours of death. erefore laboratory testing is performed after recovery has occurred. e laboratory assessments not only test for donor suitability (communicable disease and systemic infection), but a representative culture from each piece of tissue is tested for micro-biologic contamination via aerobic and anaerobic means at both room and body temperatures. Any biologic contaminant found is known as bioburden.Recoveryoccurs,whenpossible,inanoperatingroomatthehospital of the deceased donor. Many recovery programs and OPOs have dedicated recovery facilities, and the donor can betransported there for recovery. Some medical examiner oces have dedicated facilities in cooperation with the local recovery program or OPO. Recovery is done under aseptic conditionsjust like any surgical procedure. e donor is prepped, draped, and recovery occurs in a particular sequence via zones. Each zone uses new equipment. Tissue removed is swabbed for culturing, wrapped in special materials (impervious to uids), tagged with the assigned unique identier, and placed on ice for transport to the tissue processor.After arriving at the processor, tissue is placed in quarantine (−80°C)untilallserologicandmicrobiologictestsarecomplete.In addition, some donors have autopsies, the results of which must be obtained before processing. A nal chart review and additional information must be assessed and approved before processing into usable grafts. e tissue bank medical director has full responsibil-ity for release of tissue for processing.Processing occurs in clean rooms or laminar ow hoods under strict aseptic conditions. One set of technicians processes onedonor at a time—no donor pooling or cross-contamination isallowed. Dierent tissues undergo processing in dierent ways. Regardless of the tissue processor, the end product is the same: preparation and preservation of tissue without changing the bio-chemical or biological characteristics.Grafts (now in quarantine) undergo postproduction review and testing (e.g., residual moisture if freeze-dried, residual calcium if demineralized) before nal “in-package” sterilization (see earlier Sterility section). Poststerility review occurs (review of dosimetry and last look at all processing records) before approval by the quality team for release into the “bank” for distribution. Finished grafts are based on surgeon demand. Because the demand in many cases outweighs the supply, every eort is taken to maximize the donor gift. e time from receipt of donor tissue to grafts ready to distribute, at most tissue processors, takes approximately 90 days.Regardingthe“averagedonor”andthetypesofgrafts produced for dental use, my research shows that two tissue banks provide the majority (>50%) of “full-line” allograft oering to the dental implant community. ese two are Community Tissue Services (CTS)basedinDayton,Ohio,andLNHinVirginiaBeach,Virginia.Inaddition, both oer branded and private label allograft options to the two largest dental implant suppliers in the world (Nobel Biocare and Straumann, respectively). Both use similar processing technol-ogy, and both have a long history of cooperation in tissue banking (CTSwasfoundedin1986,andLNHwasfoundedin1982).esetwo nonprot organizations are innovative and active as leaders in AATB. I queried both processors regarding donors and grafts pro-duced for the dental segment in 2017 (personal communication, PaulLehner,DentalProductManager,CTS,andDavidAdamson,GeneralManagerDentalandCraniomaxillofacial,LNH).Regardingdonors,itisinterestingthatifyoulookatalldonorsreceived, the range is 12 to 80 years old (these are “musculoskel-etal” donations that can be turned into grafts for a variety of surgical specialties, including dental). An average age cannot be calculated because the distribution is bimodal, that is, with two dierent “peaks.”Onepeakoccursfromaboutage18to24years,andtheother appears from age 45 to 65 years. About 86.5% of the donors were younger than 70 years, and nearly three of every four donors are male. What is most interesting is that the donor age and gender sta-tistics have not changed since the late 2000s (compared with my data from 2007), except that the number of donors has greatly increased. e increase is due to active donation awareness programs36 and an increase in the number of individuals signing up on the national donorregistry—thatnumberiscurrently130million.37 ere were approximately 30,000 tissue donors in the United States in 2017.Finally, all dental allograft processors in the United States are AATB accredited. Accreditation is a rigorous program that requires adherence to AATB standards, membership in the national organi-zation, and periodic inspections. e AATB restricts distribution of allografts to hospitals, certain healthcare facilities, dentists, and podia-trists. Distribution intermediaries can receive and store allografts for redistribution, but they must follow AATB guidelines, be registered with the FDA, and are subject to state registration(s) and inspection(s). Xenograft Source, Production, and DistributionCliniciansandresearchershaveexperimentedwithwaysto cor-rect skeletal defects in the modern era. Nothing exemplies this 923CHAPTER 35 Bone Substitutes and Membranessearchbetterthantheearlyexperienceswithxenografts.Orell38 in 1937 reported his clinical experiences with the surgical grafting ofospurum,osnovum,andboiledbone.Ospurumwasoxboneprepared by a complicated physicochemical procedure that freed the bone of lipids, connective tissue, and some protein but still left some of the collagen matrix. It was used to ll various skeletal defects, and the author claimed that it was resorbed and replaced by host bone in 2 to 3 years.38 In 1956, Forsberg39 used nely ground, sterile os purum as an implant material for periodontal osseous defects. He reported 11 cases and claimed excellent results in 1, satisfactory results in 7, and poor results in the remaining 3 cases after a postoperative period of up to 12 months.39University studies on laboratory-produced anorganic xenograft bone began in earnest in the 1950s. Anorganic means that the organic portion (∼40% by weight) is totally removed, leaving pure HA. Scopp etal.40,41 in the 1960s reported experimental and clinical work with the rst commercially available xenogeneic implant mate-rial called Boplant (Squibb Pharmaceuticals; this product is no longer available). It was derived from calf bone, and processing consisted of detergent extraction, followed by chloroform and methanol extrac-tion to reduce the lipid content, washing with sterile deionized water, sterilization by immersions in a liquid sterilizing agent, and nally lyophilization and vacuum packaging.40,41 is work was the precur-sor to the modern xenograft. Notably, xenografts can be demineral-ized, freeze-dried, and/or deproteinized, but most distribution is of a calcied matrix form. To date, sources for xenograft material used in dentistry include bovine, porcine, equine, and species of coral. In general, bovine and porcine use cancellous bone. Both materials mimic human bone in density, porosity, and calcium content.42-44e best example of xenograft processing and use can be seenwithanorganicbovinematerialssuchasBio-Oss(GeistlichPharma North America, Inc.).45 e Geistlich Pharma website includes a searchable database for Geistlich Pharma materials and associatedclinicalstudies.OfalloftheavailableBGSmaterials,Bio-Osshasthegreatestnumber of publishedstudies. Since itsbeginnings in 1851 the company has dealt with the processing and rening of bone and collagen materials, and up to now they have the most researched biomaterials.Geistlich Pharma manufactures its biomaterials in its own production department at the company’s headquarters in Switzer-land. e entire production process is subject to the strictest safety standards and quality checks: from the selection of the raw mate-rial suppliers to the delivery of the end products. Safety during the manufacture of the products is guaranteed thanks to extensive hygiene measures in a sophisticated zone system with dierent safety levels and permanent controls.Geistlich Bio-Oss is made from the mineral part of bovinebone (and is also known as deproteinized bovine bone material). e strictly controlled manufacturing process ensures high quality and safety standards by:• adenedoriginoftherawmaterial;• arestrictedcountryoforigin,forexample,Australia,whichishistorically and currently free of bovine spongiform encepha-lopathy (a prion disease, or “mad cow”);• usingselectedandcertiedslaughterhouses;• performingofpreandpostmortemhealthinspectionforeachindividual animal;• restricting source to extremity bone (according to WorldHealthOrganizationGuidelinesontissueinfectivityclassiedas tissues with no detected infectivity or infectious prions), as opposed to axial skeleton bones that may be associated with the spinal column;• eectiveinactivationmethodswith15-hourtreatmentathightemperature and cleaning with strong alkaline solutions;• medical-gradesterilizationanddoublesterilepackaging;and• ocialcontrolsbyinternationalauthorities.Many of the controls placed on xenograft production are simi-lar to what is seen with allograft. GMP must be followed and internationalmanufacturersaresubjecttoInternationalOrgani-zation for Standardization rules.46 International materials coming to the United States must obtain FDA clearance, most often via 510(k) approvals. Distribution of most xenograft materials in den-tistry is accomplished through dental supply companies. Alloplast Production and Distributione category of alloplastic implants includes any nonosseous material placed into a bony defect for the purpose of stimulating repair or regeneration. It includes a very wide range of materi-als, both biologically and nonbiologically derived, and is limited only by the imagination of the investigator and the tolerance of living host tissue. Albee47 in 1920 reviewed the literature to date and reported that osmic acid, brin, blood, gelatin with lime salts, zinc chloride, thyroidin, glacial acetic acid, tincture of iodine, adrenaline, extract of hypophysis, copper sulfate, oil of turpentine, ammonia, lactic acid, silver nitrate solution, alcohol, carbolic acid, oak bark extract, vaccines, and sera had been used to stimulate bone growth without any appreciable success.Historically the oldest known alloplast used in medicine is cal-ciumsulfate.Calciumsulfate,alsoknownas“gypsum”or“plasterof paris,” was rst implanted in humans by Dreesman in 1892 as a void ller of tuberculous osteomyelitis.48A great many materials are used today as the basis of alloplasts. ese include (but are not limited to): HA (and its many deriva-tives), tricalcium phosphates, biphasic congurations, calcium sul-fates, bioactive glass (BG), polymer-based materials, and composite materials.49 In recent years more attention has been placed on macroporosity and microporosity (and interconnectivity of pores), interparticle spacing, mechanical qualities, and rate of resorption.Alloplast production follows the guidelines of the pharmaceutical industry. Materials are “manufactured” under GMP, subject to FDA andInternationalOrganizationforStandardizationregulations,andtheir use is restricted to medical facilities and licensed health profes-sionals. Distribution to dental practitioners in the United States is predominantly through medical and dental supply companies. Graft Descriptions50-60AllograftMineralized Cortical Particulate. Also known as freeze-dried bone allograft (FDBA) (Fig. 35.8), mineralized cortical particulate grafts still contain all of the natural bone components (inorganic and organic including BMPs hidden within the bone matrix). Even though it is called mineralized, there is no mineral added; it is simply “not” demineralized. FDBA is sourced from extrem-ity bone (femur, tibia, bula, humerus, radius, ulna, etc.). It can be processed, ground, and sieved to any desired particle range. A common range is 250 to 1000 μm. Particles smaller than 50 μm are quickly removed from the site by macrophages. Some proces-sors oer particles up to 3 mm for lling of larger defects. Because FDBA still contains its calcied portion, it has mechanical strength. For that reason it is a popular graft material in implant dentistry. FDBA is osteoconductive.  924PART VII Soft and Hard Tissue RehabilitationMineralized Cancellous Particulate. Also known as FDBA, mineralized cancellous particulate is made solely from the cancel-lous portion of bone. Graft source is the metaphyseal region of long bones. It shares many of the characteristics of mineralized cortical particulate but has greater macroporosity because of the marrow space. In addition, the trabecular surface of cancellous is covered with endosteum that, like periosteum, probably plays a role in bone regeneration. Because of the porous nature of cancel-lous bone, new vessel growth (in theory) is more rapid through the graft; therefore regeneration will be quicker. Another benet is that cancellous particles tend to lock together better than equiva-lently sized cortical particles, and this will reduce micromotion in a graft site. Mineralized cancellous grafts are osteoconductive. Mineralized Cortical/Cancellous Mix. e mineralized cortical/cancellous mix is a combination of the two previous graft types. It can be manufactured in two ways: rst is mixing together each component 50:50 (v/v), which requires cortical and cancellous material from a single donor, which requires additional quality-control measures; and the second way is to grind and sieve bone from the metaphysis of a long bone, which is known as a “natural” mix (called corticocancellous) and results in a graft with varying and undetermined proportions of cancellous component. Cortical/cancellous mix is gaining in popularity because these grafts provide good mechanical support and provide for faster incorporation.AccordingtoinformationprovidedbytissueprocessorsCTSandLNH(personal communication, Paul Lehner and DavidAdam-son), in 2017 the previous three graft types (combined) accounted for >80% of the total allograft units distributed. In contrast, in the early 1990s >80% of the grafts distributed were demineralized cortical particulate (unpublished data). is shift is directly related to the increase in graft use to support implant placement. Demineralized Cortical Particulate. Demineralized cortical particulate is also known as DFDBA or demineralized bone matrix (DBM). is historically was the rst graft produced in great numbers by tissue processors for the periodontal market. Demin-eralized grafts have osteoinductive potential because processing removes the mineralized portion of the graft, thus exposing the noncollagenous proteins (e.g., BMPs) associated with the collagen matrix. ese proteins recruit bone-forming cells to the site, thus inducing new bone growth. To be called demineralized, the AATB species that residual calcium in the nal graft cannot exceed 8% by weight. Normal cortical bone, for example, has ∼30% residual calcium. If a graft is over- or under-demineralized, it will not pos-sess its full osteoinductive potential. DFDBA is often just called demineralized. e other term, DBM, is reserved for DFDBA that is put with a carrier or is packaged in a convenience device such as a syringe. When reading market reports, DFDBA is considered nonproprietary, made by many processors; DBM is proprietary and made by few processors, usually under patent protection. Because DFDBA (and DBM) have their mineral component removed, they have little mechanical strength and often will not maintain space. Both forms, if prepared correctly, are compress-ible, have osteoinductive potential, and are osteoconductive. Mineralized Cortical/Demineralized Cortical Combination. Oneofthenewestgrafts,mineralizedcortical/demineralizedcor-tical combination (70:30 v/v) takes advantage of the best char-acteristics of its components. Unlike DFDBA, this version will maintain space and incorporates quickly compared with mineral-ized graft forms. It is both osteoconductive and osteoinductive. Laminar Bone. Laminarboneisagraftformthatwaspopularin the 1990s that has undergone a resurgence. It is made by pre-paring cortical sheets (from the diaphysis portion of long bones) and demineralizing. is exible graft not only induces new bone to grow but acts as its own membrane. Irradiated Cancellous (Vertebral Body). Vertebral bodies pos-sess marrow and cancellous with extremely dense trabeculation (per Wol’s law). Cortical particulate, cancellous particulate,and block grafts can be produced from vertebral materials. ese materials receive a 25 to 38 kGy gamma irradiation. Mineralized Cortical and Cancellous Blocks and Cubes. Blocks and cubes in almost any dimension can be made from solid sec-tions of cortical and cancellous bone. ese would be used in larger cases where missing walls must be replaced, among other cases. ese materials arrive freeze-dried, and clinicians must take care when rehydrating and during xation. In the freeze-dried state, these grafts are brittle. When properly rehydrated, these have the same biomechanics as natural bone. Most often, xation is by a lag screw method. Gel, Pastes, and Putties. Gel, pastes, and putties are all made with DBM as the main component with an inert carrier substance. e rst such graft came out in the early 1990s and used glycerol as a means of dispensing the DBM particles. is was formulated for convenience: the DBM needed no rehydration, there were no loose particles to deal with, and the graft was ready to go o the shelf. Many gels, pastes, and putties currently exist in the market. ere is overlap in the naming of these materials. In general, gel means very thin (low viscosity), with the possibility of being deliv-ered via a syringe or other device. Paste is thicker and may lend itself to delivery in an open-bore device. Putty is moldable and can be hand-delivered to a site. Many describe it like the modeling compoundPlay-Doh(HasbroCorp.,Cincinnati,Ohio)usedbychildren and artists. Putty has additional utility in that it can often be combined with other graft materials (e.g., autograft) or osteo-promotive materials. Gels, pastes, and putties also may come in various formulation. Some have only DBM and a carrier, whereas others have DBM, a mineralized component, and a carrier. ese materials would maintain space much better than a pure demin-eralized material. Rib, Mandible, Bone Pins, and Sheets. A variety of additional skeletalgraftscanbefoundatdierenttissueprocessors.Ribsareprocessed by most tissue banks. Mandibles are restricted to a few banks—these being dicult to recover and process. Bone pins• Fig. . This is a common allograft from the ilium. It is shown as a reminder that there are only two types of bone—cortical and cancellous—from which bone grafts are fashioned. Cortical has little macroporosity, whereas cancellous has much. The cancellous portion is made of inter-connecting pores and is created from trabeculae oriented as per Wolff’s law (form and function). New vessel formation can occur much quicker in cancellous bone. (From Sfasciotti GL, Trapani CT, Powers RM. Mandibular ridge augmentation using a mineralized ilium block: A case letter. J Oral Implantol 42(2):215-219, 2016.) 925CHAPTER 35 Bone Substitutes and Membranes(solid cortical) have become more popular in recent years and are usedmuchliketentingscrews.Corticalsheetsareavailable,butnot by all tissue processors. Most tissue banks, if contacted, will work to assist a clinician in nding who has the graft available. Cell-Based Materials. Osteocel(Nuvasive,Inc.,distributedbyACESurgical,Brockton,Mass.)isanexcellentexampleofacell-based material. From a single donor, DFDBA, mineralized cancel-lous, and cells from the bone marrow are processed and recombined into this specialized graft. e resulting material is osteoinductive, osteoconductive, and osteogenic. Other processors are research-ing similar solutions, and this is a fast-growing area of tissue regeneration. Placental Tissues. e placenta is the source of valuable mem-branes and cells. Human amniotic membrane is the most com-monly used and is derived from the fetal membranes. It consists of the inner amniotic membrane made of single layer of amnion cells xed to collagen-rich mesenchyme. Human amniotic membrane has low immunogenicity, antiinammatory properties, and can be isolated without the sacrice of human embryos. Amniotic mem-brane has various clinical applications in the eld of dermatology, ophthalmology, ENT (ear, nose, and throat) surgery, orthopedics, and dental surgery. Fascia Lata. Fascia lata is the deep fascia of the thigh. It invests the whole of the thigh but varies in thickness in dierent parts (the section used for dentistry is around 1 mm thick). Since the 1920s, fascia lata from deceased donors has been used in reconstructive surgery.In1993,Callan55 described cases where freeze-dried fas-cia lata was used as a membrane. Although still available from many tissue banks for orthopedic applications, it has largely been replaced by acellular dermis in dental applications. Pericardium. Pericardium is the membrane enclosing the heart, consisting of an outer brous layer and an inner double layer of serous membrane. e material resembles fascia lata and can be used in a similar fashion. Pericardium is recovered from only heart valve donors; therefore it is in short supply and not processed by most tissue banks. Xenograft pericardium can be substituted (see later discussion). Acellular Dermis. Acellular dermal matrix has been used as a soft tissue replacement since its introduction in 1994. Its rst dental use was correcting areas with insucient attached gingiva, butin 1999, Crook56 reported using it as a barrier membrane. Even though the material is in high demand, few tissue banks produce acellular dermis because most methods of production are proprietary. Acellular dermal matrices are soft tissue matrix grafts created by a process that results in decellularization but leaves the extracellular matrix intact. It starts with a full-thickness skin graft from a deceased donor. e full-thickness graft is exposed to chemicals that remove the epidermis. A secondary step exposes the remaining dermis to chemicals (detergents and endonucleases) that removed the cells and DNA. is is the “decellularization” step that renders the graft acellular. As a result, an immunologic response in the host is unlikely. Extracellular matrix is preserved, as well as biomechanical properties. ese materials have found great use in burns treatment, plastic and reconstructive surgery, podiatry, orthopedics, and dentistry. XenograftParticulate Form. Processing and production considerations for xenograft is similar to what has been described for allograft. However, cancellous (also known as spongiosa or trabecular bone) seems to be the preferred form. Xenograft is most often deproteinized (removing all immunogenic factors) by a variety of methods. What is left is a calcied matrix resembling natu-ral inorganic component (HA) in all ways. Macroporosity and microporosity are preserved. As discussed previously, both bovine and porcine bone resemble human bone from a biochemical and biomechanical perspective. Xenograft particulate is available in a variety of particle ranges. Because xenograft is pure mineral, it does resorb slowly and is an excellent material for long-term space preservation. Block Form. Solid and porous blocks of xenograft HA can be formed based on desired characteristics. ese will function to preserve space for much longer than with an allograft. Macro-porosity and microporosity can be controlled, as well as surface characteristics. As a pure HA product the grafts tend to be more brittle than natural bone, so care should be taken when modifying shape or using xation screws. Pericardium. As mentioned previously, allograft pericardium is in short supply. As a result, bovine and porcine pericardium substitutes have been developed and introduced into the dental market. Bovine has a greater collagen content than the porcine version. ey generally consist of three layers with collagen and elastic bers in an amorphous matrix. eir surface is porous, which allows for cellular attachment and proliferation, yet has an increased density for soft tissue exclusion. Pericardium mem-branes have shown a prolonged resorption in comparison with collagen membranes. Collagen-Based Products. Resorbable collagen membranes(Fig. 35.9) are manufactured from xenogeneic tendon and skin to manage oral wounds such as extraction sockets, for sinus-lift procedures and repairs, and for periodontal or endodontic surger-ies.eyactasscaoldsforbonedepositionin GBR, promoteplatelet aggregation, stabilize clots, and attract broblasts, facili-tating wound healing. ey are designed to resorb within 2 weeks to 6 months and are biocompatible, easy to manipulate, and only weakly immunogenic. For ease of use, collagen-based products are available in a wide variety of forms such as membranes, plugs, or tape. Extended collagen membranes resorb in 4 to 6 months and are used for larger bony defects that require longer healing peri-ods. ese membranes are modied by increasing the cross-link density. Coralline Grafts. Madrepore (“stone coral”) and millepora (“re coral”) are harvested and treated to become “coral-derived gran-ules”andothertypesofcorallinexenografts.Coral-basedmateri-als are mainly calcium carbonate (and an important proportion of uorides, useful in the context of grafting to promote bone development), whereas natural human bone is made of HA, along with calcium phosphate and carbonate. e coral material is transformed industrially into HA through a hydrothermal pro-cess, yielding a nonresorbable xenograft. If the process is omitted, the coralline material remains in its calcium carbonate state for better resorption of the graft by the natural bone. AlloplastHydroxyapatite. HA is a commonly used calcium phosphate biomaterial for bone regeneration applications due to having a composition and structure similar to natural bone mineral. HA-based grafts form a chemical bond directly to bone once implanted. Synthetic HA is available and used in various forms: (1) porous nonresorbable, (2) solid nonresorbable, and (3) resorbable (nonceramic, porous). HA functions as an osteo-conductive graft material. ese grafts show slow and limited 926PART VII Soft and Hard Tissue Rehabilitationresorptive potential and generally are dependent on passive dissolution in tissue uid and cell-mediated processes such as phagocytosis of particles for resorption. e degradation rate of HA depends on the method of ceramic formation, the cal-cium/phosphate ratio, crystallographic structure, and porosity. e ability of HA to resorb is also heavily dependent on the processing temperature. HA grafts synthesized at high temper-atures are very dense with very limited biodegradability. ese dense grafts are usually used as inert biocompatible llers. At lower temperatures the particulate HA is porous and undergoes slow resorption. Tricalcium Phosphates. Overthelastfewyears,TCPhasbeenusedandextensivelyinvestigatedasabonesubstitute.TCPhastwo crystallographic forms: α-TCPandβ-TCP.β-TCPexhibitsgood biocompatibility and osteoconductivity, and is used com-monly as a partially resorbable ller allowing replacement with newlyformedbone. ResorptionofTCPgrafts is thought tobedependent on dissolution by biological uids and by presence of osteoclast-mediated resorption. In terms of bone regenerative potential, β-TCPgraftshavebeenshowntobesimilartoautog-enous bone, FDBA, DFDBA, and collagen sponge. Biphasic Congurations. Biphasic congurations refer to grafts made from biphasic calcium phosphate, material composed of HA and β-TCP.ecombinationsareinterestinginthattheratioofHA to β-TCPcanbemodiedtoprovidefordesired(slowversusfast) resorption. Also, by modifying the carrier and the charac-teristics of the granules, macroporosity and microporosity can be aected. ese materials have a long history of use in orthopedics. Calcium Sulfate. Calciumsulfatecompoundshaveacompres-sivestrengthgreaterthanthatofcancellousbone.Calciumsul-fate is usually applied as a barrier material to improve the clinical outcomes of periodontal regeneration therapy. When used as a barrier, calcium sulfate materials work as an adjunct with other graft materials. Bioactive Glass. BG is a wide-open and fast-growing eld in tissue engineering. BG has been widely studied since the 1970s. Since 45S5 BGs were discovered by Hench in 1969, they have been used for interface bonding of implant, and tissue repair and regeneration of bone. Glasses are noncrystalline amorphous solids that are commonly composed of silica-based materials with other minor additives. Compared with soda-lime glass (commonlyused, as in windows or bottles), Bioglass 45S5 (trademarked by the University of Florida) contains less silica and higher amounts of calcium and phosphorus. e 45S5 name signies glass with 45 weight%ofSiO2 and a 5:1 molar ratio of calcium to phosphorus. is high ratio of calcium to phosphorus promotes formation of apatitecrystals.LowerCa:Pratiosdonotbondtobone.Bioglass45S5’s specic composition is optimal in biomedical applications because of its similar composition to that of HA, the mineral com-ponent of bone.e necessity of nding a material that forms a living bond with tissues led Hench to develop bioglass repair tissues during the Vietnam War. Bioglass oers advantages such as control of rate of degradation, excellent osteoconductivity, bioactivity, and capacity to deliver cells, but they present limitations in certain mechanical properties, such as low strength, toughness, and reli-ability. It can chemically bond with host tissue by forming a bonelike apatite layer between materials and bone tissue. Ionic dissolution products of BG can promote proliferation and dier-entiation of osteoblasts by activating a series of genes that regu-late cellular behaviors. e rst generation of BG was prepared by the melting-quenching method. Although traditional melt-ing-derived BGs have excellent bioactivity, it was red at a very high temperature (>1300°C), so it had a dense structure andsmall specic surface area, which limits its application. Com-pared with the melting-quenching method, the sol-gel method is a chemistry-based synthesis route, of which a solution con-taining the compositional precursors undergoes polymer-type reactions at room temperature to form a gel. e second genera-tion of sol-gel BGs possesses uniform composition, composed of numerous nanoparticles with microporous and mesoporous structure, and thus it has high specic surface area. ese advan-tages grant sol-gel BGs excellent bioactivity. However, up to now, there is no commercial product made of pure sol-gel BG in clinical application. Synthetic Polymers. Synthetic polymers are discussed later in the Membranes section. Titanium Mesh. Guided bone regenerative membranes can help in treating moderate-to-severe osseous defects, but the inher-ent physical property of the membrane to collapse toward the defect because of the pressure of the overlying soft tissues (thus reducing the space required for regeneration) makes the overall amount of regenerated bone questionable. e use of titanium mesh, which can maintain the space, can be a predictable and reliable treatment modality for regenerating and reconstructing a severely decient alveolar ridge. e main advantages of the tita-nium mesh are that it maintains and preserves the space to be regenerated without collapsing, and it is exible and can be bent. It can be shaped and adapted so it can assist bone regeneration in non-space-maintaining defects. Due to the presence of holes within the mesh (Fig. 35.10), it does not interfere with the blood supply directly from the periosteum to the underlying tissues and bone-grafting material. It is also completely biocompatible to oral tissues. Titanium mesh performs dual duty as a bone replacement and a barrier product. MembranesGTRandGBRmembranescanbefoundineverygraftsourcecat-egory previously listed. Each can be viewed as being either “resorb-able” or “nonresorbable” based on whether the membrane can be left in the surgical site. ere are advantages and disadvantages to each, and it is up to the clinician to understand where each type will have applicability.• Fig. . Collagen Membrane. Collagen materials come in a variety of shapes and vary in their resorption time. (Courtesy Humanus Dental AB, Malmö, Sweden: https://www.humanusdental.com/conform-resorbable-collagen-membrane-1520-mm) 927CHAPTER 35 Bone Substitutes and MembranesResorbable Membranes. ere are three types of biologically resorbable (degradable) membranes: (1) polyglycolide synthetic copolymers, (2) collagen, and (3) calcium sulfate.e most commonly used biodegradable synthetic polymers for three-dimensional scaolds in tissue engineering are saturated poly(α-hydroxy esters), including poly(lactic acid) (PLA) andpoly(glycolic acid) (PGA), as well as poly(lactic-coglycolide) copo-lymers. e chemical properties of these polymers allow hydro-lytic degradation through deesterication. Once degraded, themonomeric components of each polymer are removed by natural pathways. PGA is converted to metabolites or eliminated by other mechanisms, and PLA can be clearedthrough the tricarboxylicacidcycle.Duetotheseproperties,PLAandPGAhavebeenusedin biomedical products and devices, such as degradable sutures, which have been approved by the FDA. eir properties can be highly modied through each product’s nal material design, sur-face topography, and porosity. In addition, dissolution rates can be controlled with resorption occurring in weeks to months.Collagenmembranes,aswellasallresorbablemembranes,donot normally require a second surgery for retrieval. Patients appre-ciate the elimination of a second surgery, in addition to less mor-bidity.Collagenistheprincipalcomponentofconnectivetissueand provides structural support for tissues throughout the body. Collagenisahemostaticagent.Itpossessestheabilitytostimu-late platelet attachment and to enhance brin linkage, which may assist initial clot formation and stabilization, leading to enhanced regeneration. In addition, collagen is chemotactic for broblasts. Collagenmembranesareeasytomanipulateandadaptnicelytothe alveolar topography. Although collagen is a weak immunogen, it is very well tolerated by patients.Calciumsulfatewithitslonguseinmedicineprovidesaninex-pensive solution in a variety of clinical situations. As previously discussed, when used as a resorbable barrier, calcium sulfate mate-rials work as an adjunct with other graft materials. Nonresorbable Membranes. Materials such as cellulose acetate laboratory lters (Millipore; Merck KGaA, Darmstadt, Germany, operating as MilliporeSigma in the United States), silicone sheets, and expanded polytetrauoroethylene (e-PTFE) laboratory lters were the rst nonresorbable biomaterials used for investigating barrier membranes for regenerative therapy. Although these mate-rials demonstrated some therapeutic potential, limitations such as inability to integrate with surrounding tissue, brittleness, and the need to remove them after a certain period were observed. e function of nondegradable (nonresorbable) membranes is tem-porary, as they maintain their structural integrity on placement and are later retrieved via surgery. Although this gives the clinician greater control over the length of time the membrane will remain in place, the retrieval procedure increases the risk for surgical site morbidity and leaves the regenerated tissues susceptible to damage and postsurgery bacterial contamination. However, in situations such as alveolar ridge augmentation before placement of dental implants, it may be desirable for the membrane to retain its func-tional characteristics long enough for adequate healing to occur, and then be removed. Hence in specic situations a nonresorbable membrane provides more predictable performance.e-PTFE was originally developed in 1969, and it became the standard for bone regeneration in the early 1990s. e e-PTFE membrane was sintered (sintering is the process of compacting and forming a solid mass of material by heat or pressure without melting it to the point of liquefaction), and it had pores between 5 and 20 μm in the structure of the material. e most popular commercialtypeofe-PTFEwasGore-Tex(W.L.Gore&Associ-ates, Newark, Del.). e e-PTFE membrane acts as a mechani-cal hindrance. Fibroblasts and other connective tissue cells are prevented from entering the bone defect so that the presumably slower migrating cells with osteogenic potential are allowed to repopulate the defect.In time, clinicians discovered that e-PFTE exposed to the oral cavity resulted in migration of microorganisms through the highly porous membrane. With an average pore size of 5 to 20 μm and the diameter of pathogenic bacteria generally less than 10 μm, migration of microorganisms through the highly porous e-PTFE membrane at exposure was a common complication. A high-density polytetrauoroethylene (d-PTFE) membrane with a nominal pore size of less than 0.3 μm was developed (Cyto-plast; Osteogenics Biomedical, Lubbock, Tex.) to address thisproblem.eincreasedecacyofd-PTFEmembranesinGTRhas been proved with animal and human studies. Even when the membrane is exposed to the oral cavity, bacteria is excluded by the membrane, whereas oxygen diusion and transfusion of small molecules across the membrane is still possible. us the d-PTFE membranes can result in good bone regeneration even after expo-sure. Because the larger pore size of e-PTFE membranes allows tight soft tissue attachment, it usually requires sharp dissection at membraneremoval.Onthecontrary,removalofd-PTFEissim-plied because of lack of tissue ingrowth into the surface structure. In 1995, Bartee58 reported that the use of d-PTFE is particularly useful when primary closure is impossible without tension, such as alveolar ridge preservation, large bone defects, and the placement of implants immediately after extraction. In those cases d-PTFE membranes can be left exposed, and thus preserve soft tissue and the position of the mucogingival junction. Comparing Bone Graft Substitutes and Membrane CharacteristicsWhen comparing bone graft substitutes and membrane character-istics, the choices are many. Fortunately there are suppliers who have developed regenerative portfolios that oer a “full range” of choices, allowing the clinician latitude in providing their patient with the best treatment. Fig. 35.11 illustrates a small sample of available material diversity from one such supplier.• Fig. . Titanium Mesh. This can be used in place of a bone graft substitute when severe defects are encountered. The material acts as its own membrane. (Courtesy Salvin Dental) 928PART VII Soft and Hard Tissue Rehabilitatione clinician is faced with a wide variety of materials in the market. Many appear to be similar, whereas many others appear to be markedly superior or inferior. As pointed out in this chapter, there are but a few product categories. e products in all catego-ries are made under the strictest of regulations. Industry strives to provide safe and eective grafts for all surgical specialties.Information on any graft type is easy to nd thanks to the World Wide Web. e end-user has a responsibility to keep up to dateonmaterialsandtechniques.Companiessupplyingthespe-cialty of implant dentistry have expanded their regenerative port-folios and gained the technical knowledge to support the doctor in making grafting choices. is ensures the best treatment option for every patient.e following tables are provided as a quick summary of materials covered in this chapter. Table 35.1 examines the materials regulated as HCT/Ps(donatedhumantissue).Table 35.2 lists the materials that are on the market via the 510(k) route (xenografts and alloplasts). Looking to the FutureIn the area of hard tissue replacement, signicant advancements are being made related to milled “custom” graft materials. Most of the strides are a result of improvements in scanning, tomography, and manufacturing technologies. Also, advances in cell culture and the ability to create three-dimensional print scaolds from biologic materials provide unlimited opportunities for both hard and soft tissues. Soft tissue augmentation shows promise in several areas, mainly from improved understanding in the area of wound healingandimprovedmanufacturing.Cell-basedgraftswillplayabig part in regeneration.Milling of “custom” blocks is currently available.61 Patient selec-tion is a great part of the success and at the time of this writ-ing, only dental surgeons who have received special training on the technique can use the service. Fabrication requires cone beam computed tomography and a tissue processor with the ability to mill bone using computer-aided design and computer-aided man-ufacturing (Fig. 35.12). e technology originated in Europe, is now available in the United States, and resultant grafts:• aresourcedfromprocessedhumanallograft;• arecomposedofnaturalmineralizedcollagen(normaltrabecu-lar bone);• have65%to80%macroporosity,poresize100to1800μm (mean 600 to 900 μm);• canbeproducedtoamaximumsize:23×13×13mm;• showfastgraftincorporationandcompleteremodelingpoten-tial;• possessnoantigenicity;• resultinnodonorsitemorbidity;• heal/integratein5to6months;• canbestoredatambienttemperatureforlongperiods;and• aresafeandsterile.Cell culture technology is one of the fastest-growing areas of regenerative innovation.62 It is part of BTE, the specic eld of tis-sue engineering that mainly focuses on enhancing bone regenera-tion and repair by creating substitutes to traditional bone-grafting materials. BTE started about three decades ago and has witnessed tremendous growth ever since. Bone serves as a paradigm for gen-eral principles in tissue engineering because of its high regenerative potentialcomparedwithothertissuesinthebody.ClassicBTEparadigm includes the following three key components: biomate-rials to provide a scaold for new tissue growth, cells, and signal-ing molecules. It is quite possible that components can be made from dierent classes of materials (e.g., a xenograft combined with an alloplast), thus taking advantage of the best properties of each.Scaolds can be either acellular or cellular on implantation within this model. In the former, the architecture and geometry promote the recruitment of local stem cell and or/osteoprogenitor cells, which could be possible with attachment motifs and chemi-cal“smart”cuesplacedwithinthescaoldarchitecture.Ontheother hand, the latter strategy involves implantation of a scaold combined with stem cell and or/osteoprogenitor cells, which can be incorporated by two methods: (1) cell seeding into a “prefab-ricated” scaold, a commonly applied tissue engineering strategy; and (2) cell encapsulation during scaold fabrication made of hydrogel polymer matrix, based on the immobilization of cells within a semipermeable membrane. is technique protects cells from the immune system and permits uniform cell distribution within the construct.63• Fig. . Today’s clinician has many products to choose from. (Courtesy Salvin Dental) 929CHAPTER 35 Bone Substitutes and Membranes Allograft-Derived MaterialsHCT/P Products Function Space Maintaining Mode Time to RemodelMineralized cortical BGS particulate Yes Osteoconductive 6 monthsMineralized cancellous BGS particulate Yes Osteoconductive <6 monthsMineralized cortical/cancellous mix BGS particulate Yes Osteoconductive <6 monthsDemineralized cortical BGS particulate No Osteoconductive/osteoinductive 4–5 monthsMineralized cortical/demineralized cortical mixBGS particulate Yes Osteoconductive/osteoinductive 4–5 monthsLaminar bone BGS structural N/A Osteoconductive/osteoinductive 4–5 monthsDBM gel, putty, paste BGS particulate Varies Osteoconductive/osteoinductive VariesMineralized block, cube, rib, man-dible, pinsBGS structural Yes Osteoconductive SlowCell-based material BGS particulate Yes Osteoconductive/osteoinductive/osteogenic? VariesPlacental tissue Membrane N/A Resorbable FastFascia lata Membrane N/A Resorbable 4–6 monthsPericardium Membrane N/A Resorbable 4–6 monthsAcellular dermis Membrane N/A Resorbable 4–6 monthsBGS, Bone graft substitutes; DBM, demineralized bone matrix; N/A, not applicable. TABLE 35.1 Xenograft- and Alloplast-Derived Materials [510(k) Regulated]510(k) Products Function Space Maintaining Mode Time to RemodelXenograft mineralized cancellousBGS particulate Yes Osteoconductive SlowXenograft mineralized cancellous blockBGS structural Yes Osteoconductive SlowXenograft pericardium Membrane N/A Resorbable 4–6 monthsXenograft collagen forms Membrane N/A Resorbable Varies, weeks to monthsCoralline based BGS particulate Yes Osteoconductive Slow to mediumHydroxyapatite BGS particulate and structural Yes Osteoconductive Varies (generally slow)Tricalcium phosphates BGS particulate Yes Osteoconductive Varies (generally fast)Biphasic (hydroxyapatite + tricalcium phosphate)BGS particulate Yes Osteoconductive Varies (can be controlled by ratio of mixCalcium sulfate BGS (additive) and membrane Yes Osteoconductive (resorbable when used as membrane)FastBioactive glass BGS (mainly as particulate) Yes Osteoconductive Varies (generally slow)Synthetic polymers Membrane N/A Resorbable and nonresorbable formsVaries based on compositionTitanium mesh BGS (in severe cases) and membraneMaintains space Nonresorbable Never resorbsBGS, Bone graft substitutes; N/A, not applicable. TABLE 35.2 930PART VII Soft and Hard Tissue RehabilitationRegardingBTEcreatedscaolds,thefollowingcharacteristicsare desired:• hydrophilicity,roughness,andsurfacetopography• porosity,poresize,andinterconnectivity• mechanicalstrength close to nativetissues and a predictabledegradation rate (5 to 6 months is desired for dental use)• biocompatibleandbioactive• abilitytobindandreleasedrugsorchemicalsthatcanaectthehealing microenvironmentMuch work remains in this area, but the technologies show great promise.Soft tissue grafts and membranes will benet from a better understanding of clinical need. Manufacturers now know that collagen-based membranes can be modied by cross-linking to aect the rate of resorption. In addition, collagen-based materials can be preformed in molds to increase utility in cases with extreme anatomic variations. ickness can be modied to produce “dead soft” materials that lack any memory and adhere perfectly to local anatomy. Naturally, research is going on with all material types for membranes that can be left “predictably” exposed to the oral cav-ity in cases where primary closure cannot be achieved. Much work with d-PTFE has already occurred in this area.64Rowe et al.65 in 2016 discussed work with electrospinning. is is a process by which microbers/nanobers can be formed from a viscous polymer solution exposed to an electric eld. Although widely used in tissue engineering applications, biocom-patible PLA and poly(ε-caprolactone) electrospun meshes have displayedpropertiesthatmightenableitsapplicationasaGTR/GBRmembrane.Ininitialtestsitperformedbetterthanacurrentcommercially available product.Controllableosteoinductionmaintainedintheoriginaldefectareaisthe key to precise bone repair. In 2018, Ma etal.66 reported on research involving the development of a dual-sided (“Janus”) membrane that acts as a membrane on one side and is osteoinductive on the other.Allografts (acellular dermis, fascia lata, and pericardium) are constantly being modied. e main focus for the future will be processing innovations resulting in consistency of thickness, ste-rilityoftheproduct(toaSAL10−6—notallproductsareatthisdesired level), and an increase in the length of time materials can be stored at ambient temperature. Acellular dermis will be highly studied because of its unique makeup and utility in many surgi-cal disciplines. A unique reticular dermis is already available that retains architectural elements (open structure), mechanical prop-erties (elasticity, organized collagen, and elastin), and key matrix proteins to support physiologic cellular responses during regen-erative remodeling.67Ofcourseagreatdealofattentionisbeinggiven to placental-based materials (chorion and amnion), with many products being marketed at the time of this writing.Cell-based products for use in dentistry are currently available.68 ese feature (generally) three components: demineralized corti-cal bone, mineralized cancellous bone, and marrow cells; therefore these products mimic the biologic prole of autograft (osteoin-ductive,osteoconductive,andosteogenic).Currentlytheseprod-ucts require special shipping and storage at ultracold temperatures (not usually available in the typical dental oce). Future eorts with these grafts will be to create shipping solutions that can act as short-term storage for those oces without ultralow-tempera-ture freezers or the ability to store at lower temperatures for short periods. In addition, ways to increase the number and viability ofbone-formingcellsarebeingstudied.Cell-basedgraftsarethemost dicult of all BGS materials to produce because each com-ponent must come from a single donor, and processing must occur quickly to protect the viability of the osteogenic component. SummaryBone graft substitutes and membranes make up a signicant portion of the dental implant market. Most patients who need implant therapy will need either an autograft or a substitute. e industries and regulatory bodies responsible for the manufacture of these materials, and instruments and technologies that augment their use, have reached a high level of maturity. Patient safety and graft performance are the focus of industry while always striving for improvement. Patients are active participants in their own treatment, and the most appropriate grafting decisions can be made collaboratively through the informed consent process.e use of autografts, allografts, xenografts, or alloplasts, alone or in combination, should be based on the individual’s systemic healing capacity, the osteogenic potential of the recipient site, time availableforgraftmaturation,andthepatient’sexpectations.Cli-nicians have a responsibility to their patients to understand the manyproductsavailableforuseinGBR.Itmustalsobeunder-stood that no one ideal BGS or membrane exists.Although product comparisons may seem dicult, the pro-cess is made easier by realizing the best patient outcome. Each graft class has characteristics unique to that group, and it is up to the end-user to evaluate each characteristic against patient needs. Also, the end-user has a responsibility to handle and use each material as intended to protect his or her patients and sta.What has been presented in this chapter covers the basics of BGS and membranes. e next 10 to 15 years will see dramatic changes in biomaterials and techniques, and will provide the clini-cians even more options for successful implant treatment.AcknowledgmentseauthorthanksPaulLehner,DavidAdamson,KarenColella,William Simmons, Greg Slayton, and Jonathan Boyd for their technical assistance and their dedication to the dental specialty. HealsoexpresseshisappreciationtotheemployeesofCTS,LNH,Salvin Dental Specialties, and the AATB. ey have been friends and mentors for many years.• Fig. . A custom solution created from cone beam computed tomography and computer-aided design/computer-aided manufacturing milling (botiss.com). 931CHAPTER 35 Bone Substitutes and MembranesReferences 1. LeventhalGS. Titanium, a metal for surgery. 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