This document provides an overview of tissue engineering presented by Dr. Boris Saha. It defines tissue engineering as combining principles of life sciences and engineering to develop materials and methods to repair damaged tissues. The key elements of tissue engineering are discussed as cells, scaffolds, and signaling molecules. Various cell types, scaffold materials, and growth factors used in tissue engineering are described. Techniques for tissue engineering include both in vitro and in vivo approaches. Limitations and future perspectives of tissue engineering are also mentioned.

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2.  By Dr Boris saha
 Supervisor: Prof R.k Tiwari
Co-supervisor : Dr Sajid ali
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Tissue engineeringTISSUE ENGINEERING
Guide : PROF R.K TEWARI
CO SUPERVISOR : Dr SAJID ALI
Presented by DR BORIS SAHA ( JR1 )
Dept of conservative dentistry &
Endodontics

3.  Introduction
 Definition
 Historical background
 Need for Tissue Engineering
 Triad of TE
 Strategies of TE
 Techniques of TE
 Cells
 Scaffold
 Signalling molecules
 PRP
 PRF
 Emdogain
 Limitation of TE therapy
 Application of TE
 Future perspective
 Ethical problems
 Conclusion 3

4. In practice - term is closely associated with applications that
repair or replace portions of or whole tissues i.e.
Bone
cartilage
blood vessels
bladder
Skin
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INTRODUCTION

5. VACANTI & LANGER,:
 “A combination of the principles & methods of
life sciences with that of engineering, to develop materials &
methods to repair damaged or diseased tissues, & to create
entire tissue replacements”
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6.  In 1970 W.T. Green, an orthopedic surgeon conducted 1st research
related to TE.
 In the Mid-1980’s Dr. Vacanti and Dr. Langer devised a method that
would attempt to create scaffoldings for cell delivery instead of
using naturally occurring scaffoldings that could not be replicated.
 By 2005, TERMIS which included both Asian & European
Societies, was created
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HISTORICAL BACKGROUND

7.  Tissue engineering holds promise of producing better organs
for transplant.
 A major goal of tissue engineering is in-vitro construction of
transplantable vital tissue.
 .Correct many incurable genetic defects
 Supply of hard & soft Connective tissue
 Other application
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8. TRIAD OF TISSUE ENGINEERING
 cells
 scaffold
 signaling molecules
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10.  Characterised in 3 major classes
Conductive
Inductive
Cell transplantation approaches
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11. 11

12. 2 methods
In Vitro
In vivo
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13. Construction in laboratory of vital tissue & its subsequent
implantation into host body.
 Advantage is ability to examine tissues as they are
formed, & to perform specific tissue measurements.
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14. 14
 By in-vitro TE of tissues such as bone, need for recruitment
of specific cells to site is negotiated & predictability of
regeneration is enhanced,
overcoming many of limitations with
conventional therapies.
 Disadvantage is absence of a physiologic
environment
 Implanted tissue has to be incorporated with the
surrounding bone.

15.  Indicates obvious advantage of tissue regeneration in-vivo in
which incorporation occurs as tissues are formed.
 This has formed basis for tissue engineering, which now includes
implantation of porous matrices, seeded with appropriate cells &
signalling molecules, to facilitate tissue regeneration in-vivo.
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16.  Disadvantage of in-vivo approach
regenerating tissues may get dislodged or degraded by
mechanical forces acting normally at site, before regenerated
tissue is fully formed & incorporated
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17. CELLS
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18.  A number of criteria must be satisfied in order to
achieve effective, long-lasting repair of damaged
tissues.
 1) An adequate number of cells must be produced to fill
the defect.
 2) Cells must be able to differentiate into desired
phenotypes.
 3) Cells must adopt appropriate three-dimensional
structural support/scaffold and produce ECM.
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19. 4) Produced cells must be structurally and
mechanically compliant with the native cell.
 5) Cells must successfully be able to integrate with
native cells and overcome the risk of immunological
rejection.
6) There should be minimal associated biological risks
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20. There are two modes for supplying exogenous cells into
defect:
Cell seeding
Cell suspension
Cell incorporation into implantable matrices, which ensures
their localization at treatment site - concept being referred to
as cell seeding.
An alternative is to inject a cell suspension into sealed
compartment containing defect.
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21. Autologous cells
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22. Allogeneic cells
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23. Xenogenic cells
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24.  Characteristic Features:
They are capable of dividing &
renewing themselves for long
periods
They are unspecialized
They can give rise to specialized
cell types.
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25.  Stem cells could be:
Adult stem cells
Embryonic stem cells
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26.  Adult stem cells Also known as somatic (from Greek "of the
body") stem cells & germline (giving rise to gametes) stem cells,
they can be found in children, as well as adults.
 Pluripotent adult stem cells are rare & generally small in number
but can be found in a number of tissues including umbilical cord
blood
 Most adult stem cells are lineage-restricted & are generally
referred to by their tissue origin
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27.  Embryonic stem cell lines are cultures of cells derived
from epiblast tissue of inner cell mass of a blastocyst or
earlier morula stage embryos — approximately 4 to 5 days old
in humans & consisting of 50–150 cells. ES cells
are pluripotent & give rise during development to all
derivatives of 3 primary germ layers:
ectoderm,
endoderm &
mesoderm.
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28. Based on potency the cells are divided into:
1. Totipotent cells.
2. Pluripotent cells.
3. Multipotent cells.
4. Oligopotent cells.
5. Unipotent cells.
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29. SCAFFOLDS
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30. Used to
guide
organization,
Growth & differentiation of cells in process of forming functional
tissue
 provide both physical & chemical signals.
Tissues are composed of
cells,
insoluble extracellular matrix (E.C.M.)
soluble molecules that serve as regulators of cell function.
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31. 31
ABSORBABLE NON-ABSORBABLE
SYNTHETIC
POLYMERS
•P.L.A.
•P.G.A
NATURAL
MINERALS
•Anorganic Bone
NATURAL
POLYMERS
•Collagen
•Fibrin
•Chitosan
SYNTHETIC
CEREMICS
•Calcium Phosphate
SYNTHETIC
POLYMERS
•Polytetra
flouroethylene

32. Implemented as matrix materials for facilitating regeneration in-vivo
(Bucholtz et al 1987). 2 most widely used forms are:
 Tricalcium phosphate
 Hydroxyapatite.
1. Tricalcium Phoshphate:
Porous form of calcium phosphate
ß-TCP
Problem -physiochemical dissolution after implantation
2. Synthetic Hydroxyapatite:
development - second form of bioceramic.
Rationale - mineral naturally occurring in bone is
hydroxyapatite.
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NON-ABSORBABLE

33. PTFE – synthetic
fluoropolymer of tetrafluoroethylene that finds
numerous applications. well known brand name of
PTFE is Teflon by DuPont Co.
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NON-ABSORBABLE

34.  degradation by hydrolysis
 Polyglycolic acid - degrades fast
 Polylactic acid (L-lactide) - most stable in-vitro
 Thus, modification of poly (L-lactide) by cross-linking or
addition of D-lactide  more rapid degradation, thus
diminishing poly L-lactide disadvantage of slow degradation.
 polyglactin 910, a co-polymer of glycolide and L-lactide –
90/10 molar ratio
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ABSORBABLE

35.  Collagen - protein with 3 polypeptide chains, known as α-chains,
each containing at least 1 stretch of repeating AA sequence
 Collagen constitutes almost 1/3 of all protein in body, & accounts
for almost 60% of gingival connective tissue & 90% of total protein
in bone.
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ABSORBABLE

36.  Collagen - medical devices,
 derived from animal sources,- bovine skin, tendon,
intestine or sheep intestine.
 Collagen based sutures & hemostatic sponges have also
been used.
 Resorbable collagen barriers have been used clinically for
G.T.R. procedures, although their combination with
biologic modifiers has not been explored.
 Also, absorbable collagen sponge (ACS) has been used
as a carrier for rhBMP-2
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37.  HA skeleton (Bio-Oss®, Osteograf®) - retains
microporous & macroporous structure of cortical &
cancellous bone.
 remaining after chemical or low heat extraction of the
organic component.
 Usually bovine bone mineral is used
 Currently available - deproteinated, which supports
cell-mediated resorption.
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ABSORBABLE

38. 3-dimensional
Cross-linked
Porous
Biodegradable
Proper surface chemistry
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39.  Matching mechanical strength
 Biocompatible
 Promotes natural healing
 Commercial Feasibility
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40. SIGNALLING
MOLECULES
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41. Signalling molecules o biologic modifiers - materials or
proteins & factors that have potential to alter key
cellular events in host tissue, by stimulating or regulating
the wound healing process.
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42. 42
INTRACRINE
(PTHrp)
PARACRINE
(PDGF, TGF-β)
JUXTACRINE
(Stem cell factor)
AUTOCRINE
(BMPs, TGF-α)
SYSTEMIC
(ENDOCRINE)
(PTH,GH,LH)
LOCAL

43.  3 groups
1. Growth & Differentiation Factors
2. Extracellular Matrix Proteins & Attachment Factors
3. Mediators of Bone Metabolism
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44.  Growth factors - play important role in regeneration are:
1) Platelet derived growth factor (P.D.G.F.),
2) Insulin-like growth factor (I.G.F.),
3) Transforming Growth Factor- β (T.G.F.-β),
4) Fibroblast Growth Factor,
5) Bone Morphogenetic Proteins (B.M.P.s).
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45.  CHEMISTRY: 2 disulphide bonded poly-peptide chains that
encoded by 2 different genes-P.D.G.F.- A & P.D.G.F.-B.
 FORMS: exist either as
heterodimer (AB) or
homodimer (AA, AB).
 3 isoforms of PDGF have unique binding properties for PDGF
receptor sub-units, α & β, found on cell membrane.
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46.  PRODUCTION: Several cell types produce PDGF,
including
Degranulating platelets,
Smooth muscle cells,
Fibroblasts,
Endothelial cells,
Macrophages & keratinocytes.
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47.  Peptide growth factors with
biochemical & functional
similarities to insulin.
 Bone cells produce &
respond to IGF’s, and bone
is a storage house for these
factors in their inactive form.
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48.  Multifactorial growth factor, structurally related to B.M.P.s, but
functionally quite different.
 Chemotactic for bone cells, & may increase or decrease their
proliferation depending upon the differentiation state of the cells,
culture conditions and concentration of TGF-β applied.
 In-vivo, produces new cartilage and / or bone, if injected
in proximity to bone; however, it does not induce new
bone formation when implanted
away from a bony site.
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49.  Family of at least 9 related gene products of which 2 major members are
a-FGF or FGF-1 & b-FGF or FGF-2.
 Stimulate endothelial cells & PDL cell migration
& proliferation, as well as stimulation of bone cell
replication.
 b-FGF is more potent than a-FGF & may act via stimulation of other
growth factors like TGF-β.
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50.  Urist in 1965, reported that protein extracts from bone,
implanted into animals at non-bone sites induced formation of
new cartilage & bone tissue.
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51.  Recombinant proteins are produced from one of several cellular
expression systems:
Bacteria,
Insect cells or mammalian cells.
 rh BMP-2 is produced using mammalian cell expression system, which
allows for proficient execution of post-translational modifications that are
present in human BMPs.
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52.  Several agents which affects the growth of bone:
PROSTAGLANDINS:
Result of cyclo-oxygenation of precursors derived
from arachnoid acid. Found - variety of tissues. Effect varies
considerably from stimulating inflammation & bone resorption
to enhance bone formation
GLUCOCORTICOIDS:
Such as dexamethasone have prostaglandins, complex
direct & indirect effects on bone formation. Chronic
glucocorticoids administration results in bone loss, through
depression in osteoblast function
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53. BISPHOSPHONATES:
A class of pharmacuetical agents, which are structurally
similar to pyrophosphates, natural product of human metabolism.
Bisphosphonates binds to HA crystal of bone & prevent their
growth & dissolution
CLASSIFIED AS:
1st Generation : alkyl side chains
Eg: Endronate
2nd Generation : amino terminal grp.
Eg: Alendronate & Pamidronate
3rd Generation : cyclic side chains
Eg: Risedronate
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BISPHOSPHONATES
PYROPHOSPHATES

54.  is a concentrate of platelet-rich plasma protein derived from
whole blood, centrifuged to remove red blood cells. It has a greater
concentration of growth factors than whole blood, and has been used to
encourage a brisk healing response across several specialties, in
particular dentistry
 One of highest concentrations of PDGF & TGF-β in body are found
within α-granules of blood platelets
 Thus, concentrating platelets would result in concentration of growth
factors, enhancing wound healing on application.
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56.  Safe as it is autologous preparation.
 Promotes adhesiveness & tensile strength for clot stabilization.
 Biologically acceptable.
 Contains growth factors (PDGF & TGF-β) released by platelets.
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57.  is a 2nd-generation PRP where autologous platelets and
leucocytes are present in a complex fibrin matrix to accelerate
the healing of soft and hard tissue
 5 ml of whole venous blood two sterile vacutainer tubes
of 6 ml capacity without anticoagulant centrifugal
machine at 3000 revolutions per minute (rpm) for 10 minutes
 middle fraction containing the fibrin clot is then collected
2 mm below lower dividing line, to obtain the PRF.
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59. Enamel matrix derivative containing amelogenin
protein used for periodontal regeneration
It is available in gel form together with propylene glycol
Mechanism = attraction , attachment , proliferation ,
differention , alveolar bone growth
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60. 1) Pre-culture condition of donor cell
2) Survival of transplanted
3) Local immune response to cellular grafting
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61. Human bone marrow stromal cell lose their in vivo
osteogenic ability during in vitro expansion.
Duration of in-vitro pre-culture is critical.
Optimal pre-culture conditions remain unclear.
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62. Fate is uncertain.
Cells die quickly or migrate out of transplanted site.
Cells do not survive more than 3 weeks .
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63.  Animal models are immunocompromised.
 Tissue regeneration is partly controlled by host local microenvironment
 Immune cells and inflammatory cells affect transplanted cells
 Cross talk b/w grafted cells and host local cells play a
key role.
 Local immune response to stem cells must be
evaluated.
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64. In surgery
 Transplantation of failing tissues/organs
 Aiding tissues in the healing process
In the laboratory
 Observing immunological, pathological and healing changes in
human tissue without harming patients
 Drug therapies: efficacy and side effects of drugs
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66. Apexogenesis
Dentin regeneration
Pulp-dentinal complex regeneration using PRP
GTR used in bone defect for furcal repair in posterior
cases
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68. Bone regeneration
Cartilage tissue regeneration
Epithelial tissue regeneration
Gene therapy for repair of salivary gland function
Use of bio reactors
Future studies are now directed to development of a
composite graft (skin, bone, cartilage, and muscle),
with associated nerve and blood vessels, which is
engineered from the host stem cells.
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SFOT (surgically faciliatated orthodontic therapy )

70. Implants
Polymers & composite
Surface coated titanium
Ceramics
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71. Some tissues already in clinical use
Improvements needed to increase availability and safety
For widespread use, reduced cost is essential
Further work should focus on: vascularisation of new tissue;
maintaining nutrient supply to cells in matrix with
increasing size
Achieving full potential of stem cells to differentiate into
desired cell types
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72. FUTURE PERSEPECTIVE
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73.  A problem with current delivery of growth factors to wounds is
extremely short half-lives of these factors. This can be
attributed to:
 Proteolytic breakdown.
 Receptor mediated endocytosis.
 Solubility of delivery vehicle.
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74.  Genes are specific portions of DNA that code for proteins.
Their role in protein synthesis can be illustrated as follows:
 Activation of transcription via cell surface receptors.
 Transcription of DNA code into mRNA.
 Processing of mRNA in preparation for transportation to
cytoplasm.
 Transport of mRNA to cytoplasm.
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75.  RNA translation & peptide synthesis.
 Polypeptide elongation.
 Post-translational modifications.
 Transport to & across cell membrane.
 At each stage of gene expression, there is an opportunity for
control & regulation of protein synthesis.
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76. 2 general ways to transfer genes:
Virus mediated vectors: - ex-vivo approach
- in-vivo approach
Naked DNA using Plasmids.
Transduction (i.e. transfer of genetic fragment) to
appropriate target cells (i.e. osteoblasts) represents
first critical step in gene therapy.
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78. • Direct injection of therapeutic DNA into target cells using a gene
gun
Micro seeding
gene therapy
• Creation of artificial lipid spheres with an aqueous core
• Carries therapeutic DNA, capable of passing DNA through target
cell membrane
Cationic
Liposomes
• Therapeutic DNA gets inside target cells by chemically linked
DNA to molecule that bind to special cell receptor
Macromolecular
Conjugate
• Delivers naked DNA via polymer matrix sponges.
Gene Activated
Matrices
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79.  Most commonly used applications of tissue engineering is in
field of dermatology, where possibility of obtaining a large
amount of dermal-epidermal tissue from a small portion of skin
of same patient in a short period of time, has allowed
treatment of extensive burns.
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80. 
 A team of French scientists tested the gel on mice that had cavities.
After about a month, the cavities had disappeared
 It contains a peptide called MSH, melanocyte-stimulating hormone,
which has been shown to encourage bone regeneration
 Not only did new tooth cells grow, but they were also stronger, the
paper says.
 More work needs to be done to prove the gel technique works in
humans
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81.  Dr Sergio Canavero plans to do the first human head transplant next year ,It follows the
unveiling of virtual reality system that will 'prepare patients for life in a new body'.
 Russian wheelchair user Valery Spiridonov has volunteered to take part in the first
operation, which would see his head 'frozen' to stop brain cells from dying and tubes
connected to support key arteries and veins. The spinal cord would then be cut,
repaired and fused on to a donor body and the skin stitched back together.
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82.  Dr Canavero has described his plans to take advantage of the
'Frankenstein effect' where the muscles of a dead body can be
reanimated using electrical or magnetic stimulation. He says he hopes
this could be used to test the techniques to reconnect the spinal cord
 If successful, the process could still lead to 'unexpected
psychological reactions' from the patient as they get used to
their new life, so a virtual reality world to prepare them for a
different body is being developed.
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83. Tooth Bank is a Dental Stem Cell Storage company.
 There are several reasons; some have a family history or
higher risk that prompts them to consider different options.
Most see the future of stem cell research and don’t want to
miss the opportunity to save their own stem cells.
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84.  Using xenogenic cells. Species boundaries crossed
 Using human embryonic cells. Unethical to use human embryo
and fetus.
 Use of human embryo with large scale cultivation for profit.
 Right of tissue donors:
 profit making
 Information
 Role of cell bank: privacy of donor
 Prolonging of life through TE: Set goal for raising life span
through TE
 Organ trafficking
 Cost of using technology 84

85. Future developments in fields of molecular & cell
biology, developmental biology & tissue engineering,
will have significant impact on managing anatomic
changes due to disease process.
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