tissue engg appli'n
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PRESENTED BYASMITA BENICHATAGEMBT-09039
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Tissue engineering is an interdisciplinary field
that applies the principles of engineering andthe life sciences towards the development of
biological substitutes that restore, maintain,
or improve tissue function.Langer and J. Vacanti Tissue Engineering. Science 1993.
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Engineering at the Genetic and Molecular Level-Towards Genetically Designed Tissues for Regenerative Medicine
- Posttranscriptional Gene Silencing
- Biomolecule Use in Tissue Engineering
Engineering at the Cellular Level- Fetal Tissue Engineering: Regenerative Capacity ofFetal Stem Cells
-Embryonic Stem Cell Use
-The Unrestricted Somatic Stem Cell (USSC) From Cord Blood For Regenerative Medicine
- Mesenchymal Stem Cells: New Insights Into Tissue Engineering and Regenerative Medicine
Engineering at the Tissue Level- Cartilage Engineering
- MuscleTissue Engineering
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Bone Tissue Engineering- Tendon and Ligament Tissue Engineering: RestoringTendon/Ligament and Its Interfaces
- Neural Tissue Engineering and Regenerative Medicine
Engineering at the Organ Level-Breast Tissue Engineering
-Bioartificial Liver
-Pancreas Engineering
-Tissue-Engineered Urinary Bladder-Cell-Based Regenerative Medicine for Heart Disease
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In Neurology In Maxillofacial Surgery In Dental Implantology In General Surgery Regeneration of Renal Tissues In Plastic Surgery Cardiovascular Substitutes
Skin Substitutes In Orthopedic Surgery In Endocrinology In Hematology
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In vitro tissueengineering
In vivo tissue engineering
Ex vivo tissueengineering
Langer and J.Langer and J. VacantiVacanti TissueTissue
Engineering.Engineering. Science,Science, 1993.1993.
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(1) Start building material (e.g., extracellular matrix,biodegradable polymer)
(2) Shape it as needed(3) Seed it with living cells
(4) Bathe it with growth factors(5) Cells multiply & fill up the scaffold & grow intothree-dimensional tissue
(6) Implanted in the body(7) Cells recreate their intended tissue functions(8) Blood vessels attach themselves to the new tissue
(9) The scaffold dissolves(10)The newly grown tissue eventually blends in with itssurroundings
General Steps For Tissue Engineering
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Our skin is a major organ of the body that acts asa barrier to pathogens and trauma Largestlaminar organ.
Skin defects caused by burns, venous anddiabetic ulcers, or acute injury occasionallyinduce life-threatening situations.
Thus, the need for a functional and cost-effectivepermanent skin substitute for burn victims hasalways been garnered.
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Epidermis (5 layers)Keratinocytes provide protective properties.
Melanocytes provide pigmentation.
Langerhans cells help immune system.
Merkel cells provide sensory receptors.
Dermis (2 layers)Collagen, glycoaminoglycans, elastine, ect.
Fibroblasts are principal cellular constituent.
Vascular structures, nerves, skin appendages.
Hypodermis (fatty layer)Adipose tissue plus connective tissue.
Anchors skin to underlying tissues.
Shook absorber and insulator.
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Essential Design Properties
"The initial replacement material should provide immediatephysiologic wound closure and be eliminated once it has providedsufficient information for reconstitution of neodermis
It should protect the wound by providing a barrier to the outside
It should control water evaporation and protein and electrolyte loss
It should limit excessive heat loss
It should decrease pain and allow early mobilization
It should provide an environment for accelerated wound healing
The risk of infection must be taken into account
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Skin substitutes classified as Permanent or temporary Epidermal, dermal or composite Biological or alloplastic (synthetic)
Temporary-material designed to be placed on a fresh wound (partial
thickness) and left until healed
Semi-permanent-material remaining attached to the excised wound andeventually replaced by autogenous skin grafts
Permanent-incorporation of an epidermal analogue, dermal analogue, or
both as a permanent replacement
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Biologicals (Naturally occurring tissues) Cutaneous allografts Cutaneous xenografts Amniotic membranes
Skin substitutes Synthetic bilaminates Collagen-based composites
Collagen-based dermal analogues Deepithelized allografts
Culture-derived tissue substitues Cultured autologous keratinocytes (sheet grafts, cell suspensions) Bilayer human tissue Polyglycolic or acid mesh Fibroblast-seeded dermal analogs Collagen-glycosaminoglycan matrix Epithelial seeded dermal analogs
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Rapid and effective burn wound closure-imp aspect
Early surgical removal of heat-denatured proteinsand devitalized tissue from a wound
Lack of autograft- overcome by use of synthetic
replacements
Initial use of allografts may undergo immunogenic
rejection
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Rheinwald JG, Green H.Serial cultivation of strains of human epidermal keratinocytes
Human diploid epidermis epidermal cells were successfully grown in serial culture. To initiate colony formation, they require the presence of fibroblasts, but proliferation of
fibroblasts must be controlled so that the epidermal cell population is not overgrown.
Both conditions can be achieved by the use of lethally irradiated 3T3 cells at the correctdensity. When trypsinized human skin cells are plated together with the 3T3 cells, the growth of the
human fibroblasts is largely suppressed, but epidermal cells grow from single cells intocolonies.
Each colony consists of keratinocytes ultimately forming a stratified squamous epithelium inwhich the dividing cells are confined to the lowest layer(s).
Hydrocortisone is added to the medium, to make the colony morphology more oderly and
distinctive, and maintains proliferation at a slightly greater rate. Under given culture conditions, it is possible to isolate keratinocyte clones free of viable
fibroblasts. Like human diploid fibroblasts, human diploid keratinocytes appear to have a finite culture
lifetime. The plating efficiency and culture lifetime were lower for keratinocytes of older persons.
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Epidermis Anatomy- Here are four major layers of keratinocytes(the structural cells) in the epidermis.
The stratum basale, has cells that are shaped like columns.In this layer the cells divide and push already formed cells into
higher layers.As cells move into the higher layers, they flatten and eventually
die.
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This technique allows growth of the epidermis from a singlesmall skin biopsy within 34 weeks
2 culture of isolated colonies of epithelial cells
Approaches for skin culture substitutes in vitro
3 Cell culture ofmultilayered epithelial transplants (commonlytermed sheet grafts)
3 Growth and construction ofcompositemultilayered dermal-epidermal analogues
3 Growth and transplantation ofpre-confluent cell grafts (culturedor non-cultured)
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The growth of multilayered cultured epithelialsheets of human autologous keratinocytes-Permanent substitute
Affecting factors High cost Hospital readmissions for reconstruction of
contractures
Wound infection- fragile epithelium and partiallydeveloped dermoepidermal junction
Thus 2 devices necessary to manipulate CEA tohandle fragile cell layers
Less adherence to wound region
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Such keratinocyte-fibrin suspensions(KFSs) canbe made available for grafting within 10 days ofseeding
Thus KFSs preferred over CEA (3-4 weeks) Lack of dermal structures using keratinocyte
alone lead to combination of a preliminary wound bed preparation by
allografts, followed by subsequent KFS transplantation
together with meshed split-thickness allograftskin as an overlay
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Problems with purely epithelial cell grafts have induced thecombination of cultured autologous keratinocytes with variousalloplastic dermal regeneration templates
trypsinized whole-skin-cell suspensions (uncultured keratinocytes
and fibroblasts) centrifuged into a collagen-glycosaminoglycan (C-GAG) matrix and grafted onto guinea pigs were able to facilitatethe regeneration of a healthy epidermis
Compound materials has been propagated for reconstructive andburn surgery that consists of
(1) a well-characterized, so-called dermal portion that consists of
a porous lattice of fibres of a cross-linked bovine collagen andGAG that is supposed to be replaced by new collagensynthesized by fibroblasts that grow in on fullthickness wounds
(2) a so-called epidermal cover of synthetic polysiloxane polymer(silicone)
Collagen integrates into wound bed, silicone layer peeled off
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Pre-confluent human keratinocyte monolayergrafts have also been used in combination withbiological carrier materials
proliferating basal cells which are responsible forthe initial reformation of an epithelium can betransplanted upside down with the carrier on topas a bio-dressing
One of the most common materials has been C-GAG, with or without the cover of a gas-permeable silastic membrane that serves as abarrier to fluid loss
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Many unsolved mysteries yet to solve Research aim is to replace with a truly
functional skin inclusive of all dermalappendages that are of permanent skin-likequality
perhaps the definitive breakthrough will
come through the ability to replace worn out,defective or damaged body parts usingtechnologies that resemble naturalregeneration.
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Fundamentals of Tissue Engineering and
Regenerative Medicine, by U. Meyeret al
Essentials of Stem Cell Biology, by RobertLanza
Rheinwald JG, Green H. Serial cultivation of
strains of human epidermal keratinocytes: the
formation of keratinizing colonies from single
cells. Cell 6 (3): 33143, 1975
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ANY QUERIES???
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