Biomaterials ppt

7/21/2019 Biomaterials ppt

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Wharton’s Jelly Stem Cells: A Novel

Cell Source for

Oral Mucosa and Skin EpitheliaRegeneration

-Bhagyesha Patil- Bhagyashree Bachhav

- Harini Krishnan

- Snehal Kolhekar

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• Scientists used the stem cells taken from the umbilical cord.

• These stem cells can be stored in tissue banks, -> used

instantly when injuries are caused.

• The scientists used Wharton jelly mesenschymal stem

cells from the human umbilical cord and combined with abiomaterial made of fibrin - a protein found in the clotting of

blood - and agarose - a polymer usually extracted from

seaweed.

• The combination of the Wharton jelly mesenschymal stemcells and biomaterial led to the growth of artificial skin and

oral mucosa - a mucous membrane lining the inside of the

mouth.

THE ARTICLE

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Anatomy of skin

and oral mucosa

• Both have two thingsin common, epethiliaand the connectivetissue.

• Epithilia consists of

keratinocytes and areavascular.

• The roles of theconnective

tissue are related to thesustenance of the

differentiationstatus of the overlyingepithelium

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Embryonic

Stem Cells

Adult Stem

cells

PerinatalStem Cells

Ethical issues, human

rejection, tertomaformation

Limited proliferation

potential anddifferentiation capability

Obtained from tissue

that is normally

discarded.

low risk to the mother

and the newborn and

with few ethical

concerns.

Human

Wharton’s

jelly stemcells

(HWJSCs) are

considered

immunoprivil

eged

perinatalcells that

have high

differentiatio

n capability

and are

easily

available for

therapeutic

applications.

As mesenchymal

stem cells, HWJSCs

retain the ability

to differentiate tomesodermic

tissues (cartilage,

tendon, bone, and

ligament)

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•

A mixture of human fibrin obtained from frozen humanplasma and 0.1% agarose.

• An average of 250,000 cultured oral mucosa and skinfibroblasts were added to 5 ml of the mixture immediatelybefore inducing the polymerization of the artificial stroma.

• Once the stromas jellified, HWJSCs were seeded on top of theoral mucosa and skin artificial stromas and cultured for 7days. H-hOM (heterotypical human oral mucosa) and H-hS.

• For in vivo evaluation a segment of skin of 2.5 x 2.5 cm wasexcised from the back of the animals. Then the H-hOM and H-hS was engrafted on the surgical wounds.

DEVELOPMENT OF 3D BIOACTIVE SYSTEMS

TO INDUCE EPITHELIAL DIFFERENTIATION OF HWJSCs

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• Protein-based polymers have the advantage of mimicking many features ofextracellular matrix and thus have the potential to direct the migration, growthand organization of cells during tissue regeneration and wound healing and forstabilization of encapsulated and transplanted cells.

• Properties of fibrin:

Fibrin is a protein matrix produced from fibrinogen, which can be autologously

harvested from the patient providing an immunocompatible carrier for deliveryof active biomolecules.

In addition, fibrin naturally contains sites for cell binding, and therefore has beeninvestigated as a substrate for cell adhesion, spreading, migration andproliferation.

Fibrin provides a material that can be rapidly invaded, remodeled and replaced by

cell-associated proteolytic activity.

Moreover, due to its biomimetic and physical properties it is also widely used as a

cell carrier to many cell types, such as keratinocytes, tracheal epithelial cells ,murine embryonic stem cells, mesenchymal progenitor cells, etc.

But, problems of instability and degradation invivo , so combination with agaroseprovides improved properties resembling natural skin

FIBRIN-AGAROSE SCAFFOLD

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• These biomaterials possessed "added

resistance, firmness and elasticity to the

skin" compared to other biomaterials like

collagen, chitosan, polyglycolic acid, etc.

FIBRIN-AGAROSE SCAFFOLD

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ANALYSIS OF THE MESENCHYMAL NATURE OF HWJSCs

Aim: To confirm the differential capability of cells

Method Used: Flow Cytometry

Principle: Flow cytometry is a technology that simultaneously measures and

then analyzes multiple physical characteristics of single particles, usuallycells, as they flow in a fluid stream through a beam of light. The properties

measured include a particle’s relative size, relative granularity or internal

complexity, and relative fluorescence intensity.

Staining: 1. osteogenic – alizarin red S

2. adipogenic – Oil red O

3. chondrogenic – Alcian blue

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IN VIVO EVALUATION OF THE EPITHELIAL

DIFFERENTIATION POTENTIAL

AIM: In vivo evaluation of the Epithelial differentiation potential of HWJSCs

BRIEF PROCEDURE:

-a segment of skin 2.5x2.5cm was excised from the backs of the

immunodeficient athymic mice

- grafting of H-hOM and H-hS using absorbable suture material

- grafted tissue harvested for histological analysis

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HISTOLOGICAL ANALYSIS

AIM: Histological analysis of in vitro and in vivo samples using light

microscopy, SEM and TEM

Histology is the study of the cellular organization of body tissues and organs

OUTLINE:-Staining using hematoxylin and eosin

- for immunofluoresence labelling with epithelial markers eg. CK1, CK4,

CK8, CK13, involucrin, filaggrin, plakoglobin

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• HWJSCs :

1.

• Can be easily harvested at

low cost

2.

• Can be easily expanded and

cryopreserved

3.

• Shorter doubling time, rapid

propagation and expansion

DISCUSSIONS AND CONCLUSIONS

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• In vitro

1.

• Differentiation level of HWJSCs in vitro

is limited

2.

• Lack of well defined strata, rete ridges,

chorial papillae and surface patterns

3.

• Lack of well defined epithelial

markers.


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• In vivo

1.

• Differentiation level of HWJSCs is

efficient with high maturation level

2.

• Well defined basal, spinosum,

granulosm, and corneum cell layers

3.

• Specific differentiation patterns on

cell surface in 20 days


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• In vivo

1.

•Differentiation process requires

longer time in vivo grafting

2.

• Cell – cell contacts in H-hom and

H –hs after 30 days

3.

• Cells tend to join and form tight

epithelial barrier.


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• General

1.

•Heterotypical tissues have lower

differentiation rate in vivo

2.• Typically seen in H-hom

3.

• Potential to differentiate in to

ectodermal layers


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