This document discusses cartilage damage and potential treatments using tissue engineering. It describes how cartilage has limited ability for self-repair and can be damaged by trauma or arthritis. Two potential treatments are discussed: (1) Isolating and seeding chondrocytes cells on scaffolds to repair cartilage, and (2) Using mesenchymal stem cells (MSCs) that can differentiate into chondrocytes when implanted on scaffolds. The document evaluates studies comparing the effectiveness of these two approaches for cartilage regeneration and repair.

2.  What is cartilage?
 Cartilage function.
 Damaged Cartilage has limited
capacity for self-repair.
 Causes for cartilage damage: Dr Brian A Rothbart, 2010
 Trauma
 Degenerative disease e.g. Arthritis .

3.  Arthritis is a joint inflammation disorder.
 Inflammation happens due to
cartilage degradation.
 No cure
 All treatments are to decrease
symptoms or delay progress.
 Cartilage degradation may happens
due to collagen degradation.
o Collagen puts the ECM
together.

4.  17.1% of the disabilities in USA because of arthritis.
[Healthy people]
 350 million people
worldwide have arthritis.
 37 million people in USA.
wrongdiagnosis.com
 Health care: $81 billion annually in USA. abcnews.go.com

5.  Using chondrocytes cells for cartilage repair.
 Chondrocytes are cells that build and maintain cartilage.
 Isolating and seeding of chondrocytes cells found from porcine
knees of patient.
 The cells are mature differentiated chondrocytes or
osteochondral progenitor cells, such as MSCs. (Huckle, et al. 2003)
Chondrocytes: the cells that build and
maintain cartilage
http://saveyourself.ca/articles/shorts/2009-05-26-
carticel-good-news-bad-news.php

6.  Isolating and seeding of chondrocytes: The cartilage tissue
was harvested from porcine knees.
 Preparation of protein-loaded chitosan microsphere:
Solution mixed with FITC-BSA (fluorescein isothiocyanate
bovine serum albumin) or TGF-β1(transforming growth factor
–beta1).
 Preparation of chitosan scaffold containing MS-TGF
 SEM (scanning electron microscopy) measurement:-
Chondrocytes were growing on the scaffold.
( Sung Eun Kim, et al. 2006)

7.  Chitosan microsphere loaded with proteins, BSA (bovine
serum albumin) and TGF (transforming growth factor)in
the presence of TPP (Thiamine pyrophosphate)
 The size of microspheres was reduced by the protein.
 Diffusion of TPP into microspheres is time dependent and
critical to maintain the shape of microsphere .
 The cross-linking time should be longer than 20 min.
(Sung Eun Kim, et al. 2006)

8. Results cont’d
In vitro protein release from
microspheres:
 The proteins, BSA (bovine serum
albumin) and TGF (transforming
growth factor) were released in
biphasic manner.
 implies that initially the rate of
release is fast and later the release is
slower.
 BSA and TGF released at initial
phase, 48.8% and 14.5% respectively.
 Release rate of TGF-beta 1 from Release profiles of FITC–
microspheres was lower than BSA BSA (0) and TGF-β1 from
over the whole experiment chitosan microspheres .
 BSA (90.1%) released within 5 days.
 Release amount of TGF- beta1 only
44.9% for 7 days. ( Sung Eun Kim, et al.
2006)

9. Results cont’d
Chondrocytes growth on chitosan scaffolds without
microsphere-TGF
 SEM (scanning electron microscopy) used for evaluation cell
morphology and proliferation.
 After 4 days, chondrocytes were present in the superficial area of the
scaffold and maintain a spherical morphology.
 After 7 days, chondrocytes were slightly proliferated without spreading.
( Sung Eun Kim, et al.
2006)
Scanning electron micrographs of chondrocytes grown on chitosan scaffold for (a)
4 days and (b) 7 days after seeding.

10. Results cont’d
After 4 days , chondrocytes cells grow on chitosan scaffold
with MS-TGF, there was a significant increase in the number.
After 7 days , chondrocytes were proliferated and open pores
of scaffold occupied by the numbers of aggregates, composed
of several chondrocytes. ( Sung Eun Kim, et al. 2006)
Scanning electron micrographs of chondrocytes grown on chitosan scaffold, seeded with
chitosan microsphere containing TGF-β1. (a) Four days after seeding; the cells were
merged to form large aggregates adherent to the scaffold. (b) Magnified view of a
representative chondrocyte grown for 4 days. (c) Seven days after seeding

11.  TGF-β1, has promote the protein synthesis and cell
proliferation in articular cartilage. It acts as an
autocrine regulator of chondrocytes.
 It also inhibits the actions of matrix metalloproteinase
that play an important role in the digestion of
the ECM in both normal and degenerative articular
cartilage. ( Sung Eun Kim, et al. 2006).

12.  Using MSCs stem cells for cartilage repair.
 Implant MSCs encapsulated collagen based scaffold.
 Stem cells found in all multi cellular organisms. They
can renew themselves and differentiate to different cell
types.
 Two types of stem cells:
 Embryonic stem cells
 Adult stem cells

13.  Adult stem cells are undifferentiated cells that are
found in bone morrow, blood stream, retina and
cornea of the eyes. (Bigaria, et. al 2006)
http://www.stemcellresearch.org/testimony/prentice.h
tm

14.  MSCs is found in the bone marrow.
 MSCs can differentiate to many
different cell types: (Bobis, et. al 2006)
 Chondrocytes ( Cartilage)
 Osteoblasts (Bone)
 Myocytes (Muscle)
 MSCs don’t trigger the immune
response. (Bonfield, et. al 2010)
 MSCs are immunosuppression. (Patel, et. al 2008)

15.  Implanting MSCs in a rabbit cartilage defect.
 MSCs encapsulated in collagen hydrogel (CH) and collagen
alginate hydrogel (CAH) scaffolds.
 The scaffold encapsulated in five groups of champers:
 CH + TGF-Beta1
 CH - TGF-Beta1
 CAH + TGF-Beta1
 CAH - TGF-Beta1
 MSCs without any scaffolds nor growth factors
 The champers implanted in the rabbits for 8 weeks.
 After 8m weeks, the rabbits sacrificed and champers collected.

16.  MSCs without any scaffolds, no tissue found.
HE staining of sectioned tissue inside diffusion
chamber showed:
(A) CH –TGF-b1 group showed cells of
round shape and high cell density.
(B) CAH –TGF-b1 group showed lower
cell density of round shape.
(C) CH +TGF-b1 group showed cells of
round shape and high cell density.
(D) CAH +TGF-b1 group showed cells of
round shape and high cell density.

17. Immunohistochemical examination of sectioned tissue inside diffusion
chamber showed:
 (A, E) CH –TGF-b1 group showed intense positive staining of type II collagen.
 (B, F): CAH –TGF-b1 group showed less positive staining of type II collagen.
 (C, G) CH +TGF-b1 group showed highly positive staining of type II collagen.
 (D, H) CAH +TGF-b1 group showed highly positive staining of type II collagen.

18.  MSCs can differentiate to chondrocytes without
interact with body fluids.
 A scaffold must be used for chondrogenesis to happen.
 CH support more chondrogenesis than CAH.
 TGF-b1 is important for MSCs differentiation.

19.  Tissue engineered chondrocytes can be used to treat cartilage
damage.
 MSCs can be used to treat cartilage damage:
 Reduce the cost of healthcare for arthritis patients.
 Give arthritis patients definitive treatment.
 Open the way to treat other diseases.
 Limitation:
 TGF-β1 appears to be a powerful molecule to repair
damaged cartilage. But high dose of intra-articular injection may
induce chemotaxis and activation of inflammatory
cells, resulting in fibrosis and osteophyte formation in cartilage
defects.
 Number of surgeries that should be done to cover all the affected
areas.

20.  Which better to use chondrocytes or MSCs?
 Do you think using these methods will be
effective to treat arthritis?

21.  Bagaria, V., et al., Stem cells in orthopedics: current concepts and possible future
applications. Indian J Med Sci, 2006. 60(4): p. 162-9.
 Bobis, S., D. Jarocha, and M. Majka, Mesenchymal stem cells: characteristics and
clinical applications. Folia Histochem Cytobiol, 2006. 44(4): p. 215-30.
 Bonfield, T.L., et al., Defining human mesenchymal stem cell efficacy in vivo. J
Inflamm (Lond), 2010. 7: p. 51.
 Patel, S.A., et al., Immunological properties of mesenchymal stem cells and clinical
implications. Arch Immunol Ther Exp (Warsz), 2008. 56(1): p. 1-8.
 Zheng, L., et al., Chondrogenic differentiation of mesenchymal stem cells induced by
collagen-based hydrogel: an in vivo study. J Biomed Mater Res A, 2010. 93(2): p.
783-92.
 sung eun kim , et al.Porous chitson scaffold containing microspheres loaded with
transforming growth factors-beta1: implications for cartilage tissue engineering:
Biomedical Research Center, 19 May 2003.
 Huckle, et al. Differentiated chondrocytes for cartilage tissue engineering: 2003.