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  1. Abdullah B, Shibghatullah AH, Hamid SS, Omar NS, Samsuddin AR
    Cell Tissue Bank, 2009 Aug;10(3):205-13.
    PMID: 18975136 DOI: 10.1007/s10561-008-9111-2
    This study was performed to determine the microscopic biological response of human nasal septum chondrocytes and human knee articular chondrocytes placed on a demineralized bovine bone scaffold. Both chondrocytes were cultured and seeded onto the bovine bone scaffold with seeding density of 1 x 105 cells per 100 microl/scaffold and incubated for 1, 2, 5 and 7 days. Proliferation and viability of the cells were measured by mitochondrial dehydrogenase activity (MTT assay), adhesion study was analyzed by scanning electron microscopy and differentiation study was analyzed by immunofluorescence staining and confocal laser scanning electron microscopy. The results showed good proliferation and viability of both chondrocytes on the scaffolds from day 1 to day 7. Both chondrocytes increased in number with time and readily grew on the surface and into the open pores of the scaffold. Immunofluorescence staining demonstrated collagen type II on the scaffolds for both chondrocytes. The results showed good cells proliferation, attachment and maturity of the chondrocytes on the demineralized bovine bone scaffold. The bovine bone being easily resourced, relatively inexpensive and non toxic has good potential for use as a three dimensional construct in cartilage tissue engineering.
    Matched MeSH terms: Collagen Type II/biosynthesis
  2. Munirah S, Samsudin OC, Aminuddin BS, Ruszymah BH
    Tissue Cell, 2010 Oct;42(5):282-92.
    PMID: 20810142 DOI: 10.1016/j.tice.2010.07.002
    Monolayer culture expansion remains as a fundamental step to acquire sufficient number of cells for 3D constructs formation. It has been well-documented that cell expansion is however accompanied by cellular dedifferentiation. In order to promote cell growth and circumvent cellular dedifferentiation, we evaluated the effects of Transforming Growth Factor Beta-2 (TGF-β2), Insulin-like Growth Factor-I (IGF-I) and basic Fibroblast Growth Factor (bFGF) combination on articular chondrocytes culture and 'chondrocytes-fibrin' construct formation. Chondrocytes were serially cultured in: (1) F12:DMEM+10% Foetal Bovine Serum (FBS) with growth factors (FD10GFs), (2) F12:DMEM+2%FBS with the growth factors (FD2GFs) and, (3) F12:DMEM+10%FBS without growth factors (FD) as control. Cultured chondrocytes were evaluated by means of growth kinetics parameters, cell cycle analysis, quantitative phenotypic expression of collagen type II, aggrecan core protein sox-9 and collagen type I and, immunochemistry technique. Harvested chondrocytes were incorporated with plasma-derived fibrin and were polymerized to form the 3D constructs and implanted subcutaneously at the dorsum of athymic nude mice for eight (8) weeks. Resulted constructs were assigned for gross inspections and microscopic evaluation using standard histochemicals staining, immunochemistry technique and, quantitative phenotypic expression of cartilage markers to reassure cartilaginous tissue formation. Growth kinetics performance of chondrocytes cultured in three (3) types of culture media from the most to least was in the following order: FD10GFs>FD2GFs>FD. Following growth kinetics analysis, we decided to use FD10GFs and FD (control) for further evaluation and 'chondrocytes-fibrin' constructs formation. Chondrocytes cultured in FD10GFs preserved the normal diploid state (2c) with no evidence of aneuploidy, haploidy or tetraploidy. Expression of cartilage-specific markers namely collagen type II, aggrecan core protein and sox-9 were significantly higher in FD10GFs when compared to control. After implantation, 'chondrocytes-fibrin' constructs exhibited firm, white, smooth and glistening cartilage-like properties. FD10GFs constructs formed better quality cartilage-like tissue than FD constructs in term of overall cartilaginous tissue formation, cells organization and extracellular matrix distribution in the specimens. Cartilaginous tissue formation was confirmed by the presence of lacunae and cartilage-isolated cells embedded within basophilic ground substance. Presence of proteoglycan was confirmed by positive Safranin O staining. Collagen type II exhibited immunopositivity at the pericellular and inter-territorial matrix area. Chondrogenic properties of the construct were further confirmed by the expression of genes encoding collagen type II, aggrecan core protein and sox9. In conclusion, FD10GFs promotes the proliferation of chondrocytes and formation of good quality 'chondrocytes-fibrin' constructs which may have potential use of matrix-induced cell implantation.
    Matched MeSH terms: Collagen Type II/biosynthesis
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