Md Nazir, Noorhidayah and Zulkifly, Ahmad Hafiz and Khalid, Kamarul Ariffin and Zainol, Ismail and Zamli, Zaitunnatakhin and Sha'ban, Munirah (2019) Matrix production in chondrocytes transfected with sex determining region Y-Box 9 and telomerase reverse transcriptase genes: an in vitro evaluation from monolayer culture to three-dimensional culture. Tissue Engineering and Regenerative Medicine, 16 (3). pp. 285-299. ISSN 1738-2696 E-ISSN 2212-5469
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Abstract
Background:: This study aimed to observe the cartilaginous matrix production in SRY (sex determining region Y)-box 9 (SOX9)- and/or telomerase reverse transcriptase (TERT)-transfected chondrocytes from monolayer to three-dimensional (3D) culture. Methods:: The genes were transferred into chondrocytes at passage-1 (P1) via lipofection. The post-transfected chondrocytes (SOX9-, TERT- and SOX9/TERT) were analysed at P1, P2 and P3. The non-transfected group was used as control. The 3D culture was established using the chondrocytes seeded in a disc-shaped PLGA/fibrin and PLGA scaffolds. The resulting 3D “cells-scaffolds” constructs were analysed at week-1, -2 and -3. The histoarchitecture was evaluated using haematoxylin and eosin, alcian blue and safranin o stains. The quantitative sulphated glycosaminoglycan (sGAG) content was measured using biochemical assay. The cartilage-specific markers expression were analysed via real-time polymerase chain reaction. Results:: All monolayer cultured chondrocytes showed flattened, fibroblast-like appearance throughout passages. Proteoglycan and sGAG were not detected at the pericellular matrix region of the chondrocytes. The sGAG content assay indicated the matrix production depletion in the culture. The cartilage-specific markers, COL2A1 and ACAN, were downregulated. However, the dedifferentiation marker, COL1A1 was upregulated. In 3D “cells-scaffolds” constructs, regardless of transfection groups, chondrocytes seeded in PLGA/fibrin showed a more uniform distribution and produced denser matrix than the PLGA group especially at week-3. Both sGAG and proteoglycan were clearly visualised in the constructs, supported by the increment of sGAG content, quantitatively. Both COL2A1 and ACAN were upregulated in SOX9/TERT-PLGA and SOX9/TERT-PLGA/fibrin respectively. While, COL1A1 was downregulated in SOX9/TERT-PLGA. Conclusion:: These findings indicated that the SOX9/TERT-transfected chondrocytes incorporation into 3D scaffolds facilitates the cartilage regeneration which is viable structurally and functionally. © 2019, The Korean Tissue Engineering and Regenerative Medicine Society.
| Item Type: | Article (Journal) |
|---|---|
| Additional Information: | 2397/79713 |
| Uncontrolled Keywords: | Cartilage, Chondrocytes, Glycosaminoglycan, Proteoglycan, Tissue engineering |
| Subjects: | R Medicine > RA Public aspects of medicine |
| Kulliyyahs/Centres/Divisions/Institutes (Can select more than one option. Press CONTROL button): | Kulliyyah of Allied Health Sciences Kulliyyah of Allied Health Sciences > Department of Biomedical Science (Effective:1st July 2011) Kulliyyah of Medicine Kulliyyah of Medicine > Department of Department of Orthopaedics, Traumatology & Rehabilitation |
| Depositing User: | Prof Dr Ahmad Hafiz Zulkifly |
| Date Deposited: | 20 Mar 2020 22:37 |
| Last Modified: | 20 Mar 2020 22:37 |
| URI: | http://irep.iium.edu.my/id/eprint/79713 |
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