Noorhidayah Md Nazir1, Ahmad Hafiz Zulkifly2, Kamarul Ariffin Khalid2, Ismail Zainol3, Zaitunnatakhin Zamli1, Munirah Sha'ban4. 1. 1Department of Biomedical Science, Kulliyyah of Allied Health Sciences, International Islamic University Malaysia (IIUM), Jalan Sultan Ahmad Shah, Bandar Indera Mahkota, 25200 Kuantan, Pahang Darul Makmur Malaysia. 2. 2Department of Orthopaedics, Traumatology and Rehabilitation, Kulliyyah of Medicine, International Islamic University Malaysia (IIUM), Jalan Hospital Campus, 25100 Kuantan, Pahang Darul Makmur Malaysia. 3. 3Department of Chemistry, Faculty of Science and Mathematics, Universiti Pendidikan Sultan Idris (UPSI), 35900 Tanjong Malim, Perak Darul Ridzuan Malaysia. 4. 4Department of Physical Rehabilitation Sciences, Kulliyyah of Allied Health Sciences, International Islamic University Malaysia (IIUM), Jalan Sultan Ahmad Shah, Bandar Indera Mahkota, 25200 Kuantan, Pahang Darul Makmur Malaysia.
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.
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.
Authors: J Yang; E Chang; A M Cherry; C D Bangs; Y Oei; A Bodnar; A Bronstein; C P Chiu; G S Herron Journal: J Biol Chem Date: 1999-09-10 Impact factor: 5.157
Authors: Ronjon Paul; Rex C Haydon; Hongwei Cheng; Akira Ishikawa; Nikola Nenadovich; Wei Jiang; Lan Zhou; Benjamin Breyer; Tao Feng; Purnendu Gupta; Tong-Chuan He; Frank M Phillips Journal: Spine (Phila Pa 1976) Date: 2003-04-15 Impact factor: 3.468
Authors: Sandra Camarero-Espinosa; Barbara Rothen-Rutishauser; Christoph Weder; E Johan Foster Journal: Biomaterials Date: 2015-09-28 Impact factor: 12.479
Authors: V. Villar-Suárez; I. Calles-Venal; I. G. Bravo; J. G. Fernández-Alvarez; M. Fernández-Caso; J. M. Villar-Lacilla Journal: J Biomed Biotechnol Date: 2004