The potential of transcription factor SOX9 gene transfer in osteoarthritic chondrocytes differentiation and articular cartilage formation /
Articular cartilage has little capacity for repair, thus making it an area of focus for tissue engineering and regenerative medicine (TERM) research. The use of Sox9 gene transfer in facilitating chondrocytes differentiation is becoming more common since it plays an important role as transcription f...
Saved in:
| Main Author: | |
|---|---|
| Format: | Thesis |
| Language: | English |
| Published: |
Kuantan, Pahang :
Kulliyyah of Allied Health Sciences, International Islamic University Malaysia,
2016
|
| Subjects: | |
| Online Access: | Click here to view 1st 24 pages of the thesis. Members can view fulltext at the specified PCs in the library. |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| Summary: | Articular cartilage has little capacity for repair, thus making it an area of focus for tissue engineering and regenerative medicine (TERM) research. The use of Sox9 gene transfer in facilitating chondrocytes differentiation is becoming more common since it plays an important role as transcription factor for chondrogenesis. This study aimed to identify the potential of the transiently overexpressed Sox9 gene in chondrocytes differentiation and in vitro and in vivo tissue engineered cartilage (TEC) formation. Articular cartilages were obtained from consented patients underwent joint surgery. Chondrocytes were isolated, cultured and transfected with pcDNA3-Sox9 using lipofection technique. The TEC constructs were formed by the incorporation of the transfected or the non-transfected chondrocytes onto poly (lactic-co-glycolic acid) (PLGA) scaffold with or without fibrin. Accordingly, comparisons between four experimental groups i.e. (1) transfected chondrocytes incorporated on PLGA/fibrin [PFTC], (2) non-transfected chondrocytes on PLGA/fibrin [PFC], (3) transfected chondrocytes on PLGA [PTC] and (4) non-transfected chondrocytes on PLGA [PC] were made. Transfection efficiency was optimized and assessed by the cartilaginous genes (Sox9, collagen type II and aggrecan) expression. All constructs were evaluated at 1, 2 and 3 weeks in vitro and, at 2 and 4 weeks in vivo (post-implantation). The evaluations involved gross morphology, histology, immunohistochemistry, cell proliferation activity, gene expression, sulphated glycosaminoglycan (sGAG) production and biomechanical property. From the results, optimum transfection efficiency was achieved using 2.0 µg and 4.0 µg DNA concentration. After 3 weeks in vitro, PFTC and PFC constructs showed better histoarchitecture and extracellular matrix (ECM) distribution in concert with positive glycosaminoglycan accumulation when compared to PTC and PC. Chondrogenic properties were evidenced by the expression of cartilaginous markers i.e. collagen type II, XI and aggrecan. The 2-week post-implanted constructs indicated better cartilaginous appearance, a more notable proteoglycans and glycosaminoglycan deposition than the 4-week constructs. The expression of collagen type II, XI and aggrecan was upregulated along with positive immunohistochemistry staining against collagen type I and II. The co-expression of collagen type I in the constructs indicated the early cartilage development based on the nature of a newly construct formation in this study. Despite a few satisfactory findings, inconsistencies occurred between the four groups. Some transfected and non-transfected groups showed lack of cartilaginous properties in some aspects of examination. Down-regulation of chondrogenic markers, particularly in PC and PTC constructs at 4-week post-implantation indicated that the constructs were unable to maintain specific cartilaginous phenotype. The sGAG content and compressive strain were comparable among all 4 groups. Taken together, this study indicated that there may be more than transcription factor Sox9 gene transfer needed for cartilage regeneration. There is always room for improvement in research. Hence, it is hoped that this study provides a good ground for future cartilage TERM research and application. |
|---|---|
| Physical Description: | xxi, 177 leaves : ill. ; 30cm. |
| Bibliography: | Includes bibliographical references (leaves 128-150). |