Lentiviral-mediated multiple gene transfer to chondrocytes promotes chondrocyte differentiation and bone formation in rabbit bone marrow-derived mesenchymal stem cells.
The aim of the present study was to provide a theoretical and experimental foundation on the differentiation of stem cells through the induction of multiple genes. The lentiviral vector carrying TGF-β1 and IL-10 genes was transfected to bone marrow-derived mesenchymal stem cells (BMSCs) which differentiated into chondrogenesis. Healthy New Zealand white rabbits, 2-3 months of age were used in the present study. A 6-8 ml of bone marrow was isolated from the iliac and tibial shaft of each rabbit. The BMSCs suspension was aspired following centrifugation of the bone marrow by percoll separating medium. The BMSCs were primarily cultured and subcultured in vitro, then divided into four groups according to the difference of lentivirus vectors: group A, receiving transforming growth factor β1 (TGF‑β1); group B, receiving TGF-β1 and Interleukin-10 (IL-10); group C, empty vector transfection; and group D, receiving no cell growth factor. Fluorescence expression was detected 12 h after transfecting the lentiviral vector carrying the TGF-β1 and IL-10 gene to BMSCs. The transfection efficiency was approximately 70% with a MOI=100 after 96 h. Expression of SOX-9 aggrecan and Type Ⅱ collagen in groups A-E on day 7 and 14 was detected by RT-PCR and western blot analysis. The expression level of three genes expressed in groups A and C were higher compared to the expression in groups B, D and E. The expression level of the three genes expressed in group B was higher compared to the expression in group D. The expression level of three genes expressed in group A and C showed no statistical difference. Cytokines therefore play an important role in cell proliferation and chondrogenic differentiation. TGF-β1 has a synergistic effect in the differentiation. In addition, IL-10 may have a protective role in the restoration of cartilaginous tissue.
Department of Pharmacy, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
Ping
P
Liu
Department of Urology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.
Liang
L
Sun
Department of Orthopaedics, Chinese People's Liberation Army General Hospital, Beijing 100853, P.R. China.
Hui
H
Chen
Department of Orthopaedics, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250012, P.R. China.
Shui
S
Sun
Department of Orthopaedics, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250012, P.R. China.
Dongsheng
D
Zhou
Department of Neurosurgery, Qilu Hospital, Shandong University, Jinan, Shandong 250012, P.R. China.
Bo
B
Pang
Department of Orthopaedics, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250012, P.R. China.
Jian
J
Wang
D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000494:pharmacology; D016753:Interleukin-10 / Q000235:genetics* / Q000494:pharmacology; D016086:Lentivirus / Q000235:genetics; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D010012:Osteogenesis / Q000187:drug effects; D011817:Rabbits; D055758:SOX9 Transcription Factor / Q000235:genetics / Q000378:metabolism; D053773:Transforming Growth Factor beta1 / Q000235:genetics* / Q000494:pharmacology
D024043:Collagen Type II; D055758:SOX9 Transcription Factor; D053773:Transforming Growth Factor beta1; D016753:Interleukin-10