Co-transfection by Nell-1 and Noggin-shRNA promotes osteoblast differentiation of adipose derived mesenchymal stem cells

doi:10.3969/j.issn.2095-4344.2116

Abstract

Abstract:

BACKGROUND: Nell-1 can induce osteogenic differentiation and accelerate bone regeneration. However, little is reported about co-transfection by Nell-1 and Noggin-shRNA.

OBJECTIVE: To observe the effects of simultaneously down-regulating Noggin and up-regulating Nel-like molecule-1(Nell-1) on osteogenic differentiation of adipose derived mesenchymal stem cells (ADSCs) in vitro.

METHODS: ADSCs were isolated and cultured in vitro from adipose tissue of healthy rats, and divided into three groups. ADSCs in control group were transfected with lentiviral (Lv)-enhanced green fluorescent protein. In Nell-1 group, the cells were transfected with Lv-Nell-1. And in co-transfection group, the cells were co-transfected with Lv-Nell-1 and Lv-Noggin shRNA. At 3, 7 and 14 days after transfection, Nell-1 and osteogenesis-related genes (collagen type I, alkaline phosphatase and osteocalcin) were detected by real-time fluorescence quantitative PCR. At 14 days after transfection, ADSCs were detected by alizarin red staining.

RESULTS AND CONCLUSION: At 3 days after transfection, there was no significant difference in the Nell-1 mRNA expression between the control and Nell-1 groups (P > 0.05); and the Nell-1 mRNA expression in the control and Nell-1 groups was significantly lower than that in the co-transfection group (P < 0.05). At 7 and 14 days after transfection, the Nell-1 mRNA expression was still lower in the control and Nell-1 groups than in the co-transfection group, and moreover, the expression level in the Nell-1 group was significantly higher than that in the control group (P < 0.05). At 3, 7, and 14 days after transfection, the levels of collagen type I, alkaline phosphatase and osteocalcin mRNA in the co-transfection group were significantly higher than those in the control and Nell-1 groups, and these expression levels were also significantly higher in the Nell-group than in the control group (P < 0.05). At 14 days after transfection, there were more calcium nodules in the co-transfection group than in the Nell-1 group, while no calcium nodules were observed in the control group. The overall results indicate that Nell-1 can enhance the differentiation of ADSCs into osteoblasts, and interference with Noggin gene can upregulate the expression of Nell-1, thereby triggering a synergistic effect on osteogenic differentiation.

Key words: Nell-1, Noggin, co-transfection, adipose derived mesenchymal stem cells, osteogenic differentiation

CLC Number:

Liu Hao, Liu Jun, Rui Yongjun, Tang Fenglin, Lu Miao, Ding Tao. Co-transfection by Nell-1 and Noggin-shRNA promotes osteoblast differentiation of adipose derived mesenchymal stem cells[J]. Chinese Journal of Tissue Engineering Research, 2020, 24(31): 4966-4970.

Figures/Tables 3

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