Effect of nerve growth factor-beta on proliferation of intraspinal schwannomas

doi:10.3969/j.issn.2095-4344.1170

Abstract

Abstract:

BACKGROUND: Existing evidence has shown that that the effect of NGF/TrkA signaling pathway on proliferation and differentiation of tumor cells is closely related to PI3K/AKT signaling pathway in human benign and malignant tumors. However, there is little information on the NGF/TrkA signaling pathway in pathogenesis of intraspinal schwannomas.
OBJECTIVE: To investigate the effect of nerve growth factor-beta on the proliferation of interspinal schwannoma cells and to explore on the pathogenesis of NGF/TrkA signaling pathway in interspinal schwannoma.
METHODS: Tumor samples were collected and digested to obtain high purity tumor cells as experimental cells. Then the cells were given different concentrations of nerve growth factor-beta (15, 30, 60, 120 and 240 μg/L), K252a (100, 200, 300, 400, 500 and 600 nmol/L), LY294002 (10, 20, 30, 40, 50 and 60 μmol/L), nerve growth factor-beta (120 μg/L) plus K252a (TrkA inhibitor, 400 nmol/L), and nerve growth factor-beta (120 μg/L) plus LY294002 (P13K inhibitor, 50 μmol/L), respectively, for a certain time. The cell proliferation was detected by MTT assay. TrkA, AKT, p-AKT (Ther308), p-GSK-3 beta protein expression was detected by western blot assay. TrkA and AKT mRNA expression was detected by RT-PCR.
RESULTS AND CONCLUSION: (1) Compared with the control group, the absorbance value of cells in the nerve growth factor-beta groups was increased in a concentration-dependent manner (P < 0.05), and increased obviously at the concentration of 120 μg/L (P < 0.001). The absorbance value of cells in the K252a and LY294002 groups was decreased continuously (P < 0.05), and decreased obviously at the concentration of 400 nmol/L and 50 μmol/L, respectively (P< 0.001). (2) The expression levels of TrkA, p-AKT (Ther308), and p-GSK-3 beta protein were upregulated in the nerve growth factor-beta group (P < 0.05), and the expression level of TrkA mRNA was upregulated (P < 0.05). (3) In the nerve growth factor-beta (120 μg/L) plus K252a (400 nmol/L) group, the absorbance value of cells decreased (P < 0.001). The expression levels of TrkA, p-AKT (Ther308), and p-GSK-3 beta protein downregulated (P < 0.05), and the expression level of TrkA mRNA downregulated (P < 0.05). (4) In the nerve growth factor-beta (120 μg/L) plus LY294002 (50 μmol/L) group, the absorbance value of cells decreased (P < 0.01), and the expression levels of p-AKT (Ther308), and p-GSK-3 beta protein downregulated (P < 0.05). (5)There was no significant change in AKT protein and mRNA in each group (P > 0.05). (6) These results suggest that nerve growth factor-beta can promote interspinal schwannoma cell proliferation, which may be related to the expression of TrkA, p-AKT (Ther308) and p-GSK-3 beta protein in NGF/TrkA signaling pathway.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Spinal Canal, Neurilemmoma, Signal Transduction, Tissue Engineering

CLC Number:

R446

Chen Deping, Liu Shengze, Chen Shi, Cong Changchun, Liu Shuyi, Xiao Ying. Effect of nerve growth factor-beta on proliferation of intraspinal schwannomas[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(15): 2373-2379.

Figures/Tables 2

2.1 各组对椎管内神经鞘瘤细胞增殖的影响 ①神经生长因子β组中随其质量浓度的升高，椎管内神经鞘瘤细胞 A值逐渐升高(P < 0.05)，并呈浓度依赖性，在质量浓度为 120 μg/L时升高幅度最大(P < 0.001)，其后随神经生长因子β浓度增加，其作用与质量浓度120 μg/L无明显差异(P > 0.05)；②用最适浓度120 μg/L神经生长因子β行椎管内神经鞘瘤细胞培养，与对照组比较，培养至36 h，A值升高变化幅度最大(P < 0.001)；③K252a组随浓度增加，椎管内神经鞘瘤细胞A值逐渐降低(P < 0.05)，浓度为400 nmol/L时降低幅度最大(P < 0.001)；④LY294002组随其浓度增加，椎管内神经鞘瘤细胞 A值亦逐渐降低(P < 0.05)，浓度为50 μmol/L时下降幅度最大(P < 0.001)，见图3；⑤神经生长因子β(120 μg/L)组细胞A值升高(P < 0.001)，神经生长因子β(120 μg/L)+K252a(400 nmol/L)组(P < 0.001)和神经生长因子β(120 μg/L)+LY294002(50 μmol/L)组(P < 0.01)细胞A值下降，见图4。 2.2 Western blot及RT-PCR ①与对照组比较，神经生长因子β组中TrkA、p-AKT(Ther308)、p-GSK-3β蛋白质的表达上调(P < 0.05)，TrkA蛋白质对应的mRNA的表达亦上调(P < 0.05)，而AKT蛋白质及其对应的mRNA的表达变化不明显(P > 0.05)，见图5；②与对照组比较，神经生长因子β (120 μg/L)组TrkA、p-AKT(Ther308)、p-GSK-3β蛋白质的表达上调(P < 0.05)，TrkA蛋白质对应的mRNA的表达亦上调(P < 0.05)；③神经生长因子β(120 μg/L)+K252a (400 nmol/L)组TrkA、p-AKT(Ther308)、p-GSK-3β蛋白质的表达下调(P < 0.05)，TrkA蛋白质对应的 mRNA的表达亦下调(P < 0.05)；④神经生长因子β(120 μg/L)+LY294002 (50 μmol/L)组p-AKT(Ther308)、p-GSK-3β蛋白质的表达下调(P < 0.05)，而TrkA蛋白质及其mRNA的表达变化不明显(P > 0.05)。上述各组AKT蛋白质及其mRNA均无明显变化(P > 0.05)，见图6。"

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