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目的:研究胞内M-CSF及其受体在肝癌SMMC 7721细胞的表达与性质,探讨胞内M-CSF对SMMC 7721细胞增殖的影响及其机制。方法:以高表达M-CSF的人肝癌细胞系(SMMC 7721细胞)为模型,以免疫组化、流式细胞计数、反义技术与蛋白印迹等方法观测胞内M-CSF对SMMC 7721细胞增殖的影响及其机制。结果:M-CSF 及其受体主要在SMMC 7721细胞的胞质、胞核中表达,胞内的M-CSF 的相对分子量为20 000,M-CSFR的相对分子量为120 000;免疫共沉淀分析证明M-CSF在细胞内与M-CSFR以复合物的形式存在;M-CSF的单克隆抗体及其反义寡聚核苷酸能抑制SMMC 7721细胞的增殖、下调CYCLIND1/E的表达和上调P16的表达,且M-CSF的单克隆抗体及其反义寡聚核苷酸的联合使用能进一步加强对SMMC 7721细胞抑制作用和增加下调CYCLIND1／E和上调P16的表达幅度。结论:SMMC 7721细胞受M-CSF胞外自分泌和胞内自分泌的双重调控。
Objective: To study the expression and characterization of intracellular M-CSF and its receptor in hepatocellular carcinoma SMMC 7721 cells and explore the effect and mechanism of intracellular M-CSF on the proliferation of SMMC 7721 cells. Methods: Human hepatocarcinoma cell line (SMMC 7721) with high expression of M-CSF was used as a model to observe the effect of intracellular M-CSF on the proliferation of SMMC 7721 cells by immunohistochemistry, flow cytometry, antisense technique and Western blotting Influence and Mechanism. Results: M-CSF and its receptor were mainly expressed in the cytoplasm and nucleus of SMMC 7721 cells. The relative molecular weight of intracellular M-CSF was 20,000 and the relative molecular weight of M-CSFR was 120,000. Immunoprecipitation analysis It was demonstrated that M-CSF exists as a complex with M-CSFR in cells. M-CSF monoclonal antibody and its antisense oligonucleotide can inhibit the proliferation of SMMC 7721 cells and down-regulate the expression and up-regulation of CYCLIND1 / E P16 expression, and M-CSF monoclonal antibody and antisense oligonucleotide combination can further enhance the inhibitory effect on SMMC 7721 cells and increase down-regulation of CYCLIND1 / E and up-regulation of P16 expression amplitude. Conclusion: SMMC 7721 cells are regulated by extracellular and extracellular secretion of M-CSF.