miR-135a靶向调节SP1对人骨肉瘤细胞增殖和凋亡的影响

发布时间：2018-03-01 12:30

  本文关键词： 骨肿瘤 骨肉瘤 miR-a SP JAK/STAT　出处：《临床与实验病理学杂志》2016年10期 　论文类型：期刊论文

【摘要】：目的探讨miR-135a通过靶向调节SP1对人骨肉瘤(osteosarcoma,OS)细胞增殖和凋亡的影响。方法收集人正常成骨组织和OS组织,培养正常成骨细胞(h FOB1.19)和OS细胞(MG-63),Real-time PCR法检测miR-135a和SP1的表达,Western blot法检测SP1表达。转染miR-135a mimics和inhibitor,MTT法和Brd U-ELISA法检测OS细胞增殖变化,Western blot法检测凋亡蛋白Bax、BCL-2和Caspase 3的表达。双荧光素酶报告基因检测miR-135a与SP1的靶定关系。Western blot法检测miR-135a对OS细胞中SP1表达的影响。Western blot法检测miR-135a对OS细胞中JAK2/STAT3表达的影响。结果 OS组织和细胞中miR-135a表达均显著降低,SP1表达均显著升高。转染miR-135a mimics可降低OS细胞活性,减少Brd U阳性标记率。同时,miR-135a mimics增加OS细胞Bax和Caspase 3的表达,减少BCL-2的表达。miR-135a mimics降低荧光素酶报告基因的荧光强度,结合位点突变后荧光素酶活性升高。上调miR-135a显著降低SP1的表达并降低JAK2和STAT3的磷酸化水平。miR-135a inhibitor作用均与mimics相反。结论 miR-135a可通过靶向调节SP1抑制人OS细胞增殖,诱导OS细胞凋亡,并下调JAK2/STAT3活化。
[Abstract]:Objective to investigate the effect of miR-135a on proliferation and apoptosis of human osteosarcoma osteosarcoma cells by targeting SP1. Methods normal human osteoblasts and OS tissues were collected. The expression of miR-135a and SP1 in cultured osteoblasts were detected by Western blot, and the proliferation of OS cells was detected by Brd U-ELISA and mimics transfection. The expression of Bax-BCL-2 and Caspase 3 were detected by Western blot method and the expression of Baxchon BCL-2 and Caspase 3 were detected by Western blot method. The expression of Baxchon BCL-2 and Caspase 3 were detected by Western blot method. The expression of Baxchon BCL-2 and Caspase 3 in cultured osteoblasts were detected by Western blot. Double luciferase reporter gene was used to detect the target relationship between miR-135a and SP1. Western blot method was used to detect the effect of miR-135a on SP1 expression in OS cells. Western blot assay was used to detect the effect of miR-135a on JAK2/STAT3 expression in OS cells. Transfection of miR-135a mimics decreased the activity of OS cells. At the same time, miR-135a mimics increased the expression of Bax and Caspase 3 in OS cells, and decreased the expression of BCL-2. MiR-135a mimics decreased the fluorescence intensity of luciferase reporter gene. After binding site mutation, luciferase activity increased. Upregulation of miR-135a significantly decreased the expression of SP1 and decreased the phosphorylation level of JAK2 and STAT3. MIR-135a inhibitor had the opposite effect on mimics. Conclusion miR-135a can inhibit the proliferation of human OS cells by targeting SP1. The apoptosis of OS cells was induced and the activation of JAK2/STAT3 was down-regulated.
【作者单位】： 河南省中医院骨病一科;商丘医学高等专科学校临床系中医教研室;
【分类号】：R738.1
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本文编号：1551941