非转移性黑色素瘤糖蛋白B在增生性玻璃体视网膜病变及视网膜色素上皮细胞中的作用

发布时间：2019-01-24 16:35

【摘要】：目的研究非转移性黑色素瘤糖蛋白B(glycoprotein non-metastatic melanoma protein b,GPNMB)在增生性玻璃体视网膜病变(proliferative vitreoretinopathy,PVR)及视网膜色素上皮(retinal pigment epithelium,RPE)细胞中的作用。方法首先,采用免疫荧光法对18例PVR患者行玻璃体切除术时切除的视网膜进行定量分析;酶联免疫吸附试验检测PVR患者血清中GPNMB的含量;然后,以人RPE细胞为研究对象,构建GPNMB的siRNA以及过表达载体,转染入细胞后采用MTT法和流式细胞仪观察其对RPE细胞增殖、细胞周期的影响。结果免疫荧光实验表明,随着PVR增生膜病变程度的加重,其GPNMB的荧光强度加强,A、B、C级PVR的增生膜中GPNMB阳性荧光总量分别为5.07、11.21、19.34;酶联免疫吸附试验结果显示,PVR患者血清中GPNMB的含量远高于对照组;细胞增殖实验结果表明,GPNMB过表达可以显著促进RPE细胞的增殖,其OD值是对照组的3.7倍;然后以GPNMB过表达的人RPE细胞为研究对象,结果表明GPNMB-siRNA转染组可以显著抑制RPE细胞的增殖;流式细胞仪检测发现GPNMB对RPE细胞周期有显著影响。结论 GPNMB作为RPE细胞中的一个关键分子,通过细胞周期调控途径影响其增殖,为进一步研究GPNMB在PVR中的作用及机制奠定了基础。
[Abstract]:Objective to investigate the role of non-metastatic melanoma glycoprotein (B (glycoprotein non-metastatic melanoma protein bnmb) in proliferative vitreoretinopathy (proliferative vitreoretinopathy,PVR) and retinal pigment epithelium (retinal pigment epithelium,RPE) cells. Methods first of all, 18 patients with PVR underwent vitrectomy were quantitatively analyzed by immunofluorescence method, and the serum GPNMB content of 18 patients with PVR was detected by enzyme-linked immunosorbent assay (Elisa). Then, the siRNA and overexpression vector of GPNMB were constructed by using human RPE cells as the research object. After transfection into the cells, the effects of MTT and flow cytometry on the proliferation and cell cycle of RPE cells were observed. Results Immunofluorescence assay showed that the fluorescence intensity of GPNMB increased with the severity of PVR proliferative membrane lesion, and the total amount of GPNMB positive fluorescence in the proliferative membrane of PVR was 5.07 ~ 11.21 ~ 19.34, respectively. The results of enzyme-linked immunosorbent assay (Elisa) showed that the content of GPNMB in the serum of PVR patients was much higher than that of the control group, and the results of cell proliferation assay showed that GPNMB overexpression could significantly promote the proliferation of RPE cells, and its OD value was 3.7-fold higher than that of the control group. The results showed that GPNMB-siRNA transfection group could significantly inhibit the proliferation of RPE cells, and flow cytometry showed that GPNMB had a significant effect on the cell cycle of RPE. Conclusion as a key molecule in RPE cells, GPNMB affects its proliferation through cell cycle regulation pathway, which lays a foundation for further study on the role and mechanism of GPNMB in PVR.
【作者单位】：北京市解放军第309医院眼科;
【分类号】：R774.1

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