Department of Neurosurgery and Brain and Nerve Research Laboratory, The First Affiliated Hospital of Soochow University, 188 Shizi Street, Suzhou, China.
Cell Mol Neurobiol. 2018 Apr;38(3):669-677. doi: 10.1007/s10571-017-0525-x. Epub 2017 Aug 5.
This study was designed to examine differential expression of miR-338-5p in gliomas and the role of miR-338-5p in glioma cell invasion via its potential target gene TSHZ3 encoding Teashirt zinc finger homobox 3, predicted by bioinformatics, and matrix metallopeptidase 2 (MMP2), the key pro-invasive protease overexpressed in gliomas. Quantitative real-time reverse transcription PCR (qRT-PCR) and Spearman correlation analysis were used to determine differential expressions of miR-338-5p and TSHZ3 in astrocytic gliomas of different grades (n = 35) and glioblastoma cell lines (U87 and U251) in comparison to non-neoplastic brain (NNB) tissues (n = 6). Western blotting was used to determine the protein levels of TSHZ3 and MMP2 in glioblastoma cell lines and Matrigel invasion assay to examine the role of miR-338-5p in cell invasiveness. The results showed that the expression of miR-338-5p, normalized to hsnRNA U6, was significantly higher in grade III and IV gliomas and glioblastoma cell lines compared to that in NNB and grade II gliomas, whereas TSHZ3 expression, normalized to GAPDH, was inversely related to miR-338-5p (R = -0.636, P < 0.01). Luciferase assays showed TSHZ3 to be a target gene of miR-338-5p. In both U87 and U251 cells, miR-338-5p mimics increased MMP2 and invasiveness of the cells. Overexpression of ectopic TSHZ3 suppressed the cell invasiveness and attenuated the pro-invasive effect of miR-338-5p mimics. Overall, our results showed that miR-338-5p has a function in promoting glioma cell invasion by targeting TSHZ3 suppression on MMP2. In conclusion, miR-338-5p is a possible potential biomarker for the diagnosis and target for therapy of high-grade glioma.
本研究旨在通过生物信息学预测的潜在靶基因 TSHZ3（编码 Teashirt 锌指同源盒 3）和基质金属蛋白酶 2（MMP2），研究 miR-338-5p 在神经胶质瘤中的差异表达及其对神经胶质瘤细胞侵袭的作用，MMP2 是神经胶质瘤中过度表达的关键促侵袭蛋白酶。采用定量实时逆转录 PCR（qRT-PCR）和 Spearman 相关性分析，比较不同级别星形细胞瘤（n=35）和神经胶质瘤细胞系（U87 和 U251）与非瘤性脑组织（NNB）（n=6）中 miR-338-5p 和 TSHZ3 的差异表达。Western blot 用于检测神经胶质瘤细胞系中 TSHZ3 和 MMP2 的蛋白水平，Matrigel 侵袭实验用于检测 miR-338-5p 在细胞侵袭性中的作用。结果表明，miR-338-5p 相对于 hsnRNA U6 的表达水平在 III 级和 IV 级神经胶质瘤和神经胶质瘤细胞系中明显高于 NNB 和 II 级神经胶质瘤，而 TSHZ3 相对于 GAPDH 的表达水平与 miR-338-5p 呈负相关（R=-0.636，P<0.01）。荧光素酶报告基因实验显示 TSHZ3 是 miR-338-5p 的靶基因。在 U87 和 U251 细胞中，miR-338-5p 模拟物增加了 MMP2 和细胞的侵袭性。外源性 TSHZ3 的过表达抑制了细胞的侵袭性，并减弱了 miR-338-5p 模拟物的促侵袭作用。综上所述，miR-338-5p 通过靶向 TSHZ3 抑制 MMP2 促进神经胶质瘤细胞侵袭，具有促进神经胶质瘤细胞侵袭的功能。总之，miR-338-5p 可能是高级别神经胶质瘤的潜在诊断标志物和治疗靶点。
