反义miR-21通过RECK抑制胶质瘤细胞侵袭性生长的体内外研究

目的 探讨胶质瘤细胞LN229中RECK作为miR-21的调控靶点,在胶质瘤侵袭性生长中的作用.方法 将反义miR-21(AS-miR-21)寡核苷酸转染至人脑胶质瘤细胞LN229中.Real-time PCR检测LN229细胞中miR-21的表达量.荧光素酶实验检测miR-21对RECK的调控关系.Transwell实验评价LN229细胞侵袭能力的变化,应用Western blot检测细胞内MMP2/9和RECK蛋白水平的变化,ELISA实验检测培养基中活性MMP2/9的表达量,动物实验评价体内条件下肿瘤侵袭性的变化.结果 Real-time PCR显示转染组中miR-21的表达量与对照组相比下调60%.荧光素酶实验证明RECK是miR-21的靶点.Transwell实验证实胶质瘤细胞侵袭能力下降,Western blot和ELISA实验证实MMP2/9表达降低,动物实验及免疫荧光反映肿瘤侵袭性生长受抑制.结论 反义miR-21通过上调RECK的表达而抑制恶性胶质瘤细胞的侵袭性生长.

Abstract：

Objective To investigate the regulation of miR - 21 on invasion growth of human glioma cells by RECK.Method The human glioma LN229 cells were transfected with AS - miR - 21 or scrambled sequences by Lipofectamine2000.Real time PCR was conducted to detect the expression of miR-21.Luciferase experiment was performed to detect the relationship between miR-21 and RECK.The expression of RECK was evaluated by Western blot.The invasion ability was evaluated by transwell assay and subcutaneous models.Western Blot, ELISA and immunofluorescence were used to estimate the changes of MMP2/9.Results The expression of miR - 21 in LN229 cells decreased after transfection with AS-miR-21. It was proved that RECK was a direct target of miR -21 by luciferase experiment.Meanwhile, the high expression of RECK protein in AS - miR -21 group conformed its important function in this mechanism.Transwell assay demonstrated decreased invasion capability of LN229 cell lines transfected with AS- miR- 21.Western blot, ELISA, and immunofluorescence demonstrated the levels of MMP2/9 were down -regulated in AS -miR -21 group compared with control and scrambled group.Conclusions AS - miR -21 could depress the invasion of glioma cells owing to up - regulating the level of RECK which could inhibit MMP2/9 activities both in vitro and vivo.     

脑胶质瘤信号转导通路研究进展

脑胶质瘤是临床最为常见的原发性颅内肿瘤,约占颅内肿瘤的46%,占成年人全身肿瘤的2%;恶性胶质瘤的发病率为(5～8)/100万,在1998年     

靶向Ku70重组腺病毒shRNA在C6胶质瘤细胞的表达

目的 研究重组腺病毒Ad—Ku70shRNA对C6胶质瘤细胞系Ku70蛋白表达的影响，探讨放射治疗过程中基因增敏的新途径。方法以穿梭质粒为基础，构建重组腺病毒Ad—Ku70shRNA载体，扩增并鉴定其病毒滴度。然后转染至C6鼠脑胶质瘤细胞中，应用WestemBlot及免疫荧光观察其Ku70蛋白的表达情况，研究重组腺病毒Ad—Ku70shRNA对C6细胞Ku70表达的抑制效果。结果成功构建并扩增Ad—Ku70shRNA重组腺病毒，感染C6胶质瘤细胞后Ku70的表达明显降低。结论Ad—Ku70shRNA重组腺病毒载体可以明显降低C6胶质瘤细胞Ku70的表达。该结果为放射治疗基因增敏研究提供了有利的工具。     

中药对51例先天性小耳畸形手术患者的辅助治疗作用

2004年1月-2008年10月，笔者对先天性小耳畸形51例在手术治疗的基础上辅以中药治疗，取得了满意疗效。现报告如下。     

成人麻疹并发肝功能损害86例临床分析

成人麻疹的中毒症状明显，肝功能损害常见。现就本院收治的成人麻疹并发肝功能损害86例患者做一分析，现报告如下。     

生殖器疱疹患者超氧化物歧化酶活力及自然杀伤细胞表达的检测

目的 研究生殖器疱疹患者氧化应激状态与细胞免疫水平.方法 收集24例生殖器疱疹患者和24例正常对照者的血清,采用透射比浊法测定超氧化物歧化酶的活力,流式细胞仪检测外周血自然杀伤细胞的表达水平.结果 生殖器疱疹患者超氧化物歧化酶活力明显低于正常对照者(t=-4.14,P=0.0001),自然杀伤细胞的表达水平明显低于正常对照组(t=-2.67,P=0.0133),患者超氧化物歧化酶活力与自然杀伤细胞的表达水平无显著相关性(r=-0.02566,P= 0.9170).结论 生殖器疱疹患者存在氧化应激状态失衡,免疫功能低下.     

腹腔镜下输卵管吻合术27例临床分析

自2005年1月至2007年1月我中心对27例因"继发性不孕、输卵管梗阻"的患者行腹腔镜检查术,并在术中实施腹腔镜下输卵管吻合术27例,效果良好,报告如下。1资料与方法1.1一般资料2005年1月至2007年1月我中心收治继发性不孕症、输卵管梗阻患者27例,年龄25～39岁,平均     

三七膏加喜疗妥治疗血液透析穿刺所致血肿的护理观察

血液透析是治疗晚期尿毒症病人切实可行的有效方法,血管通路是尿毒症病人进行血液透析必不可少的前提条件,是维持血液透析病人的"生命线".     

反义miR-30a-5p抑制脑胶质瘤生长的体内实验研究

ObjectiveTo study the inhibitory effect of knocking down miR-30a-5p on the U87 human glioma xenograft growth and its possible mechanism. MethodsNude mice bearing subcutaneous U87 human glioblastoma were established and treated with miR-30a-5p antisense oligonucleotides (AS-miR-30a-5p) subcutaneous injection. Tumor size was measured every other day until the observation period ended. Researchers executed the animals after the treatment, stripped tumor tissues and extracted RNA and protein. Real-time PCR were conducted to detect the expression of miR-30a-5p. The histopathological characteristics and proliferation and apoptosis biological characters (including SEPT7, PCNA, cyclinD1, MMP-2, apoptosis related factor P53, bcl-2 and caspase3)were evaluated by HE and immunohistochemical staining, Western blot analysis respectively, and the cell apoptosis was detected by TUNEL method.ResultsIn AS-miR-30a-5p treated group, the tumor growth was delayed and the final tumor volume was smaller than that in the control and scr-ODN treated group (F=7.167, P＜0.05), and the expression of miR-30a-5p was knocked down. The expression of PCNA、cyclinD1 were significantly down-regulated while P53、SEPT7 and caspase3 up-regulated. Apoptotic index was increased significantly. ConclusionAs-miR-30a-5p suppresses the growth of U87 human gliomas xenografts significantly. Malignant phenotype of tumors are reversed to a considerable degree. Therefore, miR-30a-5p can be a candidate for targeted therapy of human glioma.