长链非编码RNA TUSC8调控miR-137对宫颈癌迁移和侵袭的影响

【摘要】 目的 探讨长链非编码RNA TUSC8调控miR 137对宫颈癌迁移和侵袭的影响及其机制。方法 qPCR检测TUSC8和miR 137在宫颈癌组织和正常宫颈组织中的表达情况及差异,分析TUSC8与宫颈癌患者临床病理学参数之间的相关关系,双荧光素酶报告基因检测TUSC8与miR 137之间的相互作用;Transwell侵袭实验检测抑制TUSC8后宫颈癌细胞侵袭行为的变化情况;划痕愈合实验检测抑制TUSC8后宫颈癌细胞迁移行为的变化情况;裸鼠体内成瘤实验抑制TUSC8后宫颈癌细胞的成瘤能力的变化。结果 与正常宫颈组织相比,宫颈癌组织中TUSC8和miR 137的表达水平均相对上调,差异有统计学意义（P＜005）;TUSC8的表达与宫颈癌的病理分期相关以及淋巴结转移情况有关,分期越高,TUSC8在宫颈癌组织中表达越高,淋巴结转移的患者中TUSC8的表达也相对较高;双荧光素酶实验证实TUSC8能与miR 137的3’ UTR特异性结合,可以调控miR 137的表达与活性;抑制TUSC8的表达后可以抑制宫颈癌细胞的迁移和侵袭能力;抑制TUSC8后宫颈癌细胞的成瘤能力受到相应的抑制。结论 TUSC8可以调控miR 137的表达影响宫颈癌细胞的迁移和侵袭行为。     

miR-34a在低氧诱导的大鼠肺动脉平滑肌细胞增殖中的作用

【目的】观察miR-34a在低氧诱导的大鼠肺动脉平滑肌细胞增殖中的作用并探讨其可能的机制。【方法】原代分离和培养大鼠肺动脉平滑肌细胞（PASMC）,并给予3%低氧处理后,用Real-time PCR法检测miR-34a和Notch1mRNA在大鼠PASMC中的表达;低氧条件下用细胞转染法过表达和抑制miR-34a、沉默Notch1基因的表达后用EDU法观察细胞增殖情况,并用Real-time PCR和Western blot法检测细胞核增殖抗原PCNA的表达。【结果】分离和培养大鼠PASMC并给予3%低氧处理后,大鼠PASMC中miR-34a表达明显降低,且低氧48h降低较明显,组间差异有统计学意义（P<0.05）。而Notch1的表达明显增高,且48h增高较明显,组间差异有统计学意义（P<0.05）。此外,过表达miR-34a和沉默Notch1后会显著抑制低氧引起的细胞增殖,抑制miR-34a的表达后则会促进细胞增殖,且组间差异有统计学意义（P<0.05）。【结论】在低氧诱导的PASMC细胞增殖过程中miR-34a参与了PASMC的增殖过程,且可能是通过上调Notch1引起了细胞增殖。     

miR-152-3p靶向IGF1基因对高糖诱导的ARPE-19细胞活性和凋亡的影响

目的：探究microRNA-152-3p(miR-152-3p)靶向胰岛素样生长因子1(IGF1)基因对高糖诱导的视网膜色素上皮ARPE-19细胞活性和凋亡的影响,并探讨其作用机制。

方法：高糖诱导ARPE-19细胞并转染miR-152-3p mimics,噻唑蓝(MTT)法检测细胞增殖活性,流式细胞术检测细胞凋亡情况,荧光定量PCR(RT-PCR)检测细胞中miR-152-3p水平,蛋白印迹(Western blot)法检测细胞中IGF1和VEGF表达水平,双荧光素酶报告基因检测IGF1和miR-152-3p靶向结合关系。

结果：高糖能够降低ARPE-19细胞活性,提高细胞凋亡率,抑制细胞中miR-152-3p的表达,提高IGF1和VEGF的表达; 而过表达miR-152-3p能够回调高糖诱导的细胞活性抑制及凋亡增加,抑制IGF1和VEGF的表达。双荧光素酶报告基因实验验证了IGF1是miR-152-3p的靶基因。

结论：miR-152-3p可通过靶向IGF1基因调节VEGF的表达抑制高糖诱导的ARPE-19细胞活性抑制和凋亡增加。     

Quaking and miR-155 interactions in inflammation and leukemogenesis

Quaking (QKI) is a tumor-suppressor gene encoding a conserved RNA-binding protein, whose expression is downregulated in several solid tumors. Here we report that QKI plays an important role in the immune response and suppression of leukemogenesis. We show that the expression of Qki is reduced in lipopolysaccharide (LPS)-challenged macrophages, suggesting that Qki is a key regulator of LPS signaling pathway. Furthermore, LPS-induced downregulation of Qki expression is miR-155-dependent. Qki overexpression impairs LPS-induced phosphorylation of JNK and particularly p38 MAPKs, in addition to increasing the production of anti-inflammatory cytokine IL-10. In contrast, Qki ablation decreases Fas expression and the rate of Caspase3/7 activity, while increasing the levels of IL-1α, IL-1β and IL-6, and p38 phosphorylation. Similarly, the p38 pathway is also a target of QKI activity in chronic lymphocytic leukemia (CLL)-derived MEC2 cells. Finally, B-CLL patients show lower levels of QKI expression compared with B cells from healthy donor, and Qki is similarily downregulated with the progression of leukemia in Eμ-miR-155 transgenic mice. Altogether, these data implicate QKI in the pathophysiology of inflammation and oncogenesis where miR-155 is involved.     

miR-224和 miR-378e在大肠癌组织中的表达及其临床意义

【摘要】 目的 探讨miR-224 和 miR-378e 在原位大肠癌癌组织和癌旁正常黏膜组织中的表达差异， 并分析其临床意义。 方法 miRNA 表达谱芯片技术检测大肠癌癌组织和癌旁组织标本中表达差异的miRNA， 运用荧光实时定量聚合酶链反应（real-time PCR）验证其结果，并分析其与临床病理资料之间的关系。 结果 miRNA 表达芯片分析发现， 与正常黏膜组织比较， miR-224在大肠癌组织中表达显著上调， miR-378e在大肠癌组织中表达显著下调， 并被 real-time PCR 所证实。 miR-224 在不同组织学类型癌组织中表达差异有统计学意义， miR-378e 在不同浸润深度癌组织中表达差异有统计学意义， miR-224 表达与组织学类型有关， miR-378e 表达与大肠癌浸润深度有关。 结论miR-224具有潜在的癌基因作用， miR-378e具有潜在的抑癌基因作用， miR-224和 miR-378e可作为     

MiR-124-3p suppresses glioma aggressiveness via targeting of Fra-2

Malignant glioma is the most common and deadly primary brain tumor in adults. However, the mechanisms underlying the malignancy of glioma remain unclear. In the present study, we found that Fos-related antigen-2 (Fra-2) was overexpressed in most glioma cells, and knockdown of Fra-2 prevented cell proliferation, migration, and invasion. Mechanistically, Fra-2 silencing led to a significant reduction in cell-cycle drivers (Cyclin D1 and Cyclin E1), one invasion-associated gene (MMP9), the mesenchymal marker (Vimentin), and induction of the epithelial marker (E-cadherin). Further study confirmed that miR-124-3p decreased the expression of Fra-2 via directly targeting the 3′-UTR, and transfection with miR-124-3p in glioma cells inhibited expression of the above cell-cycle and EMT promoters. Phenotypic experiments also showed that overexpression of Fra-2 weakened the inhibitory effects of miR-124-3p on the proliferation, migration, and invasion of glioma cells. In addition, Fra-2 knockdown impaired the malignant phenotypes enhanced by miR-124-3p inhibition, which suggested a crucial role for the miR-124-3p/Fra-2 pathway in glioma development. Consistently, high expression of Fra-2 was closely associated with low miR-124-3p level and indicated a poor prognosis in patients with glioma. In conclusion, this study indicates the existence of an aberrant miR-124-3p/Fra-2 pathway that results in glioma aggressiveness, which suggests novel therapeutic opportunities for this fatal disease.     

miR-135b reverses chemoresistance of non-small cell lung cancer cells by downregulation of FZD1

BackgroundNon-small cell lung cancer (NSCLC) chemoresistance usually limits the clinical efficacy of chemotherapeutic approaches. However, few reports have revealed the regulation of miR-135b and Frizzled-1 (FZD1) involved in NSCLC chemoresistance.MethodsTo identify the mechanism of miR-135b and FZD1 in NSCLC chemoresistance and to observe their biological functions, we detected the expression levels of miR-135b and FZD1 by conducting quantitative real-time polymerase chain reaction (RT-qPCR) and modified the expressions of miR-135b and FZD1 by transiently transfecting cells with miR-135b mimics or FZD1-siRNA. The 3′-untranslated region (3′-UTR) of FZD1 combined with miR-135b was verified through dual-luciferase reporter assay.ResultsCompared with that in A549 parental cell lines, the miR-135b expression in drug-resistant lung cancer cell lines (A549/DDP) was decreased and their FZD1 expression was increased. The increased miR-135b expression and silenced FZD1 expression enhanced the sensitivity of resistant cells to cisplatin treatment. The high expression of miR-135b in A549/DDP cells remarkably decreased the mRNA levels of FZD1. FZD1 was further identified as the functional downstream target of miR-135b by directly targeting the 3′-UTR of FZD1.ConclusionThe amplification of miR-135b suppressed NSCLC chemoresistance by directly mediating the FZD1 downregulation.     

miR-155反义寡核苷酸转染对人大肠癌Lovo细胞侵袭的影响

目的探讨miR-155对人大肠癌细胞侵袭能力的影响。方法大肠癌Lovo细胞分为3组：脂质体介导反义miR-155（AS-miR-155）组、无义寡脱氧核苷酸（ODN）组和对照组。测定荧光素酶活性验证3组细胞中miR-155的表达,采用Matrigel基质生长试验检测细胞生长情况,以Transwe Ⅱ方法检测细胞的侵袭力,结果与对照组和无义ODN组比较,转染AS-miR-155组Lovo细胞miR-2l表达水平降低;Matrigel基质生长试验显示,转染AS-miR-155组Lovo细胞体外培养克隆平均直径较小,Transwe Ⅱ细胞侵袭试验显示转染AS-miR-155组穿膜细胞数较少。结论 miR-2l高表达可促进Lovo大肠癌细胞侵袭生长,提示miR-155可以作为基因治疗大肠癌的候选靶点。     

mmu-miR-294调控MMP3靶基因对LLC细胞侵袭迁移的影响

目的 预测并验证小鼠mmu-miR-294调控的靶基因,探讨其在肺癌发生发展中的生物学功能.方法 生物信息学预测mmu-miR-294可能调控的靶基因金属蛋白酶(MMP3),双荧光素酶检测验证mmu-miR-294调控MMP3的真实性；脂质体2000介导转染mmu-miR-294模拟物进入Lewis(LLC)细胞株,通过Transwell实验检测细胞侵袭、迁移能力的改变.结果 重组质粒经XbaⅠ单酶切能获得约5000 bp和100 bp的酶切片段,阳性克隆测序,双荧光素酶报告基因检测证明合成寡核苷酸链序列插入正确；脂质体2000介导转染mmu-miR-294模拟物,过表达实验组MMP3蛋白水平较对照组明显降低.转染mmu-miR-294模拟物后LLC细胞的侵袭迁移能力显著降低(P＜0.01).结论 低表达mmu-miR-294有助于维持LLC的侵袭转移特性,增加其表达水平可以有效抑制LLC的侵袭迁移能力.mmu-miR-294可能通过调控其靶基因MMP3表达而发挥功能.