人参皂苷Rg1对心肌细胞氧化应激损伤的抑制作用

目的 通过构建H₂O₂诱导的H9c2细胞氧化应激模型,观察人参皂苷Rg₁对氧化应激的抑制作用,并探讨mi R-499c是否参与人参皂苷Rg₁抑制氧化应激的作用机制。方法 采用600μmol/L的H₂O₂诱导H9c2细胞氧化应激模型,给予人参皂苷Rg₁预处理24 h,检测乳酸脱氢酶(lactate dehydrogenase,LDH)、超氧化物歧化酶(superoxide dismutase,SOD)活性及丙二醛(malondialdehyde,MDA)、活性氧(reactive oxygen species,ROS)水平和线粒体膜电位(mitochondrial membrane potential,MMP)变化;采用Western blotting检测凋亡相关蛋白表达。采用Lipofiter 3.0将miR-499c转染至H9c2细胞再制备H₂O₂诱导的氧化应激模型,给予人参皂苷Rg     

miR-21对骨质疏松小鼠骨髓基质细胞增殖的影响

目的探讨miR-21对骨质疏松小鼠骨髓基质细胞(BMSCs)增殖的影响。方法采用双侧卵巢切除法构建骨质疏松小鼠模型(VOX),分离、培养、纯化小鼠BMSCs并采用si PORT Neo FX转染pre-miR-21、pre-miR-negative control(pre-miR-NC)、antmiR-21、ant-miR-negative control(ant-miR-NC)并进行RT-PCR验证,MTT法检测小鼠BMSCs增殖情况、茜素红与碱性磷酸酶染色法检测小鼠BMSCs成骨能力、Western-blotting检测细胞增殖、成骨分化相关蛋白水平。结果骨质疏松症小鼠BMSCs中miR-21相对表达水平低于Ctrl组(P0.05),OVX-pre-miR-21组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均高于OVX-pre-miR-NC组(P0.05),OVX-premiR-NC组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于Ctrl-pre-miR-NC(P0.05); OVX-ant-miR-21组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于OVX-ant-miR-NC组(P0.05),OVX-ant-miR-NC组BMSCs中miR-21相对表达水平、细胞增殖、PCNA水平、Ki-67水平、ALP染色程度、ALP活性、茜素红染色程度、Runx2水平、Osterix水平均显著低于Ctrl-ant-miR-NC(P0.05)。结论提高miR-21表达水平可促进骨质疏松小鼠BMSCs增殖能力与成骨分化能力。     

miR-146a-5p regulates autophagy and NLRP3 inflammasome activation in epithelial barrier damage in the in vitro cell model of ulcerative colitis through the RNF8/Notch1/mTORC1 pathway

Ulcerative colitis (UC) is a chronic inflammatory disease affecting the colon that can be influenced by microRNAs (miRNAs). This study aims to investigate the impact of miR-146a-5p on lipopolysaccharide (LPS)-induced Caco-2/HT-29 cell autophagy and NLRP3 inflammasome activation and the underlying mechanism, with the aim of identifying potential therapeutic targets. We used LPS to establish Caco-2/HT-29 cell models and measured cell viability by CCK-8. The levels of miR-146a-5p, RNF8, markers of NLRP3 inflammasome activation and autophagy, proteins involved in the Notch1/mTORC1 pathway, and inflammatory factors were assessed by RT-qPCR, Western blot, and ELISA. Intestinal epithelial barrier function was evaluated by measuring transepithelial electrical resistance. Autophagic flux was measured using tandem fluorescent-labeled LC3. miR-146a-5p was highly-expressed in LPS-induced Caco-2/HT-29 cells, and autophagy flux was blocked at the autolysosomal stage after LPS induction. Inhibition of miR-146a-5p suppressed NLRP3 inflammasome activation, reduced intestinal epithelial barrier damage, and facilitated autophagy inhibition in LPS-induced Caco-2/HT-29 cells. The autophagy inhibitor NH4Cl partially nullified the inhibitory effects of miR-146a-5p inhibition on NLRP3 inflammation activation. miR-146a-5p targeted RNF8, and silencing RNF8 partly abrogated the action of miR-146a-5p inhibition on promoting autophagy and inhibiting NLRP3 inflammasome activation. miR-146a-5p inhibition suppressed the Notch1/mTORC1 pathway activation by upregulating RNF8. Inhibition of the Notch1/mTORC1 pathway partially nullified the function of silencing RNF8 on inhibiting autophagy and bolstering NLRP3 inflammasome activation. In conclusion, miR-146a-5p inhibition may be a potential therapeutic approach for UC, as it facilitates autophagy of LPS-stimulated Caco-2/HT-29 cells, inhibits NLRP3 inflammasome activation, and reduces intestinal epithelial barrier damage by upregulating RNF8 and suppressing the Notch1/mTORC1 pathway.     

Reserpine as an uncoupler of oxidative phosphorylation and the relevance to its psychoactive properties

Many drugs differing widely in chemical structure uncouple mitochondrial oxidative phosphorylation in vitro. This observation has led to the hypothesis that in vivo uncoupling is the basis of their pharmacological activity. Serpasil, a parenteral preparation of reserpine, recently has been shown to uncouple oxidative phosphorylation in vervet monkey kidney mitochondria. Although the drug exhibits some properties of a "classical" uncoupler, our studies show that it has a dual effect on energy conservation. Reserpine released respiratory control in rat liver mitochondria only when dissolved in organic solvents (as in Serpasil) or when deprotonated. Reserpine also released the oligomycin-induced respiratory control in beef heart submitochondrial particles, and inhibited energized uptake of Ca2- by rat liver mitochondria. Reserpine had a dual effect on mitochondrial ATPase: It (a) enhanced ATP hydrolysis by intact liver mitochondria, and (b) inhibited ATP hydrolysis by submitochondrial particles of beef heart. On a molar basis, reserpine was less effective than carbonyl cyanide 3-chlorophenylhydrazone in all bioenergetic reactions examined. Homogenates and mitochondria isolated from brain and liver of rats stuporous from intraperitoneally injections of Serpasil exhibited no detectable abnormalities in respiratory states and responded to known uncouplers in the expected manner. There was no evidence of in vivo uncoupling of oxidative phosphorylation as a basis of the pharmacological activity of reserpine, although interference with energy transfer may be involved in toxic manifestations of the drug. The results indicate the need for caution in interpreting the action of drugs formulated in complex pharmaceutical preparations and based solely on in vitro experiments.     

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.