miR-18b-5p在肝癌早期筛查中的临床应用价值研究

目的 观察miR-18b-5p在肝癌早期筛查中的临床应用价值。方法 随机选择2021年1月～2021年12月确诊的肝癌患者90例,乙型肝炎患者90例,丙型肝炎50例,健康体检者90例作为研究对象。所有纳入研究患者均给予miR-18b-5p水平测定。冻存于-80℃的血液样本提取总RNA,qRT-PCR法检测miR-18b-5p表达,计算PCR的扩增的Ct,比较四组纳入研究人员的miR-18b-5p水平,肝癌组患者不同性别、年龄、分期、淋巴结转移、有无腹水患者的miR-18b-5p水平。通过ROC曲线评价PCR检测miR-18b-5p的在肝癌中诊断效能。结果 肝脏组患者miR-18b-5p水平高于乙型肝炎、丙型肝炎和对照组,乙型肝炎和丙型肝炎患者miR-18b-5p水平高于对照组,差异均具有统计学意义（P <0.05）,但乙型肝炎和丙型肝炎组患者miR-18b-5p水平无明显差异（P> 0.05）。肝癌组患者根据不同病理特征分组,肿瘤大小超过5 cm患者的miR-18b-5p水平高于肿瘤大小<5 cm,TNM分期为Ⅲ期患者的miR-18b-5p水平高于Ⅰ+Ⅱ期水平,差异均...     

藁本内酯通过调控miR-133b-5p表达对脂多糖所致心肌损伤的保护作用

目的： 探讨藁本内酯（LIG）对脂多糖（LPS）所致心肌损伤的影响及分子机制。方法： 用质量浓度为10 mg·L^-1的LPS处理心肌细胞,记为LPS组,正常培养的细胞为对照（Con）组;用浓度分别为10,20,40 μmol·L^-1的藁本内酯处理心肌细胞后再用10 mg·L^-1的LPS处理,作为藁本内酯低、中、高浓度组;将miR-NC、miR-133b-5p转染至心肌细胞中再用10 mg·L^-1的LPS处理,记为LPS+miR-NC组、LPS+miR-133b-5p组;将anti-miR-NC、anti-miR-133b-5p转染至心肌细胞中再用40 μmol·L^-1的藁本内酯、10 mg·L^-1的LPS处理,记为LPS+LIG+anti-miR-NC组、LPS+LIG+anti-miR-133b-5p组。酶联免疫吸附法（ELISA）法检测肿瘤坏死因子-α（TNF-α）、白细胞介素-6（IL-6）水平;流式细胞术检测细胞凋亡;蛋白质印迹（Western blot）法检测B细胞淋巴瘤/白血病-2（Bcl-2）、Bcl-2相关X蛋白（Bax）蛋白表达;实时荧光定量PCR （RT-qPCR）检测miR-133b-5p表达水平。结果： 藁本内酯低、中、高浓度处理后,LPS诱导的心肌细胞中TNF-α、IL-6水平显著降低,细胞凋亡率显著降低,Bcl-2表达水平显著升高,Bax表达水平显著降低,miR-133b-5p表达水平显著升高,且呈浓度依赖性（P<0.05）。miR-133b-5p过表达可抑制LPS诱导的心肌细胞中TNF-α、IL-6水平和细胞凋亡。抑制miR-133b-5p表达逆转了藁本内酯对LPS所致的心肌细胞损伤的保护作用。结论： 藁本内酯通过上调miR-133b-5p表达可抑制LPS诱导的心肌细胞凋亡和炎症反应,对LPS诱导的心肌损伤具有保护作用。     

miR-199a-5p和miR-206靶向调控SULF1并抑制卵巢上皮细胞对顺铂的敏感性

目的 SULF1是硫酸酯酶家族成员之一,主要发挥肿瘤抑制因子的作用,在多种肿瘤中低表达,本研究旨在筛选直接靶向SULF1的microRNAs,并验证候选microRNAs对SULF1表达的负调控作用。方法 利用TargetScanHuman 7.2、miRDB、DIANA和RNAhybrid数据库预测调控SULF1的microRNAs。利用实时荧光定量PCR和Western blotting实验探索microRNAs对SULF1表达的影响。双荧光素酶报告基因实验探究microRNAs和SULF1基因的直接结合能力。CCK-8实验验证microRNAs对铂类药物敏感性的影响。结果 利用数据库预测到两个microRNAs：miR-199a-5p和miR-206,二者均可靶向SULF1的3''-UTR区域。实时荧光定量PCR结果显示,与IOSE80细胞中的阴性对照相比,miR-199a-5p和miR-206对应的模拟物（mimics）分别使SULF1的mRNA表达水平降低至23.07%和20.11%,而它们的抑制剂（inhibitors）则分别使SULF1的mRNA表达水平升高3.62倍和3.09倍。Western blotting结果表明,miR-199a-5p和miR-206也可降低SULF1蛋白表达水平。双荧光素酶报告基因实验表明,miR-206和miR-199a-5p均可直接结合SULF1的3''-UTR,且miR-206具有更强的调控作用。CCK-8实验结果表明,降低miR-199a-5p和miR-206的表达水平可显著增强IOSE80细胞对顺铂的敏感性（IC₅₀： 9.287 μmol·L^-1/11.32 μmol·L^-1 vs. 16.75 μmol·L^-1）,相反,二者过表达可显著降低IOSE80细胞对顺铂的敏感性。结论 miR-199a-5p和miR-206均可直接与SULF1的3''-UTR结合,下调SULF1表达,降低卵巢细胞对顺铂的敏感性。     

miR-20b-5p与TGFBR2在宫颈癌组织中的表达及意义

目的 通过分析miR-20b-5p、TGFBR2在宫颈癌组织中的表达水平及与临床病理特征的关系,探讨miR-20b-5p与TGFBR2在宫颈癌转移中的作用。方法 选取2018年1月至2020年10月诊治的58例宫颈癌患者为宫颈癌组,选取同期32例因子宫肌瘤或子宫腺肌症行全子宫切除术患者的正常宫颈组织为对照组。RT-PCR检测2组宫颈组织中miR-20b-5p、TGFBR2 mRNA表达情况,免疫组化及Western blot检测宫颈癌组织中TGFBR2蛋白表达情况,通过在线网站预测miR-20b-5p与TGFBR2的靶向结合位点,并分析两者相关性,讨论两者其在宫颈癌组中表达差异与临床病理特征的关系。结果 宫颈癌组中miR-20b-5p表达量显著高于对照组(P<0.01),而宫颈癌组中TGFBR2 mRNA表达量显著低于对照组(P<0.01)。免疫组化及Western blot结果进一步证实TGFBR2在宫颈癌组中表达水平显著低于对照组(P<0.01)。miR-20b-5p表达水平与TGFBR2 mRNA表达水平呈负相关(r=-0.7366,P<0.01)。且mi...     

miR-98-5p靶向CCR7对乳腺癌细胞MCF-7运动能力的影响及其机制研究

目的 探究miR-98-5p靶向作用于趋化因子受体（CC chemokine receptor 7,CCR7）对乳腺癌细胞MCF-7运动能力的影响及相关机制。方法 MCF-7细胞分为对照组、miR-98-5p mimic组、miR-98-5p NC组、pc-CCR7组、miR-98-5p+pc-CCR7组,分别转染相应的miRNA和CCR7过表达载体,RT-PCR检测miR-98-5p和CCR7基因表达水平,荧光素酶报告试验检测miR-98-5p和CCR7靶向关系,Transwell法检测细胞侵袭,划痕法检测细胞迁移,Western blotting检测CCR7、基质金属蛋白酶2（MMP-2）、MMP-9、血管内皮生长因子（VEGF）、E-钙黏着蛋白（E-cadherin）、N-cadherin、波形蛋白（Vimentin）蛋白表达水平。结果 与CCR7 WT+miR-98-5p NC比较,CCR7 WT+miR-98-5p mimic荧光素酶活性显著降低。与对照组比较,miR-98-5p mimic组中miR-98-5p表达水平、E-cadherin蛋白表达水平显著升高,CCR7基因和蛋白表达水平、细胞侵袭和迁移能力、MMP-2、MMP-9、VEGF、N-cadherin、Vimentin蛋白表达水平显著降低,pc-CCR7组细胞侵袭和迁移能力、CCR7、MMP-2、MMP-9、VEGF、N-cadherin、Vimentin蛋白表达水平显著升高,E-cadherin蛋白表达水平显著降低;与pc-CCR7组比较,miR-98-5p+pc-CCR7组细胞侵袭和迁移能力、CCR7、MMP-2、MMP-9、VEGF、N-cadherin、Vimentin蛋白表达水平显著降低,E-cadherin蛋白表达水平显著升高。结论 miR-98-5p可靶向作用于CCR7,抑制乳腺癌细胞MCF-7运动能力,其作用机制与上皮细胞间充质转化有关。     

利拉鲁肽调控miR-15b-5p对高糖诱导的足细胞炎性反应和免疫功能的影响

目的 探讨利拉鲁肽通过调控miR-15b-5p对高糖诱导的足细胞炎性反应和免疫功能的影响.方法 选择小鼠肾足细胞MPC-5经过葡萄糖诱导培养后,将细胞分组为对照(Con)组、高糖(HG)组、利拉鲁肽低、中、高剂量(HG+LIR-L、HG+LIR-M、HG+LIR-H)组、高糖+利拉鲁肽(HG+LIR)组、高糖+利拉鲁肽...     

miR-135 b-5 p对IL-1β诱导软骨细胞损伤的影响及机制研究

目的 探讨微小RNA-135b-5p(miR-135b-5p)是否通过靶向血管生成素样蛋白2(ANGPTL2)影响白介素-1β(IL-1β)诱导的软骨细胞损伤.方法 使用IL-1β诱导SD大鼠膝关节软骨细胞损伤.使用10 ng/ml IL-1β处理软骨细胞48 h,论为IL-1β组,同时以未经任何处理的软骨细胞设为对照...     

LncRNA TUG1通过调节miR-181b-5p/PDCD4轴对高糖诱导的心肌细胞凋亡的影响

目的 探讨长链非编码RNA(LncRNA)牛磺酸上调基因1(TUG1)通过调节miR-181b-5p/程序性细胞死亡蛋白4(PDCD4)轴对高糖诱导的心肌细胞凋亡的影响。方法 采用高糖（25 mmol/L葡萄糖）在体外构建糖尿病心肌病（DCM）细胞模型。AC16细胞分为NG组、HG组、HG+sh-NC组、HG+sh-TUG1组、HG+miR-NC组、HG+miR-181b-5p组、HG+sh-TUG1+anti-miR-NC组、HG+sh-TUG1+anti-miR-181b-5p组、HG+miR-181b-5p+pcDNA组、HG+miR-181b-5p+pc-PDCD4组。细胞计数试剂盒-8(CCK-8)法检测细胞活力;乳酸脱氢酶（LDH）测定试剂盒检测LDH释放总量;采用实时定量聚合酶链反应（q RT-PCR）检测TUG1、miR-181b-5p和PDCD4 mRNA表达;流式细胞术检测细胞凋亡;Western blot检测B细胞淋巴瘤2-相关X(Bax)、活化的胱天蛋白酶3(cleaved caspase 3)和PDCD4蛋白表达;caspase-Glo3检测试剂盒评估casp...     

A novel and low-toxic peptide DR3penA alleviates pulmonary fibrosis by regulating the MAPK/miR-23b-5p/AQP5 signaling axis

Pulmonary fibrosis (PF) is a pathological change caused by repeated injuries and repair dysfunction of the alveolar epithelium. Our previous study revealed that the residues Asn3 and Asn4 of peptide DR8 (DHNNPQIR-NH₂) could be modified to improve stability and antifibrotic activity, and the unnatural hydrophobic amino acids α-(4-pentenyl)-Ala and d-Ala were considered in this study. DR3penA (DHα-(4-pentenyl)-ANPQIR-NH₂) was verified to have a longer half-life in serum and to significantly inhibit oxidative damage, epithelial–mesenchymal transition (EMT) and fibrogenesis in vitro and in vivo. Moreover, DR3penA has a dosage advantage over pirfenidone through the conversion of drug bioavailability under different routes of administration. A mechanistic study revealed that DR3penA increased the expression of aquaporin 5 (AQP5) by inhibiting the upregulation of miR-23b-5p and the mitogen-activated protein kinase (MAPK) pathway, indicating that DR3penA may alleviate PF by regulating MAPK/miR-23b-5p/AQP5. Safety evaluation showed that DR3penA is a peptide drug without obvious toxicity or acute side effects and has significantly improved safety compared to DR8. Thus, our findings suggest that DR3penA, as a novel and low-toxic peptide, has the potential to be a leading compound for PF therapy, which provides a foundation for the development of peptide drugs for fibrosis-related diseases.     

Role of miR-199b-5p in regulating angiogenesis in mouse myocardial microvascular endothelial cells through HSF1/VEGF pathway

Our study explored effects of miR-199b-5p on angiogenesis in mouse myocardial microvascular endothelial cells (MMVECs) and the involved working mechanisms. We applied explant culture to incubate C57/BL6 mouse MMVECs. Lipofection was used to transfect miR-199b-5p mimic, miR-199b-5p inhibitor and miR-199b-5p scramble respectively. MMVECs were divided into miR-199b-5p up-regulation, miR-199b-5p down-regulation and control groups based on above sequence. Expressions of miR-199b-5p, heat shock factor protein 1 (HSF1) mRNA were assessed by real-time quantitative polymerase chain reaction (RT-QPCR). Expressions of HSF1 and vascular endothelial growth factor (VEGF) were assessed by Western Blotting. Cell proliferation was assessed by CCK8. Tubule formation assay was conducted to assess formation of blood vessels. Results showed that miR-199b-5p up/down-regulation groups exhibited no obvious differences in the expressions of HSF1 mRNA compared to control group. However, miR-199b-5p up-regulation group recorded lower expressions of HSF1 and VEGF in the level of protein, and reduced cell proliferation and tubule formation. Whereas, miR-199b-5p down-regulation group presented the contrary results. The experiment indicated that miR-199b-5p can regulate proliferation and angiogenesis in mouse MMVECs through the pathway of HSF1/VEGF.     

微小RNA-216b-5p对骨肉瘤MG63细胞功能的影响及其与高迁移率族蛋白B1的靶向关系

目的 探讨微小RNA-216b-5p(miR-216b-5p)对骨肉瘤MG63细胞增殖、侵袭和凋亡的影响及其与高迁移率族蛋白B1(HMGB1)的靶向关系.方法 将体外培养的MG63细胞分为mimics-NC组、miR-216b-5p mimics组、inhibitor-NC组和miR-216b-5p inhibitor...     

MiR-33b-5p sensitizes gastric cancer cells to chemotherapy drugs via inhibiting HMGA2 expression

MicroRNAs (miRNAs) are internal, non-coding, and ～22?nt small RNAs that display cell- and tissue-specific expression. They play important regulatory roles in cell proliferation and chemo-sensitivity. This study focused on tumor-suppressive miR-33b-5p expression as well as its role in gastric cancer. MiR-33b-5p was found low expression in gastric cancer cell lines. Functionally, western blots and the luciferase reporter assay were used to confirm that HMGA2 was the potential target of miR-33b-5p. Next, we used CCK-8 kits to analyze the effect of miR-33b-5p combined chemotherapy drugs on cell inhibition rate, and flow cytometry to analyze cells apoptosis. Colony formation ability was determined by plating at 500 cells per well into six-well plates and culturing for 15?d. The results showed that upregulation of miR-33b-5p decreased expression of HMGA2 and inhibited gastric cancer cell growth as well as sensitized gastric cancer cells to chemotherapy drugs. MiR-33b-5p overexpression hindered luciferase activity of HMGA2,3′-untranslated region-based reporter construct in 293?T cells. These data demonstrate that miR-33b-5p may be a potential therapeutic target for gastric cancer and function as tumor-suppressive miRNA through targeting HMGA2 in gastric cancer.     

盖诺顺铂化疗加放疗对乳腺癌MCF-7细胞作用时机的实验研究

目的放化同期综合治疗应用越来越广泛，主要机制是放化疗对肿瘤组织的协同杀灭作用。该研究对乳腺癌放化同期治疗有效性、时机选择进行实验探讨。方法选用盖诺顺铂联合化疗（NP方案）对乳腺癌MCF-7细胞放化同期放射和联合化疗不同间隔时间后放射，比较单纯照射组、同期化放疗组和联合化疗后不同间隔时间放射组的细胞存活率（Survival Fraction，SF）。结果联合化疗加同期放射后SF较单独放射更低。比较联合化学药物处理后不同间隔时间放射，发现SF值最低分别是化疗处理后间隔4h，12h和36h组，联合化疗加同期放射组SF值居中，而间隔48h和72h后照射组，SF值明显上升。SF最低与最高值相差约15倍。结论联合化放疗对乳腺癌MCF-7细胞具有较强的协同杀灭作用，化放联合治疗的时机选择在肿瘤细胞杀灭过程中具有重要的意义，间隔48h和72h后照射放化疗协同杀灭效应减弱。     

MiR-199a-5p and miR-375 affect colon cancer cell sensitivity to cetuximab by targeting PHLPP1

Objectives: We aimed to analyze the differentially-expressed miRNAs in colon cancer cells in order to identify novel potential biomarkers involved in cancer cell resistance.

Design and methods: We investigated the miRNA expression profile of GEO human colon carcinoma cells, sensitive to the EGFR inhibitor Cetuximab (CTX) and their CTX-resistant counterpart (GEO CR) by using a miRNA chip.

Results: We found 27 upregulated and 10 downregulated miRNAs in GEO CR compared with GEO cells with a fold change ≥ 2. Among the upregulated miRNAs, we focused on miR-199a-5p and miR-375. We report that their enforced expression promotes CTX resistance, whereas their silencing sensitizes to the same drug. The ability of miR-199a-5p and miR-375 to target PHLPP1 (PH domain and leucine-rich repeat protein phosphatase 1), a tumor suppressor that negatively regulates the AKT pathway, accounts, at least in part, for their drug-resistance activity. Indeed, restoration of PHLPP1 increases sensitivity of the GEO cells to CTX and reverts the resistance-promoting effect of miR-199a-5p and miR-375.

Conclusion: This study proposes miR-199a-5p and miR-375 as contributors to CTX resistance in colon cancer and suggests a novel approach based on miRNAs as tools for the therapy of this tumor.     

长链非编码RNA膀胱癌相关转录本1靶向调控微小RNA-20b-5p对宫颈癌细胞自噬及顺铂敏感性的影响

目的 探讨长链非编码RNA膀胱癌相关转录本1(LncRNA BLACAT1)对宫颈癌细胞自噬及顺铂敏感性的影响及其分子机制.方法 本研究起止时间2018年6月至2019年10月,体外培养人正常宫颈上皮细胞HUCEC、宫颈癌细胞株ME180、C-33A、Hela与顺铂耐药宫颈癌细胞株Hela/DDP,实时荧光定量聚合酶链...     

微小 RNA-525-5p通过靶向 RECQ样蛋白 5调控乳腺癌放射敏感性

目的探讨微小 RNA-525-5p(miR-525-5p)对乳腺癌细胞辐射照射敏感性的影响及机制。方法该研究起止时间为 2019年 6月至 2020年 6月,人乳腺癌细胞 MDA-MB-231、MCF-7、BT474、非恶性乳腺上皮细胞 MCF-10A均购自美国菌种保藏中心。运用 qRT-PCR法检测人乳腺癌细胞 MDA-MB-231、MCF-7、BT474、非恶性乳腺上皮细胞 MCF-10A中 miR-525-5p的 mRNA的表达;将 miR-525-5p模拟物( miR-525-5p mimics)阴性对照( miR-NC)组(转染 miR-NC)、 miR-525-5p组(转染 miR-525-5p mim. ics)、 RECQL5小干扰 RNA阴性对照( si-NC)组(转染 si-NC)、 RECQL5小干扰 RNA(si-RECQL5)组(转染 si-RECQL5)、 miR-525-5p+RECQL5过表达空载体( pcDNA)组(共转染 miR-525-5p mimics和 pcDNA)、 miR-525-5p+RECQL5过表达载体( pcDNA-REC. QL5)组(共转染 miR-525-5p mimics和 pcDNA-RECQL5),均用脂质体法转染至 MDA-MB-231细胞; Western blotting检测细胞中 RECQ样蛋白 5(RECQL5)的蛋白表达;流式细胞术检测细胞凋亡;双荧光素酶报告基因检测实验检测细胞的荧光活性;克隆形成实验检测细胞的存活分数。结果与非恶性乳腺上皮细胞 MCF-10A相比,人乳腺癌细胞 MDA-MB-231、MCF-7、BT474中 miR-525-5p表达［ 0.28±0.02,0.33±0.02,0.42±0.03比 1.01±0.08］显著降低, RECQL5 mRNA和蛋白表达［ 1.68±0.15,1.58±0.12,     

微小RNA-301b-3p靶向第10号染色体缺失的磷酸酶及张力蛋白同源物对骨肉瘤细胞增殖、凋亡的影响

目的 探讨微小RNA-301b-3p(miR-301b-3p)对骨肉瘤细胞增殖、凋亡的影响及其作用机制.方法 本研究起止时间为2019年3—9月.人成骨细胞(hFOB)和骨肉瘤细胞U2OS、MG63购自美国菌种保藏中心.U2OS细胞分为miRNA抑制物阴性对照(anti-miR-NC)组、miR-301b-3p抑制物(...