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。     

Rhein attenuates renal inflammatory injury of uric acid nephropathy via lincRNA-Cox2/miR-150-5p/STAT1 axis

Rhein has protective effect on uric acid nephropathy (UAN). This article aims to demystify the mechanism of function of rhein in UAN. Mouse kidney epithelial cell line (TCMK-1) was incubated with uric acid (UA) to induce inflammatory injury. Then, the TCMK-1 cells were treated with rhein. The relationships among lincRNA-Cox2, miR-150-5p and STAT1 were evaluated by luciferase reporter assay. CCK8 and flow cytometry were performed to detect cell proliferation and apoptosis. The levels of IL-6, IL-1β and TNF-α were investigated by enzyme linked immunosorbent assay. Western blot and quantitative real-time PCR were performed to examine the expression of genes and proteins. We found that UA suppressed proliferation and enhanced apoptosis and the levels of IL-6, IL-1β and TNF-α of TCMK-1 cells, which was effectively improved by rhein treatment. Furthermore, lincRNA-Cox2 overexpression caused an increase of apoptosis and inflammatory factors in the rhein-treated TCMK-1 cells. LincRNA-Cox2 regulated STAT1 expression by sponging miR-150-5p. And lincRNA-Cox2 promoted apoptosis and inflammatory injury of TCMK-1 cells by regulating miR-150-5p/STAT1 axis. In summary, our studies demonstrate that rhein has a protective effect against UAN by inhibiting renal inflammatory injury via lincRNA-Cox2/miR-150-5p/STAT1 axis.     

2型糖尿病肾病患者血清miR-27b-3p、miR-342-3p表达特点及临床意义

目的 探讨2型糖尿病肾病（DN）患者血清miR-27b-3p、miR-342-3p表达特点及其对DN的诊断价值。方法 将157例2型糖尿病患者根据尿白蛋白与肌酐比率（UACR）分为正常白蛋白尿（NA）组49例、微量白蛋白尿（MA）组41例、大量白蛋白尿（DN）组67例。收集临床资料,采用实时定量聚合酶链反应检测血清miR-27b-3p、miR-342-3p表达,全自动生化分析仪检测血尿素氮（BUN）、血肌酐（Scr）,双抗体夹心酶联免疫吸附试验检测尿液Ⅳ型胶原（Ⅳ-C）、层粘连蛋白（LN）、Ⅲ型前胶原肽（PCⅢ）。分析DN发病的影响因素以及miR-27b-3p、miR-342-3p对DN的诊断价值。结果 与NA组比较,DN组和MA组HbA1c、UACR、尿Ⅳ-C、尿LN、尿PCⅢ水平均增加,而miR-27b-3p、miR-342-3p表达均降低（P<0.05）,DN组BUN、Scr水平增加、估算肾小球滤过率（eGFR）水平降低（P<0.05）。较高水平的尿PCⅢ是DN的危险因素,而较高水平的miR-27b-3p、miR-342-3p、eGFR是DN保护因素（P<0.0...     

LncRNA PVT1通过竞争miR-149-5p上调CHI3L1并影响变应性鼻炎进展的研究

目的　探讨长链非编码RNA（LncRNA）PVT1对变应性鼻炎（AR）进展的影响及其相关作用机制。方法　24只BALB/c小鼠,其中6只作为对照组,余18只采用卵白蛋白和氢氧化铝建立AR模型并采用随机数字表法均分为模型组、AR+NC组（尾静脉注射空载体慢病毒）、AR+shPVT1组（尾静脉注射LncRNA PVT1干扰载体慢病毒）,每组6只。采用行为学评分评估各组小鼠过敏症状;HE染色观察各组小鼠鼻黏膜组织病理学改变;实时荧光定量PCR检测各组小鼠鼻黏膜组织中LncRNA PVT1、miR-149-5p和几丁质酶-3样蛋白1（CHI3L1）mRNA的表达;Western blot检测CHI3L1蛋白表达;ELISA分析各组小鼠血清IgE、白细胞介素（IL）-5、肿瘤坏死因子（TNF）-α水平。构建LncRNA PVT1和CHI3L1的野生型（WT）和突变型（MT）荧光素酶报告质粒,单独或同时与miR-149-5p mimic共转染293T细胞,采用双荧光素酶实验分析并计算各组细胞荧光素酶相对活性。结果　与模型组相比,AR+shPVT1组小鼠行为学评分降低;血清中IgE、IL-5和TNF-α水平降低;鼻黏膜组织中LncRNA PVT1 mRNA表达水平降低,miR-149-5p mRNA表达水平升高,CHI3L1 mRNA及蛋白水平均降低,鼻黏膜组织炎症反应减轻。双荧光素酶实验证实LncRNA PVT1通过竞争性结合miR-149-5p,上调miR-149-5p靶基因CHI3L1的表达。结论　在AR模型小鼠中异常高表达的LncRNA PVT1通过发挥竞争性内源RNA作用抑制miR-149-5p表达而上调CHI3L1的表达,引发小鼠鼻黏膜组织的炎性症状并促进过敏反应。     

Differentially expressed mRNAs and their upstream miR-491-5p in patients with coronary atherosclerosis as well as the function of miR-491-5p in vascular smooth muscle cells

MicroRNAs (miRNAs) regulate gene expression and are biomarkers for coronary atherosclerosis (AS). A novel miRNA-mRNA regulation network of coronary AS still needs to be disclosed. The aim of this study was to analyze potential mRNAs in coronary AS patients and the role of their upstream miR-491-5p in vascular smooth muscle cells (VSMCs). We first confirmed top ten mRNAs according to the analysis from Gene Expression Omnibus database ({"type":"entrez-geo","attrs":{"text":"GSE132651","term_id":"132651"}}GSE132651) and examined the expression levels of them in the plaques and serum from AS patients. Five mRNAs (UBE2G2, SLC16A3, POLR2C, PNO1, and AMDHD2) presented significantly abnormal expression in both plaques and serum from AS patients, compared with that in the control groups. Subsequently, they were predicted to be targeted by 11 miRNAs by bioinformatics analysis. Among all the potential upstream miRNAs, only miR-491-5p was abnormally expressed in the plaques and serum from AS patients. Notably, miR-491-5p overexpression inhibited viability and migration, and significantly increased the expression of contractile markers (α-SMA, calponin, SM22α, and smoothelin) in VSMCs. While silencing miR-491-5p promoted viability and migration, and significantly suppressed the expression of α-SMA, calponin, SM22α, and smoothelin. Overall, miR-491-5p targeted UBE2G2, SLC16A3, and PNO1 and regulated the dysfunctions in VSMCs.     

黄芩苷通过上调miR-485-5p的表达抑制前列腺癌细胞的增殖

目的：从miRNAs的角度研究黄芩苷对前列腺癌细胞增殖的影响及其作用机制。方法：将不同浓度的黄芩苷作用22RV1细胞,应用MTT检测细胞的增殖情况,筛选最佳药物浓度;应用Real-Time PCR检测不同细胞系中miR-485-5p的表达量;检测黄芩苷处理22RV1细胞后miR-485-5p及JAK3 mRNA的表达;应用Western blot检测JAK3蛋白的表达;应用双荧光素酶基因系统验证miR-485-5p与JAK3的靶向作用关系;应用MTT法检测细胞的生存率。结果：筛选出最佳药物浓度为100 μmol·L^-1;22RV1细胞中miR-485-5p的表达量明显低于正常前列腺细胞;黄芩苷刺激细胞后miR-485-5p的表达上调;黄芩苷或miR-485-5p mimics可抑制前列腺癌细胞的JAK3 mRNA和蛋白的表达,miR-485-5p inhibitor可使JAK3 mRNA及蛋白的表达水平上调;黄芩苷或miR-485-5p均可抑制前列腺癌细胞的增殖。结论：黄芩苷通过上调miR-485-5p的表达,抑制前列腺癌细胞的增殖。     

Milk exosomes-mediated miR-31-5p delivery accelerates diabetic wound healing through promoting angiogenesis

The refractory diabetic wound has remained a worldwide challenge as one of the major health problems. The impaired angiogenesis phase during diabetic wound healing partly contributes to the pathological process. MicroRNA (miRNA) is an essential regulator of gene expression in crucial biological processes and is a promising nucleic acid drug in therapeutic fields of the diabetic wound. However, miRNA therapies have limitations due to lacking an effective delivery system. In the present study, we found a significant reduction of miR-31-5p expression in the full-thickness wounds of diabetic mice compared to normal mice. Further, miR-31-5p has been proven to promote the proliferation, migration, and angiogenesis of endothelial cells. Thus, we conceived the idea of exogenously supplementing miR-31-5p mimics to treat the diabetic wound. We used milk-derived exosomes as a novel system for miR-31-5p delivery and successfully encapsulated miR-31-5p mimics into milk exosomes through electroporation. Then, we proved that the miR-31-5p loaded in exosomes achieved higher cell uptake and was able to resist degradation. Moreover, our miRNA-exosomal formulation demonstrated dramatically improved endothelial cell functions in vitro, together with the promotion of angiogenesis and enhanced diabetic wound healing in vivo. Collectively, our data showed the feasibility of milk exosomes as a scalable, biocompatible, and cost-effective delivery system to enhance the bioavailability and efficacy of miRNAs.     

Long non-coding RNA UCA1 promotes autophagy by targeting miR-96-5p in acute myeloid leukaemia

Long non-coding RNA (lncRNA) urothelial carcinoma-associated 1 (UCA1) has been identified as an oncogene and is involved in acute myeloid leukaemia (AML). Autophagy contributes to tumourigenesis and cancer cell survival. The purpose of this study was to investigate the regulatory role and mechanism of UCA1 in AML cell viability by its effect on autophagy. The expression of UCA1, miR-96-5p, and ATG7 was determined by qRT-PCR and western blot. Cell proliferation was examined by MTT assay. The autophagy level was assessed by green fluorescent protein (GFP)-LC3 immunofluorescence and western blot. The interaction between UCA1 and miR-96-5p or ATG7 was analyzed by luciferase reporter activity. The results showed that UCA1 promoted AML cell proliferation by inducing autophagy. Mechanistically, UCA1 acted as a sponge of miR-96-5p by binding to miR-96-5p. ATG7 was a direct target of miR-96-5p and positively regulated by UCA1. Further results showed that the miR-96-5p mimic effectively counteracted the UCA1 overexpression-mediated induction of the ATG7/autophagy pathway. Collectively, UCA1 functions as a sponge of miR-96-5p to upregulate its target ATG7, thereby resulting in autophagy induction. Our findings reveal a UCA1-mediated molecular mechanism responsible for autophagy induction in AML and help to improve the understanding of the molecular mechanism of AML progression.     

Long noncoding RNA GAS5 promotes microglial inflammatory response in Parkinson's disease by regulating NLRP3 pathway through sponging miR-223-3p

Parkinson’s disease (PD) is the second most common neurodegenerative disorder. Neuroinflammation induced by microglia plays an important role in the pathogenesis of PD. Long noncoding RNA GAS5 was showed to have significant effects on regulating inflammatory response. Here, we aim to investigate the effects of GAS5 on the inflammatory response of PD, and the underlying mechanism. An in vivo model of PD was established in C57BL/6 mice by rotenone and an in vitro cell model was conducted on microglia by lipopolysaccharide (LPS). Our results indicated that GAS5 was upregulated in tissues in a mice model of PD and microglia activated by LPS. Gain- and loss- of functional experiments demonstrated that GAS5 promoted the inflammation of microglia in vitro. Besides, the knockdown of GAS5 repressed the PD progression in vivo. Mechanistically, GAS5 positively regulated the NLRP3 expression via competitively sponging miR-223-3p. Overall, our finding illuminates that GAS5 accelerates PD progression through targeting miR-223-3p/NLRP3 axis.     

IgA肾病患者的Th细胞亚群紊乱及其意义

目的探讨IgA肾病Th细胞亚群紊乱与发病机制、免疫紊乱程度、临床表现及病理类型之间的关系。方法以趋化因子CCR3和CCR5为辅助性T细胞Th2和Th1细胞特异性表面标志,用流式细胞术测定IgA肾病患者及健康对照组外周血CCR3 CD4 /CCR5 CD4 的T细胞亚群;并检验患者组的尿常规和肾功能。结果IgA肾病患者外周血CCR3 CD4 的T细胞亚群为(5.00±4.46)%,较对照组(1.56±0.73)%明显增多,CCR5 CD4 的T细胞亚群为(7.11±6.28)%,较对照组(13.85±3.58)%明显减少(P<0.01);免疫紊乱程度与临床表现及病理类型无相关性(P>0.05)。结论IgA肾病患者外周血Th2免疫应答增强、Th1免疫应答减弱,可能与该病的发病机制有关。     

长链非编码 RNA肝癌高表达转录本通过微小 RNA-134-5p促进宫颈癌细胞恶性生物学行为的机制研究

目的探讨长链非编码 RNA(lncRNA)肝癌高表达转录本( HULC)通过抑制微小 RNA-134-5p促进宫颈癌细胞增殖、袭和迁移,促进凋亡的作用及其作用机制。方法 2019年 10月至 2020年 5月,通过实时荧光定量 PCR(qRT-PCR)检测体外培侵养的人正常宫颈上皮细胞和宫颈癌细胞中 HULC的表达情况,在宫颈癌 C-33A细胞中转染特异性 HULC siRNA,qRT-PCR检测转染效果,细胞计数试剂盒( CCK-8)法分析 C-33A细胞增殖情况,采用流式细胞术分析 C-33A细胞凋亡情况, Transwell实验分析 C-33A细胞侵袭和迁移能力,双荧光素酶报告基因实验检测 HULC与 miR-134-5p的相互作用。在抑制 HULC表达的 C-33A细胞中转染 miR-134-5p抑制剂,以同样的方法检测对细胞增殖、侵袭和迁移的影响。结果 HULC在宫颈癌细胞( HeLa、Cas. ki、SiHa、C-33A)中的表达水平为(1.95±0.24、2.26±0.21、1.77±0.19、3.49±0.40),显著高于正常宫颈上皮细胞 1.00±0.11,HULC在 C-33A细胞中的表达量最高( P<0.05)。转染特异性 HULC siRNA能够抑制 C-33A细胞中 HULC的表达( 0.94±0.08比 0.18±     

miR-218-5p靶向B细胞淋巴瘤因子3表达影响白血病K562细胞的增殖和凋亡

目的 探讨微小RNA-218-5p(miR-218-5p)靶向B细胞淋巴瘤因子3(BCL3)基因对白血病细胞增殖和凋亡的影响.方法 反转录及荧光定量PCR(RT-qPCR)和蛋白质印迹法(Western blotting)检测正常人骨髓细胞和白血病骨髓细胞中miR-218-5p和BCL3的表达水平.以白血病细胞K562...     

术前血清miR-483-5p水平在肝内胆管细胞癌诊断及预后中的价值

目的:探讨术前血清miR-483-5p水平在肝内胆管细胞癌(ICC)早期诊断及预后评估中的价值。方法:选择2015年12月—2017年12月收治的91例ICC患者为ICC组,选择同期40例健康体检者为对照组,统计一般临床资料并检测血清miR-483-5p水平,采用受试者工作特征曲线,分析miR-483-5p对ICC的诊断效能。根据miR-483-5p中位值将ICC患者分为低表达组和高表达组,分析比较两组患者病理特征的差异,应用Kaplan-Meier法进行生存分析,并对生存期进行单因素和多因素Cox回归分析。结果:ICC组患者血清miR-483-5p水平明显高于对照组(P<0.05);miR-483-5p诊断ICC的曲线下面积为0.758,敏感性55.00%,特异性86.81%。高表达组和低表达组之间的肿瘤分化程度和淋巴结转移比较,差异有统计学意义(P<0.05)。Kaplan-Meier生存分析显示,高表达组患者的整体生存期明显低于低表达组(P<0.05)。单因素及多因素Cox回归分析发现,血清miR-483-5p水平与ICC患者术后生存期有关,但不是生存期的独立影响因素。结论:血清miR-483-5p水平对ICC具有一定诊断价值,且高miR-483-5p水平ICC患者的术后生存期相对较短,可作为ICC早期诊断及预后评估辅助评价指标。     

SIRT1 targeted by miR-211-5p regulated proliferation and apoptosis of Dex-treated growth plate chondrocytes via mediating SOX2

Dexamethasone (Dex) is reported to cause bone growth retardation in children, which is associated with the increased apoptosis and decreased proliferation of growth plate chondrocytes. Sirtuin 1 (SIRT1) plays an important role in chondrocyte function and homeostasis. Thus, we further explored the regulatory mechanism of SIRT1 in Dex-induced growth plate chondrocyte dysfunction. SIRT1 expression was detected in Dex-treated growth plate chondrocytes using RT-qPCR and western blot assay. The modulation of SIRT1 on SOX2 expression was evaluated. Besides, we identified that SIRT1 was targeted by miR-211-5p using TargetScan and RNA pull-down assay. A loss-of-function assay was performed to evaluate the effects of miR-211-5p on Dex-induced growth plate chondrocyte dysfunction in vitro and in vivo. We found that SIRT1 was downregulated in Dex-treated growth plate chondrocytes. The expression of SOX2 was upregulated by overexpression SIRT1. Meanwhile, downregulation of SOX2 weakened the positive function of SIRT1 overexpression on Dex-induced growth plate chondrocytes dysfunction. Subsequently, we confirmed that SIRT1 was targeted by miR-211-5p. MiR-211-5p inhibitor increased the expression levels of SIRT1 and SOX2, and restored the Dex-treated growth plate chondrocyte function. Animal assays further demonstrated that the effects of miR-211-5p on the growth plate chondrogenesis. In conclusion, our data suggest that SIRT1 exerts a protective effect on growth plate chondrocyte under Dex stimulation. MiR-211-5p/SIRT1/SOX2 axis regulates the process of Dex-inhibited growth plate chondrogenesis.     

Nardosinone suppresses monoiodoacetate-induced osteoarthritis in rats: The key role of the miR-218-5p/NUMB axis

Nardosinone is a bioactive compound with a sesquiterpenoid structure isolated from Nardostachys jatamansi. The compound has shown treatment effects against skeletal disorders. In the current study, the effects of nardosinone on osteoarthritis (OA) were first assessed and the mechanism underlying the effects was explored by detecting changes in the miR-218-5p/NUMB axis. The miR, as a potential target mediating the effects of nardosinone on OA, was first determined with microarray and RT-qPCR detections. Then, OA symptoms were induced in rats using monoiodoacetate (MIA) and treated with nardosinone. The anti-OA effects of nardosinone were assessed via the detection of the histological structure and inflammation. The role of miR-218-5p was delineated by modulating its levels in OA-affected rats. Based on the results of microarray and RT-qPCR detections, miR-218-5p was selected as the therapeutic target for nardosinone. The induction of OA resulted in tissue destruction and the production of cytokines in rat joint tissues, which was associated with the up-regulation of miR-218-5p and the downregulation of NUMB. For OA-affected rats treated with nardosinone, the joint structure was improved and the inflammatory response was suppressed, along with the restored expression levels of miR-218-5p and NUMB. The re-induced level of miR-218-5p compromised the anti-OA effects of nardosinone, indicating that the inhibition of the miR played an indispensable role in the anti-OA function of nardosinone. Collectively, the findings of our study demonstrated that nardosinone exerts treatment effects against OA by modulating the miR-218-5p/NUMB axis. Future studies will provide more detailed information on the interaction between nardosinone and miR in the attenuation of OA.     

大鼠miR-142-3p、 miR-145-3p表达模式分析

摘要： 目的 筛选并鉴定对大鼠乳腺发育和泌乳调控起关键作用的微 RNAs （miRNAs）。方法 以 U6 为内参基因, 运用实时荧光定量 PCR, 比较 miR-142-3p、 miR-145-3p 在泌乳 21 d 大鼠乳腺、 肝、 心、 脾、 肺、 肾、 卵巢、 子宫各器官的组织样品表达量的差异及不同泌乳阶段 （1、 7、 21 d） 乳腺组织 miR-142-3p、 miR-145-3p 的表达规律。结果 miR-142-3p 在泌乳 21 d 各组织的表达存在差异, 乳腺中的表达量显著高于心、 脾、 肺、 肾、 卵巢和子宫, 仅次于肝中的表达量 （P < 0.05）。miR-145-3p 在乳腺中的表达量显著高于肝、 脾和肾, 而在心、 肺、 卵巢和子宫中差异无统计学意义 （P > 0.05）。乳腺组织在泌乳 1、 7、 21 d 比较, miR-142-3p 的相对表达量持续下调, 而 miR-145-3p 的相对表达量先下调后上调。结论 miR-142-3p 和 miR-145-3p 在大鼠各组织及不同生理时期存在表达差异; miR-142-3p 可通过调节靶基因催乳素受体 （Prlr） 来调控乳腺的发育和泌乳的形成, miR-145-3p 可能具有与 miR-145、 miR-145-5p 相似的功能。     

微小RNA-182-5p调控焦亡参与肝缺血再灌注损伤

摘要：目的 探讨微小RNA-182-5p（miR-182-5p）靶向调控叉形头转录因子O亚型3a（FoxO3a）调节焦亡对肝 缺血再灌注损伤的影响。方法 建立小鼠肝脏缺血再灌注模型。按随机数字表法将40只小鼠分为5组,每组8只, 分别为假手术（sham）组,缺血再灌注（IR）各组（缺血1.5 h,按再灌注时间分为IR 2 h组、IR 6 h组、IR 12 h组和IR 24 h 组）。细胞实验分组分为两部分,（1）缺氧模型建立,分为 control 组和 IR 组。（2）缺氧/复氧模型建立,分为对照组、 mimic组,inhibitor组和inhibitor+siRNA组。HE染色观察肝组织病理变化;免疫细胞化学染色检测FoxO3a表达分布 情况;荧光实时定量PCR（qRT-PCR）和Western blot 分别从mRNA和蛋白水平检测miR-182-5p、FoxO3a、半胱天冬 酶-1（Caspase1）、白细胞介素（IL）-1β、IL-18 表达情况;分析 miR-182-5p 与 FoxO3a 基因相关性;免疫荧光检测 Caspase1表达情况;ELISA检测IL-1β和IL-18表达情况;CCK-8试剂盒检测细胞活性变化情况。结果 IR处理后小 鼠肝组织损伤增加,再灌注12 h时损伤最重,同时FoxO3a、Caspase1、IL-1β、IL-18表达增加（P＜0.05）,诱导焦亡产 生;IR处理后小鼠肝组织内miR-182-5p表达水平较sham组升高（P＜0.05）。体外培养小鼠肝细胞AML12,IR处理 后miR-182-5p表达上调,FoxO3a表达下调,同时Caspase1表达升高（P＜0.05）。过表达miR-182-5p能降低FoxO3a 表达水平;反之能增加FoxO3a表达量,进而降低Caspase1、IL-1β、IL-18的表达,增加肝细胞活性（P＜0.05）。结论 激活miR-182-5p能加重肝缺血再灌注损伤,而抑制miR-182-5p能减轻肝缺血再灌注损伤,其机制可能与miR-182- 5p通过靶向调控FoxO3a激活肝细胞焦亡、影响Caspase1、IL-1β、IL-18表达有关。     

人参皂苷Rg1通过miR-138-5p/SIRT1轴诱导骨髓间充质干细胞向髓核样细胞增殖分化

目的 探讨人参皂苷Rg1诱导大鼠骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)向髓核样细胞(nucleus pulposus cells,NPCs)分化的机制。方法 从成年SD大鼠中分离培养BMSCs。通过CCK-8法检测不同浓度人参皂苷Rg1(0,2.5,5,10,20 μmol·L^-1)对BMSCs生存活性的影响。将细胞分为空白组,5 μmol·L^-1人参皂苷Rg1干预组和10 μmol·L^-1人参皂苷Rg1干预组。培养48 h后通过流式细胞术检测细胞凋亡率;Western blotting检测NPCs标志蛋白胶原蛋白2型(collagen type 2,COL2)、Aggrecan和配对盒基因(paired box gene 1,Pax1)的表达;将细胞分为空白组、人参皂苷Rg1干预组、人参皂苷Rg1干预+NC mimic组和人参皂苷Rg1干预+miR-138-5p mimic组;通过以上方法进行进一步检测miR-138-5p对人参皂苷Rg1诱导BMSCs分化的影响;双荧光素酶报告基因和RNA结合蛋白免疫沉淀法检测miR-138-5p和sirtuin-1(SIRT1)的靶向关系;转染pcDNA-SIRT1检测其对miR-138-5p诱导BMSCs分化的影响。结果 2.5,5,10 μmol·L^-1人参皂苷Rg1呈剂量依赖性增强BMSCs活力(P < 0.05);与空白组比较,5 μmol·L^-1和10 μmol·L^-1人参皂苷Rg1干预组呈剂量依赖性抑制细胞凋亡(P < 0.05),促进NPCs标志蛋白表达(P < 0.05);与人参皂苷Rg1干预组相比,人参皂苷Rg1+miR-138-5p组BMSCs凋亡率增加(P < 0.01),NPCs标志蛋白表达减少(P < 0.01);此外,SIRT1是miR-138-5p的靶基因;与miR-138-5p mimic+pcDNA-3.1组相比,miR-138-5p mimic+pcDNA-SIRT1组细胞凋亡率降低(P < 0.01),NPCs标志蛋白表达增加(P < 0.01)。结论 人参皂苷Rg1可通过调控miR-138-5p/SIRT1轴诱导BMSCs向NPCs增殖和分化。     

过表达微小 RNA-877-5p通过靶向叉头框转录因子 M1调控胃癌细胞活力和凋亡

目的探讨微小 RNA-877-5p(miR-877-5p)对胃癌细胞活力、凋亡的影响及其分子机制。方法本研究时间为 2020年 1—7月。胃癌和正常胃黏膜上皮细胞株购自美国典型培养物保藏中心。实时定量基因扩增荧光检测( qPCR)检测胃癌细胞株 HGC-27、SUN-1、AGS和正常胃黏膜上皮细胞株 GES-1中 miR-877-5p和叉头框转录因子 M1(FOXM1)信使核糖核酸( mRNA)表达。建立 miR-877-5p过表达或抑制 FOXM1表达细胞株,观察其在 HGC-27细胞的活力、凋亡中的作用。 MTT法检测细胞的活力,流式细胞术检测细胞凋亡,蛋白质印迹法( Western blotting)检测 FOXM1、细胞周期蛋白 D1(cyclinD1)、细胞周期依赖性激酶抑制因子 p21、p27、B细胞淋巴瘤 /白血病 -2(Bcl-2)、 Bcl-2相关 X蛋白( Bax)、活化的含半胱氨酸的天冬氨酸蛋白水解酶 3(cleaved-caspase 3)蛋白表达。 TargetScan预测结合双荧光素酶报告实验分析 miR-877-5p和 FOXM1的靶向关系。共转染 miR-877-5p模拟物和 FOXM1过表达载体( pcDNA-FOXM1),研究 FOXM1过表达对 miR-877-5p过表达诱导的 HGC-27细胞增殖和凋亡的影响。结果与正常胃黏膜上皮细胞 GES-1比较,胃癌细胞 HGC-27、SUN-1、AGS中的 miR-877-5p表达下调( 1.00±0.08比 0.34±0.03,0.51±0.05,0.44±0.04,P<0.05)FOXM1 mRNA和蛋白表达上调( 1.00±0.09比 2.41±0.23,2.58±0.24,2.26±0.23,P< 0.05)。 miR-877-5p过表达显著降低 HGC-27细,胞 48 h、72 h的细胞活力( P<0.05),明显提高细胞凋亡率、 p21、p27、Bax、cleaved-caspase3蛋白的水平( P<0.05)显著减少 cyclinD1、Bcl-2蛋白表达量( P<0.05)。抑制 FOXM1表达显著降低 48 h、72 h的细胞活力( P<0.05)提高细胞凋亡率、 p,21、Bax蛋白水平( P<0.05),减少 cyclinD1、Bcl-2蛋白表达量( P<0.05)。 miR-877-5p靶向调控 FOXM1的表达,。FOXM1过表达后, miR-877-5p过表达对 HGC-27细胞增殖、 cyclinD1、Bcl-2蛋白表达的抑制作用被逆转,其对细胞凋亡、 p21、Bax蛋白表达的促进作用也被逆转。结论过表达 miR-877-5p通过靶向调控 FOXM1表达抑制胃癌细胞的活     

LncRNA CDKN2B-AS1 relieved inflammation of ulcerative colitis via sponging miR-16 and miR-195

BackgroundThis study was aimed to explore the differential expression of lncRNA CDKN2B-AS1-miR-195-5p/miR-16-5p axis in ulcerative colitis (UC) and its role in regulating UC pathogenesis.MethodsOne hundred and eighty-seven UC patients and one hundred and fifty-two healthy volunteers were recruited, and their blood samples were collected. Inflammatory cytokines in serum were determined with ELISA, and lncRNA CDKN2B-AS1, miR-195-5p and miR-16-5p levels were detected with RT-PCR. Then pcDNA3.1-CDKN2B-AS1, si-CDKN2B-AS1, miR-195-5p mimic, miR-195-5p inhibitor, miR-16-5p mimic and miR-16-5p inhibitor were transfected into HT29 cells, and proliferation and apoptosis of the cells were assessed. Dual-luciferase reporter gene assay was implemented to identify the sponging relationship between lncRNA CDKN2B-AS1 and miR-195-5p/miR-16-5p.ResultsCDKN2B-AS1 level was negatively correlated with levels of inflammatory cytokines, including TNF-α, IL-6 and sIL-2R, yet miR-16-5p and miR-195-5p levels were negatively correlated with the CDKN2B-AS1 level. The CDKN2B-AS1 combined with miR-16-5p and miR-195-5p also achieved an optimum efficacy in differentiating between light and medium UC, light and severe UC, as well as medium and heavy UC. Furthermore, pcDNA3.1-CDKN2B-AS1 depressed expressions of IFN-γ, IL-8, IL-1β and TNF-α in HT29 cells (P < 0.05), and strengthened proliferation of the cells (P < 0.05). CDKN2B-AS1 also sponged and regulated miR-16-5p and miR-195-5p in HT29 cells, and miR-16-5p and miR-195-5p could reverse the effect of CDKN2B-AS1 on inflammatory cytokine production, barrier function and apoptosis of HT29 cells (P < 0.05).ConclusionLncRNA CDKN2B-AS1 regulated inflammation of UC by sponging miR-195-5p and miR-16-5p, providing an alternative for diagnosis and treatment of UC.