荧光定量分析人皮肤瘢痕疙瘩组织中miRNA-34s的表达

背景：通过分析不同肿瘤组织与正常组织miRNA表达谱的差异,有可能筛选到特异性好、灵敏度高、可作为特定肿瘤分子标志物的miRNA。目的：采用实时荧光定量RT-PCR检测miRNA-34家族（miRNA-34s：miR-34a/b/c）在瘢痕疙瘩和正常皮肤组织中的差异表达,分析评价microRNA-34s在瘢痕疙瘩形成发展的作用及可能的机制影响。方法：收集手术切除瘢痕疙瘩组织（10例）和正常皮肤（2例）;TRIZOL法提取标本的总RNAs并进行质检,再采用 Ambion′s miRNA Isolation Kit 从总 RNAs 中分离 microRNA,采用实时荧光定量 RT-PCR 技术检测miRNA-34s（miR-34a/b/c）在瘢痕疙瘩组织和正常皮肤组织中的表达水平。结果与结论：miRNA-34s（miRNA-34a/b/c）在瘢痕疙瘩组织中均呈下调表达（P＜0.01）。表明该家族成员参与了瘢痕疙瘩的形成发展,下调表达的miRNA-34s可能导致了瘢痕疙瘩的肿瘤性生长。     

Elevated MicroRNA‐128 in Periodontitis Mitigates Tumor Necrosis Factor‐α Response via p38 Signaling Pathway in Macrophages

Background: Periodontitis is a chronic inflammatory disease resulting from an inflammatory response to subgingival plaque bacteria, including Porphyromonas gingivalis. MicroRNA (miRNA) is a current focus in regulating the inflammatory processes. In this study, the inflammatory miRNA expression in gingival tissues of patients with periodontitis and of healthy individuals is compared, and its role in regulating the inflammatory response is examined. Methods: Gingival tissues from patients with periodontitis and healthy individuals were collected for miRNA microarray. THP‐1 and CA9‐22 cells were challenged with P. gingivalis, and miRNA expression was determined by real‐time polymerase chain reaction. Target genes for miRNA were predicted using TargetScanHuman database, and miRNA gene expressions were reviewed using public databases. For the functional study, THP‐1 cells were transfected with a miRNA‐128 mimic, and target gene expression was compared with THP‐1 cells challenged with P. gingivalis. For the tolerance test, THP‐1 cells transfected with miRNA‐128 mimic were treated with phorbol 12‐myristate 13‐acetate (PMA) or paraformaldehyde (PFA)‐fixed Escherichia coli. Tumor necrosis factor (TNF)‐α production was determined by enzyme‐linked immunosorbent assay, and mitogen‐activated protein kinase (MAPK) protein phosphorylation was determined by Western blot. Results: Gingival tissues from patients with periodontitis showed increased expression of miRNA‐128, miRNA‐34a, and miRNA‐381 and decreased expression of miRNA‐15b, miRNA‐211, miRNA‐372, and miRNA‐656. THP‐1 cells and CA9‐22 cells challenged with P. gingivalis showed increased miRNA‐128 expression. Among the predicted miRNA‐128 target genes, several genes that are involved in MAPK signaling pathway showed similar gene expression pattern between P. gingivalis challenge and miRNA‐128 mimic transfection. In THP‐1 cells transfected with miRNA‐128 mimic, TNF‐α production was lower, and phosphorylation of p38 was inhibited when challenged with PMA or PFA‐fixed E. coli. Conclusion: miRNA‐128 may be involved in mitigating the inflammatory response induced by P. gingivalis in periodontitis.     

循环微小RNA对高龄急性心肌梗死的早期诊断价值

目的探讨循环微小RNA(microRNA,miR-19b、miR-223和miR-483-5p)在高龄急性心肌梗死(AMI)的早期诊断价值。方法连续入选因"胸痛或其等危症"就诊于我院且疑诊AMI的年龄≥85岁高龄患者70例,按性别和年龄1:1配对原则,分为AMI组35例和非AMI组35例。收集患者入院时首份血标本,检测循环microRNA表达水平。比较2组循环miR-19b、miR-223和miR-483-5p表达差异,绘制ROC曲线评价循环microRNA对高龄AMI的早期诊断价值。结果与非AMI组比较,AMI组miR-19b、miR-223和miR-483-5p表达显著上调(3.42±0.11 vs 1.00±0.00,P<0.01;2.25±0.12 vs 1.00±0.00,P<0.05;2.51±0.12 vs 1.00±0.00,P<0.01)。ROC曲线分析显示,miR-19b、miR-483-5p和血清肌钙蛋白I诊断高龄AMI的曲线下面积(AUC)分别为0.73(95%CI:0.64~0.83)、0.71(95%CI:0.61~0.80)和0.64(95%CI:0.53~0.74),miR-19b、miR-483-5p诊断AMI价值优于血清肌钙蛋白I;miR-19b、miR-223和miR-483-5p三者联合诊断高龄AMI的AUC为0.82(95%CI:0.74~0.90)。结论循环miR-19b和miR-483-5p表达上调有助AMI早期诊断,有望成为高龄AMI的新型诊断标志物。     

miR-197 Expression in Peripheral Blood Mononuclear Cells from Hepatitis B Virus-Infected Patients

Background/Aims

This study aimed to investigate the microRNA (miRNA) expression profiles in peripheral blood mononuclear cell (PBMC) of hepatitis B virus (HBV)-infected patients with different clinical manifestations and to analyze the function of miR-197.

Methods

PBMC miRNA expression profiles in 51 healthy controls, 70 chronic asymptomatic carriers, 107 chronic hepatitis B patients, and 76 HBV-related acute on chronic liver failure patients were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR). miR-197 mimic and inhibitor were transfected in THP-1 cells. qRT-PCR and ELISA for interleukin (IL)-18 mRNA and protein levels were performed, respectively.

Results

The microarray analysis revealed that 17 PBMC miRNA expression profiles (12 miRNAs downregulated and five miRNAs upregulated) differed significantly in HBV-induced liver disease patients presenting with various symptoms. The qRT-PCR results suggested that the PBMC miR-197 levels regularly decreased as the severity of liver disease symptoms became aggravated. IL-18, a key regulator in inflammation and immunity, was inversely correlated with miR-197 levels. Bioinformatic analysis indicated that IL-18 was a target of miR-197. Exogenous expression of miR-197 could significantly repress IL-18 expression at both the mRNA and protein levels in THP-1 cells.

Conclusions

We concluded that multiple PBMC miRNAs had differential expression profiles during HBV infection and that miR-197 may play an important role in the reactivation of liver inflammation by targeting IL-18.     

MicroRNAs途径在RAW264.7细胞中作用的初步研究

目的通过检测microRNAs生成的关键酶Dicer在RAW264.7细胞不同功能状态下的表达差异研究microRNAs途径在其中产生的作用。方法采用QRT-PCR、流式细胞术等对RAW264.7细胞在不同功能分化状态下的Dicer酶进行检测。结果GMCSF与MCSF分别刺激RAW264.7细胞诱导其增殖分化后,Dicer酶的mRNA水平显著增高,但随刺激时间呈现不同趋势(P<0.01);在诱导凋亡死亡状态下,RAW264.7中Dicer酶的表达下降30%(P<0.01);LPS与IL-4分别诱导RAW264.7细胞分化为M1和M2型,Dicer的表达与M1、M2型相关炎性因子的表达有相关性。结论 MicroRNAs调控途径参与并影响RAW264.7不同功能状态。     

微小RNA在膀胱肿瘤组织的表达及其临床意义

目的研究微小RNA（MicroRNAs/miRNAs）microRNAs在人类膀胱肿瘤组织中的表达及其作用。方法取安徽医科大学附属省立医院2009年手术切除的45例膀胱肿瘤组织标本及其肿瘤旁正常组织标本。通过microRNAs微阵点杂交法检测肿瘤组织及其癌旁正常组织的手术标本中microRNAs的表达情况,分析癌组织及其癌旁正常膀胱组织进行定性判断（癌和非癌）。结果人类膀胱肿瘤组织与正常膀胱组织相比,miR-223、miR-26b、miR-221、miR-103-1、miR-185、miR-23b、miR-203、miR-17-5p、miR-23a和miR-205表达显著上调（P〈0.005）。结论 miRNAs与膀胱肿瘤的发生发展有密切的联系,有可能成为一个有效的肿瘤标志物以进行膀胱肿瘤的早期预测和诊断。     

微小RNAs在骨与软骨组织中的调节作用

目的对微小RNAs(microRNAs,miRNAs)在骨与软骨组织中的调控作用和作用机制作一综述。方法广泛查阅近年来有关miRNAs在骨与软骨组织的调控作用及作用机制的文献,进行总结与分析。结果 miRNAs是近年来骨关节疾病研究的热点,越来越多研究显示其在骨与软骨组织形成和代谢过程中,对于细胞分化、细胞基质分泌等方面发挥重要的调控作用,但确切机制尚不清楚。结论 通过对miRNAs在骨与软骨组织中的调控作用及作用机制的研究,可能为了解退行性骨关节疾病开辟新的领域。     

microRNAs与晶状体疾病的相关性研究进展

microRNAs是一类长度约22个核苷酸的内源性非编码小RNA分子,广泛存在于真核细胞中,在细胞增殖、凋亡、分化、肿瘤发生以及机体代谢等一系列生命过程中扮演了重要角色,是近年来基因调控研究的热点。许多microRNAs广泛表达于眼部多种组织,并具有组织特异性和时序特异性,可能与眼部组织的生长、发育、功能调节、疾病发生等方面密切相关。研究microRNAs在晶状体发育和病理生理过程中的作用为进一步阐明晶状体疾病的发生机制、探索相关疾病的诊断和治疗方法提供了一种新思路。