MiRNA

miRNA
  • 文章类型: Journal Article
    背景:雄激素受体(AR)水平升高的前列腺癌(PCa)患者与较高的转移发生率相关。与器官限制的肿瘤相比,转移性前列腺肿瘤中AR及其靶基因前列腺特异性抗原(PSA)的蛋白质表达升高。雄激素治疗或AR升高促进PCa在细胞培养和小鼠模型中的转移。然而,在雄激素耗尽的条件下,AR抑制PCa细胞的细胞移动性和侵袭性。PCa患者的雄激素剥夺治疗与较高的癌症转移风险相关。因此,我们研究了AR和miRNA对PCa转移的双重作用。
    方法:PC-3AR(PC-3细胞再表达AR)和LNCaP细胞作为PCa细胞模型。Transwell迁移和侵袭分析,伤口愈合试验,斑马鱼异种移植试验,采用斑马鱼血管出口试验研究AR和雄激素在PCa转移中的作用。微西方阵列,应用免疫共沉淀和免疫荧光来剖析下面的分子机制。miRNA阵列,miRNA抑制剂或质粒,和染色质免疫沉淀法用于研究miRNAs在PCa转移中的作用。
    结果:在没有雄激素的情况下,AR抑制PCa细胞的迁移和侵袭。当雄激素存在时,AR在体外和斑马鱼异种移植模型中都能刺激PCa细胞的迁移和侵袭。雄激素增加磷酸-ARSer81和Yes相关蛋白1(YAP),PCa细胞中磷酸化YAPSer217降低,上皮间质转化(EMT)蛋白改变。Co-IP分析表明,雄激素增强了细胞核中YAP和AR之间的相互作用。敲除YAP或用YAP抑制剂治疗可消除雄激素诱导的PCa细胞迁移和侵袭,而YAP的过表达表现出相反的效果。miRNA阵列显示,雄激素在PC-3AR细胞中降低hsa-miR-5001-5p,但在PC-3细胞中增加hsa-miR-203a和hsa-miR-210-3p。用靶向hsa-miR-203a/hsa-miR-210-3p的抑制剂治疗,或hsa-miR-5001-5p的过表达降低了YAP的表达,并抑制了雄激素诱导的PCa细胞的迁移和侵袭。染色质免疫沉淀(ChIP)测定表明,在雄激素存在下,AR与has-miR-210-3p的启动子区域结合。
    结论:我们的观察表明miRNA203a/210-3p/5001-5p调节雄激素/AR/YAP诱导的PCa转移。
    BACKGROUND: Prostate cancer (PCa) patients with elevated level of androgen receptor (AR) correlate with higher metastatic incidence. Protein expression of AR and its target gene prostate-specific antigen (PSA) are elevated in metastatic prostate tumors as compared to organ-confined tumors. Androgen treatment or elevation of AR promotes metastasis of PCa in cell culture and murine model. However, under androgen depleted condition, AR suppressed cell mobility and invasiveness of PCa cells. Androgen deprivation therapy in PCa patients is associated with higher risk of cancer metastasis. We therefore investigated the dual roles of AR and miRNAs on PCa metastasis.
    METHODS: The PC-3AR (PC-3 cells re-expressing AR) and LNCaP cells were used as PCa cell model. Transwell migration and invasion assay, wound-healing assay, zebrafish xenotransplantation assay, and zebrafish vascular exit assay were used to investigate the role of AR and androgen on PCa metastasis. Micro-Western Array, co-immunoprecipitation and Immunofluorescence were applied to dissect the molecular mechanism lying underneath. The miRNA array, miRNA inhibitors or plasmid, and chromatin immunoprecipitation assay were used to study the role of miRNAs on PCa metastasis.
    RESULTS: In the absence of androgen, AR repressed the migration and invasion of PCa cells. When androgen was present, AR stimulated the migration and invasion of PCa cells both in vitro and in zebrafish xenotransplantation model. Androgen increased phospho-AR Ser81 and yes-associated protein 1 (YAP), decreased phospho-YAP Ser217, and altered epithelial-mesenchymal transition (EMT) proteins in PCa cells. Co-IP assay demonstrated that androgen augmented the interaction between YAP and AR in nucleus. Knockdown of YAP or treatment with YAP inhibitor abolished the androgen-induced migration and invasion of PCa cells, while overexpression of YAP showed opposite effects. The miRNA array revealed that androgen decreased hsa-miR-5001-5p but increased hsa-miR-203a and hsa-miR-210-3p in PC-3AR cells but not PC-3 cells. Treatment with inhibitors targeting hsa-miR-203a/hsa-miR-210-3p, or overexpression of hsa-miR-5001-5p decreased YAP expression as well as suppressed the androgen-induced migration and invasion of PCa cells. Chromatin immunoprecipitation (ChIP) assay demonstrated that AR binds with promoter region of has-miR-210-3p in the presence of androgen.
    CONCLUSIONS: Our observations indicated that miRNAs 203a/210-3p/5001-5p regulate the androgen/AR/YAP-induced PCa metastasis.
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  • 文章类型: Journal Article
    神经发育障碍(NDD)代表了在新生儿或儿童早期发作的一大组疾病;NDD包括智力障碍(ID),自闭症谱系障碍(ASD),注意缺陷多动障碍(ADHD),癫痫发作,各种运动障碍和肌肉张力异常。在这些疾病的许多潜在孟德尔遗传原因中,编码涉及基因表达途径各个方面的蛋白质的基因,从转录,拼接,翻译为最终的RNA衰变,特征相当突出。在这里,我们专注于RNA解旋酶的两个大家族(DEAD-和DExH-box解旋酶)。最近已显示几种解旋酶的编码基因中的遗传变异与NDD相关。我们解决了解旋酶的遗传限制,已发现的病理变异类型,并讨论了受影响的解旋酶蛋白所涉及的生物学途径。
    Neurodevelopmental disorders (NDDs) represent a large group of disorders with an onset in the neonatal or early childhood period; NDDs include intellectual disability (ID), autism spectrum disorders (ASD), attention deficit hyperactivity disorders (ADHD), seizures, various motor disabilities and abnormal muscle tone. Among the many underlying Mendelian genetic causes for these conditions, genes coding for proteins involved in all aspects of the gene expression pathway, ranging from transcription, splicing, translation to the eventual RNA decay, feature rather prominently. Here we focus on two large families of RNA helicases (DEAD- and DExH-box helicases). Genetic variants in the coding genes for several helicases have recently been shown to be associated with NDD. We address genetic constraints for helicases, types of pathological variants which have been discovered and discuss the biological pathways in which the affected helicase proteins are involved.
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  • 文章类型: Case Reports
    儿童多系统炎症综合征(MIS-C)是一种与COVID-19密切相关的紧迫儿科炎症,自大流行以来,COVID-19引起了广泛关注。像川崎病一样,这种情况的特征是过度活跃的免疫反应,导致包括发热在内的症状,心脏和肾脏并发症。为了阐明MIS-C的发病机制并确定潜在的生物标志物,我们对特定的细胞因子(IL-6,IL-1β,IL-6R,IL-10和TNF-α)和microRNA(miRNA)表达谱在不同的间隔(3至20天)在严重受影响的MIS-C患者的外周血样品中。我们的调查显示,IL-6,IL-1β的循环水平逐渐下降,静脉注射免疫球蛋白(IVIG)治疗后的IL-10和TNF-α。值得注意的是,IL-6表现出从74.30至1.49yg的显著减少。/mL,而IL-6R水平在整个病程中始终保持稳定。此外,我们观察到hsa-miR-596和hsa-miR-224-5p的表达与上述细胞因子之间呈负相关。我们的发现强调了血液细胞因子和miRNA浓度与MIS-C的严重程度之间的紧密关联。这些见解增强了我们对MIS-C发病机制的遗传调控机制的理解,通过miRNA分析为早期生物标志物检测和治疗监测提供了潜在的途径。
    Multisystem inflammatory syndrome in children (MIS-C) is an imperative pediatric inflammatory condition closely linked to COVID-19, which garners substantial attention since the onset of the pandemic. Like Kawasaki illness, this condition is characterized by an overactive immune response, leading to symptoms including pyrexia, cardiac and renal complications. To elucidate the pathogenesis of MIS-C and identify potential biomarkers, we conducted an extensive examination of specific cytokines (IL-6, IL-1β, IL-6R, IL-10, and TNF-α) and microRNA (miRNA) expression profiles at various intervals (ranging from 3 to 20 days) in the peripheral blood sample of a severely affected MIS-C patient. Our investigation revealed a gradual decline in circulating levels of IL-6, IL-1β, IL-10, and TNF-α following intravenous immune globulin (IVIG) therapy. Notably, IL-6 exhibited a significant reduction from 74.30 to 1.49 pg./mL, while IL-6R levels remained consistently stable throughout the disease course. Furthermore, we observed an inverse correlation between the expression of hsa-miR-596 and hsa-miR-224-5p and the aforementioned cytokines. Our findings underscore a robust association between blood cytokine and miRNA concentrations and the severity of MIS-C. These insights enhance our understanding of the genetic regulatory mechanisms implicated in MIS-C pathogenesis, offering potential avenues for early biomarker detection and therapy monitoring through miRNA analysis.
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  • 文章类型: Journal Article
    远处转移是乳腺癌患者死亡的主要原因。上皮-间质转化(EMT)有助于乳腺癌的转移。G蛋白信号调节因子(RGS)蛋白调节各种癌症的转移。这项研究确定了RGS10在乳腺癌EMT和转移中的新作用。RGS10蛋白水平在乳腺癌组织中显著低于正常乳腺组织,RGS10蛋白的缺乏预示着乳腺癌患者的预后较差。高侵袭性细胞系MDA-MB-231中的RGS10蛋白水平低于低侵袭性细胞系MDA-MB-231,侵袭性较小的细胞系MCF7和SKBR3。在SKBR3细胞中沉默RGS10可增强EMT并引起SKBR3细胞迁移和侵袭。RGS10抑制乳腺癌EMT和转移的能力取决于脂质运载蛋白2和MIR539-5p。这些发现将RGS10确定为肿瘤抑制因子,预后生物标志物,和潜在的乳腺癌治疗靶点。
    Distant metastasis is the major cause of death in patients with breast cancer. Epithelial-mesenchymal transition (EMT) contributes to breast cancer metastasis. Regulator of G protein-signaling (RGS) proteins modulates metastasis in various cancers. This study identified a novel role for RGS10 in EMT and metastasis in breast cancer. RGS10 protein levels were significantly lower in breast cancer tissues compared to normal breast tissues, and deficiency in RGS10 protein predicted a worse prognosis in patients with breast cancer. RGS10 protein levels were lower in the highly aggressive cell line MDA-MB-231 than in the poorly aggressive, less invasive cell lines MCF7 and SKBR3. Silencing RGS10 in SKBR3 cells enhanced EMT and caused SKBR3 cell migration and invasion. The ability of RGS10 to suppress EMT and metastasis in breast cancer was dependent on lipocalin-2 and MIR539-5p. These findings identify RGS10 as a tumor suppressor, prognostic biomarker, and potential therapeutic target for breast cancer.
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  • 文章类型: Journal Article
    磷酸酶和Tensin同源基因(PTEN)在调节多种细胞过程中至关重要,包括增长,分化,扩散,和细胞存活,主要通过调节PI3K/AKT/mTOR通路。PTEN基因表达的改变与表观遗传机制有关。特别是小的非编码RNA的调控,如miRNA。已显示控制PTEN的miRNA表达水平的修饰导致其表达不足。这种减压,反过来,影响PI3K/AKT/mTOR通路,从而影响增殖和凋亡等关键机制,在前列腺癌(PCa)的发生和发展中起着重要作用。因此,我们旨在系统回顾有关miRNA在PCa中介导的PTEN调控的现有信息。
    搜索电子数据库以确定通过PCamiRNA评估PTEN调节的研究,搜索包括microRNA的组合,PTEN和前列腺肿瘤。所包括的文章的质量评估是使用SYRCLE和CASP工具的改编版本进行的。
    我们包括39篇文章,这些文章测量了PCa中miRNA的相对基因表达及其与PTEN调控的关系。据报道,共有42种miRNA通过PTEN失调参与PCa的发展和进展(34种miRNA上调,8种miRNA下调)。16个miRNAs被证明是导致癌症发生的遗传相互作用的主要调节因子。作为与PTEN下调相关的PCa中报道最多的miR-21。我们显示PTEN的沉默可以通过miR-200b和DNMT1之间的环或通过microRNA直接靶向PTEN来促进,导致PI3K/AKT/mTOR的组成型激活和与中间基因的相互作用支持凋亡抑制,扩散,入侵,和PCa的转移。
    根据我们的评论,PTEN的失调主要由miR-21、-20a、-20b,-93,-106a,和-106b上调在PCa的发展中起着核心作用,可能是诊断的潜在生物标志物,预后,和治疗目标。
    UNASSIGNED: The Phosphatase and Tensin Homolog gene (PTEN) is pivotal in regulating diverse cellular processes, including growth, differentiation, proliferation, and cell survival, mainly by modulating the PI3K/AKT/mTOR pathway. Alterations in the expression of the PTEN gene have been associated with epigenetic mechanisms, particularly the regulation by small non-coding RNAs, such as miRNAs. Modifications in the expression levels of miRNAs that control PTEN have been shown to lead to its underexpression. This underexpression, in turn, impacts the PI3K/AKT/mTOR pathway, thereby influencing crucial mechanisms like proliferation and apoptosis, playing an important role in the initiation and progression of prostate cancer (PCa). Thus, we aimed to systematically reviewed available information concerning the regulation of PTEN mediated by miRNA in PCa.
    UNASSIGNED: Electronic databases were searched to identify studies assessing PTEN regulation via PCa miRNAs, the search included combination of the words microRNAs, PTEN and prostatic neoplasms. The quality assessment of the articles included was carried out using an adapted version of SYRCLE and CASP tool.
    UNASSIGNED: We included 39 articles that measured the relative gene expression of miRNAs in PCa and their relationship with PTEN regulation. A total of 42 miRNAs were reported involved in the development and progression of PCa via PTEN dysregulation (34 miRNAs up-regulated and eight miRNAs down-regulated). Sixteen miRNAs were shown as the principal regulators for genetic interactions leading to carcinogenesis, being the miR-21 the most reported in PCa associated with PTEN down-regulation. We showed the silencing of PTEN could be promoted by a loop between miR-200b and DNMT1 or by direct targeting of PTEN by microRNAs, leading to the constitutive activation of PI3K/AKT/mTOR and interactions with intermediary genes support apoptosis inhibition, proliferation, invasion, and metastasis in PCa.
    UNASSIGNED: According to our review, dysregulation of PTEN mediated mainly by miR-21, -20a, -20b, -93, -106a, and -106b up-regulation has a central role in PCa development and could be potential biomarkers for diagnosis, prognostic, and therapeutic targets.
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  • 文章类型: Journal Article
    一些研究表明,EB病毒(EBV)感染通过感染B淋巴细胞增加了患阿尔茨海默病(AD)的可能性。当前研究的目的是评估EBV感染与AD之间的可能关联。
    通过使用GEO平台的GEO2R工具,利用微阵列数据集GSE49628、GSE126379、GSE122063和GSE132903来提取DEG。STRING工具用于确定DEG之间的相互作用,和Cytoscape用于可视化结果。发现的DEG进行了功能分析,包括通路和GO,使用DAVID2021和ClueGo/CluePedia。通过使用MNC,MCC,学位,和cytoHubba的放射性,我们确定了七个常见的关键基因。通过GeneMANIA网络工具进行基因共表达分析。此外,通过GTEx软件对关键基因进行表达分析,已经在人脑的各个区域被发现。通过miRNetv2.0工具进行miRNA-基因相互作用。Enrichr平台上的DsigDB用于提取与关键基因相关的治疗药物。
    在具有|log2FC|≥0.5和p值<0.05的数据集的GEO2R分析中,鉴定了8386、10,434、7408和759个基因。通过组合不同数据集的提取基因,总共鉴定了141个常见的DEG。在PPI分析期间共发现141个节点和207条边。具有实质性改变的DEGGO分析揭示了它们与分子功能和生物过程有关,比如神经元死亡的正向调节,线粒体的自噬调节,细胞对胰岛素刺激的反应,钙信号调节,细胞器沿着微管运输,蛋白激酶活性,和磷酸丝氨酸结合。京都基因百科全书和基因组分析发现了神经变性途径中DEGs之间的相关性:多种疾病,细胞周期,和cGMP-PKG信号通路。最后,是啊,YWHAG,YWHAB,YWHAZ,MAP2K1,PPP2CA,和TUBB基因被鉴定为与EBV和AD密切相关。三个miRNA,即,hsa-mir-15a-5p,hsa-let-7a-5p,还有hsa-mir-7-5p,被鉴定为调节大多数与EBV和AD相关的hub基因。进一步预测了前10名重要治疗药物。
    我们发现了AD的新生物标志物和治疗靶点,以及EBV感染可能首次参与AD易感性的可能生物学机制。
    UNASSIGNED: Several studies have revealed that Epstein-Barr virus (EBV) infection raised the likelihood of developing Alzheimer\'s disease (AD) via infecting B lymphocytes. The purpose of the current investigation was to assess the possible association between EBV infection and AD.
    UNASSIGNED: The microarray datasets GSE49628, GSE126379, GSE122063, and GSE132903 were utilized to extract DEGs by using the GEO2R tool of the GEO platform. The STRING tool was used to determine the interaction between the DEGs, and Cytoscape was used to visualize the results. The DEGs that were found underwent function analysis, including pathway and GO, using the DAVID 2021 and ClueGo/CluePedia. By using MNC, MCC, Degree, and Radiality of cytoHubba, we identified seven common key genes. Gene co-expression analysis was performed through the GeneMANIA web tool. Furthermore, expression analysis of key genes was performed through GTEx software, which have been identified in various human brain regions. The miRNA-gene interaction was performed through the miRNet v 2.0 tool. DsigDB on the Enrichr platform was utilized to extract therapeutic drugs connected to key genes.
    UNASSIGNED: In GEO2R analysis of datasets with |log2FC|≥ 0.5 and p-value <0.05, 8386, 10,434, 7408, and 759 genes were identified. A total of 141 common DEGs were identified by combining the extracted genes of different datasets. A total of 141 nodes and 207 edges were found during the PPI analysis. The DEG GO analysis with substantial alterations disclosed that they are associated to molecular functions and biological processes, such as positive regulation of neuron death, autophagy regulation of mitochondrion, response of cell to insulin stimulus, calcium signaling regulation, organelle transport along microtubules, protein kinase activity, and phosphoserine binding. Kyoto Encyclopedia of Genes and Genomes analysis discovered the correlation between the DEGs in pathways of neurodegeneration: multiple disease, cell cycle, and cGMP-PKG signaling pathway. Finally, YWHAH, YWHAG, YWHAB, YWHAZ, MAP2K1, PPP2CA, and TUBB genes were identified that are strongly linked to EBV and AD. Three miRNAs, i.e., hsa-mir-15a-5p, hsa-let-7a-5p, and hsa-mir-7-5p, were identified to regulate most of hub genes that are associated with EBV and AD. Further top 10 significant therapeutic drugs were predicted.
    UNASSIGNED: We have discovered new biomarkers and therapeutic targets for AD, as well as the possible biological mechanisms whereby infection with EBV may be involved in AD susceptibility for the first time.
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  • 文章类型: Journal Article
    最近对阿尔茨海默病(AD)涉及的调控网络的研究表明,长链非编码RNA(lncRNAs)是关键的调控参与者,尽管对机制的理解很差。分析来自死后AD脑海马的RNA-seq数据中的差异基因表达,我们根据k-mer谱将一系列AD失调的lncRNA转录本分类为功能相似的群落。使用基于机器学习的算法,他们的亚细胞定位被映射。我们通过AD失调的miRNA进一步探索了每个社区的功能相关性,RNA结合蛋白(RBP)相互作用物,和途径富集分析。对来自每个社区的miRNA-lncRNA和RBP-lncRNA网络的进一步调查显示,miRNA,和每个簇的lncRNAs。实验验证社区产生ELAVL4和miR-16-5p作为主要的RBP和miRNA,分别。五个lncRNA作为来自RBP/miRNA-lncRNA网络的顶级候选物出现。对这些网络的进一步分析揭示了多个调节三联体的存在,其中RBP-lncRNA相互作用可以通过增强的miRNA-lncRNA相互作用来增强。我们的结果通过其相互作用的伙伴促进了对lncRNA介导的AD调节机制的理解,并证明了这些功能分离但重叠的调节网络如何从整体上调节疾病。
    Recent studies on the regulatory networks implicated in Alzheimer\'s disease (AD) evince long non-coding RNAs (lncRNAs) as crucial regulatory players, albeit a poor understanding of the mechanism. Analyzing differential gene expression in the RNA-seq data from the post-mortem AD brain hippocampus, we categorized a list of AD-dysregulated lncRNA transcripts into functionally similar communities based on their k-mer profiles. Using machine-learning-based algorithms, their subcellular localizations were mapped. We further explored the functional relevance of each community through AD-dysregulated miRNA, RNA-binding protein (RBP) interactors, and pathway enrichment analyses. Further investigation of the miRNA-lncRNA and RBP-lncRNA networks from each community revealed the top RBPs, miRNAs, and lncRNAs for each cluster. The experimental validation community yielded ELAVL4 and miR-16-5p as the predominant RBP and miRNA, respectively. Five lncRNAs emerged as the top-ranking candidates from the RBP/miRNA-lncRNA networks. Further analyses of these networks revealed the presence of multiple regulatory triads where the RBP-lncRNA interactions could be augmented by the enhanced miRNA-lncRNA interactions. Our results advance the understanding of the mechanism of lncRNA-mediated AD regulation through their interacting partners and demonstrate how these functionally segregated but overlapping regulatory networks can modulate the disease holistically.
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  • 文章类型: Journal Article
    腮腺炎是由腮腺炎病毒(MuV)引起的儿童常见感染。无菌性脑膜炎和脑炎是腮腺炎以及睾丸炎和卵巢炎的常见症状,可在男性和女性中出现,分别。我们已经使用计算工具:RNA22,miRanda和psRNATarget来预测microRNA-mRNA结合位点,以发现推定的microRNA在宿主对腮腺炎病毒感染的反应中起作用。我们的计算研究表明hsa-mir-3155a最有可能参与腮腺炎感染。通过预测hsa-mir-3155a与MuV基因组的结合位点进一步研究了这一点。此外,使用MC-Fold和MC-Sym进行结构预测,分别用于预测miRNA和mRNA的3D结构。通过分子对接模拟研究证实了miRNA-mRNA之间的相互作用谱。一起来看,已发现推定的miRNA(hsa_miR_6794_5p)最有可能参与MuV感染中转录活性的调节。
    Mumps is a common childhood infection caused by the mumps virus (MuV). Aseptic meningitis and encephalitis are usual symptoms of mumps together with orchitis and oophoritis that can arise in males and females, respectively. We have used computational tools: RNA22, miRanda and psRNATarget to predict the microRNA-mRNA binding sites to find the putative microRNAs playing role in the host response to mumps virus infection. Our computational studies indicate that hsa-mir-3155a is most likely involved in mumps infection. This was further investigated by the prediction of binding sites of hsa-mir-3155a to the MuV genome. Additionally, structure prediction using MC-Fold and MC-Sym, respectively has been applied to predict the 3D structures of miRNA and mRNA. The miRNA-mRNA interaction profile between has been confirmed through molecular docking simulation studies. Taken together, the putative miRNA (hsa_miR_6794_5p) has been found to be most likely involved in the regulation of transcriptional activity in the MuV infection.
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  • 文章类型: Journal Article
    肌萎缩侧索硬化症(ALS)是一种致命的神经退行性疾病,其特征是运动神经元逐渐耗尽。RNA结合基序蛋白5(RBM5),一种大量表达的RNA结合蛋白,在细胞死亡过程中起着至关重要的作用。然而,关于RBM5在ALS发病机制中的作用知之甚少。这里,我们发现,由于miR-141-5p的减少,RBM5在ALShSOD1G93A-NSC34细胞模型和hSOD1G93A小鼠中上调.RBM5的上调通过抑制Rac1介导的神经保护作用增加了运动神经元的凋亡。相比之下,RBM5的基因敲除通过激活Rac1信号从hSOD1G93A诱导的变性中拯救了运动神经元。Rac1抑制剂显著抑制RBM5敲低的神经保护作用,NSC23766。这些发现表明RBM5可能通过激活Rac1信号传导作为ALS的治疗靶标。
    Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disorder distinguished by gradual depletion of motor neurons. RNA binding motif protein 5 (RBM5), an abundantly expressed RNA-binding protein, plays a critical role in the process of cellular death. However, little is known about the role of RBM5 in the pathogenesis of ALS. Here, we found that RBM5 was upregulated in ALS hSOD1G93A-NSC34 cell models and hSOD1G93A mice due to a reduction of miR-141-5p. The upregulation of RBM5 increased the apoptosis of motor neurons by inhibiting Rac1-mediated neuroprotection. In contrast, genetic knockdown of RBM5 rescued motor neurons from hSOD1G93A-induced degeneration by activating Rac1 signaling. The neuroprotective effect of RBM5-knockdown was significantly inhibited by the Rac1 inhibitor, NSC23766. These findings suggest that RBM5 could potentially serve as a therapeutic target in ALS by activating the Rac1 signalling.
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  • 文章类型: Journal Article
    精液冷冻保存是用于牛人工授精和体外受精的精液生产中最流行的做法。精浆含有细胞外囊泡(spEV),可在卵母细胞繁殖过程中调节精子活力和功能。冻融精液剂量中的spEV研究可能会产生预测公牛生育能力的新指标,但是精液补充剂的存在可能会阻碍spEV的分子谱分析。这项研究的目的是提供在冷冻保存过程之前和之后从精浆中分离出的EV的广泛表征,并添加了商业动物无蛋白精液补充剂,以了解源自补充剂的EV在阻碍使用中的潜在影响SpEV衍生的生物标志物用于评估公牛生育力。
    从精浆中分离出EV(有或没有延伸剂),从没有精子的冷冻保存的稻草中,并使用两种不同的方法从扩展器,超速离心(UC)和尺寸排阻色谱(SEC),并以其结构和组成为特征。
    电动汽车的物理表征表明,大小和颗粒数与分离方法有关。spEV较大但较少(UC:168.9nm,n=2.68×109;SEC:197.0nm,n=6.42×109)与扩展器电动汽车(UC:129.0nm,n=2.68×1011;SEC:161.8nm,n=6.47×1011)。Western印迹分析(WB)证实spEVS中存在典型的EV标志物:膜结合CD9(25kDa)和腔内标志物Alix(96kDa)和TSG101(48KDa)。尽管透射电子显微镜证实了所有制剂中都存在脂质双层结构,当使用单分子阵列(SiMoa)时,在从延伸剂分离的囊泡中没有检测到特异性EV标记。在至少一种制剂中鉴定了总共724个Bos金牛miRNA。在来自延伸体的EV中鉴定的miRNA的百分比(总读段的0.05%-0.49%)低于含有spEV的制备物(总读段的10.56%-63.69%)。Edge-R通过两种方法鉴定了从延伸剂分离的EV之间的总共111个DE-miRNA。其中,11DE-miRNAs(bta-miR-11980,bta-miR-11987,bta-miR-12057,bta-miR-1246,bta-miR-125b,bta-miR-181b,bta-miR-2340,bta-miR-2358,bta-miR-2478,bta-miR-2898和bta-miR-345-3p)在从具有延伸剂的精浆制剂中分离的EV中也很丰富。
    这项研究清楚地表明,延伸剂的存在并不能阻止冷冻保存的精液中spEV的表征。然而,spEV的分子谱分析可能受到所用的分离方法和来自延伸剂的一些miRNA的存在的影响。因此,在这样的研究中,建议同时表征spEV和从延伸剂分离的囊泡。
    UNASSIGNED: Semen cryopreservation is the most popular practice for semen production for artificial insemination and in vitro fertilization in cattle. The Seminal plasma contains extracellular vesicles (spEVs) which modulate sperm viability and function during oocyte fecundation. The study of spEVs in frozen-thawed semen doses may yield novel indicators for predicting bull fertility, but the presence of the semen extender may hinder molecular profiling of spEVs. The aim of this study was to provide extensive characterization of EVs isolated from seminal plasma before and after the cryopreservation process and the addition of a commercial animal protein-free semen extender to understand the potential influence of EVs originating from the extender in hindering the use of spEVs derived biomarkers for assessment of bull fertility.
    UNASSIGNED: EVs were isolated from the seminal plasma (with or without the extender), from the cryopreserved straw devoid of spermatozoa, and from the extender using two different methods, ultracentrifugation (UC) and size exclusion chromatography (SEC), and characterized for their structure and composition.
    UNASSIGNED: Physical characterization of EVs showed that size and particle numbers were related to the method of isolation. spEVs were larger but less abundant (UC: 168.9 nm, n = 2.68 × 109; SEC: 197.0 nm, n = 6.42 × 109) compared to extender EVs (UC: 129.0 nm, n = 2.68 × 1011; SEC: 161.8 nm, n = 6.47 × 1011). Western blotting analysis (WB) confirmed the presence of typical EV markers in spEVS: the membrane bound CD9 (25 kDa) and the luminal markers Alix (96 kDa) and TSG101 (48 KDa). Although Transmission Electron Microscopy confirmed the presence of a lipid bilayer structure in all preparations, no specific EV markers were detected in the vesicles isolated from extender when the Single Molecule Array (SiMoa) was used. A total of 724 Bos taurus miRNAs were identified in at least one preparation. The percentage of miRNAs identified in EVs from the extender (0.05%-0.49% of the total reads) was lower than in the preparation containing spEVs (10.56%-63.69% of the total reads). Edge-R identified a total of 111 DE-miRNAs between EVs isolated from the extender by two methods. Among them, 11 DE-miRNAs (bta-miR-11980, bta-miR-11987, bta-miR-12057, bta-miR-1246, bta-miR-125b, bta-miR-181b, bta-miR-2340, bta-miR-2358, bta-miR-2478, bta-miR-2898, and bta-miR-345-3p) were also abundant in EVs isolated from seminal plasma preparations with extender.
    UNASSIGNED: This study clearly demonstrates that the presence of the extender does not prevent the characterization of spEVs in cryopreserved semen. However, the molecular profiling of spEVs can be influenced by the isolation method used and by the presence of some miRNAs from the extender. Therefore, in such studies, it is advisable to characterize both spEVs and the vesicles isolated from the extender.
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