外泌体miR-145和miR-21在非小细胞肺癌中的研究进展

非小细胞肺癌(non-small cell lung cancer,NSCLC)是全球发病率最高的恶性肿瘤之一,大多数患者在确诊时已是中晚期,预后极差。外泌体是细胞主动分泌的直径为40～100 nm的膜小体,包含丰富的蛋白质、microRNAs(miRNAs)等物质,参与细胞间的物质交换和信息交流。非小细胞肺癌源性的外泌体miRNA参与调节肿瘤细胞的发生发展、侵袭及转移等过程,在肿瘤细胞的生理、病理过程中扮演了重要角色。本文将系统阐述外泌体的生物学特性、生物学功能以及外泌体miR-145和miR-21在调控非小细胞肺癌进展、早期诊断、治疗靶点筛选及预后中的作用,为外泌体miRNA基础研究与临床应用提供参考。     

外泌体miRNA生物标志物在肝癌中的研究进展*

外泌体广泛存在于多种体液中,携带有大量活性物质,如mRNA、miRNA、蛋白和脂质等。其中的miRNA是一类短非编码RNA,在转录后水平调节基因的表达,广泛参与个体生长发育等各生命活动。外泌体miRNA有多种生物学功能,在肿瘤的发生发展、侵袭转移、机体耐药及免疫调控等多方面发挥着重要作用。目前的研究表明,无论是作为肿瘤早筛早诊和预后评估标志物还是用于肿瘤治疗,外泌体miRNA都有很好的应用前景。本文就近年来外泌体miRNA在肝癌中的研究进展和临床应用进行综述。     

胃癌相关miRNA表达的表观遗传学调控机制

胃癌是人类最常见的肿瘤之一,其发病机制尚不完全清楚.微小RNA(microRNA,miRNA)是一组最近发现的长度为22个核苷酸左右的非编码RNA,具有负性调控基因表达的功能.本文对miRNA在胃癌发生中的作用及其表达调控机制进行综述.不断有文献显示,miRNA在多种肿瘤(包括胃癌)的发生过程中发挥着重要作用.作者和其他研究人员发现,miRNA的表达异常(如:miR-421和miR-21的上调或/和miR-31和miR-218的下调等)与胃癌的发生相关,提示miRNA是胃癌发生的重要因素.目前,miRNA表达的分子机制尚未完全明了.最近研究较清楚地显示,miRNA的表达受到DNA甲基化和组蛋白修饰等机制的调控.这说明,胃癌相关miRNA的表达水平受到表观遗传机制的调控。     

大熊猫脐带间充质干细胞外泌体的分离鉴定及miRNAs富集分析

大熊猫脐带间充质干细胞(Umbilical cord mesenchymal stem cells, UC-MSCs)通过旁分泌所释放的外泌体在大熊猫保健与疾病治疗方面具有一定的应用前景。本研究旨在建立大熊猫UC-MSCs外泌体分离方法,开展生物学特征分析和分子鉴定,并研究UC-MSCs外泌体中miRNAs的种类与功能。采用超速离心法从大熊猫UCMSCs培养上清中成功分离外泌体,通过透射电子显微镜进行形态学观察,纳米颗粒跟踪分析仪检测粒径大小,蛋白免疫印迹法检测特异性分子标记表达。采用Small RNA测序技术对UC-MSCs外泌体中的miRNAs进行鉴定,并对其靶基因进行了预测与功能分析。结果显示,大熊猫UC-MSCs外泌体呈圆形杯托状结构,直径为(79.15±4.81) nm,外泌体标志蛋白CD81与TSG101呈阳性表达而CALNEXIN呈阴性表达。大熊猫UC-MSCs外泌体中的miRNA主要为miR-148-3p (30.28%)与miR-21-5p (21.72%)。本研究首次从大熊猫UC-MSCs培养上清中分离出外泌体,并对其所含的miRNAs进行富集分析及功能预测,为大熊...     

外泌体和miRNA在川崎病中的研究进展

外泌体(exosome)是一类径在30～150 nm之间、具有脂质双层膜结构的微小囊泡。生物体内多数细胞都可以分泌外泌体,外泌体可在细胞之间起传递遗传信息的作用,其携带的蛋白质、DNA、RNA等可被受体细胞所摄取并发挥作用。MiRNA作为内源基因编码单链核苷酸分子,可参与体内基因的表达调控,同时在外泌体中也广泛存在。川崎病(Kawasaki disease)是一种急性自身免疫性疾病,目前miRNA在川崎病中的研究已被广泛认可。外泌体在各类疾病中的研究越来越多,但在川崎病的研究中仍然处于初级阶段,外泌体中可以包含多种miRNA而不受破坏,所以外泌体中的miRNA与川崎病的关联性可以进一步探讨。本综述讨论了外泌体的作用和临床意义以及在川崎病诊断与治疗中的优点和缺陷,并提出了外泌体及miRNA可能在其中发挥的作用。     

肝细胞癌患者血清外泌体microRNA表达鉴定

目的:探讨肝癌细胞外泌体中差异表达的microRNAs(miRNAs)在肝细胞癌(HCC)诊断中的应用价值。方法:通过高通量测序筛选肝癌细胞外泌体中差异表达的miRNAs。实时定量PCR验证差异表达分子;检测差异表达的miRNAs在健康人(Health)、慢性乙型肝炎患者(CHB)、肝硬化患者(LC)及乙型肝炎病毒阳性的肝细胞癌患者(HCC)血清外泌体中的表达。结果:高通量测序筛选到肝癌细胞外泌体中差异表达的miRNA共88种,其中58种表达上调,30种表达下调。选择其中8种差异表达的miRNAs进行q RT-PCR验证,结果显示,此8种miRNAs在细胞上清外泌体、细胞内、癌与癌旁组织中的表达趋势与测序结果一致。miR-221-3p和miR-224-5p在HCC组外泌体中的表达水平显著高于Health组、CHB组和LC组(P0.01),miR-124-3p和let-7a-5p在HCC组外泌体中的表达水平显著低于其他各组(P0.05)。四个组中,miR-21-5p、miR-191-5p、miR-34a-5p和miR-122-5p的表达水平不存在显著性差异(P0.05)。结论:血清外泌体中的miR-221-3p、miR-224-5p、miR-124-3p和let-7a-5p可能成为肝细胞癌的候选标志物。     

高通量测序技术分析急性髓系白血病血浆外泌体微RNA表达谱差异

急性髓系白血病（acute myeloid leukemia,AML）是一类造血干细胞的恶性克隆性疾病,目前的诊断方法不利于疾病的早期发现,且诊断结果重复性较差。已有大量研究显示,细胞外microRNA（miRNA或miR）富集在外泌体（exosome）中,且受其表面膜的保护而具有很好的稳定性,是理想的分子标志物。目前,多种实体肿瘤均已检测到肿瘤特异性外泌体miRNA(exosomal miRNA)。然而,在AML患者中未见此外泌体miRNA报道。本研究探讨急性髓系白血病血浆外泌体miRNA表达谱差异及新miRNA序列。采用solexa高通量测序技术对7例AML患者（AML组）及7例健康对照者（对照组）血浆外泌体miRNAs进行测序,利用Mireap预测软件进行新miRNAs分析,通过edger差异分析软件筛选组间差异miRNA,获得211个已知的差异表达miRNAs以及2个新miRNAs,选择4个差异表达的miRNAs：miR-155-5p、miR-335-5p、miR-451a及xxx-m0038 5p（新miRNA）,在两组（各23例）的血浆外泌体样本中,进行实时荧光定量PCR（qRT-PCR）验证,验证结果与测序结果一致。对差异表达的外泌体miRNA进行靶基因预测及其GO（Gene Ontology）和信号通路富集分析,发现靶基因聚集的生物学功能多数参与生物进展过程的调控。靶基因主要富集在FoxO、MAPK、Hippo信号通路以及HTLV-I感染等。结果显示,AML患者与健康对照者的血浆外泌体miRNA存在着差异性表达。差异性表达的miRNA特异性很高,对进一步阐明AML白血病发生与发展的分子机制、研发新的无创诊断方法、新的诊断标记物和有效治疗AML的方法具有十分重要和深远的意义。     

miRNA调控成肌分化的研究进展

成肌分化过程包括成肌细胞的增殖,然后分化为肌细胞,最后融合形成肌管;microRNA(miRNA)是一类在转录后水平调控基因表达的微小非编码RNA,它通过靶向靶基因mRNA的3'UTR,抑制其翻译或诱导其降解。已有研究表明,miRNA在成肌分化中起重要调控作用。根据表达方式的不同,分为肌肉特异表达的miRNA,有miR-1,miR-133,miR-206,miR-208,miR-499和miR-486;和非肌肉特异表达的miRNA,其中miR-27,miR-29,miR-128,miR-199a和miR-431在成肌分化过程中具有重要的调控功能。另外,阐述了几个与miRNA相互作用从而调控成肌分化的lncRNA的功能。通过介绍两类miRNA的靶基因及调控机制,阐述了最新的研究进展。     

外泌体miRNA的生物学功能及其在肺纤维化疾病中的调控作用

外泌体是一种微型纳米级细胞外囊泡,由于能够直接参与细胞间信息的传递和物质的运输,被认为是细胞间通讯、免疫调节、疾病诊断和预后循环生物学标志物的重要载体,其携带的核酸和蛋白质等内含物能够影响受体细胞的生理状态.作为一种内源性非编码微小RNA,microRNA (miRNA)对疾病诊断和治疗有着重要的研究价值,有大量证据表明该类分子对肺部疾病的发病进程起着控制调节作用.本文聚焦于近年来细胞外泌体来源miRNA的生物学特性和功能领域,综述了近年来生物医学研究中的热点分子外泌体miRNA在肺部疾病尤其是肺纤维化中调控功能和机制的研究,因此不仅能为肺纤维化疾病的诊断提供新的标志物分子,并且还能够为肺纤维化的外泌体干预治疗建议新的干预策略.     

MicroRNAs与肿瘤

microRNA(miRNA)是一类对基因具有调控功能的内源性非编码小分子RNA,通过与靶mRNA完全或不完全互补配对,引起mRNA降解或翻译抑制,从而对基因转录后水平进行调控.目前认为miRNA在多种生物学过程中起着至关重要的作用,包括细胞增殖、分化、凋亡等.研究显示miRNA的表达异常能导致疾病甚至肿瘤的发生,有类似于抑癌基因或癌基因的功能.本文就miRNA在肿瘤发生和诊断方面的研究进展作一综述.     

miR-425-5p as an exosomal biomarker for metastatic prostate cancer

Prostate cancer-related deaths are mostly caused by metastasis, which indicates the importance of identifying clinical prognostic biomarkers. In this study, we evaluated the expression profile of exosomal microRNAs (miRNAs) derived from metastatic prostate cancer (mPCa) cell lines (LNCaP and PC-3). miRNA signatures in exosomes and cells were evaluated by miRNA microarray analysis. Fourteen miRNAs were identified as candidates for specific noninvasive biomarkers. The expression of five miRNAs was validated using RT-qPCR, which confirmed that miR-205-5p, miR-148a-3p, miR-125b-5p, miR-183-5p, and miR-425-5p were differentially expressed in mPCa exosomes. Bioinformatic analyses showed that miR-425-5p was associated with residual tumor, pathologic T and N stages, and TP53 status in PCa samples. Gene ontology analysis of negatively correlated and predicted targeted genes showed enrichment of genes related to bone development pathways. The LinkedOmics database indicated that the potential target HSPB8 has a significant negative correlation with miR-425-5p. In conclusion, this study identified a panel of exosomal miRNAs with potential value as prognostic biomarkers for prostate cancer.     

Small RNA deep sequencing reveals a distinct miRNA signature released in exosomes from prion-infected neuronal cells

Prion diseases are transmissible neurodegenerative disorders affecting both humans and animals. The cellular prion protein, PrP^C, and the abnormal infectious form, PrP^Sc, are found associated with exosomes, which are small 50–130 nm vesicles released from cells. Exosomes also contain microRNAs (miRNAs), a class of non-coding RNA, and have been utilized to identify miRNA signatures for diagnosis of disease. While some miRNAs are deregulated in prion-infected brain tissue, the role of miRNA in circulating exosomes released during prion disease is unknown. Here, we investigated the miRNA profile in exosomes released from prion-infected neuronal cells. We performed the first small RNA deep sequencing study of exosomes and demonstrated that neuronal exosomes contain a diverse range of RNA species including retroviral RNA repeat regions, messenger RNA fragments, transfer RNA fragments, non-coding RNA, small nuclear RNA, small nucleolar RNA, small cytoplasmic RNA, silencing RNA as well as known and novel candidate miRNA. Significantly, we show that exosomes released by prion-infected neuronal cells have increased let-7b, let-7i, miR-128a, miR-21, miR-222, miR-29b, miR-342-3p and miR-424 levels with decreased miR-146 a levels compared to non-infected exosomes. Overall, these results demonstrate that circulating exosomes released during prion infection have a distinct miRNA signature that can be utilized for diagnosis and understanding pathogenic mechanisms in prion disease.     

Exosomal microRNAs from mesenchymal stem/stromal cells: Biology and applications in neuroprotection

Mesenchymal stem/stromal cells (MSCs) are extensively studied as cell-therapy agents for neurological diseases. Recent studies consider exosomes secreted by MSCs as important mediators for MSCs’ neuroprotective functions. Exosomes transfer functional molecules including proteins, lipids, metabolites, DNAs, and coding and non-coding RNAs from MSCs to their target cells. Emerging evidence shows that exosomal microRNAs (miRNAs) play a key role in the neuroprotective properties of these exosomes by targeting several genes and regulating various biological processes. Multiple exosomal miRNAs have been identified to have neuroprotective effects by promoting neurogenesis, neurite remodeling and survival, and neuroplasticity. Thus, exosomal miRNAs have significant therapeutic potential for neurological disorders such as stroke, traumatic brain injury, and neuroinflammatory or neurodegenerative diseases and disorders. This review discusses the neuroprotective effects of selected miRNAs (miR-21, miR-17-92, miR-133, miR-138, miR-124, miR-30, miR146a, and miR-29b) and explores their mechanisms of action and applications for the treatment of various neurological disease and disorders. It also provides an overview of state-of-the-art bioengineering approaches for isolating exosomes, optimizing their yield and manipulating the miRNA content of their cargo to improve their therapeutic potential.     

Loss of exosomal miR-148b from cancer-associated fibroblasts promotes endometrial cancer cell invasion and cancer metastasis

Cancer-associated fibroblasts (CAFs) play crucial roles in tumor progression, given the dependence of cancer cells on stromal support. Therefore, understanding how CAFs communicate with endometrial cancer cell in tumor environment is important for endometrial cancer therapy. Exosomes, which contain proteins and noncoding RNA, are identified as an important mediator of cell–cell communication. However, the function of exosomes in endometrial cancer metastasis remains poorly understood. In the current study we found that CAF-derived exosomes significantly promoted endometrial cancer cell invasion comparing to those from normal fibroblasts (NFs). We identified a significant decrease of miR-148b in CAFs and CAFs-derived exosomes. By exogenously transfect microRNAs, we demonstrated that miR-148b could be transferred from CAFs to endometrial cancer cell through exosomes. In vitro and in vivo studies further revealed that miR-148b functioned as a tumor suppressor by directly binding to its downstream target gene, DNMT1 to suppress endometrial cancer metastasis. In endometrial cancer DNMT1 presented a potential role in enhancing cancer cell metastasis by inducing epithelial–mesenchymal transition (EMT). Therefore, downregulated miR-148b induced EMT of endometrial cancer cell as a result of relieving the suppression of DNMT1. Taken together, these results suggest that CAFs-mediated endometrial cancer progression is partially related to the loss of miR-148b in the exosomes of CAFs and promoting the transfer of stromal cell-derived miR-148b might be a potential treatment to prevent endometrial cancer progression.     

Expression of exosomal microRNAs during chondrogenic differentiation of human bone mesenchymal stem cells

The aim of the current study was to compare the expression of microRNAs (miRNAs) in exosomes derived from human bone mesenchymal stem cells (hBMSCs) with and without chondrogenic induction. Exosomes derived from hBMSCs were isolated and identified. Microarray analysis was performed to compare miRNA expression between exosomes derived from hBMSCs with and without chondrogenic induction, and quantitative real-time polymerase chain reaction (qRT-PCR) was used to verify the differentially expressed miRNAs. hBMSCs were transfected with miRNA mimic to extract miRNA-overexpressed exosomes. The results showed that most exosomes exhibited a cup-shaped or round-shaped morphology with a diameter of approximately 50-200 nm and expressed CD9 and CD63. We detected 141 miRNAs that were differentially expressed with and without chondrogenic induction by over a twofold change, including 35 upregulated miRNAs, such as miR-1246, miR-1290, miR-193a-5p, miR-320c, and miR-92a, and 106 downregulated miRNAs, such as miR-377-3p and miR-6891-5p. qRT-PCR analysis validated these results. Exosomes derived from hBMSCs overexpressing miR-320c were more efficient than normal exosomes derived from control hBMSCs at promoting osteoarthritis chondrocyte proliferation, down-regulated matrix metallopeptidase 13 and up-regulated (sex determining region Y)-box 9 expression during hBMSC chondrogenic differentiation. In conclusion, we identified a group of upregulated miRNAs in exosomes derived from hBMSCs with chondrogenic induction that may play an important role in mesenchymal stem cell-derived exosomes in cartilage regeneration and, ultimately, the treatment of arthritis. We demonstrated the potential of these modified exosomes in the development of novel therapeutic strategies.     

Comparative profile of exosomal microRNAs in postmenopausal women with various bone mineral densities by small RNA sequencing

To explore the role of plasma miRNAs in exosomes in early postmenopausal women. Small RNA sequencing was implemented to clarify the expression of miRNA in plasma exosomes obtained from 15 postmenopausal women, divided into groups of osteoporosis, osteopenia, and normal bone mass based on bone mineral density. Differentially expressed miRNAs (DEMs) were identified by comparing miRNA expression profiles. Five putative miRNAs, miR-224-3p, miR-25-5p, miR-302a-3p, miR-642a-3p, and miR-766-5p were confirmed by real-time PCR; miRNA target genes were obtained from 4 databases: miRWalk, miRDB, RNA22, and TargetScan. The miRNA-mRNA- Kyoto Encyclopedia of Genes and Genomes (KEGG) networks were analyzed, and the DEMs' potential role was investigated by gene ontology terms and KEGG pathway annotation. The results suggest that characterizing plasma exosomal miRNA profiles of early postmenopausal women by small RNA sequencing could identify novel exo-miRNAs involved in bone remodeling, and miR-642a-3p maybe contribute to the prediction and diagnosis of early postmenopausal osteoporosis.     

miR-125a/b inhibits tumor-associated macrophages mediated in cancer stem cells of hepatocellular carcinoma by targeting CD90

Cancer stem cells promote tumorigenesis and progression of hepatocellular carcinoma (HCC). Recently, emerging evidence indicates tumor-associated macrophages (TAMs) play an important role in tumor progression. However, TAMs often occurs with unknown mechanisms. As an important mediator in intercellular communications, exosomes secreted by host cells mediate the exchange of genetic materials and proteins, which involves tumor aggressiveness. The aim of the study was to investigate whether exosomes derived from TAMs mediate stem cell properties in HCC. TAMs were isolated from the tissues of HCC. microRNA (miRNA) expression profiles of TAMs were analyzed using miRNA microarray. In vitro cell coculture was further conducted to investigate the crosstalk between TAMs and tumor cells mediated by TAMs exosomes. In this study, we showed that TAMs exosomes promote HCC cell proliferation and stem cell properties. Using miRNA profiles assay, we identified significantly lower levels of miR-125a and miR-125b in exosomes and cell lysate isolated from TAMs. Functional studies revealed that the HCC cells were treated with TAM exosomes or transfected with miR-125a/b suppressed cell proliferation and stem cell properties by targeting CD90, a stem cell marker of HCC stem cells. The study indicated that miR-125a/b targeting CD90 played important roles in cancer stem cells of HCC.     

尿液外泌小体中miRNA的含量与肾纤维化的关系

目的:探讨尿液外泌小体(exosomes)中微小RNA(miRNA,miR)的变化与肾纤维化的关系,以寻找早期诊断肾纤维化的生物标志物。方法:以行肾穿刺活检术并诊断为原发性肾脏病的患者为研究对象,其中,肾活检未发现肾纤维化的患者作为对照组,而存在轻到中度肾纤维化的患者作为纤维化组。收集20 m L晨尿,用超速离心方法分离尿液exosomes,用电镜观察其形态,用定量PCR方法检测其中miRNA的含量,并分析其与肾纤维化的关系。结果:超速离心获得的尿液沉淀物呈现exosomes的形态学特征。miR-21、miR-29b、miR-29c、miR-30e、miR-192、miR-200a、miR-200c和miR-429可在所有患者的尿液exosomes中被检出,但含量存在较大差异。与对照组相比,纤维化组患者尿液exosomes中,miR-21、miR-29b、miR-30e和miR-200c的含量显著增高,miR-29c的含量显著下降,而miR-192、miR-200a和miR-429的含量无显著变化。尿液exosomes中miRNA含量与纤维化肾组织中miRNA表达量的变化并不完全一致。结论:尿液exosomes中miR-29c和miR-21的含量在肾纤维化的病变中发生显著改变,可能成为早期诊断肾纤维化的生物标志物。