弥漫性皮肤系统性硬皮病患者外周血中性粒细胞外泌体中 miRNAs 和 lncRNAs 的特征。
The profiles of miRNAs and lncRNAs in peripheral blood neutrophils exosomes of diffuse cutaneous systemic sclerosis.
机构信息
Department of Rheumatology and immunology, Xiangya Hospital of Central South University, Changsha, China; The Institute of Rheumatology and Immunology, Central South University, Changsha, China.
出版信息
J Dermatol Sci. 2020 May;98(2):88-97. doi: 10.1016/j.jdermsci.2020.02.009. Epub 2020 Feb 28.
BACKGROUND
Diffuse cutaneous systemic sclerosis (dSSc) is a systemic autoimmune disease with skin fibrosis. Neutrophils display important roles in autoimmunity, inflammation, vasculopathy and fibrosis. Exosomes (EXOs) are cell-derived vesicles contained various noncoding RNAs, mRNA and proteins with biological roles.
OBJECTIVE
To investigate the roles of miRNAs and lncRNAs from dSSc neutrophils EXOs.
METHODS
EXOs were isolated from cultured neutrophils supernatants and identified by transmission electron microscopy. Global expression of miRNAs and lncRNAs in neutrophils EXOs were sequenced by Illumina HiSeq 3000 and bioinformatic analyses were performed by R/Bioconductor. Genes were validated by real-time quantitative PCR.
RESULTS
In profiles of neutrophils EXOs, we identified 22 dysregulated miRNAs and 281 dysregulated lncRNAs. Predicted target genes of them were enriched in GO, KEGG and Reactome pathways, Wnt, AMPK, IL-23 and NOTCH signaling pathways were selected for further analysis. Widely interactions among them were also found. Human dermal microvascular endothelial cells and human primary skin fibroblasts were stimulated with dSSc neutrophils EXOs, these fibrosis related genes were detected and some changes were found, such as ENST00000533886.1-hsa-miR-1268a-CAMK2G in Wnt and IL-23 signaling pathways, ENST00000610091.1-hsa-miR-299-3p, 512-3p-CPT1A in IL-23 and AMPK signaling pathways, NR_001564.2, ENST00000520562.1, ENST00000596567.1-hsa-miR-299-3p, 512-3p -TFDP2 in IL-23, AMPK and NOTCH signaling pathways.
CONCLUSIONS
The profiles of miRNAs and lncRNAs of neutrophils EXOs provided novel clues for dSSc pathogenesis. We identified several gene pairs in the Wnt, AMPK, IL-23 and NOTCH signaling pathways, which could be potential biomarkers and therapeutic targets in dSSc.
背景
弥漫性皮肤系统性硬皮病（dSSc）是一种伴有皮肤纤维化的系统性自身免疫性疾病。中性粒细胞在自身免疫、炎症、血管病变和纤维化中发挥重要作用。外泌体（EXOs）是含有各种非编码 RNA、mRNA 和具有生物学功能的蛋白质的细胞衍生小泡。
目的
研究 dSSc 中性粒细胞 EXOs 中 miRNAs 和 lncRNAs 的作用。
方法
通过透射电子显微镜分离培养中性粒细胞上清液中的 EXOs，并进行鉴定。通过 Illumina HiSeq 3000 对中性粒细胞 EXOs 中的 miRNAs 和 lncRNAs 进行全基因组表达谱测序，并通过 R/Bioconductor 进行生物信息学分析。通过实时定量 PCR 验证基因。
结果
在中性粒细胞 EXOs 的图谱中，我们鉴定出 22 个失调的 miRNAs 和 281 个失调的 lncRNAs。它们的预测靶基因富集在 GO、KEGG 和 Reactome 途径中，选择 Wnt、AMPK、IL-23 和 NOTCH 信号通路进行进一步分析。还发现它们之间存在广泛的相互作用。用 dSSc 中性粒细胞 EXOs 刺激人真皮微血管内皮细胞和人原代皮肤成纤维细胞，检测这些纤维化相关基因，发现一些变化，如 Wnt 和 IL-23 信号通路中的 ENST00000533886.1-hsa-miR-1268a-CAMK2G，IL-23 和 AMPK 信号通路中的 ENST00000610091.1-hsa-miR-299-3p、512-3p-CPT1A，IL-23、AMPK 和 NOTCH 信号通路中的 NR_001564.2、ENST00000520562.1、ENST00000596567.1-hsa-miR-299-3p、512-3p-TFDP2。
结论
中性粒细胞 EXOs 的 miRNAs 和 lncRNAs 图谱为 dSSc 的发病机制提供了新的线索。我们在 Wnt、AMPK、IL-23 和 NOTCH 信号通路中鉴定了几个基因对，它们可能是 dSSc 的潜在生物标志物和治疗靶点。
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