以次世代定序及生物資訊鑑別退化性關節炎之成纖維樣滑膜細胞的小分子核醣核酸及其影響標靶

Next-generation sequencing and bioinformatics identifies miRNA and its targets in osteoarthritic fibroblast-like synoviocytes

黃靖雰(Ching-Fen Huang)

指導教授 ： 黃阿梅

高雄醫學大學/醫學院/臨床醫學研究所碩士班/碩士(2020年)

摘要

退化性骨關節炎之滑膜炎(Synovitis in osteoarthritis, OA) 是一低度發炎疾病，其經由軟骨分解產物刺激成纖維樣滑膜細胞(FLS, fibroblast-like synoviocytes)產生發炎介質前驅物所致。FLS 參與關節恆定(joint homeostasis)且關節微環境下的低度發炎會引起FLS轉化。本研究旨在利用次世代定序與生物資訊分析，來探討新穎的miRNA(小分子核糖核酸)如何調控人類OA FLS的機制。將人類正常和OA的 FLS進行RNA萃取，並進一步進行RNA和small RNA定序分析。結果共有14個失調基因具有潛在miRNA調控作用，其中5個失調基因，包括SKAP2 (src kinase associated phosphoprotein 2)、AP1S2 (adaptor related protein complex 1 sigma 2 subunit)、PHF21A (PHD finger protein 21A)、LPP (lipoma preferred partner) 和TFAP2A(transcription factor AP-2 alpha) ，在OA FLS 和 OA滑膜組織(synovial tissue)與GEO資料庫(Gene Expression Omnibus database) 顯示類似的表現情形。IPA(Ingenuity Pathway Analysis)分析發現，可能經由miR-141-3p的調控導致失調的LPP參與OA FLS的細胞遷移和細胞擴散。因此，本研究結果推論出一個新的觀點，提出FLS在OA發病機制新發現的分子特徵，可能是未來的治療目標。

關鍵字

骨關節炎 ； 滑膜炎 ； 成纖維樣滑膜細胞 ； 次世代定序

並列摘要

Synovitis in osteoarthritis (OA) is the consequence of low grade inflammatory process caused by cartilage breakdown products that stimulated the production of pro-inflammatory mediators by fibroblast-like synoviocytes (FLS). FLS participate in joint homeostasis and low grade inflammation in the joint microenvironment triggers FLS transformation. In the current study, we aimed to identify differentially expressed genes and potential miRNA regulations in human OA FLS through deep sequencing and bioinformatics approaches. The 245 differentially expressed genes in OA FLS were identified, and pathway analysis using various bioinformatics databases indicated their enrichment in functions related to altered extracellular matrix organization, cell adhesion and cellular movement. Moreover, among the 14 dysregulated genes with potential miRNA regulations identified, src kinase associated phosphoprotein 2 (SKAP2), adaptor related protein complex 1 sigma 2 subunit (AP1S2), PHD finger protein 21A (PHF21A), lipoma preferred partner (LPP), and transcription factor AP-2 alpha (TFAP2A) showed similar expression patterns in OA FLS and OA synovial tissue datasets in Gene Expression Omnibus database. Ingenuity Pathway Analysis identified the dysregulated LPP participated in cell migration and cell spreading of OA FLS, which was potentially regulated by miR-141-3p. The current findings suggested new perspectives into understanding the novel molecular signatures of FLS involved in the pathogenesis of OA, which may be potential therapeutic targets.

並列關鍵字

osteoarthritis ； synovitis ； fibroblast-like synoviocyte ； next-generation sequencing

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