Li Yu-Xi, Liu Ting, Liang Yu-Wei, Huang Jia-Jun, Huang Jun-Shen, Liu Xiang-Ge, Cheng Zi-Ying, Lu Shi-Xin, Li Ming, Huang Lin
Department of Orthopaedics, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, China.
Department of Anaesthesia, Sun Yat-sen Memorial Hospital, Sun Yat-sen University, Guangzhou, China.
Ann Transl Med. 2021 Oct;9(20):1563. doi: 10.21037/atm-21-5020.
The precise pathogenesis of ankylosing spondylitis (AS) is still largely unknown at present. Our previous study found that toll-like receptor 4 (TLR4) downregulated and performed immunoregulatory dysfunction in mesenchymal stem cells from AS patients (AS-MSCs). The aim of this study was to explore the expression profiles of long non-coding RNAs (lncRNAs) and messenger RNAs (mRNAs) in TLR4-primed AS-MSCs, and to clarify the potential mechanisms.
The immunoregulatory effects of MSCs were determined after TLR4 activation. Next, the differentially-expressed (DE) lncRNAs and mRNAs between AS-MSCs and TLR4-primed AS-MSCs [stimulated by lipopolysaccharide (LPS)] were identified via high-throughput sequencing followed by quantitative real-time PCR (qRT-PCR) confirmation. Finally, bioinformatics analyses were performed to identify the critical biological functions, signaling pathways, and associated functional networks involved in the TLR4-primed immunoregulatory function of AS-MSCs.
A total of 147 DE lncRNAs and 698 DE mRNAs were identified between TLR4-primed AS-MSCs and unstimulated AS-MSCs. Of these, 107 lncRNAs were upregulated and 40 were downregulated (fold change ≥2, P<0.05), while 504 mRNAs were upregulated and 194 were downregulated (fold change ≥2, P<0.05). Five lncRNAs and five mRNAs with the largest fold changes were respectively verified by qRT-PCR. Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses demonstrated that the DE mRNAs and lncRNAs were highly associated with the inflammatory response, such as NOD-like receptor (NLR) signaling pathway, the TNF signaling pathway and the NF-κB signaling pathway. Cis-regulation prediction revealed eight novel lncRNAs, while trans-regulation prediction revealed 15 lncRNAs, respectively. Eight core pairs of lncRNA and target mRNA in the lncRNA-transcription factor (TF)-mRNA network were as follows: PACERR-PTGS2, LOC105378085-SOD2, LOC107986655-HIVEP2, MICB-DT-MICB, LOC105373925-SP140L, LOC107984251-IFIT5, LOC112268267-GBP2, and LOC101926887-IFIT3, respectively.
TLR4 activation in AS can enhance the immunoregulatory ability of MSCs. Eight core pairs of lncRNA and target mRNA were observed in TLR4-primed AS-MSCs, which could contribute to understanding the potential mechanism of AS-MSC immunoregulatory dysfunction.
目前，强直性脊柱炎（AS）的确切发病机制仍不清楚。我们之前的研究发现，Toll样受体4（TLR4）在AS患者的间充质干细胞（AS-MSCs）中表达下调并表现出免疫调节功能障碍。本研究旨在探讨TLR4预处理的AS-MSCs中长链非编码RNA（lncRNAs）和信使RNA（mRNAs）的表达谱，并阐明潜在机制。
TLR4激活后测定MSCs的免疫调节作用。接下来，通过高通量测序鉴定AS-MSCs与TLR4预处理的AS-MSCs[由脂多糖（LPS）刺激]之间的差异表达（DE）lncRNAs和mRNAs，随后进行定量实时PCR（qRT-PCR）验证。最后，进行生物信息学分析，以确定参与TLR4预处理的AS-MSCs免疫调节功能的关键生物学功能、信号通路和相关功能网络。
在TLR4预处理的AS-MSCs和未刺激的AS-MSCs之间共鉴定出147个DE lncRNAs和698个DE mRNAs。其中，107个lncRNAs上调，40个下调（倍数变化≥2，P<0.05），而504个mRNAs上调，194个下调（倍数变化≥2，P<0.05）。通过qRT-PCR分别验证了倍数变化最大的5个lncRNAs和5个mRNAs。基因本体（GO）和京都基因与基因组百科全书（KEGG）分析表明，DE mRNAs和lncRNAs与炎症反应高度相关，如NOD样受体（NLR）信号通路、TNF信号通路和NF-κB信号通路。顺式调控预测分别揭示了8个新的lncRNAs，而反式调控预测揭示了15个lncRNAs。lncRNA-转录因子（TF）-mRNA网络中的8对核心lncRNA和靶mRNA分别如下：PACERR-PTGS2、LOC105378085-SOD2、LOC107986655-HIVEP2、MICB-DT-MICB、LOC105373925-SP140L、LOC107984251-IFIT5、LOC112268267-GBP2和LOC101926887-IFIT3。
AS中TLR4激活可增强MSCs的免疫调节能力。在TLR4预处理的AS-MSCs中观察到8对核心lncRNA和靶mRNA，这有助于理解AS-MSC免疫调节功能障碍的潜在机制。
