Department of Clinical Laboratory, Kobe University Hospital, Kobe, Japan.
Department of Clinical Laboratory, Kobe University Hospital, Kobe, Japan Division of Laboratory Medicine, Kobe University Graduate School of Medicine, Kobe, Japan.
Ann Rheum Dis. 2016 Mar;75(3):601-8. doi: 10.1136/annrheumdis-2014-206417. Epub 2015 Jan 16.
MicroRNAs (miRNAs) are small endogenous, non-coding RNAs that act as post-transcriptional regulators. We analysed the in vivo effect of miRNA-124 (miR-124, the rat analogue of human miR-124a) on adjuvant-induced arthritis (AIA) in rats.
AIA was induced in Lewis rats by injecting incomplete Freund's adjuvant with heat-killed Mycobacterium tuberculosis. Precursor (pre)-miR-124 was injected into the right hind ankle on day 9. Morphological changes in the ankle joint were assessed by micro-CT and histopathology. Cytokine expression was examined by western blotting and real-time RT-PCR. The effect of miR-124 on predicted target messenger RNAs (mRNAs) was examined by luciferase reporter assays. The effect of pre-miR-124 or pre-miR-124a on the differentiation of human osteoclasts was examined by tartrate-resistant acid phosphatase staining.
We found that miR-124 suppressed AIA in rats, as demonstrated by decreased synoviocyte proliferation, leucocyte infiltration and cartilage or bone destruction. Osteoclast counts and expression level of receptor activator of the nuclear factor κB ligand (RANKL), integrin β1 (ITGB1) and nuclear factor of activated T cells cytoplasmic 1 (NFATc1) were reduced in AIA rats treated with pre-miR-124. Luciferase analysis showed that miR-124 directly targeted the 3'UTR of the rat NFATc1, ITGB1, specificity protein 1 and CCAAT/enhancer-binding protein α mRNAs. Pre-miR-124 also suppressed NFATc1 expression in RAW264.7 cells. Both miR-124 and miR-124a directly targeted the 3'-UTR of human NFATc1 mRNA, and both pre-miR-124 and pre-miR-124a suppressed the differentiation of human osteoclasts.
We found that miR-124 ameliorated AIA by suppressing critical prerequisites for arthritis development, such as RANKL and NFATc1. Thus, miR-124a is a candidate for therapeutic use for human rheumatoid arthritis.
MicroRNAs（miRNAs）是一种小的内源性非编码 RNA，作为转录后调控因子发挥作用。我们分析了 miRNA-124（miR-124，大鼠与人 miR-124a 的类似物）对大鼠佐剂诱导关节炎（AIA）的体内作用。
用加热杀死的结核分枝杆菌的不完全弗氏佐剂向 Lewis 大鼠诱导 AIA。在第 9 天向右侧后踝关节注射前体（pre）-miR-124。通过 micro-CT 和组织病理学评估踝关节的形态变化。通过 Western blot 和实时 RT-PCR 检查细胞因子表达。通过荧光素酶报告基因检测分析 miR-124 对预测靶信使 RNA（mRNA）的影响。通过抗酒石酸酸性磷酸酶染色检查 pre-miR-124 或 pre-miR-124a 对人破骨细胞分化的影响。
我们发现 miR-124 抑制了大鼠的 AIA，表现为滑膜细胞增殖、白细胞浸润和软骨或骨破坏减少。用 pre-miR-124 治疗的 AIA 大鼠中破骨细胞计数和核因子κB 受体激活物配体（RANKL）、整合素β1（ITGB1）和激活 T 细胞核因子细胞质 1（NFATc1）的表达水平降低。荧光素酶分析表明，miR-124 直接靶向大鼠 NFATc1、ITGB1、特异性蛋白 1 和 CCAAT/增强子结合蛋白α mRNA 的 3'UTR。Pre-miR-124 还抑制了 RAW264.7 细胞中的 NFATc1 表达。miR-124 和 miR-124a 均直接靶向人 NFATc1 mRNA 的 3'-UTR，并且 pre-miR-124 和 pre-miR-124a 均抑制人破骨细胞的分化。
我们发现 miR-124 通过抑制关节炎发展的关键前提条件，如 RANKL 和 NFATc1，改善了 AIA。因此，miR-124a 是治疗人类类风湿关节炎的候选药物。
