Kim Seong-Kyu, Choe Jung-Yoon, Kim Ji-Won, Park Ki-Yeun
Division of Rheumatology, Department of Internal Medicine, Catholic University of Daegu School of Medicine, Daegu 42472, Republic of Korea.
Arthritis and Autoimmunity Research Center, Catholic University of Daegu, 33, Duryugongwon-ro 17-gil, Nam-gu, Daegu 42472, Republic of Korea.
Pharmaceuticals (Basel). 2023 Mar 16;16(3):446. doi: 10.3390/ph16030446.
Histone deacetylase (HDAC) has been found to play a crucial role in the regulation of osteoclast differentiation and formation. This study was designed to identify the effect of the HDAC6 inhibitor CKD-WID on the receptor for the activation of nuclear factor-κB ligand (RANKL)-mediated osteoclast formation in the presence of monosodium urate (MSU) in RAW 264.7 murine macrophage cells. The expression of osteoclast-specific target genes, calcineurin, and nuclear factor of activated T-cells cytoplasmic 1 (NFATc1) was evaluated in RAW 264.7 murine macrophages treated with MSU, RANKL, or CKD-WID by real-time quantitative polymerase chain reaction and Western blot assay. The effect of CKD-WID on osteoclast formation was measured by tartrate-resistant acid phosphatase (TRAP) staining, F-actin ring formation staining, and assays for bone resorption activity. RANKL in the presence of MSU significantly induced HDAC6 gene and protein expression in RAW 264.7 cells. CKD-WID markedly suppressed the expression of osteoclast-related markers such as c-Fos, TRAP, cathepsin K, and carbonic anhydrase II induced by co-stimulation with RANKL and MSU in RAW 264.7 cells. Transcription factor NFATc1 mRNA expression and nuclear NFATc1 protein expression induced by co-stimulation with RANKL and MSU were significantly inhibited by CKD-WID treatment. CKD-WID also decreased the number of TRAP-positive multinuclear cells and F-actin ring-positive cells and attenuated bone resorption activity. Co-stimulation with RANKL and MSU increased calcineurin gene and protein expression, which was significantly blocked by CKD-WID treatment. The HDAC6 inhibitor CKD-WID suppressed MSU-induced osteoclast formation through blocking the calcineurin-NFAT pathway in RAW 264.7 cells. This suggests that HDAC6 is considered a therapeutic target in uric acid-mediated osteoclastogenesis.
已发现组蛋白脱乙酰酶（HDAC）在破骨细胞分化和形成的调节中起关键作用。本研究旨在确定HDAC6抑制剂CKD-WID对RAW 264.7小鼠巨噬细胞中尿酸单钠（MSU）存在下核因子-κB配体（RANKL）介导的破骨细胞形成受体的影响。通过实时定量聚合酶链反应和蛋白质印迹分析，评估了用MSU、RANKL或CKD-WID处理的RAW 264.7小鼠巨噬细胞中破骨细胞特异性靶基因、钙调神经磷酸酶和活化T细胞核因子细胞质1（NFATc1）的表达。通过抗酒石酸酸性磷酸酶（TRAP）染色、F-肌动蛋白环形成染色和骨吸收活性测定，测量CKD-WID对破骨细胞形成的影响。在MSU存在下，RANKL显著诱导RAW 264.7细胞中HDAC6基因和蛋白表达。CKD-WID显著抑制RAW 264.7细胞中由RANKL和MSU共同刺激诱导的破骨细胞相关标志物如c-Fos、TRAP、组织蛋白酶K和碳酸酐酶II的表达。用CKD-WID处理可显著抑制RANKL和MSU共同刺激诱导的转录因子NFATc1 mRNA表达和核NFATc1蛋白表达。CKD-WID还减少了TRAP阳性多核细胞和F-肌动蛋白环阳性细胞的数量，并减弱了骨吸收活性。RANKL和MSU共同刺激增加了钙调神经磷酸酶基因和蛋白表达，而CKD-WID处理可显著阻断这种增加。HDAC6抑制剂CKD-WID通过阻断RAW 264.7细胞中的钙调神经磷酸酶-NFAT途径，抑制MSU诱导的破骨细胞形成。这表明HDAC6被认为是尿酸介导的破骨细胞生成中的一个治疗靶点。
