Antibodies against ClC7 inhibit extracellular acidification-induced Cl^-currents and bone resorption activity in mouse osteoclasts
Antibodies against ClC7 inhibit extracellular acidification-induced Cl^-currents and bone resorption activity in mouse osteoclasts
复制标题
ClC7 抗体抑制小鼠破骨细胞中细胞外酸化诱导的 Cl^电流和骨吸收活性
DOI：
10.1007/s00210-010-0576-8
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发表时间：
2011
期刊：
影响因子：
--
通讯作者：
Okabe K
中科院分区：
文献类型：
--
作者：
Ohgi K;Okamoto F;Kajiya H;Sakagami R;Okabe K
CL−通道/转运体ClC7在破骨细胞性骨吸收中起关键作用，可能成为骨质疏松的治疗靶点。在这项研究中，我们针对常染色体显性遗传性骨化病II型患者中发现的三个不同的多肽序列G215、P249和R286制备了抗ClC7多克隆抗体，并利用全细胞膜片钳技术检测了这些抗体对细胞外酸化诱导的ClC7Cl−电流(酸激活的Cl−电流)和小鼠破骨细胞的骨吸收活性的影响。破骨细胞胞内透析抗G215抗体(Ab-G215)和胞外应用抗P249抗体(Ab-P249)或抗R286抗体(Ab-R286)可抑制酸激活的Cl-−电流。这些抗体也抑制高表达的ClC7细胞的酸激活的Cl-−电流，但不影响低张刺激诱发的Cl-−电流和小鼠破骨细胞固有的内向整流性K+电流。此外，细胞外应用抗体-P249和抗体-R286也降低了骨吸收活性。我们的结果表明，这些抗体特异性地阻断了小鼠破骨细胞中的ClC7。因此，抗ClC7抗体在治疗骨质疏松症方面具有潜在的应用前景。
The Cl−channel/transporter ClC7 is crucial for osteoclastic bone resorption and might become a therapeutic target for osteoporosis. In this study, we raised anti-ClC7 polyclonal antibodies against three different peptide sequences, including G215, P249, and R286, which are the mutation regions found in autosomal dominant osteopetrosis type II patients and examined the effects of these antibodies on the ClC7 Cl−current induced by extracellular acidification (acid-activated Cl−current) using the whole-cell patch clamp technique and bone resorption activity in mouse osteoclasts. Intracellular dialysis of osteoclasts with antibodies to intracellular G215 (Ab-G215) and extracellular application of antibodies to extracellular P249 (Ab-P249) or R286 (Ab-R286) inhibited the acid-activated Cl−current. These antibodies also suppressed the acid-activated Cl−current in ClC7 overexpressing Raw264.7 cells; however, Cl−currents evoked by hypotonic stimulation and the inherent inwardly rectifying K+currents in mouse osteoclasts were unaffected by these antibodies. Furthermore, extracellularly applied Ab-P249 and Ab-R286 also reduced bone resorption activity. Our results demonstrate that these antibodies specifically block ClC7 in mouse osteoclasts. Thus, anti-ClC7 antibodies have potential promise for treatment of osteoporosis.