Details

Autor(en) / Beteiligte

• Kajiya, Hiroshi
• Okamoto, Fujio
• Li, Jing‐Ping
• Nakao, Akihiro
• Okabe, Koji

Titel

Expression of Mouse Osteoclast K‐Cl Co‐Transporter‐1 and Its Role During Bone Resorption

Ist Teil von

• Journal of bone and mineral research, 2006-07, Vol.21 (7), p.984-992

Ort / Verlag

Washington, DC: John Wiley and Sons and The American Society for Bone and Mineral Research (ASBMR)

Erscheinungsjahr

2006

Quelle

MEDLINE

Beschreibungen/Notizen

• To assess the role of Cl− transport during osteoclastic bone resorption, we studied the expression and function of K+/Cl− co‐transporters (KCCs). KCC1 and chloride channel‐7 were found to be expressed in mouse osteoclasts. The KCC inhibitor, R(+)‐butylindazone (DIOA), KCC1 antisense oligo‐nucleotides, and siRNA suppressed osteoclastic pit formation. DIOA also decreased Cl− extrusion and reduced H+ extrusion activity. These results show that KCC1 provides a Cl− extrusion mechanism accompanying the H+ extrusion during bone resorption. Introduction: Mice with deficient chloride (Cl−) channels, ClC7, show severe osteopetrosis, resulting from impairment of Cl− extrusion during osteoclastic bone resorption. However, the expression and functional role of Cl− transporters other than ClC7 in mammalian osteoclasts is unknown. The aim of this study was to determine expression of K+/Cl− co‐transporters (KCCs) and their functional role for bone resorption in mouse osteoclasts. Materials and Methods: Mouse osteoclasts were derived from cultured bone marrow cells with macrophage‐colony stimulating factor (M‐CSF) and RANKL or from co‐culture of bone marrow cells and primary osteoblasts. We examined the expression of Cl− transporters using RT‐PCR, immunochemical, and Western blot methods. The effects of Cl− transport inhibitors on H+ and Cl− extrusion were assessed by measuring intracellular H+ ([H+]i) and Cl− ([Cl−]i). The effects of inhibitors, antisense oligo‐nucleotides, and siRNA for Cl− transporters on bone resorption activities were evaluated using a pit formation assay. Results and Conclusions: Mouse osteoclasts express not only ClC7 but also K+/Cl− co‐transporter mRNA. The existence of KCC1 in the cell membrane of mouse osteoclasts was confirmed by immunochemical staining and Western blot analysis. KCC inhibitors and Cl− channels blockers increased [Cl−]i and [H+]i in resorbing osteoclasts, suggesting that the suppression of Cl− extrusion through KCC and Cl− channels leads to reduced H+ extrusion activity. The combination of both inhibitors greatly suppressed these extrusion activities. KCC inhibitors and Cl− channel blockers also decreased osteoclastic bone resorption in our pit area essay. Furthermore, KCC1 antisense oligo‐nucleotides and siRNA suppressed osteoclastic pit formation as well as treatment of ClC7 inhibitors. These results indicate that K+/Cl− co‐transporter‐1 expressed in mouse osteoclasts acts as a Cl− extruder and plays an important role for H+ extrusion during bone resorption.