Swelling-activated ClC-3 activity regulates prostaglandin E₂ release in human OUMS-27 chondrocytes

Articular chondrocytes are exposed to dynamic osmotic environments during normal joint loading, and thus, require effective volume regulatory mechanisms. A regulatory volume decrease (RVD) is one of the mechanisms for protecting chondrocytes from swelling and damage. Swelling-activated Cl^- currents (I_Cl,swell) are responsible for the RVD, but the molecular identity in chondrocytes is largely unknown. In this study, we reveal that in human OUMS-27 chondrocytes, I_Cl,swell can be elicited by hypoosmotic stimulation (180 mOsm) and be inhibited by classical Cl^- channel blockers, 4,4'-diisothiocyano-2,2'-stilbenedisulfonic acid (DIDS) and niflumic acid, and be attenuated by siRNA knockdown of ClC-3. Our molecular analyses revealed that ClC-3A is expressed as a major splice variant in both human articular chondrocytes and OUMS-27 cells. The onset and early phase of RVD following hypoosmotic stress in OUMS-27 cells were affected by DIDS and ClC-3 knockdown. Hypoosmotic stimulation caused Ca²⁺ influx and subsequent release of prostaglandin E₂ (PGE₂) in OUMS-27 cells, and both of these responses were reduced by DIDS and ClC-3 knockdown. These results strongly suggest that ClC-3 is responsible for I_Cl,swell and RVD under the hypoosmotic environments. It is likely that ClC-3 is associated with the pathogenesis of cartilage degenerative diseases including osteoarthritis via PGE₂ release.