title: 17β-Oestradiol inhibits doxorubicin-induced apoptosis via block of the volume-sensitive Cl(-)  current in rabbit articular chondrocytes.
abstract: BACKGROUND AND PURPOSE Chondrocyte apoptosis contributes to disruption of cartilage integrity in osteoarthritis. Recent evidence suggested that the volume-sensitive organic osmolyte/anion channel [volume-sensitive (outwardly rectifying) Cl(-)  current (I(Cl,vol) )] plays a functional role in the development of cell shrinkage associated with apoptosis (apoptotic volume decrease) in several cell types. In this study, we investigated the cellular effects of 17β-oestradiol on doxorubicin-induced apoptotic responses in rabbit articular chondrocytes. EXPERIMENTAL APPROACH Whole-cell membrane currents and cross-sectional area were measured from chondrocytes using a patch-clamp method and microscopic cell imaging, respectively. Caspase-3/7 activity was assayed as an index of apoptosis. KEY RESULTS Addition of doxorubicin (1 µM) to isosmotic bath solution rapidly activated the Cl(-)  current with properties similar to those of I(Cl,vol)  in chondrocytes. Doxorubicin also gradually decreased the cross-sectional area of chondrocytes, followed by enhanced caspase-3/7 activity; both of these responses were totally abolished by the I(Cl,vol)  blocker DCPIB (20 µM). Pretreatment of chondrocytes with 17β-oestradiol (1 nM) for short (approximately 10 min) and long (24 h) periods almost completely prevented the doxorubicin-induced activation of I(Cl,vol)  and subsequent elevation of caspase-3/7 activity. These effects of 17β-oestradiol were significantly attenuated by the oestrogen receptor blocker ICI 182780 (10 µM), as well as the phosphatidyl inositol-3-kinase (PI3K) inhibitors wortmannin (100 nM) and LY294002 (20 µM). Testosterone (10 nM) had no effect on the doxorubicin-induced Cl(-)  current. CONCLUSIONS AND IMPLICATIONS 17β-Oestradiol prevents the doxorubicin-induced cell shrinkage mediated through activation of I(Cl,vol)  and subsequent induction of apoptosis signals, through a membrane receptor-dependent PI3K pathway in rabbit articular chondrocytes.
authors.affiliation: Department of Orthopaedic Surgery, Shiga University of Medical Science, Otsu, Shiga, Japan.
authors.forename: Kousuke
authors.initials: K
authors.lastname: Kumagai
authors.forename: Shinji
authors.initials: S
authors.lastname: Imai
authors.forename: Futoshi
authors.initials: F
authors.lastname: Toyoda
authors.forename: Noriaki
authors.initials: N
authors.lastname: Okumura
authors.forename: Eiji
authors.initials: E
authors.lastname: Isoya
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Matsuura
authors.forename: Yoshitaka
authors.initials: Y
authors.lastname: Matsusue
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D002358:Cartilage, Articular / Q000166:cytology; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D002478:Cells, Cultured; D002712:Chlorides / Q000502:physiology*; D019902:Chondrocytes / Q000187:drug effects* / Q000502:physiology; D004317:Doxorubicin / Q000494:pharmacology; D004958:Estradiol / Q000494:pharmacology*; D004967:Estrogens / Q000494:pharmacology*; D008297:Male; D019255:NADPH Oxidases / Q000378:metabolism; D018408:Patch-Clamp Techniques; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D011817:Rabbits; D017382:Reactive Oxygen Species / Q000378:metabolism
chemical_list: D002712:Chlorides; D004967:Estrogens; D017382:Reactive Oxygen Species; D004958:Estradiol; D004317:Doxorubicin; D019255:NADPH Oxidases; D019869:Phosphatidylinositol 3-Kinases; D053148:Caspase 3; D053179:Caspase 7