Mechanosensitive synoviocytes:: A Ca2+-PKCα-MAP kinase pathway contributes to stretch-induced hyaluronan synthesis in vitro

Hyaluronan (HA) is central to joint function, contributing to synovial fluid retention, lubrication, matrix organisation and joint embryogenesis. HA synthesis by intimal synoviocytes is stimulated by stretch (SSHA), linking HA production to joint usage; but the signal transduction paths are unknown. Low passage rabbit synoviocytes (RS), cultured from micro dissected synovial intima, were subjected to 10min of 10% static stretch followed by 170-min relaxation, or to sustained stretch for 180min in a Flexcell 2000 apparatus. Medium HA content was analysed by a HA-binding assay. The roles of protein kinase C (PKC) isoforms, extracellular signal-regulated kinases (ERK1/2) and Ca2+ signalling in SSHA were tested using kinase inhibitors, Ca2+ chelators and Ca2+ channel activators combined with Western blots for activated kinases. Stretch increased HA secretion by 57%, independently of stretch duration. PKC alpha translocated from cylosol to membrane and triggered the phosphorylation of ERK1/2. The PKC inhibitor bisindolylmaleimide (BIM) blocked both SSHA and ERK phosphorylation, as did Go 6976, a specific inhibitor of Ca2+- dependent PKC. The Ca2+ channel activator Bay K stimulated HA secretion and ERK phosphorylation. Extra- and intra-cellular Ca2+ chelation by EGTA and BAPTA-AM (respectively) inhibited SSHA. SSHA was also blocked by the partially selective protein kinase A inhibitor, H-89. Connective tissue growth factor, CTGF, was not involved in SSHA. Thus, stimulation of synoviocyte HA secretion by static stretch is due at least in part the o activation of a Ca2+ influx-dependent activation of the PKC alpha-MEK-ERK1/2 cascade. This is functionally important because it links joint lubrication to joint use. (c) 2006 Elsevier B.V./Anternational Society of Matrix Biology. All rights reserved.