Role Of ClC-3 Chloride Channel In Regulating The Expression Of Collagen Type ? In Chondrocytes

Background: Osteoarthritis(OA)is a degenerative joint disease characterized by progressive loss of cartilage matrix.The osmotic pressure,inflammation and various physical and chemical changes in the joint cavity affect the functional status of the chondrocytes,including various ion channels distributed in the chondrocytes: sodium channels,potassium channels,and chloride channels.In recent years,studies had shown that Cl C-3 chloride channel plays an important role in the regulation of inflammation and bone mineralization,but its role in regulating extracellular matrix of cartilage has not been reported.Objective: This study focuses on the expression level of Cl C-3 chloride channel in the OA chondrocytes in comparison normal chondrocytes.Besides analyzing the effect of Cl C-3 chloride channel expression on chondrocyte functions in normal chondrocytes.We also aim to explore the effect of IL-1? on the expression of Cl C-3 chloride channel in normal chondrocytes.Methods: The cartilage tissue samples of patients with femoral neck fracture and osteoarthritis were collected during the joint replacement patients of Shenzhen Second People's Hospital,thereafter chondrocytes were isolated and cultured.Primary normal chondrocytes and OA chondrocytes were used for experiments.A chloride channel blocker has been used to block chloride channels.Cl C-3-si RNA transfection and lentiviral transfection have been performed.Then Cl C-3 gene overexpression and/or low expression was determined.IL-1? cytokine has been added to the chondrocytes culture.Later,real-time quantitative PCR and immunofluorescence staining were used to observe COL II gene m RNA relative expression and protein levels in the chondrocytes.Results:(1)Cl C-3 chloride channel inhibition significantly down-regulated COL II gene expression in the chondrocytes.(2)The lentivirus transfection to overexpress Cl C-3 chloride channel in normal chondrocytes significantly enhanced COL II gene m RNA and protein expression.(3)The inflammatory factor IL-1? significantly inhibited the m RNA and protein levels of COL II,but did not affect the expression of Cl C-3 chloride channel m RNA.Conclusion:(1)The expression of Cl C-3 chloride channel in normal chondrocytes is found significantly associated with collagen type II expression in chondrocytes;(2)The inflammatory factor IL-1? could down-regulate COL II expression in the normal chondrocytes without affecting Cl C-3 chloride channel expression.(3)The specific mechanism of action and signaling pathway of Cl C-3 chloride channel overexpression in the OA chondrocytes remain unclear,which need further investigation.