SOCS1 and SOCS3 are the best characterized, and SOCS1 is considered more potent than SOCS3. Although IL 1B is not a main inducer of SOCS family proteins or a potent activator of signal transducer and activator of transcription, IL 1B has been reported to induce SOCS1 or SOCS3 in several types of cells including chondrocytes. NSC 737664 Further more, SOCS1 may inhibit IL 1B signaling pathways, SOCS1null T cells were found to be hypersensitive to IL 1B. When HEK293 cells transfected with SOCS1 were stimulated with IL 1B, SOCS1 bound to NF ��B p65 and regulated NF ��B signaling in the nucleus. However, the mechanisms of SOCS1 mediated inhibition of IL 1B signaling pathways have not been fully studied. Here, we demonstrated that the SOCS1 is present in OA cartilage, especially in the area of severe cartil age damage, and is inducible by IL 1B in primary human articular chondrocytes.
Furthermore, SOCS1 sup presses the production of proteolytic matrix metallopro teinases and aggrecanase Inhibitors,Modulators,Libraries 1 in human SW1353 chondrocytic cell lines and HACs by inhi biting c Jun N terminal kinase and p38 mitogen activated protein kinases activation, by preventing the degradation of the inhibitor of NF ��B, and by accelerating degradation of TGF B activated protein kin ase 1. Methods Plasmids and reagents A PINCO retroviral vector expressing myc tagged hu man SOCS1 was kindly provided Inhibitors,Modulators,Libraries by William E. Carson. pShuttle2 and pBABE retroviral vectors were purchased from Addgene. SOCS1 small hairpin RNA and copGFP Control Lentivirus particles came from Santa Cruz Biotechnology.
The Platinum A retroviral packing cell line was obtained from Cell BioLabs. NF ��B mediated luciferase activity was assayed by using Inhibitors,Modulators,Libraries pGL luc based 3X ��B Inhibitors,Modulators,Libraries L plasmid. Recombinant Inhibitors,Modulators,Libraries IL 1B was purchased from Peprotech. ELISA kits for MMP 1, MMP 3, MMP 13, and TIMP 1 were obtained from R D Systems. Anti SOCS1 was purchased from LifeSpan Bioscience for immunohistochemistry, and Chemi con International, for immuno blot. Anti TAK1 was purchased from Novus Biologicals for immunoprecipitation and from Santa Cruz for immunoblot. Anti phospho NF ��B p65 and anti myc were obtained from ABcam, and anti I��B was from Santa Cruz. Anti ADAMTS4 was from Calbiochem. The other antibodies were pur chased from Cell Signaling Technology. An ERK inhibitor U0126 was obtained from Promega, and JNK inhibitor SP600125 was from BioMol International.
A p38 MAP kinase inhibitor SB202190 U0126 MEK and NF ��B inhibitor SN50 were purchased from Alexis Biochemicals. MG132 was from Sigma Aldrich. SW1353 chondro sarcoma cell line was obtained from American Type Culture Collection. Patients and cartilage samples OA cartilage was obtained from 14 patients with pri mary knee OA who underwent total knee replacement arthroplasty. Control healthy cartilage specimens were obtained from four patients with femur neck fractures who had no history of hip OA. A written informed con sent was obtained from all study participants.