Arthritis & Rheumatism, Volume 63,
November 2011 Abstract Supplement
Abstracts of the American College of
Rheumatology/Association of Rheumatology Health Professionals
Annual Scientific Meeting
Chicago, Illinois November 4-9, 2011.
Chondrocytes in a Subset of Osteo-Arthritic Patients with Reduced Nuclear FactorkappaB-p65 Levels Are Vulnerable to Apoptosis Induced by Tumor Necrosis Factor Alpha, Which Could Be Released by the Synovium.
Arntz, Onno J., Vermeij, Eline A., Davidson, Esmeralda N. Blaney, Abdollahi-Roodsaz, Shahla, Bennink, Miranda B., van der Kraan, Peter M., van den Berg, Wim B.
Chondrocytes play a central role in cartilage pathology as seen in rheumatoid arthritis (RA ) and osteoarthritis (OA) patients by a deranged synthesis of extracellular matrix (ECM) components and the enhanced release of ECM destructive metalloproteinases (MMPs). Nuclear factor-kappaB (NF-kappaB) is an important transcription factor in the regulation of MMPs, but is also regarded as a survival factor in cells. We studied the levels of NFkappaB-p65(P65) in articular chondrocytes of osteoarthritis(OA), rheumatoid arthritis(RA), and mouse models and study the biological consequence regarding cell viability.
Levels of P65 were detected by immunohistochemistry and RT-qPCR levels in human cartilage of RA and OA patients, in murine cartilage of the spontaneous osteoarthritis mouse model (STR/ort) and cartilage from young (14 weeks) and old (>12 months) C57Black/6 mice. To study the functional consequences of decreased level of P65 in chondrocytes the murine H4 chondrocyte-cell line was stably transduced with a lentivirus expressing a short-hairpin RNA against P65 to reduce the P65 protein levels by a RNA interference approach. We selected several cell lines that expressed different amounts of P65 protein. To study the biological consequences, IL-1b, TNFa or conditioned medium of OA synovium was added to the murine chondrocyte-cell line with the lowest P65 level. Cell death was measured by Facs analysis of 7-AAD positive cells. In conditioned medium of OA synovium, levels of TNFa were measured by Luminex. P65 and an apoptotic marker (Caspase-3 active form) were detected by immunohistochemistry in serial paraffin sections of human OA cartilage explants from 6 different patients (410 section each patient). To study a relation between P65 in chondrocytes, synovial TNFa production and chondrocyte cell death, in 6 OA patients levels were detected in the same joint.
A subset of OA patients showed low P65 levels (5 out of 10) compared to RA. In STR/ort mice levels were diminished more than 75% when joints became affected. Levels of P65 in normal young and old mice were equal. In vitro, adding TNFa to selected chondrocyte cell lines caused only cell death in the cell-line with low levels of P65. IL-1 had no effect on cell viability. Challenging the cell line with low P65 levels with conditioned medium from synovial tissue resulted in more than 60% chondrocyte death in 3 of the 5 OA patients. Synovial TNFa production was detected in the same samples that caused cell death. In human OA cartilage, chondrocyte P65 showed a significant negative correlation with Caspase-3 (an apoptotic marker) staining. Analyzing all parameters of a single joint in 6 OA patients showed that the combination of low endogenous P65 levels in cartilage and synovial TNF production caused chondrocyte cell death.
This study clearly demonstrated that low levels of P65 makes chondrocytes more vulnerable for TNFa, a cytokine which can be produced by the synovial tissue during OA, and that this transcription factor is downregulated in chondrocytes in murine OA and a considerable portion of OA patients.
To cite this abstract, please use the following information:
Arntz, Onno J., Vermeij, Eline A., Davidson, Esmeralda N. Blaney, Abdollahi-Roodsaz, Shahla, Bennink, Miranda B., van der Kraan, Peter M., et al; Chondrocytes in a Subset of Osteo-Arthritic Patients with Reduced Nuclear FactorkappaB-p65 Levels Are Vulnerable to Apoptosis Induced by Tumor Necrosis Factor Alpha, Which Could Be Released by the Synovium. [abstract]. Arthritis Rheum 2011;63 Suppl 10 :1785