Disruption of rhythms of molecular clocks in primary synovial fibroblasts of patients with osteoarthritis and rheumatoid arthritis, role of IL-1β/TNF

Introduction: Circadian rhythms play an important role in the body and in single cells. Rhythms of molecular clocks have not been investigated in synovial fibroblasts (SF) of patients with osteoarthritis (OA) and rheumatoid arthritis (RA). The study was initiated to fill this gap and to study effects of interleukin (IL)-1 beta/tumor necrosis factor (TNF) on rhythmicity in synovial fibroblasts of RA and OA patients. Methods: The presence of BMAL-1, CLOCK, Period 1 and Period 2 proteins in synovial tissue was investigated by immunofluorescence. The presence of mRNA of molecular clocks was studied during 72 h by qPCR. Characteristics of rhythms were studied with time series analysis. Results: BMAL-1, CLOCK, Period 1 and Period 2 proteins were abundantly present in synovial tissue of OA, RA and controls. Receiving synovial tissue at different operation time points during the day (8:00 am to 4:00 pm) did not reveal a rhythm of BMAL-1 or Period 1 protein. In OASF and RASF, no typical rhythm curve of molecular clock mRNA was observed. Time series analysis identified a first peak between 2 and 18 hours after synchronization but a period was not detectable due to loss of rhythm. TNF inhibited mRNA of CLOCK, Period 1 and Period 2 in OASF, while IL-1 beta and TNF increased these factors in RASF. This was supported by dose-dependently increased levels in MH7A RA fibroblasts. In RASF, IL-1 beta and TNF shifted the first peak of BMAL-1 mRNA to later time points (8 h to 14 h). Conclusion: Rhythmicity is not present in primary OASF and RASF, which is unexpected because fibroblasts usually demonstrate perfect rhythms during several days. This might lead to uncoupling of important cellular pathways.