gms | German Medical Science

Objectives: Surface-active phospholipids (PLs), hyaluronan and lubricin were reported to provide boundary lubrication within human articular joints. The PLs are thought to be synthesized and released by fibroblast-like synoviocytes (FLS). We recently published two lipidomic studies about 103 PLs species with altered levels in synovial fluid of late osteoarthritis (OA) patients when compared to normal. Thus, we hypothesize that the biosynthesis of PLs by FLS during OA is regulated by cytokines and growth factors. The goal of our study was to develop an in vitro model to investigate the biosynthesis of PLs by osteoarthritic FLS as modulated by cytokines and growth factors.

Methods: FLS from 6 OA patients were cultured in supplemented D-MEM with 5% human lipoprotein deficient serum. [D9]-choline chloride and [D4]-ethanolamine were used to label PLs. In order to analyze sufficiently labelled PLs and to minimize re-uptake, short labelling periods between 4 to 48 hours were first evaluated. The effects of IL-1β , TNFα , TGF-β , IGF-I, dexamethasone and quinacrine were screened. Lipids were extracted according to Bligh and Dyer, quantified using electrospray ionization tandem mass spectrometry, and normalized to protein content. Data were analysed using paired t-test. Statistical significance was assumed at p<0.05. The present study was approved by the ethical review committee of our university.

Results and Conclusion: Our new experimental model allows the determination of the following labelled PLs classes: Phosphatidylcholine (PC), lysophosphatidylcholine (LPC), sphingomyelin (SPM), phosphatidylethanolamine (PE), and plasmalogen (Plasm). Increasing the time of labelling markedly elevated the percentage of labelled PLs. 10 ng/ml IL-1β and 100 ng/ml TNFα as well as 10 ng/ml TGF-β and 100 ng/ml IGF-I each significantly stimulated the biosynthesis of individual PL classes by 1.2 to 1.4-fold. Interestingly, 10µM dexamethasone decreased the biosynthesis of individual PL classes by 0.7 to 0.9-fold whereas 5µM of the PLA₂-inhibitor quinacrine elevated the synthesis of individual PL classes by 1.4 to 2.0-fold.

Our data allowed us to establish a novel biosynthesis model to study the effects of various agents on the de novo biosynthesis of PLs by FLS. Cytokines and growth factors were found to enhance the biosynthesis of PLs by 1.2 to 2-fold. We previously reported an elevated level of PLs in synovial fluid during late stage OA compared to normal with PC being increased by 5.4-fold, LPC by 4.8-fold, SPM by 4.4-fold, PE by 4.2-fold, and Plasm by 3.9-fold. Our data suggest that these elevated levels of PLs are not only locally produced by FLS but might also origin from blood by diffusion. Our ongoing research is designed to focus on experiments to thoroughly investigate the origin of PLs in SF as well as the metabolic control of PL biosynthesis and release by FLS.