Dataset Information

Interaction between bovine IL-1-stimulated cartilage and human serum, LC-MS/MS

ABSTRACT: Joint diseases are often characterized by inflammatory processes resulting in pathological changes in joint tissues, including cartilage degradation and release of components to the synovial fluid. The complement system plays a central role in promoting the inflammation. Since several cartilage proteins are known to interact with complement, causing either activation or inhibition of the system, we aimed to investigate these interactions comprehensively. Bovine cartilage explants were cultured with interleukin-1alpha (IL-1a) to induce cartilage degradation, followed by incubation with human serum to allow interactions with complement. Label-free selected reaction monitoring (SRM) mass spectrometry (MS) was then used to specifically quantify complement proteins interacting with the cartilage explant. In parallel, the time-dependent degradation of cartilage was detected using tandem MS (MS/MS). Complement proteins resulting from activation of the classical and alternative pathway as well as the terminal pathway were detected on IL-1a stimulated cartilage at time points when clear alterations in the extracellular matrix composition had occurred. To confirm SRM results indicating complement activation, increased levels of the complement activation product C4d were detected by ELISA in serum after incubation with IL-1a stimulated cartilage. Further, typical activated (cleaved) C3 fragments were detected by western blotting of urea extracts of IL-1a stimulated cartilage. No complement activation was triggered by cartilage cultured in the absence of IL-1a. Components released from IL-1a stimulated cartilage during culture had an inhibitory effect on complement activation. These were released after a longer incubation period with IL-1a and may represent a feedback reaction to cartilage-triggered complement activation observed after a shorter incubation period.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human) Bos Taurus (bovine)

TISSUE(S): Cartilage, Blood Serum

SUBMITTER: Camilla Melin Fürst