gms | German Medical Science

Objectives: Sublytic deposition of the terminal complement complex (TCC) has previously been associated with an inflammatory and catabolic response in chondrocytes and thus osteoarthritis (OA) progression. As chondrocyte senescence is considered as one of the major phenotypical alterations in OA, we investigated whether TCC deposition contributes to stress-induced premature senescence (SISP) of chondrocytes after ex vivo injury or during in vivo aging.

Methods: Human cartilage was obtained from OA patients undergoing surgical knee replacement. Explants were isolated from macroscopically intact (OARSI grade ≤1) or highly degenerated cartilage (OARSI grade≥3). TCC was detected by means of IHC. After mechanical impact (0.59 J; applied by a drop tower), OARSI grade≤1 explants were cultured with/ without 30% human serum (HS). TCC deposition was prevented by heat-inactivation of HS (HI) or addition of its inhibitor clusterin (CLU). After 4 d, gene expression of senescence-associated (SA) and SA secretory phenotype (SASP) markers (CDKN1A, CDKN2A, IL6, MMP13) was assessed by qRT-PCR. Unimpacted cartilage served as control (C). Isolated chondrocytes were used to analyze ß-galactosidase (ß-gal) activity. To study age-dependent OA, knee joints of 80-week-old CD59-KO mice (lack of TCC inhibitor CD59) and C6-deficient mice (insufficient TCC formation) were analyzed by qRT-PCR. Cartilage integrity was assessed by Safranin-O stained knee sections. Data sets (n≥ 5 donors/group) were analyzed by means of one-way ANOVA or t-test.

Results and conclusion: Compared to macroscopically intact tissue, gene expression of CDKN1A and CDKN2A was significantly higher in OARSI grade≥3 tissue (3.1-fold, p=0.002 and 4.7-fold, p=0.02). Moreover, TCC deposition was remarkably enhanced. Exposure to HS after trauma revealed additive effects on the mRNA levels of CDKN1A (2.9-fold, p ≤0.0001), CDKN2A (5.7-fold, p=0.001), MMP13 (4.1-fold, p≤ 0.0001), and IL6 (23-fold, p=0.003) as compared to unimpacted tissue. HS-mediated effects could be significantly reduced by CLU. Further, HS exposure significantly increased the number of ß-gal-positive chondrocytes ([vs C] +79.2%, p≤ 0.0001), which was less pronounced in case of HI ([vs HS] -30%, p=0.01). In the murine aging model, we found significantly lower gene expression of cdkn1a (-3.3-fold, p=0.006) and cdkn2a (-1.8-fold, p=0.05) in C6-deficient mice as compared to the CD59-KO group. Gene expression of mmp13 was decreased by trend (-30%, p=0.33). First histological analysis confirmed that OA-changes, such as PG loss and hypocellularity, were more pronounced in CD59-KO mice.

The present results imply a clear association between TCC deposition and a SISP-like phenotype in chondrocytes in the human ex vivo cartilage trauma model as well as in aged CD59-KO mice. We suggest that sublytic TCC deposition leads to cellular stress, thus inducing SIPS in chondrocytes. Therefore, TCC might represent a potential target, in order to reduce phenotypical alteration of surviving chondrocytes.