Predominant activation of MAP kinases and pro-destructive/pro-inflammatory features by TNF alpha in early-passage synovial fibroblasts via TNF receptor-1: failure of p38 inhibition to suppress matrix metalloproteinase-1 in rheumatoid arthritis.

OBJECTIVE: To examine the relative importance of tumour necrosis factor-receptor 1 (TNF-R1) and TNF-R2 and their signalling pathways for pro-inflammatory and pro-destructive features of early-passage synovial fibroblasts (SFB) from rheumatoid arthritis (RA) and osteoarthritis (OA). METHODS: Cells were stimulated with tumour necrosis factor (TNF)alpha or agonistic anti-TNF-R1/TNF-R2 monoclonal antibodies. Phosphorylation of p38, ERK and JNK kinases was assessed by western blot; proliferation by bromodesoxyuridine incorporation; interleukin (IL)6, IL8, prostaglandin E(2) (PGE(2)) and matrix metalloproteinase (MMP)-1 secretion by ELISA; and MMP-3 secretion by western blot. Functional assays were performed with or without inhibition of p38 (SB203580), ERK (U0126) or JNK (SP600125). RESULTS: In RA- and OA-SFB, TNFalpha-induced phosphorylation of p38, ERK or JNK was exclusively mediated by TNF-R1. Reduction of proliferation and induction of IL6, IL8 and MMP-1 were solely mediated by TNF-R1, whereas PGE(2) and MMP-3 secretion was mediated by both TNF-Rs. In general, inhibition of ERK or JNK did not significantly alter the TNFalpha influence on these effector molecules. In contrast, inhibition of p38 reversed TNFalpha effects on proliferation and IL6/PGE(2) secretion (but not on IL8 and MMP-3 secretion). The above effects were comparable in RA- and OA-SFB, except that TNFalpha-induced MMP-1 secretion was reversed by p38 inhibition only in OA-SFB. CONCLUSION: In early-passage RA/OA-SFB, activation of MAPK cascades and pro-inflammatory/pro-destructive features by TNFalpha is predominantly mediated by TNF-R1 and, for proliferation and IL6/PGE(2) secretion, exclusively regulated by p38. Strikingly, RA-SFB are insensitive to p38 inhibition of MMP-1 secretion. This indicates a resistance of RA-SFB to the inhibition of pro-destructive functions and suggests underlying structural/functional alterations of the p38 pathway, which may contribute to the pathogenesis or therapeutic sensitivity of RA, or both.

Genetic profile of bone metastases in renal cell carcinoma.

OBJECTIVE: The aim of this study was to define specific genetic alterations which are common in bone metastases in renal cancer patients. METHODS: Tumor DNA from 31 metastases and 13 related primary tumors was extracted from paraffin embedded tissue sections. DOP-PCR was performed to amplify the whole DNA. After labelling by PCR, CGH was performed according to standard protocols. RESULTS: The mean number of aberrations per metastasis was 6.3 (1-13). Losses of chromosomes 3p (76%), 6 (20%), 8p (20%), 9 (34%), 14q (27%) and 18 (20%) as well as gains of chromosomes 5 (45%), 8q (34%) and 17 (27%) were detected frequently. Thirteen related primary tumors were also investigated. In 7 cases, at least one identical alteration was found in both primary tumor and metastases. In these cases, the number of alterations was mostly higher in primary tumors than in metastases without statistical significance. However, in general, the frequency of alterations was higher in metastases. CONCLUSIONS: Bone metastases from renal cell carcinoma are characterized by typical genetic alterations. Changes leading to metastasizing happen early in tumor pathogenesis. However, further accumulation of genetic changes occurs in metastases leading to a more complex genetic pattern which might be necessary for progression to clinically relevant metastases.

Synovial fibroblasts and synovial macrophages from patients with rheumatoid arthritis and other inflammatory joint diseases show chromosomal aberrations.

Chromosomal aberrations were investigated in nuclei extracted from synovial tissue and first-passage synovial fibroblasts (P-1 SFB, 98% enrichment) or macrophages (P-1 Mphi) from patients with rheumatoid arthritis (n=10). The findings were compared with those in other rheumatic diseases (osteoarthritis, n=14; reactive arthritis, n=1), as well as with those in chronic obstructive pulmonary disease (n=8). Controls were paired peripheral blood lymphocytes from arthritic patients, synovial tissue or SFB/Mphi from joint trauma/normals (n=9), and peripheral blood monocytes from normal donors (n=10). GTG banding of metaphase chromosomes and interphase fluorescence in situ hybridization with centromere-specific probes were used. Comparable chromosomal aberrations were observed in synovial tissue and P-1 SFB of patients with rheumatoid arthritis, osteoarthritis, and reactive arthritis (polysomy 7 and aneusomies of chromosomes 4, 8, 9, 12, and 18). Notably, aneusomies of chromosomes 4, 6, 7, 8, 9, 11, 12, and/or X were also detected in P-1 synovial Mphi from rheumatoid arthritis (90% of the cases), osteoarthritis (93%), and reactive arthritis (1/1), as well as bronchial Mphi from chronic obstructive pulmonary disease (25%). No aberrations were detected in paired peripheral blood lymphocytes (except for one osteoarthritis case with a karyotype 45,X[10]/46,XX[17]), or in peripheral blood monocytes and synovial tissue of normals/joint trauma. Because Mphi aberrations were common to chronic joint and pulmonary disease, chronic inflammatory stress may induce chromosomal aberrations with potential functional relevance in local mesenchymal cells and infiltrating leukocytes in an organ-independent fashion.