PMC

The synovium in RA is marked by intimal lining hyperplasia, infiltrating mononuclear cells in the sublining, and occasional lymphoid aggregates. From Reference ().

Immunohistochemistry was performed on synovial tissue from patients with (A) rheumatoid arthritis (RA) and (B) osteoarthritis (OA) to detect p53 protein. p53 was detected in the intimal lining and sublining infiltrating leukocytes. The p53 protein expression is significantly higher in the RA synovial tissue compared with OA synovium. From Reference ().

Cultured FLS were cultured in an artificial matrix and cultured for 3 weeks. The three-dimensional structure was fixed with 2% paraformaldehyde, embedded in paraffin and sections were stained with hematoxylin and eosin. FLS migrated to the surface of the matrix to establish a lining layer at the matrix-liquid interface (A). Immunohistochemistry was performed with cadherin-11 specific antibody and isotype control (B). Cadherin-11 specific staining was localized in the lining layer. From Reference ().

FLS derived from normal or rheumatoid synovial tissue were transduced with human papilloma virus type 18 (HPV-18) encoding E6 gene, which inactivates p53. The FLS were co-implanted with cartilage into SCID mice and harvested 60 days later. RA FLS (A) adhered to and invaded the matrix compared with normal FLS (C) which did not invade. Cartilage invasion was increased when RA FLS was transduced with HPV-18 encoding E6 gene (B). The normal FLS (C) acquired an RA-like invasive phenotype when p53 protein was inactivated with E6 (D). From Reference ().

(A) RA FLS were transfected with pCEP4, hemagglutinin-tagged, full-length PUMA expression vector (HA-PUMA), or HA-tagged PUMA expression vector with a deletion of the Bcl-2 homology 3 domain (HA-PUMAdBH3). Data are presented as the percentage of nonviable cells. HA-PUMA-transfected FLS showed significantly more dead cells compared with pCEP4- or HA-PUM-AdBH3-transfected cells (*P < 0.01). (B) Comparison of apoptosis induced by pCEP4, HA-PUMA, or HA-PUMAdBH3 in RA and OA FLS lines. The extent of PUMA-induced apoptosis was the same in both cell lines. *P < 0.05 compared with controls. (C) RA FLS were transfected with pCEP4, HA-PUMA, or HA-PUMAdBH3 and DNA fragmentation, another measure of apoptosis, was measured by ELISA. Significant induction of DNA fragmentation was noted in HA-PUMA-transfected cells compared with pCEP4-transfected cells (*P < 0.05). (D) FLS transfected with pCEP4, HA-PUMA, or HA-PUMAdBH3 were cultured in chamber slides. The cells were then immunostained for activated caspase 3. HA-PUMA-transfected FLS showed significantly more activated caspase 3-positive cells compared with pCEP4- or HA-PUMAdBH3-transfected cells (P < 0.01). From reference ().

MMPs gene expression: After TLR-3 receptor stimulation by synthetic ligand poly (I:C), two independent pathways are activated, MKK7/JNK and IKKε/TBK1. Both of these complexes phosphorylate c-Jun, leading to increased AP-1 activation. JNK-mediated c-Jun activation and AP-1-mediated gene expression of MMPs require MKK7. IKKε, independent of JNK can also phosphorylate c-Jun after forming a complex with TBK1 and NAP1. Type I IFN gene expression: Transcriptional activation of type I IFN genes require formation of an enhanceosome, an interaction between transcription factors ATF2/c-Jun and IRF3. In FLS, in response to TLR-3 activation, IRF3 serves as a substrate for both JNK and IKKε, followed by phosphorylation, dimerization and translocation to the nucleus. JNK-mediated IRF3 activation and IFN gene expression requires MKK7. MKK4 in complex with JNK also contributes to type I IFN gene expression through ATF2 phosphorylation and formation of ATF2/c-Jun complex. TLR3, Toll-like receptor; MKK4, MAPK kinase 4; MKK7, MAPK kinase 7; JNK, c-Jun N-terminal kinase; TBK1, TANK-binding kinase; NAP1, NAK-associated protein-1; IKKe, IKK-related kinase; IRF3, interferon regulatory factor 3; c-JUN, component of AP-1 transcription factor; ATF2, activated transcription factor 2; MMPs, matrix metalloproteinases; Type I IFNs, interferon α, β and interferon-stimulated genes.