IntroductionThe purpose of this study was to investigate the profile of histone deacetylase (HDAC) expression in the synovial tissue of rheumatoid arthritis (RA) compared with that of normal control and osteoarthritis (OA), and to examine whether there is a link between HDAC activity and synovial inflammation.MethodsHDAC activity and histone acetyltransferase (HAT) activity were determined in nuclear extracts of total synovial tissue surgically obtained from normal, OA and RA joints. The level of cytoplasmic tumor necrosis factor a (TNFα) fraction was measured by ELISA. Total RNA of synovial tissue was used for RT-PCR of HDAC1-8. In synovial fibroblasts from RA (RASFs), the effects of TNFα on nuclear HDAC activity and class I HDACs (1, 2, 3, 8) mRNA expressions were examined by quantitative real-time PCR. The protein expression and distribution of class I HDACs were examined by Western blotting.ResultsNuclear HDAC activity was significantly higher in RA than in OA and normal controls and correlated with the amount of cytoplasmic TNFα. The mRNA expression of HDAC1 in RA synovial tissue was higher than in OA and normal controls, and showed positive correlation with TNFα mRNA expression. The protein level of nuclear HDAC1 was higher in RA synovial tissue compared with OA synovial tissue. Stimulation with TNFα significantly increased the nuclear HDAC activity and HDAC1 mRNA expression at 24 hours and HDAC1 protein expression at 48 hours in RASFs.ConclusionsOur results showed nuclear HDAC activity and expression of HDAC1 were significantly higher in RA than in OA synovial tissues, and they were upregulated by TNFα stimulation in RASFs. These data might provide important clues for the development of specific small molecule HDAC inhibitors.

The cellular mechanism by which TNF-alpha inhibits osteoblastic differentiation induced by BMPs was investigated using mouse myoblast C2C12 cells expressing functional BMP receptors and Smad signaling molecules except ALK-6. Osteoblast transformation in response to BMP-2 was morphologically suppressed by TNF-alpha. Expression of biological markers for osteoblasts including Runx2 and osteocalcin, alkaline phosphatase activity, and parathyroid hormone (PTH) responsiveness shown by PTH-induced cAMP production were readily activated by BMP-2, -4, -6, and -7. The BMP-induced osteoblastic phenotype was dose-dependently inhibited by TNF-alpha. BMP-induced Smad1,5,8 phosphorylation of C2C12 cells was suppressed by TNF-alpha signaling. In addition, cDNA array analysis showed an increased expression of inhibitory Smad6 by TNF-alpha. MAP kinase analysis showed that ERK1/ERK2 and SAPK/JNK phosphorylation were selectively activated by TNF-alpha regardless of the presence of BMP ligands. BMPs had no effect on expression levels of TNF type 1 and 2 receptors. Notably, inhibition of SAPK/JNK restored TNF-alpha effects on BMP-induced osteoblast differentiation demonstrated by Id-1-promoter activity as well as Runx2 and osteocalcin mRNA levels. Collectively, TNF-alpha elicits BMP-induced osteogenic inhibition by suppressing BMP-Smad signaling pathway, at least in part, through SAPK/JNK activation and Smad6 upregulation.

Objective. The accumulation of advanced glycation end products (AGEs), S100A12, and high mobility group box chromosomal protein 1 has been associated with joint inflammation in rheumatoid arthritis (RA). This study was undertaken to determine the induction of the receptor for these proteins, termed receptor for AGEs (RAGE), in synovial tissue (ST) macrophages from RA patients.Methods. RAGE and CD68 expression in ST were determined by 2-color immunofluorescence labeling. Cell surface and messenger RNA (mRNA) expression of RAGE were examined by flow cytometry and reverse transcriptase-polymerase chain reaction (PCR) or realtime PCR, respectively.Results. CD68؉ lining macrophages, like the vasculature, expressed high levels of RAGE in inflamed ST from RA patients. RAGE mRNA expression was significantly higher in RA ST than in ST from patients with osteoarthritis. RAGE mRNA levels were significantly higher in ST macrophages and normal endothelial cells than in ST CD4؉ T cells and synovial fibroblasts stimulated with tumor necrosis factor ␣ and interleukin-1␤ (IL-1␤). Cell surface RAGE was highly induced on normal monocytes after a 24-hour incubation with a 20% concentration of RA ST cell culture supernatants. RAGE mRNA expression in adherent monocytes was augmented by various cytokines, most potently by IL-1␤.Conclusion. These results indicate that RAGE overexpression in lining macrophages may be induced, at least in part, by cytokines such as IL-1, leading to the amplification of inflammatory responses mediated by RAGE ligands that are abundant in RA joints.

C-reactive protein (CRP) is one of the strongest independent predictors of cardiovascular disease. We have previously reported that oxidized LDL (oxLDL) interacts with b2-glycoprotein I (b2GPI), implicating oxLDL/b2GPI complexes as putative autoantigens in autoimmune-mediated atherosclerotic vascular disease. In this study, we investigated the interaction of CRP with oxLDL/b2GPI complexes and its association with atherosclerosis in patients with diabetes mellitus (DM). CRP/oxLDL/b2GPI complexes were predominantly found in sera of DM patients with atherosclerosis. In contrast, noncomplexed CRP isoforms were present in sera of patients with acute/chronic inflammation, i.e., various pyrogenic diseases, rheumatoid arthritis (RA), and DM. Immunohistochemistry staining colocalized CRP and b2GPI together with oxLDL in carotid artery plaques but not in synovial tissue from RA patients, strongly suggesting that complex formation occurs during the development of atherosclerosis. Serum levels of CRP correlated with soluble forms of intercellular adhesion molecule-1 and vascular cell adhesion molecule-1, and oxLDL/b2GPI complexes correlated with total cholesterol and hemoglobin A1c. Thus, the generation of CRP/oxLDL/b2GPI complexes seems to be associated with arterial inflammation, hyperglycemia, and hypercholesterolemia. CRP/oxLDL/b2GPI complexes can be distinguished from pyrogenic noncomplexed CRP isoforms and may represent a more specific and predictive marker for atherosclerosis.-Tabuchi, M., K.

Despite its potent ability to inhibit proinflammatory cytokine synthesis, interleukin (IL