inositol-1-4-5-trisphosphate and Arthritis--Rheumatoid

Peripheral blood (PB) T cells from rheumatoid arthritis (RA) patients proliferate poorly to mitogen, a change that is related to decreased intracellular Ca2+ ([Ca2+]i) signaling after T cell receptor (TCR) stimulation. We hypothesized that this was, in part, due to the effect of mediators of inflammation and predicted that greater changes in [Ca2+]i signaling would be seen in synovial fluid (SF) T cells. We also examined the mechanisms underlying the altered [Ca2+]i signals.. Paired PB and SF T cells from patients with chronic inflammatory arthritis were stimulated with mitogen to assess the magnitude of the [Ca2+]i signal in cell populations by fluorometry, the pattern of the [Ca2+]i signal in individual cells in a single-cell ion-imaging system, and the spatial distribution of Ca2+ within intracellular organelles.. There was a significantly smaller [Ca2+]i signal after phytohemagglutinin protein stimulation of SF T cells (peak rise in [Ca2+]i signal PB versus SF 200 nM versus 180 nM; P < 0.05). In single SF T cells, a change in the pattern of the [Ca2+]i signal and a reduction in the number of responding cells was seen. These changes were a magnification of those seen in RA PB compared with control PB T cells. The contribution of Ca2+ release from intracellular stores to the final [Ca2+]i signal in PB and SF T cells was equal, but there was a significant increase in the Ca2+ remaining in the endoplasmic reticulum (ER) in SF T cells after TCR activation (PB versus SF 6 nM versus 19 nM; P < 0.05). Non-ER Ca2+ stores were not similarly affected.. We found abnormalities in the magnitude, pattern, and spatial distribution of [Ca2+]i signaling in T cells from SF of patients with chronic inflammatory arthritis. A reduction in the number of responding SF T cells may partly explain some of our observations. However, we propose that the observed redistribution of SF Ca2+ stores may underlie the altered [Ca2+]i signaling, thus making these cells hyporesponsive to mitogen. The inflammatory environment of the joint and the late stage of differentiation of SF T cells are both likely to contribute to these changes in [Ca2+]i signaling, resulting in aberrant T cell function and promotion of disease chronicity.

Topics: Arthritis, Rheumatoid; Calcium; Calcium Signaling; Chronic Disease; Endoplasmic Reticulum; Enzyme Inhibitors; Humans; Inositol 1,4,5-Trisphosphate; Intracellular Membranes; Ionomycin; Ionophores; Organelles; Reference Values; Synovial Fluid; T-Lymphocytes; Thapsigargin

In the present study, we demonstrate that the alpha2-macroglobulin (alpha2M) signaling receptor is up-regulated on rheumatoid synovial fibroblasts. In rheumatoid cells, 125I-alpha2M-methylamine bound to two sites; namely, one of high affinity (Kd approximately 52 pM) and the second of lower affinity (Kd approximately 9.7 nM). In normal synovial fibroblasts only one site for 125I-alpha2M-methylamine (Kd approximately 5.36 nM) was present. Receptor-associated protein did not inhibit the binding of alpha2M-methylamine to the high affinity binding sites, but it caused a 70-80% reduction in its binding to low affinity binding sites establishing its identity as the low density lipoprotein receptor-related protein/alpha2M receptor. Binding of alpha2M-methylamine to rheumatoid but not normal synovial fibroblasts caused a rapid rise in inositol 1,4,5-trisphosphate synthesis with a peak reached within 10 s of ligand exposure. Concomitantly, rheumatoid but not normal cells showed a rise in intracellular Ca2+. Pretreatment of rheumatoid cells with Receptor-associated protein or pertussis toxin did not affect the alpha2M-methylamine-induced increase in intracellular Ca2+. These are characteristic properties of ligation by alpha2M-methylamine of the alpha2M signaling receptor but not the lipoprotein receptor-related protein/alpha2M receptor. Binding of alpha2M-methylamine to rheumatoid synovial fibroblasts significantly increased the synthesis of DNA compared with normal synovial fibroblasts treated similarly.

Topics: alpha-Macroglobulins; Arthritis, Rheumatoid; Calcium; Cell Division; Cells, Cultured; Fibroblasts; Humans; Inositol 1,4,5-Trisphosphate; Low Density Lipoprotein Receptor-Related Protein-1; Platelet Activating Factor; Receptors, Immunologic; Signal Transduction; Synovial Membrane; Up-Regulation