Proinflammatory cytokines (IL-15, TNFα, IL-6 and IL-1β) in rheumatoid arthritis bone marrow preferentially promote activation of T-cells

Abstract

Objective 1. To compare the real T cell numbers in bone marrow isolated from RA and OA patients; 2. To measure the levels of soluble IL-15, TNFα, IL-6, IL-1β and IL-8 in bone marrow plasma; 3. To analyze the expression of IL-15Rα, CD25 and CD69 on T cells in bone marrow; 4. To measure the levels of soluble TNFα, IL-6 and IL-17 in cultured bone marrow mononuclear cells stimulated by IL-15; 5. To measure levels of IL-15 in bone marrow stromal cells. Methods Bone marrow samples were obtained from patients undergoing joint replacement surgery. Levels of IL-15, TNFα, IL-6, IL-1β, IL-17 and IL-8 were measured using specific ELISAs. The real number of lymphocytes stained for CD3 +, CD4 +, CD8 + were counted in the presence of TruCount beads using FACS. Surface expression of IL-15Rα, CD25, CD69 and intracellular IL-15 was evaluated by FACS. Results Bone marrow from RA patients contained double CD3 + T-cells in comparison to OA (6.1±2.8 vs. 3.2±1.6 x10 6cells/ml, p=0.008). There were elevated levels of IL-15, TNFα, IL-6, IL-1β in bone marrow plasma from RA in comparison to OA patients. OA (pg/ml) RA (pg/ml) p IL-15 760±238.7 1304.5±956.3 0.008 TNFα 435.6±125.9 828.6±567 0.04 IL-1 388±148.8 2787±2838 0.03 IL-6 257.5±178 1335.4±1211 0.04 Lymphocytes obtained from RA patients expressed higher levels of IL-15R alpha (p=0.01). Expression of CD69 was significantly higher on both CD4 + (p<0.04) and CD8 + (p<0.02) T cells isolated from RA bone marrow. The percentage of CD4+CD25+ cells was higher in RA bone marrow (p<0.02). RA BMMC produced higher level of IL-17 after stimulation by IL-15 than OA BMMC. Bone marrow stromal cells produce high levels of IL-15. Conclusion It is likely that locally overproduced IL-15 (and other proinflammatory cytokines) are responsible for activation and proliferation of T-cells reflected by significantly increased number of activated T-cells in RA bone marrow.