UAH - CSIC Study of the activation and proliferation of T lymphocytes from patients with Rheumatoid Arthritis by enumeration microparticles San Juan, Carlos; Elkhader, Ismail; Martín, Mercedes INTRODUCTION Rheumatoid arthritis (RA) is an autoimmune polifactorial disease whose characteristic presentation is a chronic polyarthritis that can evolve towards destruction and often joint crippling deformity. Rheumatoid synovitis is characterized by infiltration of CD4 T lymphocytes (especially CD45RO), CD8 T and macrophages. During the last decade there has been investigations about the hypothesis that it gives CD4 T cells and antigen presenting cells (CPA) a preponderant role. We use the technique of flow cytometry cell enumeration, which is based on the coadquisición of a known number of microparticles with cells that we want to study, setting a ratio of both events, in order to determine the concentration of cells in real time acquisition. HYPOTHESIS: There is a worse response to the stimuli of proliferation (TCE, IL2) of CD4 T cells in the patient than in the control because in the patient there is an increased number of activated T cells previously MATERIALS AND METHODS: For the study PBMCs was isolated from a control and from a patient by Ficoll, then a study immunophenotype by flow cytometry using four-color markers CD45ROFITC, CD25PE, CD3PerCp and CD4APC. (figure 1) RESULT: We note that in general TCE and IL2 stimuli respectively produce a low and relative activation and proliferation of lymphocytes, however, the combination of the two stimuli produces a strong response in both the control and the patient, without showing some differences between both. As for the level of activation of T lymphocytes we can appreciate a larger number of CD4+CD25+ and CD8+CD25+ T lymphocytes at basal time in RA patients, compared to the controls. In turn the behavior of these two subpopulations to stimuli differs between both study groups. In the case of CD4+CD25+ T lymphocytes, there is a better proliferative response to stimuli in patients than in controls. While unlike the CD8+CD25 + T lymphocytes have a more difficult response to the same stimuli. CONCLUSION: Patients of RA have a high degree of activation in T lymphocytes, which may explain the different proliferative response to stimuli polyclonal "in vitro“ especially is lower in limphocytes T8 CD25+ for the prompts IL-2 and IL-2 + TCE. The study was carried out by the technique of cell enumeration in basal time and after 5 days of culture. The cells are arranged in culture in a 250,000 cells / ml in a 96-well plate with U bottom, at 37 º C and 5% CO2, with the following experimental conditions: complete medium, TEC, TEC + IL2 and IL2. In this paper we show the results of the flow cytometer for counting and enumeration for linfocytes, T4 cells and lymphocytes T8. Graphics total lynfocytes, in the graphics for complet medium and TEC can see a downward trend in the proliferation in control and patient Similarly, the graphs TEC + IL2 and IL2 can obserbar an upward trend in the proliferation, the trend remains lower Upward in the patient's lymphocytes Graphic 2 T4 CD25 + lymphocytes, the graphs for complet medium and TCE show a trend towards a decline in control and patient, in contrast to TCE + IL-2 and IL-2 can be seen a sharp upward trend Graphics 3 T8 CD25 + lymphocytes, in graphic medium complet and TCE are declining trend more pronounced for the patient, for TCE + IL-2 and IL2 there is a big change in control proliferation in the proliferation is lower for the patient CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT CONTROL PATIENT Figure 1: FSC- SSC blot plot; Gabe Statistic table and CD3-CD4 blot plot