Vitamin D Effect in Rheumatoid Arthritis.

Regulatory T (Tregs) cells play an important role in the maintenance of immunological tolerance. It decrease in the peripheral blood of rheumatoid arthritis patients. Vitamin D has an immunomodulatory and anti-inflammatory effect in rheumatoid arthritis. Vitamin D supplementation significantly enhances Tregs percentage in the peripheral blood of RA patients. So supplementation of Vit D improves rheumatoid arthritis disease activity.

Background: Regulatory T cells (Tregs) play an important role in the maintenance of immunological tolerance, so Tregs deficiency or decrease suppressor functions may be associated with development of autoimmune diseases. Vitamin D is essential for normal bone mineralization and growth, prevention of osteopenia, osteoporosis, and nonspecific painful musculoskeletal conditions . Vitamin D is thought to have an immunomodulatory and anti-inflammatory actions, as its receptors are widely expressed in peripheral mononuclear blood cells, also its deficiency associated with several autoimmune disorders, including rheumatoid arthritis Methods: 40 patients with active RA were randomly assigned into two groups. Group I received MTX plus hydroxychloroquine, group II received MTX and hydroxychloroquine plus vitamin D supplementation for 3 months, in addition to 30 healthy volunteers as control group. Peripheral blood Tregs were measured by Flow Cytometry. Statistical Analysis. The collected data analyzed by SPSS software (version 16). The range, mean and standard deviation were calculated for quantitative variables. Categorical variables were expressed as number and percentages; Chi square was used as a test of their significance. Skewness, kurtosis; Shapiro-Wilk, and The Kolmogorov-Smirnov tests were used to test the normality for the data. The difference between two means was analyzed using the students (t) test (paired and unpaired samples- T tests). Significance was considered at p<0.05.

4 ml of heparinized blood from the subjects' peripheral blood was collected. Human peripheral blood natural regulatory T cells (Tregs) were surface stained with Mouse Anti-Human CD4 FITC (Fluorescein) conjugated Monoclonal Antibody and Mouse Anti-Human IL-2 Rα/CD25 APC (allophycocyanin) conjugated Monoclonal Antibody, followed by intracellular staining using Rabbit Anti-Human/ Mouse FoxP3 PE (phycoerythrin) conjugated Antigen Affinity-purified Monoclonal Antibody, cells were fixed and permeabilized with FoxP3 Fixation & Permeabilization Buffer Kit. Cells were gated on lymphocytes.

Inclusion Criteria: disease duration less than 3 years from the time of diagnosis, age more than 18 years, a diagnosis of RA according to the 2010 ACR/ EULAR Classification criteria for RA diagnosis , active disease according to DAS-28, for those receiving nonsteroidal anti-inflammatory drugs and corticosteroids, the dosage had to be stable for at least 2 weeks before screening. Exclusion Criteria: Patients with allergic, and infectious diseases, patients receiving steroids for the first time within 2 weeks before the study, patients with hypercalcemia, hypercalciuria, nephrolithiasis, or neoplastic diseases were excluded from the study. We also excluded patients on any biologic therapy and patients with prior use of leflunomide, hydroxychloroquine, or sulphasalazine for more than 2 months.

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