Frequencies of HLA-A, B and DRB1 alleles in a large normal population living in the city of Mashhad, Northeastern Iran.

OBJECTIVES: The population in Iran is a genetic admixture of the ancestral Aryan and other populations neighboring Iran. Different ethnic groups in Iran show wide regional distributions for many human leukocyte antigen (HLA) alleles. Therefore, it is necessary and sensible to study the differences in HLA allele distribution in different area. We studied the HLA class I and II allele frequencies in a large unrelated healthy Iranian population from Mashhad in the Northeast region. MATERIALS AND METHODS: Five hundred unrelated healthy adult individuals borne and living in Mashhad, Northeast of Iran, were genotyped for HLA-A, B and HLA-DRB1 alleles using PCR with low resolution sequence specific primers (SSP-PCR) technique. RESULTS: A total of 14 HLA-A, 24 HLA-B and 10 HLA-DRB1 alleles were spread throughout the studied population with distinct allele frequencies. At the HLA-A locus, HLA-A*02 was found to be the most frequent allele, with a frequency of 20.9%. The most common HLA-B alleles was B*35 (16.4%). The two most common observed alleles in HLA class II alleles were DRB1*15 (20.0%) followed by DRB1*13 (16.2%). CONCLUSION: This study is the first on the HLA class I and II allele frequencies in Northeastern Iranian population living in Mashhad. Distribution of HLA-A and B loci showed some similarities with those of other Iranians. Some difference in HLA-DRB1 polymorphisms however was observed. Considering the highly mixed population of Mashhad, the finding was not unexpected.

Effects of 1,25-dihydroxyvitamin D3 on IL-17/IL-23 axis, IFN-γ and IL-4 expression in systemic lupus erythematosus induced mice model.

OBJECTIVES: Systemic lupus erythematosus (SLE) is a multi-factorial autoimmune disease which may be characterized by T lymphocytes dysfunctions. Th17 cells have been identified as new effector cells, which play an important role in the pathogenesis. In recent years, immunomodulatory effect of vitamin D3 has been noticed. In the present experiment, the effect of vitamin D3 on the expression of IL-17, IL-23, IL-4 and IFN-γ were assessed in activated chromatin-induced mouse model for SLE. MATERIALS AND METHODS: Five groups of mice were included in this study; Group one received active chromatin +CFA + PBS; Group 2 received vitamin D3 starting 2 weeks before disease induction; Group 3 received vitamin D3 (50 ng/day) starting with the disease establishment; Group 4 received non active chromatin +CFA + PBS; Group 5 received CFA + PBS. On day 56 splenocytes were isolated and gene expression of interleukin IL-17, IL-23, IL-4 and IFN-γ were analyzed by Real-Time PCR method. Proteinuria and serum anti-dsDNA and Th17 levels were measured using commercial kits. RESULTS: The results showed that IL-17, IL-23, and IFN-γ mRNA expression, and IL-17 titers were decreased remarkably and that of IL-4 increased in mice which received vitamin D3 before SLE induction. Administration of vitamin D3 after the establishment of SLE failed to affect the IL-17 or IL-23 mRNA levels. Lastly, pre-treatment of mice with vitamin D3 decreased the anti-ds DNA antibody titer. CONCLUSION: Our findings showed that vitamin D3 supplementation in lupus induced mice through modulating the expression rate of some inflammatory cytokines diminished the inflammatory conditions in SLE.

Anti-inflammatory effect of garlic 14-kDa protein on LPS-stimulated-J774A.1 macrophages.

CONTEXT: Garlic 14-kDa protein is purified from garlic (Allium sativum L.) which is used in traditional medicine and exerts various immunomodulatory activities. OBJECTIVE: The present study investigated the suppressive effect of garlic 14-kDa protein on LPS-induced expression of pro-inflammatory mediators and underlying mechanism in inflammatory macrophages. MATERIALS AND METHODS: J774A.1 macrophages were treated with 14-kDa protein (5-30 µg/ml) with/without LPS (1 µg/ml) and the production of inflammatory mediators such as prostaglandin E2 (PGE2), TNF-α, and IL-1ß released were measured using ELISA. Nitric oxide (NO) production was determined using the Griess method. The anti-inflammatory activity of 14-kDa protein was examined by measuring inducible nitric oxide synthase and cyclooxygenase-2 proteins using western blot. The expression of nuclear NF-κB p65 subunit was assessed by western blot. RESULTS: Garlic 14-kDa protein significantly inhibited the excessive production of NO, PGE, TNF-α, and IL-1ß in lipopolysaccharide (LPS)-activated J774A.1 macrophages in a concentration-related manner without cytotoxic effect. Western blot analysis demonstrated that garlic 14-kDa protein suppressed corresponding inducible NO synthase expression and activated cyclooxygenase-2 protein expression. The inhibitory effect was mediated partly by a reduction in the activity and expression of transcription factor NF-κB protein. CONCLUSION: Our results suggested, for the first time, garlic 14-kDa protein exhibits anti-inflammatory properties in macrophages possibly by suppressing the inflammatory mediators via the inhibition of transcription factor NF-κB signaling pathway. The traditional use of garlic as anti-inflammatory remedy could be ascribed partly to 14-kDa protein content. This protein might be a useful candidate for controlling inflammatory diseases and further investigations in vivo.

Influence of vitamin D on cell cycle, apoptosis, and some apoptosis related molecules in systemic lupus erythematosus.

OBJECTIVES: Genetic and environmental factors are involved in the pathogenesis of systemic lupus erythematosus (SLE). Autoreactive lymphocytes are cleared through apoptosis and any disturbance in the apoptosis or clearance of apoptotic cells may disturb tolerance and lead to autoimmunity. Vitamin D has anti-proliferative effects and controls cell cycle progression. In this study we investigated the effects of vitamin D on cell cycle and apoptosis induction in lupus patients. MATERIALS AND METHODS: Isolated peripheral blood mononuclear cells (PBMCs) from 25 SLE patients were cultured in the presence of 50 nM of 1,25(OH)2D3; then one part of the cells were stained with FITC labeled Annexin V and PI and were analyzed for apoptosis determination. For gene expression assessment of FasL, Bcl-2 and Bax, RNA was extracted from one another part of the cells, cDNA was synthesized and gene expression analysis was performed using Real time PCR. An additional part of the cells were treated with PI and the cell cycle was analyzed using flowcytometer. RESULTS: The mean number of early apoptotic cells in vitamin D treated cells decreased significantly (18.48±7.9%) compared to untreated cells (22.02±9.4%) (P=0.008). Cell cycle analysis showed a significant increase in G1 phase in vitamin D treated cells (67.33±5.2%) compared to non treated ones (60.77±5.7%) (P =0.02). Vitamin D up-regulated the expression levels of Bcl-2 by (18.87 fold increase), and down-regulated expression of Bax (23%) and FasL (25%). CONCLUSION: Vitamin D has regulatory effects on cell cycle progression, apoptosis and apoptosis related molecules in lupus patients.

Ursolic acid induced apoptotic cell death following activation of caspases in isolated human melanoma cells.

Ursolic acid (UA) is plentifully present in fruits, foods, and medicinal plants, and has anti-cancer activity on many cancer cell lines. However, the effects of UA on some melanoma cells and the mechanisms of action have not been reported. The effect of UA on isolated human melanoma and fibroblast cell lines has been investigated using the MTT assay, and cell death was determined using Annexin-V/PI staining. To explore whether the activation of caspases was required for apoptosis induction, cells were treated with pan-caspase inhibitor and UA. Changes in apoptosis pattern were analyzed by flow cytometry. Activation of caspases was detected by western blot analysis. A significant concentration-dependent suppression of cell proliferation was seen after 24 and 48 h in the presence of UA. Apoptosis was considerably increased by UA, indicated by enhancement of an Annexin-V positive population and a sub-G1 peak. UA induced proteolytic processing of caspase-3. The results confirm the anti-cancer properties of UA on cultured human melanoma cells, possibly mediated through the induction of apoptosis following activation of caspases.