Screening and molecular characterization of Trichomonas vaginalis genotypes isolated from married women in northern Iran.

Trichomonas vaginalis is an anaerobic protozoan parasite that causes trichomonosis in human. It is one of the most common non-viral sexually transmitted infections. It has been found to be most prevalent in patients referred to sexually transmitted disease clinics. In recent years, molecular methods have been used to identify genotypes of this parasite in different parts of the world and so far 6 types of T. vaginalis have identified. The aim of this study was to investigate the prevalence and genotype identification of T. vaginalis from married women in northern Iran. A total of 450 vaginal specimens were taken from married women, referring to health centers in northern Iran. Demographic information of women was collected through a questionnaire. The samples were first examined microscopically and then monitored in Dorsch culture medium for up to 10 days. Actin genes of positive samples were amplified by PCR. Finally, PCR products were used to determine the sequence and genotype of the parasite. Overall, 0.7% (3/450) samples were positive for T. vaginalis. All of the three infected women were housewives. After sequencing, the genotype of these parasites were type H (66.7%) (Accession no; MW414672-MW414673) and type E (33.3%) (Accession no: MW414671). Low prevalence of T. vaginalis in north of Iran indicate high level of hygiene in sexual intercourse and avoiding from high risk sexual behaviors, and also it seems that genotype H is dominant type of the parasite in the study area.

Cloning and expressing of interleukine 2 in amniotic membrane-derived mesenchymal stem cells, as a potent feeder layer.

The application of mesenchymal stem cells (MSCs) is rapidly expanding due to their unique properties in cell therapy, especially as the feeder layer in the ex-vivo expansion of immune cells. Also, Interleukin 2 (IL-2) is an essential human cytokine in the expansion of hematopoietic precursors and progenitors, i.e., NK cells and T cells, while there is no endogenous expression of IL-2 in MSCs. This study aimed to examine the potency of amniotic membrane (AM)-MSCs as the IL-2 secretory cells. IL-2-containing pCMV3-C-GFPspark shuttle vector was transformed in E.coli DH5-alpha. After cloning, the plasmid DNA was extracted and transfected in isolated AM-MSCs, by lipofectamine-2000. Then, the RNA and protein expression levels of exogenous IL-2 were evaluated 3 to 15 days after transfection, using ELISA and qRT-PCR. Fluorescent microscopy and flowcytometry assays were used for evaluating the GFP-positivity of transfected AM-MSCs, as IL-2 expression control. There was a significant increase in RNA expression of exogenous IL-2 in transfected AM-MSCs in 3 to 15 days after transfection. (p<0.001) Also, IL-2 concentration released in the medium was increased in 3rd day after transfection (611 pg/ml). However, the RNA and protein expression of IL-2 was reduced through passing the time. The results show AM-MSC is a suitable host for the expression and secretion of IL-2 as a critical cytokine in the ex-vivo expansion of hematopoietic precursors and progenitors, i.e., NK cells and T cells. Also, the survival time of IL-2 expression in AM-MSCs was long enough for use as a feeder layer.

Mutations of the phenylalanine hydroxylase gene in Iranian patients with phenylketonuria.

Phenylketonuria (PKU) is an autosomal recessive disease which results from mutations in the phenylalanine hydroxylase (PAH) gene. The aim of this study was the identification of sixteen different mutations in Iranian patients with hyperphenylalaninemia. The mutations were detected during the characterization of PAH genotypes of 39 PKU patients from Qazvin and Zanjan provinces of Iran. PAH mutations have been analyzed by PCR and direct sequencing of PCR products of the promoter region and all 13 exons of PAH gene, including the splicing sites. A mutation detection rate of 74.3 % was realized. Two mutations were found at high frequencies: R176X (10.25 %) and p.P281L (10.25 %). The frequencies of the other mutations were: IVS2+5G>A (2.56 %), IVS2+5G>C (2.56 %), p.L48S (2.56 %), p.R243Q (2.56 %), p.R252Q (5.12 %), p.R261Q (7.69 %), p.R261X (5.12 %), p.E280K (2.56 %), p.I283N (2.56 %), IVS9+5G>A (2.56 %), IVS9+1G>A (1.28 %), IVS11+1G>C (1.28 %), p.C357R (1.28 %), c.632delC (2.56 %). The present results confirm the high heterogeneity of the PAH locus and contribute to information about the distribution and frequency of PKU mutations in the Iranian population.