Simultaneous regulation of miR-451 and miR-191 led to erythroid fate decision of mouse embryonic stem cell

Objective(s): Various microRNAs (miRNAs) are expressed during development of mammalian cells, when they aid in modulating gene expression by mediating mRNA transcript cleavage and/or regulation of translation rate. miR-191 and miR-451 have been shown to be critical regulators of hematopoiesis and have important roles in the induction of erythroid fate decision. So, the aim of this study is investigation of the miR-191 and miR-451 roles in the controlling mouse embryonic stem cell (mESC) differentiation toward the erythroid lineage. Materials and Methods: mESCs were infected with either pCDH-miR-Off-191 viruses in pCDH-miR-Off-191 group or simultaneously with pCDH-miR-Off-191 and pCDH-miR-451 lentiviruses in simultaneous group. Then, the expression profiles of erythroid specific transcription factors and globin genes were analyzed using QRT-PCR on day 14 and 21 of differentiation. Flow cytometry analysis was used to evaluate of TER119 and CD235a erythroid specific surface markers. Results: Gata-1, Klf-1, Epor and globin chains were found to be expressed in pCDH-miR-Off-191 and in simultaneous groups. The majority of globin chains showed changes in their expression levels with progression of differentiation from day 14 to day 21. Flow cytometry results showed that miR-451 upregulation and miR-191 down-regulation is associated with the expression of TER119 and CD235a. Of these two groups analyzed, simultaneous group was most significantly potent in stimulation of erythroid fate decision of mESCs. Conclusion: Together, present data demonstrate that down-regulation of miR-191 alone can enhance the differentiation of mESCs. However, the simultaneous effect of miR-451up-regulation and miR-191 down-regulation is much stronger and can have more practical use in artificial blood production

The association between common C677T mutation in methylenetetrahydrofolate reductase gene and the risk of venous thrombosis in an Iranian population

Background: Venous thrombosis is a multicausal disease involving acquired and genetic factors. The prevalence of methylenetetrahydrofolate reductase (MTHFR) C677T genotypes and its association with venous thrombosis is not established in the Iranian population. In this study we investigated a possible association between fasting hyperhomocysteinemia and C677T mutation in the MTHFR gene with venous thrombosis. Materials and Methods: We studied 200 venous thrombotic patients and 100 healthy controls, of similar age and sex. Mutation analysis was carried out by PCR-RFLP, and the homocysteine level was measured by EIA. Results: No significant differences in the frequency of C677T genotypes were observed between patients and controls (P=0.2). The frequency of the T allele was 21 and 27.2 in controls and patients, respectively (odds ratio, 1.27; 95 CI, 0.83-1.94, P=0.15). Fasting homocysteine level was significantly higher in patients than controls (P=0.001). Conclusions: We concluded that hyperhomocysteinemia, but not MTHFR C677T mutation, is a significant risk factor for venous thrombosis in the Iranian population, and measuring the level of homocysteine is less expensive and more useful than the genetic test for the MTHFR mutation

The Cytotoxicity Pathway Of Natural Killer Cells In Cord Blood Compared To Peripheral Blood

( The cytotoxic activity of natural killer cells is usually tested by radioactive assay 51Cr release assay), which detects the release of cytoplasmic contents after plasma membrane disintegration of dying cells. In contrast to this indirect evaluation of cytotoxicity, the assessment of cell damage by flow cytometry aims to provide a more exact characterization of the death pathway via detection of the percentage of apoptosis and necrotic cells. Annexin V-FITC (Axv -FITC) can be used to label cells in the early apoptotic state, while propidium iodide (PI) indicates late apoptosis or necrosis. The NK cytotoxicity of cord blood (CB) and peripheral blood (PB) was determined after 4 hours of incubation in the absence of cytokines. After 4 hours in vitro incubation, co-staining with Annexin V-FITC (Axv-FITC) and propidium iodide (PI) permitted discrimination between viable, early apoptotic and necrotic cells. As we would expect, the cytotoxicity pathway in PB mononuclear cells (MNCs) consists of both apoptosis and necrosis pathways but in CB MNCs it almost consists of early apoptosis; and necrosis is negligible. With escalating E: T (effector: target) ratio changes in the percentage of apoptotic cells in PB samples were significantly higher than CB samples. The mechanism(s) of the low cytotoxicity of resting cord NK cells is not well understood. Complementary research in this field is recognized to elucidate the phenotypical and functional properties of CB cells and how they relate to maturational stages. CB studies are important for transplantation research and ma! y provide insight to the suppressive mechanism by which the host -recipient could evade GVHD and rejection

The reactivity of human serum natural autoantibodies with certain autoantigens increases along with aging

Introduction Natural autoantibodies (NAAs) that recognize autoantigens are mainly generated in the absence of any apparent immunization with foreign antigens (Ags). These antibodies maintain the body homeostasis through functions, such as inhibition of tumor angiogenesis, detection of structural changes in autoantigens, and exertion of anti-inflammatory impacts. The human body is shown to go through immunological and physiological changes during aging. However, data regarding the potential variations in the binding activity of NAAs is still scarce. Therefore, in this study, we were about to explore the trend through which the reactivity of serum NAAs with several autoantigens varies with advancing age. Materials and methods Serum samples were prepared from healthy individuals of seven age intervals: (0) cord blood, (1) infancy, (2) childhood, (3) adolescence, (4) early adulthood, (5) middle adulthood, and (6) late adulthood (the elderly). The mean immune reactivity (MIR) of the sera with 24 human autoantigens that were obtained from Immunculus research center (Russia) was determined using ELISA and inter-group comparisons were also performed. Results In general, the MIR of serum natural antibodies with the autoantigens was shown to follow an upward trend with advancing age so that the lowest and highest MIRs were detected in the cord blood and late adulthood samples, respectively. Moreover, the results of the inter-group comparisons indicated that the MIRs of the first, second, fourth, and sixth groups were significantly higher than those of their previous groups, i.e. zero, first, third, and fifth groups, respectively. Conclusion This study showed that the reactivity of human tissue-specific and non-specific NAAs with autoantigens varied along with aging. Regarding the crucial roles NAAs play in maintaining the body homeostasis, the variation in their concentration at different age intervals might account for the immunological and pathological changes that occur in the elderly. © 2013 Published by Elsevier Masson SAS

Frequency rate of active hepatitis G virus infection among human immunodeficiency virus infected patients in Tehran

Background : Hepatitis virus infections are common in patients suffering from human immunodeficiency virus (HIV) infection due to similar transmission routes of these viruses. Hepatitis virus infections lead to impaired survival in HIV positive patients. In contrast to the expectations, some studies indicate that HGV infection slows down the progression and eventually death of AIDS patients. The aim of this study was to estimate the frequency of active HGV infection among HIV infected patients in Tehran.Material and Methods: Flow cytometric analysis of lymphocyte subpopulations was performed on blood samples of 103 patients with HIV infection referring to the central laboratory of Iranian blood transfusion organization. Their plasma samples were analyzed for the presence of HGV RNA by RT-PCR. HGV Positive cases were confirmed by PCR-ELISA. Results: Of 103 HIV-positive patients, 16 (15.5) were HGV RNA-positive. It was apparent that the frequency of HGV in patients with CD4 count above 500 cell/μl is higher. Conclusion: Sucha frequency is not high compared with those found in other studies. The high frequency of active HGV infection in patients with asymptomatic HIV infection may be due to the loss of active HGV infection in patients with advanced HIV infection