Alteration of peripheral blood T-Reg cells and cytokines production in angiography personnel exposed to scattered X-rays

Angiocardiography is an X-ray examination of the blood vessels or chambers of the heart. Cardiologists and staff members applying this procedure are exposed to high levels of scattered radiation. In our previous study the incidence of unstable chromosomal aberrations and cytokinesis-blocked micronuclei were found to be significantly higher in exposed individuals than the age and sex matched controls. In the present study we assessed cytokine production by peripheral blood mononuclear cells of the above cases and the percentage of Treg cells. According to film dosimeter analysis, personnels received 0.25-15 mSv during the previous year [average of 3 mSv/y]. Isolated PBMCs from the test and control groups were stimulated with Phorbol Myristate Acetate/Ionomycin [PMA/I]. Cytokine production was measured in the supernatants of cultured lymphocytes. The percentage of Treg cells was studied by flow cytometry. The production of IL-10 and IL-5 was significantly down-regulated in the test group compared to the control group. In contrast, IL-12 was up-regulated. Yet, no statistically significant difference was found for IFN-gamma between two groups. In addition, we found higher percentage of CD4+CD25+bright Treg cells in the study group compared to the controls. Taken together, it was shown that low doses of scattered X-rays could skew cytokine profile of peripheral blood mononuclear cells in favour of inflammatory response causing the increase of Treg cells

Purification and biochemical characterization of two novel antigens from Leishmania major promastigotes

The identification and characterization of antigens that elicit human T cell responses is an important step toward understanding of Leishmania major infection and ultimately in the development of a vaccine. Micropreparative SDS-PAGE followed by electrotransfer to a PVDF membrane and elution of proteins from the PVDF, was used to separate 2 novel proteins from L. major promastigotes, which can induce antibodies of the IgG2a isotype in mice and also are recognized by antisera of recovered human cutaneous leishmaniasis subjects. Fractionation of the crude extract of L. major revealed that all detectable proteins of interest were present within the soluble Leishmania antigens (SLA). Quantitation of these proteins showed that their expression in promastigotes is relatively very low. Considering the molecular weight, immunoreactivity, chromatographic and electrophoretic behavior in reducing and non-reducing conditions, these proteins are probably 2 isoforms of a single protein. A digest of these proteins was resolved on Tricine-SDS-PAGE and immunoreactive fragments were identified by human sera. Two immunoreactive fragments (36.4 and 34.8 kDa) were only generated by endoproteinase Glu-C treatment. These immunoreactive fragments or their parent molecules may be ideal candidates for incorporation in a cocktail vaccine against cutaneous leishmaniasis.

Production of nitric oxide by murine macrophages induced by lipophosphoglycan of Leishmania major

Protozoan parasites of the genus Leishmania cause a number of important human diseases. One of the key determinants of parasite infectivity and survival is the surface glycoconjugate lipophosphoglycan (LPG). In addition, LPG is shown to be useful as a transmission blocking vaccine. Since culture supernatant of parasite promastigotes is a good source of LPG, we made attempts to characterize functions of the culture supernatant, and membrane LPG isolated from metacyclic promastigotes of Leishmania major. The purification scheme included anion-exchange chromatography, hydrophobic interaction chromatography and cold methanol precipitation. The purity of supernatant LPG (sLPG) and membrane LPG (mLPG) was determined by SDS-PAGE and thin layer chromatography. The effect of mLPG and sLPG on nitric oxide (NO) production by murine macrophages cell line (J774.1A) was studied. Both sLPG and mLPG induced NO production in a dose dependent manner but sLPG induced significantly higher amount of NO than mLPG. Our results show that sLPG is able to promote NO production by murine macrophages.

Identification of novel Leishmania major antigens that elicit IgG2a response in resistant and susceptible mice

Experimental murine models with high, intermediate and low levels of genetically based susceptibility to Leishmania major infection reproduce almost entire spectrum of clinical manifestations of the human disease. There are increasing non-comparative studies on immune responses against isolated antigens of L. major in different murine strains. The aim of the present study was to find out whether there is an antigen that can induce protective immune response in resistant and susceptible murine strains. To do that, crude antigenic extract of procyclic and metacyclic promastigotes of L. major was prepared and subjected to SDS-PAGE electrophoresis. Western-blotting was used to search for antigen(s) capable of raising high antibody level of IgG2a versus IgG1 in the sera of both infected resistant and susceptible strains. Two novel antigens from metacyclic promastigotes of L. major (140 and 152 kDa) were potentially able to induce specific dominant IgG2a responses in BALB/c and C57BL/6 mice. The 2 antigens also reacted with IgG antibody of cutaneous leishmaniasis patients. We confirm that 140 and 152 kDa proteins of L. major promastigotes are inducing IgG production in mice and humans.

Effects of combined therapy with thalidomide and glucantime on leishmaniasis induced by Leishmania major in BALB/c mice

For treating Leishmania major infection in BALB/c mice, we used thalidomide in conjunction with glucantime. Groups of mice were challenged with 5 x 10(3) metacyclic promastigotes of L. major subcutaneously. A week after the challenge, drug treatment was started and continued for 12 days. Thalidomide was orally administrated 30 mg/kg/day and glucantime was administrated intraperitoneally (200 mg/kg/day). It was shown that the combined therapy is more effective than single therapies with each one of the drugs since the foot pad swelling in the group of mice received thalidomide and glucantime was significantly decreased (0.9 +/- 0.2 mm) compared to mice treated with either glucantime, thalidomide, or carrier alone (1.2 +/- 0.25, 1.4 +/- 0.3, and 1.7 +/- 0.27 mm, respectively). Cytokine study showed that the effect of thalidomide was not dependent on IL-12; however, it up-regulated IFN-gamma and down-regulated IL-10 production. Conclusively, thalidomide seems promising as a conjunctive therapy with antimony in murine model of visceral leishmaniasis.

effect of vitamin E on splenocytes apoptosis of gamma-irradiated BALB/c mice

Apoptosis, a physiologic mechanism to eliminate unwanted cells, is also induced by ionizing irradiation, through production of free radicals. It has been demonstrated that antioxidants such as vitamin E are able to protect cells from damage caused by free radicals. Taken together we found it reasonable to make an attempt to evaluate the protective effect of vitamin E against apoptosis. The irradiated mice received 1 Gy/day gamma radiation for one day [low dose] or for three successive days [high dose, 3Gy]. The splenocytes were then isolated at 6, 14 and 24 h after exposure. DNA gel electrophoresis and DNA fragmentation assay were done in addition to the evaluation of splenocytes cytology. Our results showed that Vitamin E can reduce apoptosis against low dose irradiation. However it is not able to completely block programmed cell death in high dose irradition

[Evaluation of cellular immunity of patients with brucellosis by measuring interferon gamma [IFN-gamma] and interleukin-13 [IL-13] by flow cytometry and Elisa]

Introduction: The analysis of cytokine production is a valuable component of studies of immune response to stimulation such as pathogens, vaccines, and other immunological challenges. The animal findings Show that brucella infection induces cell-mediated responses. Some cytokines have an Important role in resistance to brucella infection

Objective: This Study is don't to Poesent a orecise an useful unseparated whole blood of healthy normal and patients with acute and chronic brucellosis as the source of cells evaluation of interferon gamma [IFN-gamma] and interleukin-13 expression

Materials and Methods: Diluted whole blood samples of 27 patients with acute [14] and chronic brucellosis [n13], and sex and age-matched healthy volunteers [n=22] With Meanage of 35-33+21 were cultured in the presence of either mitogen; heat inactivated bacteria or medium alone. Intracellular IL-13 and IFN-gamma were measured by specific sandwich ELISA and flow Cytometry was detected by the number of cytokine-producing CD3+ cells

Results: Findings indicated that extrandintracellular specific IFN-Y in creased Cansiderably [P<0.001] in cute brucelles patients. not only IFN-gamma production but also the number of IFN-gamma- producing CD3 cells were significantly decreased in response to antigen in chronic group of patients. There was a reverse correlation between the number of IFN-gamma-producing and IL-13-producing CD3 cells only in acute group which shows polarization of immune responses to Th1 in them

Conclusion: Although the percentage of CD3 IL-13-producing cells was dramatically high in the chronic group of patients, no correlation was found between the number of IFN-g-producing and IL-13-producing CD3 cells. In conclusion, the correlation of Th2 cytokines production and progression of chronic human brucellosis was not demonstrated. Nevertheless, diminished production of Th1 cytokines production in chronic group may suggest T cells unresponsiveness to Brucella antigen which helps prolongation of brucellosis in chronic patients

Restorative effect of vitamin E on some immunological parameters of sub-lethal gamma-irradiated BALB/c mice

Elevated amount of the free radicals due to ionizing radiation causes deteriorating damage on immune system. Therefore, we made attempts to investigate the protective effect of vitamin E [vit-E], a biological antioxidant in BALB/c mice, so as to find an affordable prophylactic supplementation for individuals who are at risk. Several groups of mice were selected and exposed to sub-lethal gamma-irradiation with or without vit-E supplementation. At the end of exposure, mice were immunized by either live attenuated Brucella melitensis vaccine or sheep red blood cell [SRBC]. Consequently, the following parameters were assayed: specific antibody response, delayed type hypersensitivity and lymphocyte proliferation. We showed that vit-E supplementation restored the immune response in gamma-irradiated mice. These findings might have implications for individuals who are at risk of exposure to ionizing radiation