[Synergistic effect of rCagA and LPS of H. pylori O2 serotype in induction of proper immune response against H. pylori]

Background: H. pylori is one of gastric bacterial infectious agents that reported from world whole. This organism is heterogen and have hyper variable regions in its genome that to cause organism escape from immune response. H. pylori cagA+ strains will report that is stimulating factor for adenocarcinoma, gastritis in infected persons. H. pylori LPS have less toxicity, mitogenicity and pyrogenicity than Entrobactericeae. In this study we investigated the LPS and rCagA senergistic effect on stimulation of Th1 immune response in mice model

Materials and Methods: LPS of H. pylori O[2] serotype was extracted by hot phenol-water method. Proper conserved fragment of cagA was expressed in proper vectors. These antigens were injected to Balb/c mice and immune response was assayed by ELISA

Results: the IgG1/IgG2a ratio in the immunized mice with rCagA and rCagA plus CpG was <1, indicating a Th1 type response, while the control group was >1, indicating a strong Th2 response. In mice immunized with LPS and rCagA, the immune response elevated which indicated synergistic effect of this antigen on stimulating of strong immune response against H. pylori infection

Conclusion: effective immunizations against H. pylori will possible affected treatment in next future. Protective immune response in H. pylori is balance between Th1/Th2.These data suggest that immunization with rCagA and LPS promoted a Th1 immune response. H. pylori rCagA and LPS serve as an excellent antigen for immunization. In conclusion, we recommended multicomponent vaccine contain of rCagA and LPS for vaccine formulation against H. pylori infection

Cloning and sequencing of Leishmania major thiol-specific-antioxidant antigen [TSA] gene

Leishmaniasis is caused by parasitic protozoa of the genus Leishmania which, in the infected host are obligate intracellular parasite. TSA is the immuno-dominant antigen of Leishmania major which is considered as the most promising molecule for a recombinant or DNA vaccine against leishmaniasis. Genomic DNA of TSA protein was extracted and amplificated as a template. Then the PCR product was cloned into pTZ57R/T vector. Finally, the recombinant plasmid was extracted from transformed Escherichia coli [TG1 strain] and sequenced. MRHO/IR/75/ER [an Iranian strain] of L. major and TSA gene [Accession number LmjF15.1080] were used. Sequence analysis of cloned TSA gene into pTZ57R/T vector showed high homology of 90% with LmjF15.1080 [TSA gene] and strain "LV39" [Accession no. AF069386] and strain "Friedlin" [Accession no.AF044679]. We cloned TSA gene of L. major successfully. Recombinant plasmid was confirmed. It is ready to express recombinant protein for further studies

immunomedulatory effect of Ganoderma Lucidum polysachharide extract on BALB/c peritoneal macrophage function

Ganoderma Lucidum has been regarded as a natural immunomodulator. The exact carbohydrate epitope responsible for the immunomodulatory activity and its receptor have not been identified, but it seems likely that it is the receptor CR3 [complement receptor 3] which can bind to beta-glucan polysachharide. Because glucose-6-phosphate dehydrogenase [G6PD] activity has a critical role in the regulation of macrophage functions such as nitric oxide [NO] production, we assessed the immunomodulatory effect of GL-PS in BALB/c peritoneal macrophages. For this purpose, BALB/c mice peritoneal macrophages were isolated and treated with various concentrations of GL-PS [0.001, 0.01, 0.1, 1, 10, 100 microg/ml]. After 24 hours, the viability of treated macrophages was measured by MTT assay at 540 nm and the effective dose was determined to be 0.1 microg/ml. Then, macrophages were sonicated and special activity of G6PD was measured in the cell extracts by measuring the alterations in NADPH absorption at 339nm and protein concentration by Bradford method. Also, NO production was determined by use of Griess-reagent after 18 hours. Results of this study showed that 0.1 microg/ ml of GL-PS had the maximal effect on cell viability [stimulation Index] in comparison to other doses [0.05

[Protective effect of vitamin D3 and GP63 conjugated with tetanus toxoid on outcome of cutaneous leishmaniasis lesions in balb/c mice]

GP63 is a major surface protease of Leishmania promastigotes that plays an important role in its virulance. As GP63 on its own can not develop an effective protection against leishmaniasis, the goal of this study was to evaluate the protective effect of GP63 conjugated with tetanus toxoid [TT] and Vitamin D3 in susceptible BALB/c mice against cutaneous leishmaniasis. This study was a basic-applied experimental study performed in Tarbiat Modarres University from September 2002 to April 2005. Cloned virulant Leishmania [L.] major [MRHO/IR/75/ER] strain was cultured and 5x10[9] cells were harvested. GP63 Molecule was purified and conjugated with TT and conjugated molecule was used for immunization of 8 groups of female BALB/c mice. Results showed that the group of mice receiving conjugated molecule with Vitamin D3 had significant differences from other groups regarding lesion progression [P

[ role of 6-aminonicotinamide in the resistance of peritoneal macrophages against Leishmania major infection in BALB/c mouse]

The objective of this study was to investigate the relationship between glucose-6-phosphate dehydrogenase inhibition in macrophages treated with 6-Aminonicotinamide, the amount of nitric oxide [NO] production and the resistance of infected macrophages against Leishmania major infection. Peritoneal macrophages of BALB/c mice were isolated and treated with different concentrations [1.25, 2.5, 5, 10 mM] of 6-aminonicotinamide. After 24 hours, the viability of treated macrophages was measured by MTT assay at 540 nm. G6PD activity was measured in the cell extracts 24 hours later. Macrophages were then infected with leishmanial amastigotes and after 18 hours NO production was determined using Griess-reagent. In order to study the inhibition of macrophage activity, 5 mM concentration of 6-AN was used and number of leishmanial amastigotes was recorded in these cells from day 1 to7. Different concentrations of 6-AN were shown to cause a significant increase in cell death and decrease in G6PD activity and NO production in macrophages. Also, the number of amastigotes in macrophages was increased significantly [p < 0.05]. He concentration of 6-aminonicotinamide and G6PD activity affect the viability of BALB/c mice peritoneal macrophages through production of NO. Inhibition of G6PD activity leads to decreased leishmani-cidal activity of mouse peritoneal macrophages

[Effects of strain, temperature and media composition on the production of catalase antigen in Aspergillus fumigatus]

To evaluate The effect of some physicochemical factors such as strain, temperature and media composition on the production of catalase Antigen. Iterventional. Four media [AMM, YED, S and CDA] which were different in kind, and amount of glucose and other ionic and aminoacid components were used at 28°c and 37°c. Also Two pathogenic human and animal isolates were used for mass Production of fungi, the mycelia were being filtered after 96 hours, were disrupted using lysing buffer, and then centrifuged at 150000g, The water soluble suppernatant, was used to purify the antigen. The amount of proteins water - Soluble were measurered by Brodford method. Then electrophorised and the sample was specificaully stained with ferocyanid. The water - sdubles were loaded on the anionic and cathionic columns, respectively. The amounts of their proteins were measured follewed by reading their O.Ds eluents of cathionic calumn were electrophorised and their electrophoretic patterns were compared. Results showed that the amount of the proteins in two S and CDA was 1/3 +/- 0/1mg/ml and in the other two YED and AMM it was 1/1 +/- 0/1mg/ml. After chromatography, still the protein amountof the first two water - soluble's [S and CDA] were more than that of the second two water - soluble's [YED and AMM]. Electrphoresis of the eluents showed that the eluents of S and CDA had stronger bands than the eluents of YED and AMM. It is concluded that strain and temperature has no effects on the protein amounts of water -solubles as well as eluents. While themedia composition had positive effect on their protein amounts. This study also showed that in contrast to the media composition, strain and tempreature has no effects on the electrophoretic Pattern of The water - solubles and eluents

Nitric oxide production following vaccination with killed mycobacterium tuberculosis [H37Rv] and BCG

Protective immune response induced by viable BCG has been suggested by several investigators. Both killed BCG and Mycobacterium tuberculosis are able to induce response. Nitric oxide [NO] is one of the non- specific responses produced against these agents. To survey the effect of alive, killed BCG and also killed Mycobacterium tuberculosis [H37Rv strain] on NO Production. 6-8- week-old female BALB/c mice were used. Three groups were vaccinated with viable, killed BCG and killed Mtb, respectively. One group received PBS as a control. After 5-8 weeks of vaccination, peritoneal cells of all groups were collected in usual manner and plated out in 96-well plates. Cells were treated with killed H37Rv, killed BCG and viable BCG alone or with rIFN gamma and NO inhibitors [aminoguanidine and NGMA]. Supernatant of each well was collected after 24h. NO level was estimated by Griess method by ELISA reader at 540nm absorbance. Results indicated that NO induction level in vaccinated groups were higher than control [P