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@#A series of new 2,5-disubstituted-1,3,4-oxadiazole derivatives (5a-j and 6a-j) have been designed and synthesized in four-steps. Sixteen compounds among the twenty compounds are reported for the first time. The compounds were characterized and confirmed by the FTIR, 1D- and 2D-NMR and HRMS analyses, and were tested against Mycobacterium smegmatis and Mycobacterium tuberculosis H37Ra. Compound 5d was the most active against M. smegmatis with MIC value of 25 µM, and exhibited cidal activity with MBC of 68 µM, respectively. The time-kill assay showed the good killing rate at 77% with the combination of isoniazid (INH). In addition, checkboard assay confirmed the interaction of compound 5d was categorised as additive. Docking simulation has been performed to position 5d into the pantothenate synthetase active site with binding free energy value –8.6 kcal mol-1. It also occupied the same active site as that of standard native ligand with similar interactions, which clearly indicate their potential as pantothenate synthetase inhibitor.
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Despite recent economic prosperity, Korea still has high prevalence of tuberculosis. Molecular biologic characterization of Korean Mycobacterium tuberculosis strains might provide a deeper understanding of the forces contributing to the spread of tuberculosis in Korea. Therefore, we analyzed the cell lysate proteome of a representative Korean Mycobacterium tuberculosis isolate (K01) in comparison with laboratory reference strains H37Rv and H37Ra. Seven spots were strongly expressed only in K01 strain compared with M. tuberculosis H37Rv and H37Ra. Through continuous MALDI-MS analysis, these spots were identified as hypothetical protein Rv3849, secreted immunogenic protein Mpt64, Acetyl/propionyl-CoA Carbpxylase (AccD1), alkyl hydroperoxide reductase C (AhpC), N-acetylmuramyl-L-alanine amidase, a putative UDP glucose epimerase, and a transposase. A deeper study of these proteins may provide a clue in the development of effective new anti-tuberculosis vaccines against Korean M. tuberculosis isolates.
Subject(s)
Korea , Mycobacterium tuberculosis , Mycobacterium , Peroxiredoxins , Prevalence , Proteome , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Transposases , Tuberculosis , UDPglucose 4-Epimerase , VaccinesABSTRACT
Objective To study the location of mycobacterium tuberculosis(MTB) and whether the cellular immunity is induced after immunization of H37Ra in mice.Methods Totally 72 BALB/C mice were included.Thirty mice were intracutaneously injected of 0.1 ml H37Ra solution(about 10~(6) bacteria) at caudal region;thirty mice were injected of BCG of same quantity instead;twelve were free from immunization as control.Viable MTB were detected in spleen and lungs on day 15,30 and 60 after intracutaneous vaccination of H37Ra.The proliferation of T lymphocytes stimulated with purified protein derivative(PPD) was measured by MTT assay and the production of interleukin-2(IL-2) and soluble interleukin-2 receptor(sIL-2R) in the cultural supernatants of T lymphocytes was also determined by ELISA on day 30 and 60 after intracutaneous vaccination of H37Ra.Results Viable H37Ra or BCG could be located in the spleen and lungs in mice for at least 60 d.After H37Ra immunization,the stimulation index(SI) of T lymphocyte proliferation on day 30 and 60 was(2.81?0.63),(2.16?0.52) respectively,which was of no significant difference with that of BCG immunization,but of significant difference with that of non-immunized mice(P
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Objective:To investigate the cellular immune responses induced by Mycobacterium tuberculosis H37Ra and examine its protective efficacy against MTB in mice.Methods:Balb/c mice were divided into H37Ra,BCG and NS groups randomly,and intracutaneously injected at caudal region with H37Ra,BCG and NS respectively.At 8 weeks after immunization,some mice were sacrificed,and the percentages of CD3~+T,CD4~+T and CD8~+T-lymphocyte in spleen were detected by flow cytometry.The spleen lymphocytes of mice were cultured with PPD in vitro;the proliferation of splenocytes was detected by MTT assay.The productions of IFN-?in the supematant of culture were determined by ELISA.After 8 weeks of immunization,Balb/c mice were challenged via intraperitoneal route with MTB H37Rv strain.Four weeks later,mice were sacrificed,then the bacteria load in the lung were determined.Results:After 8 weeks of immunization,the percentages of CD3~+T and CD4~+T-lymphocyte in mice of H37Ra vaccination were significantly higher than those of the NS control subjects(41.63?1.68)%,(27.08?0.58)% vs (38.34?0.74)%,(24.37?1.60)%(P0.05).The proliferation of antigen-specific lymphocytes,the productions of IFN-?in the H37Ra group were significantly higher than those of the NS group(P0.05).At 4 weeks after challenge,compared with the NS group, the mean numbers of viable bacteria in the lung of the H37Ra group reduced 0.954log_(10)CFU(P0.05).Conclusion:Mycobacterium tuberculosis H37Ra could induce specific cellular immune responses in mice.The immunity effect of H37Ra is similar to BCG.
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Objective:To investigate the immune efficacy after sequential immunization with Mycobacterium tuberculosis DNA vaccine encoding mature form of Ag85B(pTB30m)and Mycobacterium tuberculosis H37Ra in mice.Methods:Much of highly pure plasmid DNA(pTB30m)extracted by alkaline lysis method was confirmed by restriction endonuclease digestion.Then,its DNA concentration and purity were determined by UV spectrophotometry.At various intervals(4weeks,8weeks)after sequential immunization,ELISA was used to detect the level of the serum antibody against PPD.Also,the spleen lymphocytes of mice were cultured with PPD in vitro.Lymphocyte transformation was detected by MTT assay.Results:Prepared pTB30m was highly pure and came to the needed concentration.Compared with Group Naive control,the specific antibody levels against PPD and the stimulation index(SI)of spleen lymphocytes were all statistically higher in Group DNA-85B/H37Ra(P0.05),as compared with Group DNA-85B/BCG,Group H37Ra and Group BCG,However,compared with Group H37Ra and Group BCG,the SI of mice was significantly larger in Group DNA-85B/H37Ra(P
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Objective:To research the proliferation of lymphocyte with stimulation of purified protein derivative (PPD) after immunization with H37Ra in mice.Methods:The rate of lymphocyte proliferation with stimulation of PPD was determined at day 30 and day 60 after subcutaneous immunization and celiac immunization with H37Ra in mice.Results:After immunization with H37Ra in mice,the rate of lymphocyte proliferation by celiac immunization route and subcutaneous immunization route was 21.98% and 24.85% respectively at day 30;and it was 25.50% and 25.02% respectively at day 60.The rate of lymphocyte proliferation immunized with H37Ra at day 30 and day 60 were much higher than that of non-immunization( P 0.05);there were no significant difference between subcutaneous immunization route and celiac immunization route of H37Ra in mice.Conclusion:H37Ra can induce anti-tuberculosis immune response.
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Aim To establish and evaluate the simple and effective animal model of rheumatoid arthritis in SD rats induced by heat-killed mycobacterium tuberculosis H37Ra(Mtb).Methods SD rats were immunized s.c.at the base of the tail with Mtb(1 mg/rat) in mineral oil,and then body weight,blood routine,clinical signs of arthritis were observed regularly.And also hind paw volume in SD rats with AA was tested.Meanwhile,T-lymphocyte subset was determined by flow cytometry in peripheral blood,the level of TNF-?in serum and IL-1 and IL-6 and vascular endothe-lial growth factor(VEGF) in joint tissue were tested by ELISA.The ankle pathological change and radiographic evidence of the hind paws of SD rats were observed to evaluate the AA model effects and availability in SD rats.Results When compared with normal control rats,the total number of white blood cells,platelets and monocytes in peripheral blood and ratio of CD4/CD8 was markedly raised after immunization with Mtb.But body weight of model rats was significantly low from day 14 to 28 after injection of Mtb.The inflammatory arthritic lesions such as edema and erythema in the paws appeared after 9～10 d.The peak of inflammation was 14～16 d.The level of TNF-?in serum and IL-1,IL-6 and VEGF in joint tissue was significantly higher in SD rats with Mtb than that in normal control rats at d 28.The histopathological examination revealed cellular infiltration,synovial and cartilage hyperplasia in ankle joints of SD rats.Conclusions The model induced by Mtb in SD rats is more similar in clinical characterization of rheumatoid arthritis in human being.It is a more efficient and simpler manipulative procedure for screening of new anti-arthritis drugs and experimental studying of anti-rheumatoid arthritis.
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0.05).But at sixtieth day after immunization,the number of H37Ra growing in the spleen or lung was significantly larger than those of BCG(P
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Objective:To explore the activity of tumor necrosis factor-?(TNF-?)from celiac macrophage after immunization of H37Ra in mice.Methods:The activity of TNF-? from celiac macrophage was measured by biological method at 30 th day and 60 th day after subcutaneous immunization and celiac immunization of H37Ra in mice.Results:After immunization of H37Ra in mice,the activity of TNF-? by celiac immunized route and subcutaneous immunized route was 22.06U/ml and 22.63U/ml respectively at 30 th day;and it was 36.76U/ml and 33.78U/ml respectively at 60 th day.The activity of TNF-? immunized with H37Ra at 30 th day and 60 th day was much higher than that of non-immunization group( P 0.05),however,the activity of TNF-? in H37Ra immunization group was much higher than that in BCG immunization group at 60 th day( P