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Mycobacterium smegmatis enhances shikonin-induced immunogenic cell death—an efficient in situ tumor vaccine strategy 被引量:1
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作者 Zhaoye Qian Zhe Zhang +4 位作者 Lanqi Cen Yaohua Ke Jie Shao Manman Tian Baorui Liu 《Journal of Biomedical Research》 CAS CSCD 2024年第4期369-381,共13页
Tumor vaccines are a promising avenue in cancer immunotherapy.Despite the progress in targeting specific immune epitopes,tumor cells lacking these epitopes can evade the treatment.Here,we aimed to construct an efficie... Tumor vaccines are a promising avenue in cancer immunotherapy.Despite the progress in targeting specific immune epitopes,tumor cells lacking these epitopes can evade the treatment.Here,we aimed to construct an efficient in situ tumor vaccine called Vac-SM,utilizing shikonin(SKN)to induce immunogenic cell death(ICD)and Mycobacterium smegmatis as an immune adjuvant to enhance in situ tumor vaccine efficacy.SKN showed a dose-dependent and time-dependent cytotoxic effect on the tumor cell line and induced ICD in tumor cells as evidenced by the CCK-8 assay and the detection of the expression of relevant indicators,respectively.Compared with the control group,the in situ Vac-SM injection in mouse subcutaneous metastatic tumors significantly inhibited tumor growth and distant tumor metastasis,while also improving survival rates.Mycobacterium smegmatis effectively induced maturation and activation of bone marrow-derived dendritic cells(DCs),and in vivo tumor-draining lymph nodes showed an increased maturation of DCs and a higher proportion of effector memory T-cell subsets with the Vac-SM treatment,based on flow cytometry analysis results.Collectively,the Vac-SM vaccine effectively induces ICD,improves antigen presentation by DCs,activates a specific systemic antitumor T-cell immune response,exhibits a favorable safety profile,and holds the promise for clinical translation for local tumor immunotherapy. 展开更多
关键词 MYCOBACTERIUM smegmatis SHIKONIN IMMUNOGENIC cell death tumor vaccines IMMUNOGENICITY CYTOTOXICITY
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无细胞M.Smegmatis菌苗的研究与应用
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作者 徐苗 陈保文 +2 位作者 沈小兵 苏城 王国治 《中国防痨杂志》 CAS 北大核心 2002年第z1期1-2,共2页
  结核病是世界卫生组织(WHO)公布的危害人类健康的十大疾病之一.我国目前同样面临结核病的严峻挑战,2000年全国结核病流行病学调查结果表明,我国目前有近半人口(约5.5亿)感染了结核菌,其中有5000多万人为结核菌素实验强反应者(硬结...   结核病是世界卫生组织(WHO)公布的危害人类健康的十大疾病之一.我国目前同样面临结核病的严峻挑战,2000年全国结核病流行病学调查结果表明,我国目前有近半人口(约5.5亿)感染了结核菌,其中有5000多万人为结核菌素实验强反应者(硬结直径≥15m或有水泡、坏死),因其结核发病率极高,被称为结核感染高危人群.如何控制这些人潜在的内源性复发、外源性再感染和发病,是结核病控制的重点.…… 展开更多
关键词 菌苗 动物模型 免疫功能 疫苗 M.smegmatis 卡介苗 结核菌苗 无细胞 亚细胞 结核菌感染 结核病 豚鼠 荷兰猪
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Recombinant Mycobacterium smegmatis expressing Hsp65-hIL-2 fusion protein and its influence on lymphocyte function in mice 被引量:1
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作者 Xiao-Qing Guo Yan-Ming Wei Bo Yu 《Asian Pacific Journal of Tropical Medicine》 SCIE CAS 2012年第5期347-351,共5页
Objective:To coastruct a strain of recombinant Mycobacterium smegmatis expressing the heat shock protein 65(Hsp65) and human interleukin 2(IL-2) fusion protein(rMS-Hsp65/IL-2) and to explore the effect of this constru... Objective:To coastruct a strain of recombinant Mycobacterium smegmatis expressing the heat shock protein 65(Hsp65) and human interleukin 2(IL-2) fusion protein(rMS-Hsp65/IL-2) and to explore the effect of this construct on lymphocyte function in mice.Methods:The fusion gene encoding Hsp65-hIL-2 was cloned into shuttle vector pSMT3.The recombinant plasmid pSMT3- Hsp65-hIL-2 was transferred to Mycobacterium smegmatis by electroporation.Positive clones were selected by hygromycin and identified by PCR.The expression of fusion protein Hsp65- hIL-2 was verified using indirect immunofluorescence staining.Mice were immunized for two times by subcutaneously injection with 1×10~6 CFU rMS-Hsp65/IL-2 at a three-week interval.Two weeks after the second immunization,mice were sacrificed and the serum samples were collected for determination of anli-Hsp65 specific IgG.Splenic lymphocytes were isolated and treated with the rMS-Hsp65/IL-2 to determine lymphocytic proliferation activity by MTT assay.IFN-γand IL-2 in the medium of the treated cells were also determined by ELISA.Results:Successful construction of rMS-Hsp65/IL-2 was verified by PCR and immunofluorescence staining.Compared to the splenic lymphocytes isolated from mice immunized with Bacille Calmette-Guerin or mice immunized with Mycobacterium smegmatis alone,the splenic lymphocytes isolated from mice immunized with rMS-Hsp65/IL-2 showed a marked increase in the proliferation of lymphocytes, together with an increased production of important cytokines such as IFN- 7 and IL-2. Conclusions:rMS-Hsp65/IL-2 markedly enhances lymphocyte function.Therefore,the fusion protein generated by rMS-Hsp65/lL-2 may be of potential value in generating an effective vaccine against tuberculosis. 展开更多
关键词 MYCOBACTERIUM tuberculosis MYCOBACTERIUM smegmatis HSP65 IL-2 Fusion protein Lymphocytes MICE
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Cloning and Expression of a Novel Phytase Gene (phyMS) from <em>Mycobacterium smegmatis</em>
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作者 Tamrin Nuge Yumi Zuhanis Has-Yun Hashim +1 位作者 Abd-El Aziem Farouk Hamzah Mohd Salleh 《Advances in Enzyme Research》 2014年第1期27-38,共12页
Phytase, also known as phytate-degrading enzyme, catalyzes the hydrolysis of phytate (inositol hexakisphosphate) with sequential release of phosphate and lower inositol phosphate. We report here a new plasmid construc... Phytase, also known as phytate-degrading enzyme, catalyzes the hydrolysis of phytate (inositol hexakisphosphate) with sequential release of phosphate and lower inositol phosphate. We report here a new plasmid construct designated as pMSuia from pBAD-TOPO that harbors a 1.1 kb phytase gene (phyMS) from Mycobacterium smegmatis, and expression as well as characterization of the purified recombinant M. smegmatis phytase. DNA sequencing analysis and multiple alignment exercise indicated that the M. smegmatis phytase is different from both known acid and alkaline phytases. The active ~45 kDa recombinant enzyme was expressed and confirmed by enzyme assay and Western blot analyses. Ni-NTA affinity purified recombinant M. smegmatis phytase exhibited specific activity of 233.51 U/mg, optimal pH of 3 and 7 and optimal temperature of 60°C. The purified enzyme retains almost 30% of the initial activity after incubation at 90°C for 60 min. The enzyme showed broad substrate specificity with Km and Vmax of the recombinant enzyme for sodium phytate substrate of 0.105 ± 0.016 mM and 26.93 ± 1.21 mM min-1, respectively. 展开更多
关键词 MYCOBACTERIUM smegmatis Thermostability PHYTASE BROAD pH Activity BROAD Substrate
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Screening of Several Anti-Infectives for in Vitro Activity against Mycobacterium smegmatis
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作者 Grace Lovia Allotey-Babington Henry Nettey +6 位作者 Philip Debrah Ofosua Adi-Dako Clement Sasu Anastasia Antwi Yvonne Darko Newriza Nartey Jida Asare 《Advances in Microbiology》 2014年第16期1197-1203,共7页
Aim: To evaluate in vitro the effectiveness of several anti-infective agents alone or in combination against Mycobacterium smegmatis. Method: A convenient stratified sampling method was used to obtain selected anti-in... Aim: To evaluate in vitro the effectiveness of several anti-infective agents alone or in combination against Mycobacterium smegmatis. Method: A convenient stratified sampling method was used to obtain selected anti-infective agents. For individual drug samples, Minimum Inhibitory Concentrations (MIC) were obtained using the agar-well plate diffusion technique. Fractional Inhibitory Concentration Indices (FICI) were calculated for drug combinations using their MIC as obtained from the broth dilution method. Results: Of the thirty (30) anti-infective agents analyzed, ten (10) had MIC equivalent to or better than rifampicin (reference TB drug). Seven (7) drugs had MIC higher than rifampicin, while twelve (12) showed no growth inhibition of M. smegmatis. Analysis of the effect of drug combinations on M. smegmatis indicated that four (4) combinations, including rifampicin/ethambutol showed synergism. One (1) was additive, two (2) were indifferent and one (1) combination showed antagonism. Conclusion: Notable in the results obtained was the high effectiveness of the carbapenems in inhibiting the growth of M. smegmatis. Carbapenems, though not indicated for TB treatment, has a potential of playing a significant role in the treatment of tuberculosis. Also the drug combinations which showed synergism, especially those that involved the macrolide antibiotics, should further be investigated. These results have to be confirmed by in vivo clinical studies to define their roles in tuberculosis treatment. 展开更多
关键词 Antimicrobials Minimum INHIBITORY Concentration M. smegmatis TUBERCULOSIS
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Disruption of <i>Mycobacterium smegmatis</i>Biofilms Using Bacteriophages Alone or in Combination with Mechanical Stress
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作者 Brendan Kiefer John L. Dahl 《Advances in Microbiology》 2015年第10期699-710,共12页
Environmental mycobacteria are capable of forming biofilms in low-nutrient environments, and these biofilms may act as reservoirs for opportunistic infections. The purpose of this study was to determine if bacteriopha... Environmental mycobacteria are capable of forming biofilms in low-nutrient environments, and these biofilms may act as reservoirs for opportunistic infections. The purpose of this study was to determine if bacteriophages could disrupt existing biofilms of acid-fast staining Mycobacterium smegmatis. Using the MBEC 96-well plastic peg assay system, M. smegmatis biofilms were created and then tested for their stability in the presence of mycobacteriophages isolated from a Minnesota sphagnum peat bog. All phages tested were lytic and were observed to have weak, intermediate, and strong abilities to disrupt M. smegmatis biofilms. The formation of biofilms was severely impaired in the presence of mycobacteriophages. Phage treatment was also shown to augment?M. smegmatis biofilm disruption by mechanical forces of sonication or water flow. Our study shows that, as with biofilms of Gram-positive and Gram-negative bacteria, mycobacterial biofilms are also susceptible to destruction by bacteriophages. 展开更多
关键词 Mycobacterium smegmatis Biofilm MYCOBACTERIOPHAGE BACTERIOPHAGE
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Characteristics of Mycobacterium tuberculosis serine protease Rv1043c in enzymology and pathogenicity in mice
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作者 TANG Yang-yang CUI Ying-ying +4 位作者 JIANG Yan-yan SHAO Ming-zhu ZANG Xin-xin DANG Guang-hui LIU Si-guo 《Journal of Integrative Agriculture》 SCIE CAS CSCD 2023年第12期3755-3768,共14页
The serine proteases of Mycobacteria tuberculosis(Mtb)are important contributors to the process of bacterial invasion and its pathogenesis.In the present study,we systematically characterized the role of the Rv1043c p... The serine proteases of Mycobacteria tuberculosis(Mtb)are important contributors to the process of bacterial invasion and its pathogenesis.In the present study,we systematically characterized the role of the Rv1043c protein in Mycobacterium infection by purifying the Rv1043c protein in Escherichia coli and constructing a Mycobacterium smegmatis(Msg)strain overexpressing Rv1043c(Msg_Rv1043c).We found that Rv1043c had serine protease activity and localized to the surface of Mtb.We determined that the optimal pH and temperature for the Rv1043c serine protease were 9.0 and 45°C,respectively.Moreover,the serine protease activity of Rv1043c was enhanced by divalent metal ions of Ca^(2+)and Mg^(2+).Site-directed mutagenesis studies demonstrated that the serine 279 residue in Rv1043c plays a catalytic role.Additionally,mouse model studies confirmed that Rv1043c significantly enhanced the survival of Msg in vivo,induced pulmonary injury and lung cell apoptosis,and promoted the release of pro-inflammatory cytokines interleukin-1βand interleukin-6 in mice.This study presents novel insights into the relationship between mycobacterial serine protease and the pathogenesis of the disease. 展开更多
关键词 Mycobacterium tuberculosis Mycobacterium smegmatis serine protease Rv1043c PATHOGENICITY
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基于MspA蛋白质纳米孔传感器的主客体化学研究 被引量:4
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作者 段静 卓莎 +2 位作者 姚付军 张亚妮 亢晓峰 《分析化学》 SCIE EI CAS CSCD 北大核心 2016年第12期1801-1807,共7页
以大肠杆菌原核系统表达的方法制备了MspA蛋白质纳米孔,并利用表面活性剂高温提取获得高活性MspA蛋白纳米孔。实验结果表明,0.5%的提取剂90℃提取30 min为最佳提取条件。利用制备的MspA纳米孔在单分子水平上研究了其与多种环糊精的相互... 以大肠杆菌原核系统表达的方法制备了MspA蛋白质纳米孔,并利用表面活性剂高温提取获得高活性MspA蛋白纳米孔。实验结果表明,0.5%的提取剂90℃提取30 min为最佳提取条件。利用制备的MspA纳米孔在单分子水平上研究了其与多种环糊精的相互作用,结果表明,在众多环糊精分子中,全-6-氨基-β-环糊精(am_7-β-CD)与MspA蛋白纳米孔相互作用较强且阻塞平台较一致,其滞留时间随电压增大而逐渐减小,但阻塞电流受电压影响不大。筛选使用am_7-β-CD作为适配体非共价键装配MspA纳米孔,在单分子水平上研究主客体化学反应。加入am_7-β-CD后,当盐酸金刚烷胺小分子浓度为50 nmol/L时,即出现二级阻塞平台,其滞留时间随电压增大而逐渐减小。这种新型蛋白质纳米孔单分子传感器可用于有机小分子的检测识别,极大拓宽了MspA纳米孔分析技术的的应用范围。 展开更多
关键词 MYCOBACTERIUM smegmatis PORIN A 纳米通道 单分子检测 环糊精 盐酸金刚烷胺
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The Construction of Schistosoma Japonicum Vaccine BCG-Sj26GST and Its Identification 被引量:1
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作者 HUANGFU Yongmu , ZHENG Bo , CHENG Jizhong , LIANG Juqing , FENG Zuohua Department of Medical Molecular Biology, Research Center of Experimental Medicine, Tongji Medical University,Wuhan 430030 《Journal of Huazhong University of Science and Technology(Medical Sciences)》 SCIE CAS 1999年第3期161-165,169,共6页
Summary: The expression of foreign gene, Schistosoma Japonicum 26 ku antigen (Sj26GST), in Bacillus Calmette Guerin (BCG), Mycobacterium ( M. smegmatis ) and Escherichia coli ( E. coli ) were stud... Summary: The expression of foreign gene, Schistosoma Japonicum 26 ku antigen (Sj26GST), in Bacillus Calmette Guerin (BCG), Mycobacterium ( M. smegmatis ) and Escherichia coli ( E. coli ) were studied. The cDNA fragment encoding Sj26GST was amplified by PCR using plasmid pGEX, which could express Sj26GST in E. coli as template. The Sj26GST cDNA was cloned into the downstream of human M. tuberculosis heat shock protein (hsp) 70 promoter with correct reading frame, and then the DNA fragment containing hsp70 promoter and Sj26GST gene were subcloned together into E. coli Mycobacteria shuttle plasmid pBCG 2000 to construct the expression shuttle plasmid pBCG Sj26. The recombinant BCG and M. smegmatis mc 2 155, which were electroplated with pBCG Sj26, could express Sj26GST and the recombinant Schistosoma Japonicum vaccine BCG Sj26GST was made. The recombinant Sj26GST (rSj26GST) were soluble and could be observed on SDS PAGE at molecular weight of 26 ku. The content of rSj26GST accounted for 15 % and 10 % of total bacterial protein in BCG and M. smegmatis respectively. The results of Western blot showed the combination of rSj26GST with antibody of GST. 展开更多
关键词 BCG M. smegmatis shuttle plasmid Schistosoma Japonicum 26ku antigen gene VACCINE gene expression
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GP30 of the mycobacteriophage CASbig impairs mycobacterial adaptation during acidic stress and in macrophages
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作者 WU LI JIANGZHE SI +10 位作者 SHUAI QIU JING LUO HUIQIONG SUN XIAOQING LI ZHIBIN WAN WEI GAO HANLU ZOU LEI ZHANG XIAOHONG XIANG YANZHANG LI TIESHAN TENG 《BIOCELL》 SCIE 2020年第4期695-701,共7页
The rapid emergence of multidrug-resistant and extensively drug-resistant Tuberculosis retrieved intense interest in phage-based therapy.This old approach,which was abandoned in the west in the 1940s but is generating... The rapid emergence of multidrug-resistant and extensively drug-resistant Tuberculosis retrieved intense interest in phage-based therapy.This old approach,which was abandoned in the west in the 1940s but is generating renewed interest,has stimulated fresh research on mycobacteriophages and their lytic efficiency against their hosts.GP30 is a novel protein of the mycobacteriophage CASbig with undiscovered function.In this study,we analyzed the role of CASbig gp30 in the host Mycobacterium smegmatis.Overexpression of gp30 in the host led to reduced growth in acidic medium and attenuated the intracellular survival rate of M.smegmatis inside the THP-1 macrophages,which may be linked to the altered lipid profile of the recombinant bacterial cell wall.In a word,this study suggested that gp30,a novel gene from a mycobacteriophage,modulated lipid composition and content to hamper the survivability of bacteria under stress conditions. 展开更多
关键词 MYCOBACTERIUM smegmatis gp30 CASbig ACIDIC STRESS Macrophage
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Comparative Study on the Immunogenicity between Recombinant MS-Sj26GST Vaccine and Recombinant BCG-Sj26GST Vaccine in Schistosoma japonicum
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作者 戴五星 高红 +3 位作者 黄海浪 袁野 胡佳杰 皇甫永穆 《Journal of Huazhong University of Science and Technology(Medical Sciences)》 SCIE CAS 2003年第3期213-215,218,共4页
The BALB/c mice were immunized with rMS Sj26GST and rBCG Sj26GST vaccine in Schistosoma japonicum by subcutaneous injection After they were immunized for 8 weeks, the eyeballs were removed to get blood and macroph... The BALB/c mice were immunized with rMS Sj26GST and rBCG Sj26GST vaccine in Schistosoma japonicum by subcutaneous injection After they were immunized for 8 weeks, the eyeballs were removed to get blood and macrophages of abdominal cavity and spleen cells were harvested The lymphocytic stimulating index (SI) was used to measure the cellular proliferating ability and NO release was used to measure the phagocytic activity of the macrophages By using ELISA kit, the levels of interleukin 2 (IL 2) and interferon γ (IFN γ) in serum and the splenic lymphocytic cultured supernatant were detected The results showed that after the mice were immunized with 10 6 CFU of rMS Sj26GST and rBCG Sj26GST vaccine separately by subcutaneous injection, proliferating ability of splenic lymphocytes in the mice showed no difference ( P >0 05), but both were significantly increased as compared with that in the control group( P <0 05); The contents of NO in the intraperitoneal macrophages of rMS Sj26GST vaccine group were significantly lower than in the control group ( P <0 001) and rBCG Sj26GST vaccine group ( P <0 01); The levels of serum IL 2 in the rMS Sj26GST vaccine group were significantly increased as compared with that in the control group ( P <0 001), vector group ( P <0 01) and rBCG Sj26GST vaccine group ( P <0 05); The contents of serum IFN γ in the rMS Sj26GST vaccine group were significantly increased as compared with that in the control group ( P <0 01) and rBCG Sj26GST vaccine group ( P <0 05) The contents of IFN γ in the cultured supernatant were significantly lower than those of rBCG Sj26GST vaccine group ( P <0 001), but were significantly increased as compared with that in the control group ( P <0 01) It was indicated that both vaccines could enhance the immune response of the mice, but rMS Sj26GST vaccine had stronger immunogenicity than rBCG Sj26GST vaccine 展开更多
关键词 Schistosoma japonnicum Mycobacterium smegmatis mc 2155(MS) BCG VACCINE IMMUNOGENICITY
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Siderochelins with anti-mycobacterial activity from Amycolatopsis sp. LZ149 被引量:1
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作者 LU Chun-Hua YE Fang-Wen SHEN Yue-Mao 《Chinese Journal of Natural Medicines》 SCIE CAS CSCD 2015年第1期69-72,共4页
Three new compounds, namely siderochelins D (2), E (3), and F (4), together with one known siderochelin A (1), were isolated from Amycolatopsis sp. LZ149 and elucidated by spectroscopic analyses includinglD- a... Three new compounds, namely siderochelins D (2), E (3), and F (4), together with one known siderochelin A (1), were isolated from Amycolatopsis sp. LZ149 and elucidated by spectroscopic analyses includinglD- and 2D-NMR and X-ray single crystal diffraction. Compounds 1-3 showed antibacterial activity against Mycobacterium smegmatis. 展开更多
关键词 Amvcolatopsis sp. LZ149 Siderochelin Bioassay-guided Mycobacterium smegmatis X-ray single crystal diffraction
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