去甲斑蝥素抑制肺癌细胞增殖的体外研究

为分析去甲基斑蝥素二钠盐对人和小鼠肺癌细胞增殖的抑制作用,体外培养人非小细胞肺癌A549细胞和小鼠肺癌Lewis细胞,用WST-1还原法检测不同浓度去甲斑蝥素(norcantharidin,NCTD)对肿瘤细胞增殖的抑制作用,并用流式细胞术分析细胞周期;免疫印迹技术分析MAPK激酶表达。NCTD抑制A549和Lewis增殖的IC50值分别为13.5和9.5μmol/L;0、6.25、12.50和25.00μmol/L NCTD处理24h,诱导A549细胞G2/M期阻滞率分别为26.30%、29.44%、49.83%和81.35%。结果表明,抑制肿瘤细胞增殖可能是去甲基斑蝥素抗肿瘤作用的分子机制之一。     

蒲公英黄酮与多糖对DPPH清除能力比较研究

考察不同质量浓度蒲公英黄酮和多糖对DPPH(1,1-二苯基-2-三硝基苯肼)的清除能力并进行比较。试验利用醇沉法超声提取蒲公英黄酮,水提法超声提取蒲公英多糖,采用紫外分光光度计法测定此黄酮及多糖提取物对DPPH自由基的清除效果,利用统计学方法分析黄酮及多糖清除DPPH自由基的差异性。结果表明:当黄酮质量浓度为1.6 mg/m L时,DPPH清除效果最佳,清除率达96.11%,IC50(抑制剂的半抑制质量浓度)值为0.38 mg/m L;当多糖质量浓度为1.4 mg/m L时,DPPH清除效果最佳,清除率达89.33%,IC50值为0.51 mg/m L;统计学结果显示蒲公英黄酮及多糖对DPPH自由基清除效果差异显著(P0.05)。研究首次对蒲公英黄酮与多糖清除DPPH自由基效果进行比较,蒲公英黄酮及多糖均对DPPH自由基清除效果较好,但黄酮的清除效果显著高于多糖的清除效果(P0.05)。试验结论为进一步开展蒲公英黄酮研究奠定理论基础,也为开发利用蒲公英资源提供理论依据。     

桑黄粗多糖和蛹虫草粗多糖协同增效诱导肺癌细胞A549 S期阻滞及凋亡的研究

通过细胞抑制试验研究桑黄子实体粗多糖与蛹虫草子实体粗多糖混合后的粗多糖对肺癌细胞A549的抑制作用,为利用药用真菌开发抗肿瘤药物提供试验依据。采用MTT法检测0.8 mg/m L的桑黄粗多糖和蛹虫草粗多糖对肺癌细胞A549增殖均有一定的抑制作用,作用48 h后的抑制率分别为32.95%、41.93%;将0.8 mg/m L桑黄粗多糖和0.8 mg/m L蛹虫草粗多糖按1∶1体积比混合的粗多糖则表现出协同增效抑制A549细胞增殖作用,抑制率可达54.71%。用流式细胞计数法测试分析的结果是:0.8 mg/m L蛹虫草粗多糖可影响A549细胞周期各时相的分布,而0.8 mg/m L桑黄粗多糖则对细胞周期无明显作用,但0.8 mg/m L混合粗多糖对A549细胞周期各时相分布的影响显著增强,G0/G1期细胞比率从75.39%±2.49%降低至64.43%±2.12%(P0.01),S期细胞比率从16.61%±0.87%升高至27.97%±2.82%(P0.01);混合粗多糖作用48 h后,A549细胞的晚期凋亡率从正常组的1.92%±0.55%提高至20.75%±0.24%,坏死细胞率由正常组的1.88%±0.43%提高至9.64%±0.42%,表现出明显的协同增效诱导A549细胞凋亡的作用。荧光定量PCR检测结果表明,A549细胞分别用0.8mg/m L的桑黄粗多糖和蛹虫草粗多糖及其混合粗多糖处理后,细胞内周期调控相关蛋白基因Cyclin A、CDK4的mRNA转录水平均显著下调,Cyclin D1、CDK2的mRNA转录水平显著上调,凋亡相关基因Bid、Bak、Cyt-C、Caspase3的mRNA转录水平也显著上调。研究结果提示桑黄粗多糖和蛹虫草粗多糖混合使用,具有协同增效抑制肺癌细胞A549增殖的作用,推测其通过调控细胞周期和细胞凋亡相关基因的表达,导致Cyt-C释放及启动caspase级联反应,从而诱导细胞凋亡。     

Undifilum oxytropis次级代谢产物GC-MS分析及抗肿瘤活性研究

研究疯草内生真菌(Undifilum oxytropis),次级代谢产物成分与活性,通过GC-MS对菌丝体及发酵液进行了分析,并通过MTT法测定抗肿瘤活性。结果表明,菌丝体和发酵液中均以烷烃和酯类物质为主,菌丝体中检出活性物质角鲨烯(0.863%);菌丝体对HCT-8人回盲肠癌细胞和OS-RC-2人肾癌细胞抑制率均较高,发酵液对HCT-8人回盲肠癌细胞抑制率相对较弱。     

莪术油注射液体外抗猪细小病毒的试验

制备莪术油注射液,评价莪术油注射液体外对猪细小病毒(PPV)7909标准毒株的作用效果。在PK-15细胞上,通过不同加药方式观察PK-15细胞病变(CPE),用MTT法测定细胞活力。结果在PK-15细胞上,药物最大安全浓度(TD0)为86.4μg/mL,半数感染浓度(TD50)为162.2μg/mL;在药物安全浓度范围内,莪术油注射液对PPV 7909标准毒株的半数抑制浓度(IC50)为1.865μg/mL,半数阻断浓度(IC50)为3.75μg/mL,半数直接杀灭作用浓度(IC50)为81.2μg/mL。结果表明莪术油对PPV 7909标准毒株有明显的体外抑制作用,但直接杀灭作用不明显。     

中药牛皮消多糖体外抗新城疫病毒作用的研究

为探讨牛皮消多糖(CAP)对体外抗新城疫病毒(NDV)能力的影响,本试验用水提醇沉法提取牛皮消多糖,通过苯酚硫酸法和红外光谱对牛皮消多糖进行检测,采用MTT法测定牛皮消多糖对鸡胚成纤维细胞(CEF)的安全浓度,多糖的安全浓度统一为625μg/mL,选择安全浓度范围内的(39.06～625μg/mL)5种浓度的各级多糖,用3种加药方式(先加多糖后接种病毒、先接种病毒后加多糖、多糖和病毒感作后加药)检测牛皮消多糖对NDV的阻断、抑制及直接杀灭作用。结果显示,先接病毒后加多糖组和多糖与病毒感作后加药组CAP_(50)、CAP_(30)、CAP_t、CAP_(80)和CAP_(70)均能显著抑制NDV感染CEF的能力,两组各级多糖抑制率分别为131.23%、110.12%、108.15%、100.24%、93.07%和61.76%、43.73%、44.51%、29.02%、37.45%;先加多糖后接病毒组CAP_(50)、CAP_(30)能显著抑制NDV感染CEF的作用(P0.05),阻断作用下对NDV的抑制率可达到15.82%、8.33%,其余各组在安全浓度范围内对NDV无抑制作用。各组中药牛皮消多糖均有较好的抗NDV活性,其中对NDV的抑制作用和直接杀灭作用强于对其阻断作用。CAP_(50)及CAP_(30)的抗NDV活性较强,可作为进一步研究材料并具备一定的研究价值。     

苦树侧枝不同极性部位的抗氧化活性及对肺癌细胞A549的生长抑制作用

为研究苦树侧枝不同极性部位体外抗氧化、抗肿瘤活性，依次用二氯甲烷、乙酸乙酯、正丁醇和水对苦树侧枝的乙醇提取物进行萃取得到不同极性部位萃取物，采用ABTS 法、DPPH法和普鲁士蓝法测定4种不同极性部位的体外抗氧化能力，用噻唑兰（MTT）法检测A549细胞活力。结果显示：乙酸乙酯部位对2，2-连氮（3-乙基苯并噻唑咻-6磺酸）二氨盐（ABTS）自由基、二苯代苦味酰基（DPPH）自由基的清除率效果及对铁的还原能力均最强，其IC50（或A0.5）值分别为0.015、0.057、1.115 mg/mL。正丁醇部位对A549细胞的增殖抑制作用最高，抑制率为69.764%，IC50为100.642 μg/mL，其余部位对A549细胞的增殖抑制作用从大到小顺序依次为二氯甲烷部位、乙酸乙酯部位和水部位。结果说明苦树侧枝的乙醇提取物不同极性部位的抗氧化和抗肿瘤活性存在差异，部分极性部位效果较好，研究结论可为苦树的进一步深入研究和合理应用提供参考。 [关键词] 苦树| 侧枝|抗氧化|抗肿瘤     

构建稳定转化的BmN细胞表达人白介素-28A及表达产物的体外抗肿瘤活性

为建立能表达人白介素-28A(hIL-28A)基因的稳定转化家蚕卵巢细胞(BmN)系,构建了重组表达载体pIZT/V5-His-hIL-28A并转染BmN细胞,采用终浓度为300～400μg/mL的博莱霉素(zeocin)筛选2个月后,获得了稳定转化BmN细胞系。SDS-PAGE和Western blotting检测显示在稳定转化BmN细胞中表达的重组hIL-28A蛋白分子质量约为26kD;用ELISA试剂盒测定hIL-28A的表达水平约为2.035×10-5ng/个细胞。用噻唑蓝法(MTT法)检测hIL-28A的体外抗肿瘤活性,结果显示其对肺癌细胞A549、急性早幼粒细胞白血病细胞HL60、肝癌细胞BEL-7402和乳腺癌细胞M231的半数抑制浓度(IC50)分别为3.21、2.84、6.29和9.32ng/mL。研究结果表明hIL-28A可在稳定转化BmN细胞中表达,表达产物具有体外抗肿瘤活性。     

蛹虫草多糖对鸡脾脏淋巴细胞的影响

试验将7种浓度蛹虫草多糖(CMP)单独或与刀豆蛋白A(ConA)、脂多糖(LPS)同时加入鸡脾脏淋巴细胞的培养体系中,培养48 h时用MTT法测定淋巴细胞增殖(A570值)的变化;将5种浓度的CMP 40%乙醇提取物(CMP40)、CMP 50%乙醇提取物(CMP50)分别与ConA加入到鸡脾淋巴细胞的培养体系中,培养24 h后收集细胞培养上清液,测定白细胞介素-2(IL-2)和γ-干扰素(IFN-γ)的含量。结果显示,CMP在31.25~2 000μg/mL时,CMP40+ConA组的A570值显著高于ConA对照组;CMP在31.25~500μg/mL时,CMP50+ConA组A570值显著高于ConA对照组,CMP40+LPS组、CMP50+LPS组的A570值显著高于LPS对照组;CMP40在125~1 000μg/mL时、CMP50在500μg/mL时,各多糖组的A570值显著高于细胞对照组;CMP40和CMP50在250~1 000μg/mL时,各多糖组的IL-2和IFN-γ浓度显著高于对照组。结果表明,CMP40和CMP50在合适的剂量能单独或协同ConA、LPS刺激鸡的T、B淋巴细胞增殖,促进淋巴细胞分泌IL-2和IFN-γ,从而增强鸡的细胞免疫能力。     

蒲公英内生真菌PG2分子鉴定及抑菌物质的研究

药用植物内生真菌是新型抗菌药物的微生物来源。试验以清热解毒类中草药蒲公英中分离的高抑菌活性的内生真菌PG2为研究对象,通过ITS序列分析确定其种属地位;对其发酵液中高效抑菌活性物质的类别、热稳定性及抑菌物质的极性进行了初步研究,并对高抑菌活性的物质通过柱层析分离,以红外光谱分析其结构。结果表明:PG2经分子生物学鉴定为半知菌门、丛梗孢科、尾孢霉属;活性物质为多糖类物质,具有一定的热稳定性。抑菌活性物质主要分布于乙酸乙酯和正丁醇的萃取相中。通过对正丁醇萃取相的抑菌活性物质硅胶柱层析,分离出活性成分SP1,经红外光谱法测定其含有羰基、羟基等官能团结构。结论:PG2为产高效抑菌物质的内生真菌,分离出的活性物质SP1为含有羰基、羟基等官能团的多糖类物质。     

Preventive and Therapeutic Effects of Clostridium butyricum-longan Polysaccharide Fermentation Broth on Mouse Model with Ulcerative Colitis

The aim of the study was to observe the preventive and therapeutic effect of Clostridium butyricum-longan polysaccharide fermentation broth on mouse model with ulcerative colitis (UC).30 SPF male BALB/c mice were randomly divided into blank group,model group and fermentation broth group,10 mice/group.According to the dosage standard of 10 mL/kg body weight,sterile deionized water was administered to the blank group and model group,and Clostridium butyricum-longan polysaccharide fermentation liquid was administered to the fermentation broth group for 7 days.From the 8th day,the model group and the fermentation broth group were allowed to drink 5% dextran sulfate sodium (DSS) solution for 7 consecutive days to induce the UC model,and the blank group was free to drink sterile water.On the 14th day,all mice were weighed,and DAL scores and clinical scores were scored.Then the mice were dissected after eyeball collection and blood sampling.The length of the colon was measured and scored for histological damage of the colon,sections were made after formalin fixation.The content of IL-6,IL-10 and TNF-α in serum were detected by ELISA.The results showed that UC model was established successfully and symptoms were obvious in the model group.Compared with the model group,the weight loss of the mice in the fermentation broth group was significantly reduced (P<0.05),DAL score,clinical score,colon shortening degree and histological injury score were extremely significantly reduced (P<0.01).The colon tissue of mice in the fermentation broth group showed a relatively complete epithelial structure and crypts,and only part of the inflammatory cells invaded the epithelial tissue.ELISA results showed that the IL-6 and TNF-α levels in the model group were significantly higher than those in the fermentation broth group (P<0.05),and the IL-10 content was significantly lower than that in the fermentation broth group (P<0.05).It showed that Clostridium butyricum-longan polysaccharide fermentation broth had a certain preventive effect on DSS-induced UC in mice,and its mechanism might be related to reducing the content of IL-6 and TNF-α,increasing the content of IL-10,and inhibiting inflammation.     

丁酸梭菌-龙眼多糖发酵液对小鼠溃疡性结肠炎的防治作用

试验旨在观察丁酸梭菌-龙眼多糖发酵液预防给药对溃疡性结肠炎（ulcerative colitis，UC）模型小鼠的防治作用。30只SPF级雄性BALB/c小鼠随机分成空白组、模型组和发酵液组，10只/组。按10 mL/kg体重的剂量标准，空白组和模型组灌胃无菌去离子水，发酵液组灌胃丁酸梭菌-龙眼多糖发酵液，灌胃持续7 d。第8天开始，模型组和发酵液组连续7 d自由饮用5%葡聚糖硫酸钠（dextran sulfate sodium，DSS）溶液诱导UC模型，空白组自由饮用无菌水。试验第14天，称量所有小鼠体重，进行疾病活动指数（disease activity index，DAL）评分、临床评分，摘眼球采血后处死并解剖小鼠，取结肠肠段测量长度并进行组织学损伤评分，同时用甲醛固定后制作切片。ELISA检测血清中白细胞介素-6（interleukin-6，IL-6）、IL-10及肿瘤坏死因子-α（tumor necrosis factor-α，TNF-α）含量。结果表明，UC模型造模成功，模型组发病症状明显。与模型组相比，发酵液组小鼠体重下降程度显著降低（P<0.05），DAL评分、临床评分、结肠缩短程度及组织学损伤评分均极显著下降（P<0.01）。发酵液组小鼠结肠组织表现出相对完整的上皮层结构及隐窝，仅有部分炎症细胞侵入上皮组织。ELISA检测结果显示，发酵液组IL-6和TNF-α含量显著低于模型组（P<0.05），IL-10的含量显著高于模型组（P<0.05）。说明丁酸梭菌-龙眼多糖发酵液对DSS诱导的小鼠溃疡性结肠炎具有一定的防治作用，其作用机制可能与降低促炎因子IL-6和TNF-α含量、提升抗炎因子IL-10含量抑制炎症有关。     

桑枝皮多糖的提取及组成成分与体外生物活性分析

为了更好地发掘桑枝皮的药用价值,采用热水浸提法从桑枝皮中分离提取多糖,其得率达到28.5%。采用薄层层析(TLC)法和非衍生化高效液相色谱(HPLC)法分析桑枝皮多糖的组成成分,初步推定桑枝皮多糖为一种杂多糖,由鼠李糖、木糖、阿拉伯糖、甘露糖、葡萄糖和半乳糖等单糖组成。桑枝皮多糖的体外生物活性试验显示其具有较强的对DPPH自由基的清除能力(IC50为1.7mg/mL)和对α-葡萄糖苷酶的抑制活性(IC50为0.298mg/mL)。上述结果提示桑枝皮多糖是降血糖和抗氧化的天然药物资源。     

葫芦多糖体外抗鸡新城疫病毒作用的研究

为研究葫芦多糖抗新城疫病毒（Newcastle disease virus,NDV） 活性,本试验采用水提醇沉法提取葫芦总多糖及4种分级多糖,通过苯酚-硫酸法及红外光谱对葫芦多糖进行检测,MTT法测定各级多糖对鸡胚成纤维细胞（CEF）的安全浓度,多糖的安全浓度统一定为78.125 μg/mL,以3种加药方式（先加多糖后接种病毒、先接种病毒后加多糖、多糖和病毒感作后加药）,检测葫芦多糖对NDV的阻断、抑制及直接杀灭作用。结果表明,葫芦总多糖及4种分级多糖均具有抗NDV活性,其中对NDV的抑制作用及直接杀灭作用强于对NDV的阻断作用。70%、80%梯度醇沉的葫芦多糖（LSP₇₀、LSP₈₀）及总多糖（LSP_t）,抑制作用下的病毒抑制率分别为40.41%、44.54%、61.85%,杀灭作用下的病毒抑制率分别为44.74%、58.76%和59.38%,阻断作用下对NDV的抑制率,其中LSP₈₀组最高达37.14%,其余各组均低于25%。综上可知,70%、80%梯度醇沉的葫芦多糖及葫芦总多糖的抗NDV活性较好,可作为进一步研究的材料。     

Study on Antiviral Effect of Polysaccharides from Lagenaria siceraria (Molina) Standl.on NDV in vitro

To research on polysaccharides from Lagenaria siceraria (Molina) Standl.against Newcastle disease virus (NDV).Total polysaccharide and four different solvent extractions of Lagenaria siceraria (Molina) Standl.were extracted by the methods of hot water extraction and ethanol precipition in this study.The content of polysaccharide was measured by phenol-sulfuric acid method and infrared spectroscopy method.The safe concentration and growth of chick embryo fibroblast (CEF) were assayed by MTT method,in order to facilitated the comparison under the same level,the safe concentration was united as 78.125 μg/mL.Under the safety range of concentration,detected the block-virus activity,anti-virus activity and virus-killing activity of polysaccharides through the ways of pre-adding polysaccharides,post-adding polysaccharides and adding polysaccharides with NDV.The results showed that the direct inactivation and propagation inhibition activity of total polysaccharide and four different solvent extractions were stronger than anti-absorption function.Anti-virus inhibition rate of 70%,80% gradient alcohol precipitation of polysaccharides from Lagenaria siceraria (Molina) Standl.(LSP₇₀,LSP₈₀) and total polysaccharide (LSP_t)were 40.41%,44.54% and 61.85%,virus-killing inhibition rate were 44.74%,58.76% and 59.38%.LSP₈₀ had the highest virus inhibition rate as 37.14% in the block-virus activity of those five polysaccharides.In summary,70%,80% gradient alcohol precipitation and total polysaccharide in polysaccharides possessed better activity and would be as the materials for further research.     

鱼腥草多糖对MA104和MARC145细胞活力影响的比较研究

研究鱼腥草多糖(houttuynia cordata polysaccharide,HCP)对MA104和MARC145细胞活力的影响,筛选梯度浓度的鱼腥草多糖对MA104和MARC145细胞促生长最佳浓度剂量范围和毒性剂量.通过水提醇沉法提取HCP,得糖率为8.18％;分别采用MTT法和细胞平板计数法绘制MA104和...     

金龟子绿僵菌发酵液中苦马豆素含量及催化酶基因mRNA表达分析

旨在探明金龟子绿僵菌发酵液中苦马豆素含量及与苦马豆素生物合成有关的催化酶基因mRNA表达的关系,将金龟子绿僵菌接种于察氏培养基进行发酵培养,运用Q Exactive高分辨质谱仪检测培养第1、3、5、7天发酵液中苦马豆素的含量,采用qRT-PCR法检测培养第1、3、5、7天发酵液中苦马豆素生物合成通路上的催化酶基因SwnA、SwnH₁、SwnH₂、SwnK、SwnN、SwnR和SwnT的mRNA转录水平。结果显示,根据苦马豆素质量浓度和峰面积的变化测算出回归方程为y=31 302.5x-45 910.5（R²=0.996 7）,在培养第3天发酵液中苦马豆素含量最高为174.014 μg·mg^-1;SwnA、SwnH₁、SwnH₂、SwnK、SwnN、SwnR和SwnT基因在不同发酵时期mRNA表达差异较大,其中SwnR基因mRNA表达与发酵液中苦马豆素含量变化相一致,而SwnH₂基因mRNA表达与发酵液中苦马豆素含量变化相反。结果表明,金龟子绿僵菌SwnR基因mRNA表达与苦马豆素合成关系密切,可为后续开展苦马豆素微生物发酵工艺及其生物合成通路研究提供重要参考。     

重组结核分枝杆菌CFP10蛋白对A549细胞TLR信号途径介导的炎症反应的影响

本研究旨在检测重组结核分枝杆菌10 ku培养滤液蛋白（culture filtrate protein 10,CFP10）对肺泡Ⅱ型上皮细胞系A549细胞Toll样受体（TLRs）信号途径及其介导的炎症反应的影响。PCR扩增cfp10目的基因片段,构建重组质粒载体pCzn1-CFP10。将重组质粒pCzn1-CFP10转入BL21（DE3）大肠杆菌感受态细胞,IPTG诱导表达重组蛋白CFP10（rCFP10）并鉴定,纯化rCFP10,去除内毒素及脱盐备用。设置对照组,利用MTT检测rCFP10对A549细胞的存活率的影响;通过qRT-PCR、Western blot及ELISA等技术,分别在转录和翻译水平检测rCFP10对A549细胞TLRs信号途径关键分子及其下游炎症因子表达的影响。结果显示,成功构建重组表达载体pCzn1-CFP10并表达纯化出高纯度的rCFP10。MTT结果显示,随着rCFP10浓度增加和处理时间延长,A549细胞存活率显著降低。与空白对照组相比,rCFP10分别从转录和翻译水平显著（P<0.05）或极显著（P<0.01）上调A549细胞中TLRs途径关键分子TLR2、TLR4、MyD88、TRAF6和NF-κB p65及下游炎症因子IL-6、TNF-α的表达水平。rCFP10蛋白可以通过激活A549细胞TLRs受体信号途径促进细胞炎症因子的分泌,这将为进一步加深理解结核病发病机制提供理论依据。     

The effects of the Recombinant Mycobacterium Tuberculosis CFP10 Protein on the TLR Signal-mediated Inflammatory Responses in A549 Cells

The purpose of this study was to investigate the effects of recombinant 10 kDa culture filtrate protein (CFP10) of Mycobacterium tuberculosis on the inflammatory responses mediated by Toll-like receptor (TLR) signaling in A549 cells. The recombinant plasmid pCzn1-CFP10 was obtained by amplifying the cfp10 gene fragment using PCR and cloning it into the prokaryotic expression vector pCzn1. The obtained recombinant plasmid pCzn1-CFP10 was transformed into Escherichia coli BL21(DE3) and induced by IPTG to express the recombinant protein CFP10 (rCFP10). The purified rCFP10 protein was preserved after endotoxin was removed and desalted before use. The effect of rCFP10 treatment on the survival rate of A549 cells was detected by MTT assay. A549 cells were treated with rCFP10 to detect changes of key molecules of TLR signaling pathway and downstream inflammatory factors in A549 cells by qRT-PCR,Western blot and ELISA. The results showed that the recombinant expression vector pCzn1-CFP10 was successfully constructed and the high-purity rCFP10 protein was expressed and purified in this study. MTT assay showed that rCFP10 could inhibit the survival rate of A549 cells in a time- and concentration-dependent manner. In addition, rCFP10 could significantly (P<0.05) up-regulate the key molecules of TLR pathways TLR2, TLR4, MyD88, TRAF6, NF-κBp65 and downstream inflammatory cytokines IL-6 and TNF-α in A549 cells as compared to the control group. The recombinant Mycobacterium tuberculosis protein CFP10 could promote the secretion of cytokines by activating TLR receptor signaling pathway in A549 cells, which will provide a theoretical basis for further understanding of the pathogenesis of Mycobacterium tuberculosis.