本文主要研究内容
作者占萌（2019）在《高黏附乳酸菌的筛选及其对RAW264.7细胞的免疫调节作用》一文中研究指出：乳酸菌是一类有益于宿主健康的微生物,主要通过在肠道中的定植发挥益生作用。乳酸菌对肠道上皮细胞的黏附、定植作用有助于恢复或增强肠道稳态,提高机体免疫力。因而乳酸菌的黏附能力是评价其益生性的重要指标。然而,有些具有益生功能的乳酸菌作用效果和稳定性却不尽如人意。所以筛选出一株黏附能力高,又具有免疫调节潜力的乳酸菌,对开发高效的免疫调节益生菌制剂尤为重要。本课题旨在从实验室保藏的12株乳杆菌和2株耐久肠球菌中筛选出高黏附性乳酸菌,并分析影响该菌株黏附性的因素,对黏附机制进行初步探究。最后评价了该株高黏附乳酸菌对巨噬细胞RAW264.7的免疫调节作用。选取具有良好黏附性能的嗜酸乳杆菌NCFM作为对照,通过对14株不同种乳酸菌进行表面疏水性、自聚集能力以及体外黏附Caco-2细胞能力的测定,筛选出高黏附性乳酸菌菌株。并分析了表面性质与黏附性之间的关系。结果发现嗜酸乳杆菌KLDS 1.0901对Caco-2细胞的黏附率是受试的15株菌中最高的,为12.73%,且KLDS 1.0901具有仅次于耐久肠球菌KLDS6.0933的疏水率,为13.88%。鼠李糖乳杆菌KLDS1.0205的黏附能力最低,为0.13%。不同乳酸菌的表面疏水性与对Caco-2细胞的黏附能力呈现一定的相关性。采用体外实验探究了高黏附性嗜酸乳杆菌KLDS1.0901的浓度、生长阶段、化学和酶处理对其黏附Caco-2细胞能力的影响。试验结果表明,KLDS1.0901的生长阶段显著影响其黏附性。菌体浓度为1081010 CFU/mL时,对黏附性影响不显著。KLDS1.0901对Caco-2细胞的黏附可能是以表层蛋白为主的多种黏附素共同作用的结果。结合透射电镜和SDS-PAGE技术,初步分离、鉴定了KLDS1.0901的表层蛋白。结果表明KLDS1.0901表面具有丰富的绒毛状表层蛋白,含量约为15.6%,分子量大约为43 kDa。并证实表层蛋白粗提物能显著抑制KLDS1.0901对Caco-2的黏附,且浓度越大,抑制作用越强。采用流式细胞术分析了未处理、去除表层蛋白和热致死的KLDS1.0901在小鼠肠道中的定植与分布。结果表明,KLDS1.0901在灌胃前期主要黏附于十二指肠和空肠,后期KLDS1.0901趋向于黏附结肠。而去除表层蛋白和热灭活的KLDS1.0901虽然在第一天能大量黏附于十二指肠,但是只能短暂停留于肠道各部位,随着时间推移,这两组在肠道各部位的黏附量显著减少。从体内证实了KLDS1.0901也能在小鼠肠道内长时间黏附定植,并且其表面的表层蛋白同样能显著影响其在体内的黏附能力。体外探究了高黏附菌株KLDS1.0901对巨噬细胞RAW264.7的免疫调节作用。测定了不同剂量的未处理KLDS1.0901（LLA）、去除表层蛋白KLDS1.0901（RSLA）和热致死的KLDS1.0901（HKLA）对巨噬细胞增殖能力及吞噬作用的影响;以及刺激巨噬细胞后IL-10和TNF-α分泌水平及其对应mRNA的表达量。试验结果表明:当菌体与巨噬细胞以10:1和1:1共同孵育时,LLA和HKLA组均能显著促进巨噬细胞的增殖,但只有LLA能显著促进巨噬细胞的吞噬活性。在所有浓度下,RSLA组均显著抑制了巨噬细胞活性。并证实KLDS1.0901表面的表层蛋白是增强巨噬细胞活性的重要物质,当以500μg/mL的表层蛋白粗品刺激巨噬细胞时,其增殖和吞噬活性最强。KLDS1.0901与巨噬细胞以1000:1共同孵育时,与空白对照组相比,LLA显著抑制了IL-10和TNF-的分泌。其他浓度条件下:LLA,RSLA和HKLR组IL-10的分泌水平均显著高于对照组,但远低于LPS组;LLA、RSLA和HKLA能够诱导细胞分泌与LPS组相同水平的TNF-α。与空白对照组相比,当菌体与巨噬细胞以100:1共培养时,LLA和HKLA均能显著上调IL-10和TNF-mRNA表达水平。这与细胞因子水平的测定结果相佐证。结论为筛选得到一株表层蛋白含量丰富、黏附性高和具有免疫调节潜力的嗜酸乳杆菌KLDS1.0901。该菌株有作为粘膜疫苗载体使用的潜能,亦可开发成高效调节机体免疫的益生菌。
Abstract
ru suan jun shi yi lei you yi yu su zhu jian kang de wei sheng wu ,zhu yao tong guo zai chang dao zhong de ding zhi fa hui yi sheng zuo yong 。ru suan jun dui chang dao shang pi xi bao de nian fu 、ding zhi zuo yong you zhu yu hui fu huo zeng jiang chang dao wen tai ,di gao ji ti mian yi li 。yin er ru suan jun de nian fu neng li shi ping jia ji yi sheng xing de chong yao zhi biao 。ran er ,you xie ju you yi sheng gong neng de ru suan jun zuo yong xiao guo he wen ding xing que bu jin ru ren yi 。suo yi shai shua chu yi zhu nian fu neng li gao ,you ju you mian yi diao jie qian li de ru suan jun ,dui kai fa gao xiao de mian yi diao jie yi sheng jun zhi ji you wei chong yao 。ben ke ti zhi zai cong shi yan shi bao cang de 12zhu ru gan jun he 2zhu nai jiu chang qiu jun zhong shai shua chu gao nian fu xing ru suan jun ,bing fen xi ying xiang gai jun zhu nian fu xing de yin su ,dui nian fu ji zhi jin hang chu bu tan jiu 。zui hou ping jia le gai zhu gao nian fu ru suan jun dui ju shi xi bao RAW264.7de mian yi diao jie zuo yong 。shua qu ju you liang hao nian fu xing neng de shi suan ru gan jun NCFMzuo wei dui zhao ,tong guo dui 14zhu bu tong chong ru suan jun jin hang biao mian shu shui xing 、zi ju ji neng li yi ji ti wai nian fu Caco-2xi bao neng li de ce ding ,shai shua chu gao nian fu xing ru suan jun jun zhu 。bing fen xi le biao mian xing zhi yu nian fu xing zhi jian de guan ji 。jie guo fa xian shi suan ru gan jun KLDS 1.0901dui Caco-2xi bao de nian fu lv shi shou shi de 15zhu jun zhong zui gao de ,wei 12.73%,ju KLDS 1.0901ju you jin ci yu nai jiu chang qiu jun KLDS6.0933de shu shui lv ,wei 13.88%。shu li tang ru gan jun KLDS1.0205de nian fu neng li zui di ,wei 0.13%。bu tong ru suan jun de biao mian shu shui xing yu dui Caco-2xi bao de nian fu neng li cheng xian yi ding de xiang guan xing 。cai yong ti wai shi yan tan jiu le gao nian fu xing shi suan ru gan jun KLDS1.0901de nong du 、sheng chang jie duan 、hua xue he mei chu li dui ji nian fu Caco-2xi bao neng li de ying xiang 。shi yan jie guo biao ming ,KLDS1.0901de sheng chang jie duan xian zhe ying xiang ji nian fu xing 。jun ti nong du wei 1081010 CFU/mLshi ,dui nian fu xing ying xiang bu xian zhe 。KLDS1.0901dui Caco-2xi bao de nian fu ke neng shi yi biao ceng dan bai wei zhu de duo chong nian fu su gong tong zuo yong de jie guo 。jie ge tou she dian jing he SDS-PAGEji shu ,chu bu fen li 、jian ding le KLDS1.0901de biao ceng dan bai 。jie guo biao ming KLDS1.0901biao mian ju you feng fu de rong mao zhuang biao ceng dan bai ,han liang yao wei 15.6%,fen zi liang da yao wei 43 kDa。bing zheng shi biao ceng dan bai cu di wu neng xian zhe yi zhi KLDS1.0901dui Caco-2de nian fu ,ju nong du yue da ,yi zhi zuo yong yue jiang 。cai yong liu shi xi bao shu fen xi le wei chu li 、qu chu biao ceng dan bai he re zhi si de KLDS1.0901zai xiao shu chang dao zhong de ding zhi yu fen bu 。jie guo biao ming ,KLDS1.0901zai guan wei qian ji zhu yao nian fu yu shi er zhi chang he kong chang ,hou ji KLDS1.0901qu xiang yu nian fu jie chang 。er qu chu biao ceng dan bai he re mie huo de KLDS1.0901sui ran zai di yi tian neng da liang nian fu yu shi er zhi chang ,dan shi zhi neng duan zan ting liu yu chang dao ge bu wei ,sui zhao shi jian tui yi ,zhe liang zu zai chang dao ge bu wei de nian fu liang xian zhe jian shao 。cong ti nei zheng shi le KLDS1.0901ye neng zai xiao shu chang dao nei chang shi jian nian fu ding zhi ,bing ju ji biao mian de biao ceng dan bai tong yang neng xian zhe ying xiang ji zai ti nei de nian fu neng li 。ti wai tan jiu le gao nian fu jun zhu KLDS1.0901dui ju shi xi bao RAW264.7de mian yi diao jie zuo yong 。ce ding le bu tong ji liang de wei chu li KLDS1.0901（LLA）、qu chu biao ceng dan bai KLDS1.0901（RSLA）he re zhi si de KLDS1.0901（HKLA）dui ju shi xi bao zeng shi neng li ji tun shi zuo yong de ying xiang ;yi ji ci ji ju shi xi bao hou IL-10he TNF-αfen bi shui ping ji ji dui ying mRNAde biao da liang 。shi yan jie guo biao ming :dang jun ti yu ju shi xi bao yi 10:1he 1:1gong tong fu yo shi ,LLAhe HKLAzu jun neng xian zhe cu jin ju shi xi bao de zeng shi ,dan zhi you LLAneng xian zhe cu jin ju shi xi bao de tun shi huo xing 。zai suo you nong du xia ,RSLAzu jun xian zhe yi zhi le ju shi xi bao huo xing 。bing zheng shi KLDS1.0901biao mian de biao ceng dan bai shi zeng jiang ju shi xi bao huo xing de chong yao wu zhi ,dang yi 500μg/mLde biao ceng dan bai cu pin ci ji ju shi xi bao shi ,ji zeng shi he tun shi huo xing zui jiang 。KLDS1.0901yu ju shi xi bao yi 1000:1gong tong fu yo shi ,yu kong bai dui zhao zu xiang bi ,LLAxian zhe yi zhi le IL-10he TNF-de fen bi 。ji ta nong du tiao jian xia :LLA,RSLAhe HKLRzu IL-10de fen bi shui ping jun xian zhe gao yu dui zhao zu ,dan yuan di yu LPSzu ;LLA、RSLAhe HKLAneng gou you dao xi bao fen bi yu LPSzu xiang tong shui ping de TNF-α。yu kong bai dui zhao zu xiang bi ,dang jun ti yu ju shi xi bao yi 100:1gong pei yang shi ,LLAhe HKLAjun neng xian zhe shang diao IL-10he TNF-mRNAbiao da shui ping 。zhe yu xi bao yin zi shui ping de ce ding jie guo xiang zuo zheng 。jie lun wei shai shua de dao yi zhu biao ceng dan bai han liang feng fu 、nian fu xing gao he ju you mian yi diao jie qian li de shi suan ru gan jun KLDS1.0901。gai jun zhu you zuo wei nian mo yi miao zai ti shi yong de qian neng ,yi ke kai fa cheng gao xiao diao jie ji ti mian yi de yi sheng jun 。
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论文作者分别是来自东北农业大学的占萌，发表于刊物东北农业大学2019-08-27论文，是一篇关于嗜酸乳杆菌论文,黏附论文,表层蛋白论文,免疫调节论文,东北农业大学2019-08-27论文的文章。本文可供学术参考使用，各位学者可以免费参考阅读下载，文章观点不代表本站观点，资料来自东北农业大学2019-08-27论文网站，若本站收录的文献无意侵犯了您的著作版权，请联系我们删除。
标签：嗜酸乳杆菌论文; 黏附论文; 表层蛋白论文; 免疫调节论文; 东北农业大学2019-08-27论文;