肠道菌群通过TLRs减轻电离辐射诱发DNA损伤的机制研究

发布时间：2018-06-10 01:02

本文选题：肠道内源性共生菌 + TLRs　；参考：《安徽医科大学》2012年硕士论文

【摘要】：一般认为，肠道内源性菌群与宿主存在共生关系，肠道内源性菌群参与宿主的营养和能量平衡，对宿主免疫系统的进化和功能的维持发挥着积极的作用。反过来，宿主为微生物提供一个能够维持其生存的环境，从这个环境中微生物能够获得其必须的营养物质，保证其在宿主不同个体间的传播和隐匿。已有研究表明，肠道内源性菌群可以通过其组份激活Toll样受体（Toll-like receptors，TLRs），减轻硫酸葡聚糖钠（dextran sulfate sodium，DSS）对肠道粘膜造成的损伤，维持肠道的自稳平衡。文献报道发现，TLR4和TLR5受体激活具有防护电离辐射损伤作用。TLRs的配体来源主要为细菌的不同组分。作为电离辐射的特异性损伤之一，DNA损伤的修复在正常机体生命活动中处于一种动态变化过程。肠道菌群作为机体的共生物，伴随生命的进程。本研究利用电离辐射诱发小鼠造血细胞DNA损伤为研究模型，探索肠道菌群中TLRs家族对DNA损伤和修复过程的影响和相关分子机制。首先，利用广谱抗生素清除小鼠肠道内源性共生菌群，而后对小鼠进行2Gyγ射线照射，与单纯照射小鼠相比，肠道菌群清除后进行照射的小鼠骨髓细胞染色体畸变率明显增加；外周血细胞彗星电泳实验显示，反映DNA损伤的彗星尾距明显增加，表明清除肠道菌群可以促进电离辐射导致的DNA损伤。为了进一步分析肠道菌群对DNA损伤的防护作用是否与TLRs受体相关，我们分析了肠道菌群清除小鼠血清中TLRs配体的变化。通过Elisa方法对肠道菌群清除小鼠体内的TLRs含量进行检测，发现血清中三种TLRs（TLR2/6、TLR4和TLR5）的配体Lipopeptede、LPS和Flagellin均有不同程度的下降，因此推测：TLRs配体的减少及由此而导致TLRs功能激活的减弱可能是导致其DNA损伤增加的直接原因。为了确定这些来源于细菌组份的TLRs配体是否具有减轻辐射诱发DNA损伤的作用，我们将这些配体分别回转正常小鼠和肠道内源性菌群清除小鼠，分别用CBLB502（Flagellin的功能同源物）、LPS、Lipopeptide激活TLR5、TLR4和TLR2/6后，发现能够明显减轻电离辐射诱发的DNA损伤（骨髓细胞染色体畸变率降低，外周血细胞彗星电泳彗星尾距缩短）。上述研究表明，肠道菌群通过激活TLRs家族（TLR2/6、TLR4和TLR5），减轻电离辐射诱发的小鼠造血细胞DNA损伤。为了进一步探索肠道菌群通过TLRs受体激活调节电离辐射诱发造血细胞DNA损伤的分子机制，采用基因芯片方法筛选TLR5激活后骨髓细胞基因表达谱的变化情况。结果表明，与对照组相比，TLR5激活后，骨髓细胞中共有85个基因表达上调，2个基因表达下调。进一步挑选出4个参与DNA损伤修复的基因Apex2、Gadd45β、Sod2和Rad21，用Real Time-PCR方法对TLR5、TLR4和TLR2/6激活后4种基因的表达进行了验证，发现这些基因的表达均有不同程度的上调。对肠道菌群清除小鼠骨髓细胞分析发现，上述4个基因表达均出现明显下调。本研究发现，肠道内源性菌群可以通过其组份激活TLRs家族，，通过调节Apex2、Gadd45β、Sod2和Rad21等DNA损伤修复相关基因的表达，减轻电离辐射诱发的造血细胞DNA损伤。提示肠道内源性共生菌群通过其组份对TLRs的识别和激活，或参与稳态下机体自身的DNA损伤修复过程，从而降低宿主由DNA损伤所导致的癌症等疾病风险。
[Abstract]:In this study , it is found that the intestinal flora can activate Toll - like receptors ( TLRs ) by activating Toll - like receptors ( TLRs ) to reduce the damage to intestinal mucosa and maintain the self - stable balance of intestinal tract .

Firstly , the mouse intestinal endosymbionts were cleared by broad - spectrum antibiotics , and then the mice were irradiated with 2Gy 纬 - ray , and the chromosome aberration rate of bone marrow cells irradiated by intestinal flora was significantly increased compared with the simple irradiated mice .
In order to determine whether the TLRs ligands derived from the bacterial group were associated with TLRs receptors , it was speculated that the decrease of TLRs ligands and the decrease of TLRs ligands in mice induced by ionizing radiation could be a direct cause of increased DNA damage . The study indicated that the intestinal flora could reduce the DNA damage of hematopoietic cells induced by ionizing radiation by activating TLRs family ( TLR2 / 6 , TLR 5 ) and TLR5 .

In order to further explore the molecular mechanism of regulating the DNA damage of hematopoietic cells induced by ionizing radiation by TLRs receptor activation , the expression profile of bone marrow cells after activation of TLR5 was investigated by gene chip method . The results showed that the expression of four genes was up - regulated in bone marrow cells after TLR5 was activated . Four genes involved in DNA damage repair were selected to be up - regulated .

In this study , it was found that endogenous bacteria in intestinal tract can activate TLRs family by regulating the expression of related genes by regulating DNA damage such as Apex2 , Gadd45尾 , Sod2 and Rad21 , and reduce the DNA damage of hematopoietic cells induced by ionizing radiation . It is suggested that endogenous symbiotic bacteria in intestinal tract can recognize and activate TLRs by their components or participate in the process of DNA damage repair of organism under steady state , thus reducing the risk of diseases such as cancer caused by DNA damage .
【学位授予单位】：安徽医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R142

【共引文献】