INTERRELATIONSHIPS BETWEEN DIGESTIVE PROTEOLYTIC ACTIVITIES AND PRODUCTION AND QUANTITATION OF TOXINS IN PSEUDOMEMBRANOUS COLITIS INDUCED BY CLOSTRIDIUM-DIFFICILE IN GNOTOBIOTIC MICE
INTERRELATIONSHIPS BETWEEN DIGESTIVE PROTEOLYTIC ACTIVITIES AND PRODUCTION AND QUANTITATION OF TOXINS IN PSEUDOMEMBRANOUS COLITIS INDUCED BY CLOSTRIDIUM-DIFFICILE IN GNOTOBIOTIC MICE
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DOI：
10.1128/iai.57.12.3922-3927.1989
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发表时间：
1989-12-01
影响因子：
3.1
通讯作者：
DUBOSRAMARE, F
中科院分区：
文献类型：
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作者：
CORTHIER, G;MULLER, MC;DUBOSRAMARE, F
艰难梭菌的致病性与体内毒素的产生有关，检测生物样品中产生的毒素具有很大的意义。几份报告表明，粪便中的蛋白酶会干扰毒素A的免疫学定量方法。本工作的目的是估计蛋白酶和C.艰难梭菌毒素在伪膜性盲肠炎的无菌小鼠模型中产生。盲肠蛋白水解活性水解毒素A和免疫球蛋白G结合到用于免疫测定的微量滴定板。这种干扰可以通过向样品中加入胰蛋白酶抑制剂来阻断。可溶性毒素A在酶联免疫吸附测定中与结合抗体结合的能力不受蛋白酶的影响，但生物活性降低了100倍。毒素B的细胞毒性不被蛋白水解活性处理改变。小鼠接种低毒素A产生的C.艰难梭菌没有死亡，并且没有发生蛋白水解活性的调节。接种C. difficle，毒素A和B在感染后12 h达到最高水平。此时，蛋白水解活性未从零时观察到的水平降低。小鼠在2天内死亡。在这个时候（约32感染后），蛋白水解活性急剧下降，在较低的部分消化道。血清抑制蛋白酶的研究结果表明，有一个100倍的增加，血清来源的小鼠免疫球蛋白在管腔中的C。difficle感染的进展表明下消化道中蛋白酶活性的降低可能与炎症过程中血清的渗出有关。
Clostridium difficile pathogenicity is related to in vivo production of toxins, and it is of great interest to detect toxins produced in biological samples. Several reports have shown that proteases in stools interfere with immunological methods for quantitation of toxin A. The purpose of this work was to estimate the relationship between the proteases and the C. difficile toxins produced in a gnotobiotic mouse model of pseudomembraneous cecitis. Cecal proteolytic activities hydrolyzed toxin A, and immunoglobulin G bound to the microtiter plate used in immunoassays. This interference could be blocked by the addition of trypsin inhibitor to the samples. The ability of soluble toxin A to bind to bound antibodies in an enzyme-linked immunosorbent assay was not affected by the proteases, but the biological activity was reduced 100-fold. The cytotoxicity of toxin B was not modified by proteolytic activity treatment. Mice inoculated with a low toxin A-producing strain of C. difficile did not die, and no modulation of proteoytic activities occurred. After inoculation with the lethal VPI strain of C. difficle, toxins A and B reached maximum levels in the ceca at 12 h postinfection. At this time, the proteolytic activities did not decrease from the levels seen at zero time. Mice died within 2 days. At this time (about 32 postinfection), proteolytic activities were sharply decreased in the lower parts of the digestive tracts. The findings that serum inhibited the proteases and that there was a 100-fold increase in serum-derived mouse immunoglobulins in the lumen as the C. difficle infection progressed suggest that decrease in protease activity in the lower digestive tract may be related to the exudation of serum from the inflammation process.