Embaye H, Hart C A, Getty B, Fletcher J N, Saunders J R, Batt R M
Department of Small Animal Medicine and Surgery, Royal Veterinary College, University of London.
Gut. 1992 Sep;33(9):1184-9. doi: 10.1136/gut.33.9.1184.
This study examines the effects of an enteropathogenic Escherichia coli on microvillar membrane proteins during organ culture of rabbit ileal explants. Explants maintained with enteropathogenic E coli showed brush border effacement affecting approximately 50% of enterocytes, and where enteropathogenic E coli were closely adherent to the enterocyte surface microvilli were apparently being shed as vesicles. The microvillar membrane enzymes alkaline phosphatase, aminopeptidase N and alpha-glucosidase were released into the culture medium during organ culture, and this process was significantly enhanced by enteropathogenic E coli. This increased loss of microvillar membrane enzymes into the culture medium was associated with decreased tissue activities of microvillar membrane enzymes in enteropathogenic E coli infected ileal explants compared with control. For aminopeptidase N in particular, however, total enzyme activities in the tissue plus culture medium were increased comparing enteropathogenic E coli with control, suggesting that there might be an increase in the rate of synthesis of certain microvillar membrane proteins. Reorientating sucrose density gradient centrifugation of culture medium showed that alkaline phosphatase, aminopeptidase N and alpha-glucosidase were predominantly associated with particles of peak modal density 1.19 g/ml in both groups, confirming that enteropathogenic E coli accelerate release of microvillar membrane enzymes as vesicles. Analytical fractionation of ileal explants showed that enteropathogenic E coli resulted in a loss of microvillar membrane enzyme activities from the main brush border peak of modal density 1.21 g/ml present in controls. The density of the remaining smaller and lighter peak increased from 1.19 g/ml to 1.23 g/ml after homogenisation in digitonin, confirming association of these proteins with cholesterol containing membranes and not endoplasmic reticulum. These findings suggest that enteropathogenic E. coli accelerate the normal shedding of microvillar membrane proteins as vesicles, and may stimulate a compensatory increase in microvillar membrane protein synthesis.
本研究检测了肠道致病性大肠杆菌对兔回肠外植体器官培养过程中微绒毛膜蛋白的影响。用肠道致病性大肠杆菌培养的外植体显示刷状缘消失，约50%的肠细胞受到影响，且肠道致病性大肠杆菌紧密附着于肠细胞表面时，微绒毛明显以小泡形式脱落。在器官培养过程中，微绒毛膜酶碱性磷酸酶、氨肽酶N和α-葡萄糖苷酶释放到培养基中，且该过程被肠道致病性大肠杆菌显著增强。与对照相比，肠道致病性大肠杆菌感染的回肠外植体中，微绒毛膜酶向培养基中的流失增加，这与微绒毛膜酶的组织活性降低有关。然而，特别是对于氨肽酶N，与对照相比，肠道致病性大肠杆菌组组织和培养基中的总酶活性增加，这表明某些微绒毛膜蛋白的合成速率可能增加。对培养基进行蔗糖密度梯度离心重定向显示，两组中碱性磷酸酶、氨肽酶N和α-葡萄糖苷酶主要与峰模式密度为1.19 g/ml的颗粒相关，证实肠道致病性大肠杆菌加速了微绒毛膜酶以小泡形式的释放。对回肠外植体进行分析分级分离显示，肠道致病性大肠杆菌导致对照中存在的峰模式密度为1.21 g/ml的主要刷状缘峰的微绒毛膜酶活性丧失。在用洋地黄皂苷匀浆后，剩余较小且较轻峰的密度从1.19 g/ml增加到1.23 g/ml，证实这些蛋白与含胆固醇的膜相关，而非内质网。这些发现表明，肠道致病性大肠杆菌加速了微绒毛膜蛋白以小泡形式的正常脱落，并可能刺激微绒毛膜蛋白合成的代偿性增加。
