Biological and Clinical Studies on the Differnece between Pathogenic and Nonpathogenic Entamoeba histolytica

(1)异巴内阿米巴的体外培养：过氧化氢处理的麻瓜明显促进异巴内阿米巴的体外生长；然而，最近的核磁共振分析表明，鞭毛虫仍然有氨基酸和一些有机酸的信号，表明这些鞭毛虫仍然保持着代谢活性。随后的实验中，将鞭毛虫在56°C的温度下处理30分钟，然后再用H_2O_2处理，导致NRM上的信号完全丧失。这种无菌的、无代谢活性的鞭毛对e.d dispar的体外生长有良好的促进作用，这与体外培养的定义是一致的。此外，我们最近开发了不添加任何培养物的完整的axic培养基，使我们能够在这种条件下维持3株菌株。(2)超微结构差异：在dispar中，糖原颗粒主要分布在未发育的细胞器周围的细胞质周围，而在相同条件下生长的溶组织芽胞杆菌中，糖原颗粒均匀分布在细胞质中。(3)生化研究：最显著的差异是糖原的含量。每毫克蛋白质中含有的糖原是溶组织芽孢杆菌的8^-9倍。此外，果糖能很好地促进e.d dispar的生长，而葡萄糖则能更好地支持e.d dispar和e.d histolytica的生长，这使我们能够分别表征e.d dispar和e.g histolytica的己糖激酶和葡萄糖激酶。(4)遗传学研究：目前主要研究溶组织芽孢杆菌的硫代谢。我们能够克隆到仅在植物和细菌中检测到的l -半胱氨酸合成酶cDNA。该基因位于NADP^+依赖性醇脱氢酶基因旁边。这两种蛋白被认为是与慢性结肠炎相关的主要抗原。原因正在调查中。此外，我们没有检测到l -胱氨酸合成酶的m-RNA，其原因也在研究中。(5)异差e.p.a的致病性和毒力：将异差e.p.a毒株灌胃新生仓鼠肝脏，评价其毒力；但未见病理改变。通过对仓鼠反复注射类固醇诱导免疫功能低下的动物，没有导致e.d dispar在肝脏形成脓肿病变，表明e.d dispar的毒力可以忽略不计。少

(1) Axnic cultivation of Entamoeba dispar : Hydrogen peroxide treated Crithidia fasciculata cartainly promoted the growth of E.dispar in vitro ; however, recent NMR analysis indicated that the flagellate still had signals due to amino acids and some organic acids suggesting that such flagellates still maintained metabolic activities. Subsequent trial using the flagellates treated at 56ﾟC,30 min followed by H_2O_2 treatment resulted in complete loss of the signals on NRM.Such sterilized and metabolically inactive flagellates had a favorale promoting activity for the in vitro growth of E.dispar, which is consistent with the definition of axnic cultivation. Moreover, we have recently developed the complete axnic medium without any culture supplement, which enabled us to mainatain 3 strains under such conditions.(2) Ultrastructural difference : In E.dispar, numerous glycogen particles were detected primarily on the periphery of cytoplasm surrounding undeveloped cellular organelles at the c … More enter, while the particles were dispersed evenly throughout the cytoplasm in E.histolytica grown under the same conditions.(3) Biochemical studies : The mst striking difference was the amount of glycogen. E.dispar trophozoite contained 8^-9 times as much glycogen per mg protein as E.histolytica. In addition, fructose well promoted the growth of E.dispar, while glucose was better to support the growth of E.histolytica, which enabled us to characterize hexokinase and glucokinase in E.dispar and E.histolytica respectiely.(4) Genetic studies : Now we arefocusing our efforts on sulfur metabolism of E.histolytica. We were able to clone cDNA for L-cystein synthase, which has been detected only in plants ans bacteria. This gene was located next to NADP^+-dependent alcohol dehyrogenase gene. Both of these proteins have been considered as major antigens related with chronic colitis. The reason for this is under current investigation. Moreover, we did not detect m-RNA for L-cysten synthase, the reason of which is also being studied.(5) Pathogenicity and virulence of E.dispar : Axnic strains of E.dispar were inculated into liver of newborn hamsters to assess the virulence ; however, no nistopathological changes could be found. Immunocompromized animals were induced by injecting steroid repeatedly into hamsters, which did not lead to formation of abscess lesion in the liver by E.dispar, sugegsting virulence of E.dispar should be negligible. Less