Chen X M, Levine S A, Tietz P, Krueger E, McNiven M A, Jefferson D M, Mahle M, LaRusso N F
Center for Basic Research in Digestive Diseases, Division of Gastroenterology and Internal Medicine, Rochester, MN 55905, USA.
Hepatology. 1998 Oct;28(4):906-13. doi: 10.1002/hep.510280402.
While the clinical features of sclerosing cholangitis secondary to opportunistic infections of the biliary tree in patients with acquired immunodeficiency syndrome (AIDS) are well known, the mechanisms by which microbial pathogens such as Cryptosporidium parvum associated with this syndrome actually cause disease are obscure. We established an in vitro model of biliary cryptosporidiosis employing a human biliary epithelial cell line. Using morphological and biochemical techniques, we examined the interaction of C. parvum with cultured human cholangiocytes. When the apical plasma membrane of polarized, confluent monolayers of human biliary epithelial cells was exposed to C. parvum oocysts that had been excysted in vitro, sporozoites attached to and invaded the cells in a time-, dose-, temperature-, and pH-dependent manner. The infectious process was both plasma membrane domain- and cell-specific, because no attachment or invasion occurred when the basolateral membrane of cholangiocytes was exposed to the parasite, or when a human hepatocyte cell line (HepG2) was used. Time-lapse video microscopy and scanning electron microscopy (SEM) showed that sporozoite attachment was rapid, involved extensive cholangiocyte membrane ruffling, and culminated in parasite penetration into a tight-fitting vacuole formed by invagination of the plasma membrane similar to those found in naturally occurring infection in vivo. Transmission electron microscopy (TEM) showed that C. parvum organisms formed parasitophorus vacuoles and were able to undergo a complete reproductive cycle, forming both asexual and sexual reproductive stages. Unexpectedly, direct cytopathic effects were noted in infected monolayers, with widespread programmed cell death (i.e., apoptosis) of biliary epithelial cells as assessed both morphologically and biochemically beginning within hours after exposure to the organism. The novel finding of specific cytopathic invasion of biliary epithelia by C. parvum may be relevant to the pathogenesis and possible therapy of the secondary sclerosing cholangitis seen in AIDS patients with biliary cryptosporidiosis.
虽然获得性免疫缺陷综合征（AIDS）患者继发于胆道树机会性感染的硬化性胆管炎的临床特征已为人熟知，但诸如微小隐孢子虫等与该综合征相关的微生物病原体实际导致疾病的机制仍不清楚。我们利用人胆管上皮细胞系建立了胆道隐孢子虫病的体外模型。运用形态学和生物化学技术，我们研究了微小隐孢子虫与培养的人胆管细胞的相互作用。当极化的、汇合的人胆管上皮细胞单层的顶端质膜暴露于体外脱囊的微小隐孢子虫卵囊时，子孢子以时间、剂量、温度和pH依赖方式附着并侵入细胞。感染过程具有质膜结构域和细胞特异性，因为当胆管细胞的基底外侧膜暴露于寄生虫时，或者当使用人肝细胞系（HepG2）时，均未发生附着或侵入。延时视频显微镜和扫描电子显微镜（SEM）显示，子孢子附着迅速，涉及广泛的胆管细胞膜褶皱，并最终导致寄生虫穿透由质膜内陷形成的紧密贴合的液泡，类似于在体内自然发生的感染中所见。透射电子显微镜（TEM）显示，微小隐孢子虫形成了寄生泡，并能够经历完整的生殖周期，形成无性和有性生殖阶段。出乎意料的是，在感染的单层细胞中观察到直接的细胞病变效应，在暴露于该生物体数小时后，通过形态学和生物化学评估发现胆管上皮细胞普遍发生程序性细胞死亡（即凋亡）。微小隐孢子虫对胆管上皮细胞进行特异性细胞病变侵袭这一新颖发现可能与AIDS患者胆道隐孢子虫病中所见的继发性硬化性胆管炎的发病机制及可能的治疗方法有关。
