Pathophysiology of Biliary Cryptosporidiosis

我们的长期目标仍然是应用基本概念和广泛的细胞技术， 分子生物学，以了解胆管细胞的作用，上皮细胞内衬肝内胆管， 健康和疾病。我们继续关注胆管细胞和隐孢子虫之间的相互作用 parvum（C. parvum），一种引起肠道和胆道隐孢子虫病的新兴病原体。最近的证据 从我们的实验室表明：（i）细胞表达和调节的关键受体[Toll样受体 （TLR）]和特异性细胞内信号传导途径（NF-κ B激活）参与胆管细胞C。 小孢子识别和战略防御;和（ii）内源性microRNA（miRNA）介导的后 转录基因调控参与胆管细胞对C.细小病毒感染因此，在本发明中， 我们将测试中心假设，C。微小胆管细胞相互作用激活宿主细胞 TLR/NF-kB信号级联启动胆管细胞防御反应，包括上调 TLR和抗菌肽（防御素）通过转录和转录后 内源性miRNA介导的）调节途径。在我们的三个综合具体目标中，我们将测试 （i）胆管细胞TLRs识别C.并调节NF-κ B活化， 通过NF-κ B介导的抗微生物肽（人β-防御素[HBD]）和TLR的上调 转录调控;（ii）miRNA介导的TLR表达的转录后抑制， 存在于胆管细胞中，受TLR/NF-κ B信号调节，并参与C.小孢子诱导的 TLR的上调;和（iii）胆管细胞中TLR和HBD的表达是根除所需的 梭在体外和体内实验模型中胆道中的细小病毒感染。的创新方面 我们的项目包括：新的方法[小干扰RNA（siRNA）]，动物模型（胆汁 野生型、TLR和MyD 88敲除小鼠中的隐孢子虫病）和新概念（miRNA介导的基因 调节胆管细胞防御）。因此，这些实验将确定分子机制， C.微小胆管细胞的相互作用触发宿主上皮防御，重点是miRNA的作用， 介导的转录后调节TLR相关的防御，一个新的概念， 对健康和疾病中先天免疫和适应性免疫的重要概括性。

Our long-term OBJECTIVES remain to apply the fundamental concepts and broad technologies of cell and molecular biology to understand the role of cholangiocytes, the epithelial cells lining intrahepatic bile ducts, in health and disease. We continue to focus on the interactions between cholangiocytes and Cryptosporidium parvum (C. parvum), an emerging pathogen causing intestinal and biliary cryptosporidiosis. Recent evidence from our lab indicates that: (i) the cellular expression and regulation of key receptors [Toll-like receptors (TLRs)] and specific intracellular signaling pathways (NF-kB activation) are involved in cholangiocyte C. parvum recognition and strategic defense; and (ii) endogenous microRNA (miRNA)-mediated post- transcriptional gene regulation is involved in cholangiocyte defense responses to C. parvum infection. Thus, we will test the CENTRAL HYPOTHESIS that C. parvum-cholangiocyte interactions activate host-cell TLR/NF-kB signaling cascades initiating cholangiocyte defense responses including upregulation of both TLRs and anti-microbial peptides (defensins) through both transcriptional and post-transcriptional endogenous miRNA-mediated) regulatory pathways. In our three integrated SPECIFIC AIMS, we will test the hypotheses that: (i) cholangiocyte TLRs recognize C. parvum and modulate NF-kB activation resulting in upregulation of anti-microbial peptides (human beta-defensins [HBDs]) and TLRs through NF-kB mediated transcriptional regulation; (ii) miRNA-mediated post-transcriptional repression of TLR expression normally exists in cholangiocytes, is regulated by TLR/NF-kB signals, and is involved in C. parvum-induced upregulation of TLRs; and (iii) expression of TLRs and HBDs in cholangiocytes are required for eradication of C. parvum infection in the biliary tract in in vitro and in vivo experimental models. Innovative aspects of our program include: novel methodologies [small-interfering RNAs (siRNAs)], animal models (biliary cryptosporidiosis in wild type, TLR and MyD88 knock out mice), and new concepts (miRNA-mediated gene regulation in cholangiocyte defense). Thus, the experiments will define the molecular mechanisms by which C. parvum-cholangiocyte interactions trigger host epithelial defense, with an emphasis on the role of miRNA- mediated post-transcriptional regulation in TLR-associated defense, a NEW CONCEPT that may have important generalizability to both innate and adaptive immunity in health and disease.