BLADDER GLYCOSPHINGOLIPIDS IN THE PATHOGENESIS OF URINARY TRACT INFECTION

虽然膀胱是迄今为止UTI最常见的部位，但很少 知道感染细菌附着在这个分子 组织与肾脏相比，其中球系GSLs的作用 作为大肠杆菌的细菌附着位点已经被充分研究。 由于GSL的碳水化合物部分在不同组织中不同，因此通常 在宿主基因的控制下，这些分子作为两者的决定因素 患者对感染易感性和病原体的组织嗜性 细菌本提案的总体目标是利用原电池 膀胱的培养物作为模型系统来确定其作用和 调节膀胱GSL作为尿路致病菌的附着位点， 集中在女性膀胱炎的两个最常见的原因，E.杆菌和 S.腐生菌，在以下拟定研究中： (1)评价球细胞系（Gb）和神经节细胞系 (Gg)GSLs存在于膀胱移行上皮原代培养物中。 上皮和结合物E. coli和革兰氏阳性菌S.腐生菌，我们将 从细胞中纯化GSLs，鉴定E.大肠杆菌和S.腐生菌结合 GSL，并使用特异性单克隆抗体确认GSL的身份 （2）对单克隆抗体（MAbs）和糖结构进行了分析; 评价特定目标1中确定的GSL为 表面暴露在原代膀胱细胞培养物中，并在功能上 相关的E. coli和革兰氏阳性菌S.腐生菌附着，我们将研究 细胞培养物预处理前后的细菌粘附 在腐生菌附着之前，我们将研究细菌附着 并且在用GSL抑制剂预处理细胞培养物后 （3）评价GSLs的生物学活性，以确定GSLs的生物学活性。 推测分泌基因和ABO基因影响Gb的表达 和Gb GSL的容量，并由此确定E.大肠杆菌结合，我们 将从已知膀胱癌的患者中建立原代膀胱培养物 分泌型/ABO状态，并确定预测的 培养细胞中存在糖基转移酶和GSL产物; （4）评价原发性肝癌细胞因子释放的假说。 尿上皮细胞培养物并调节大肠杆菌和链球菌的表达。 腐生菌结合的GSLs在膀胱中，我们将检查的影响 E. coli和革兰氏阳性菌S.腐生菌对细胞因子释放 IL-1、IL-6、IL-8对人肝癌细胞系H5 N1表达的影响 相关GSLs在原代尿路上皮细胞培养物中的表达 细菌粘附到这些细胞上。这些研究的表达和 膀胱GSLs的调节与感染的发病机制有关， 炎症和分化，最终可能导致 涉及防止细菌附着和感染的新方法。

Although the bladder is by far the most common site of UTI, little is known about the molecules to which infecting bacteria attach in this tissue as compared with the kidney, in which the role of globoseries GSLs as bacterial attachment sites for Escherichia coli has been well studied. Since the carbohydrate portion of GSLs varies in different tissues, often under host genetic control, these molecules serve as determinants of both patient susceptibility to infection and tissue tropism for pathogenic bacteria. The overall goal of this proposal is to utilize primary cell cultures from the bladder as a model system to define the role and regulation of bladder GSLs as attachment sites for uropathogenic bacteria, focusing on the two most common causes of cystitis in women, E. coli and S. saprophyticus, in the following proposed studies: (1) To evaluate the hypothesis that globoseries (Gb) and ganglioseries (Gg) GSLs are present in primary cultures of bladder transitional epithelium and bind E. coli and S. saprophyticus, respectively, we will purify GSLs from the cells, identify E. coli-and S. saprophyticus-binding GSLs, and confirm the identities of the GSLs using specific monoclonal antibodies (MAbs) and formal carbohydrate structural analysis; (2) To evaluate the hypothesis that the GSLs identified in Specific Aim 1 are surface exposed in primary bladder cell cultures and are functionally relevant in E. coli and S. saprophyticus attachment, we will study bacterial adherence before and after pre-treatment of the cell cultures with an saprophyticus attachment, we will study bacterial adherence before and after pre-treatment of the cell cultures with an inhibitor of GSL synthesis or MAbs directed against relevant GSLs; (3) To evaluate the hypothesis that the secretor and ABO genes influence the expression of Gb and Gb GSLs in the bladder and thus the capacity for E. coli binding, we will establish primary bladder cultures from patients of known secretor/ABO status and determine whether the predicted glycosyltransferases and GSL products are present in the cultured cells; and (4) To evaluate the hypothesis that cytokines are release from primary uroepithelial cell cultures and modulate the expression of E.coli-and S. saprophyticus-binding GSLs in the bladder, we will to examine the effects of adherence by E. coli and S. saprophyticus on cytokine release from the cells and the effects of tumor necrosis factor and IL-1, IL-6 and IL-8 on the expression of relevant GSLs in primary urothelial cell cultures and on bacterial adherence to these cells. These studies on the expression and regulation of bladder GSLs are relevant to the pathogenesis of infection, inflammation and differentiation in this organ and could eventually lead to novel approaches to prevention of bacterial attachment and infection.