Role of CD14 and TLR4 in Urothelial Response to LPS.

描述（由申请人提供）：尿路感染导致每年约960万医生就诊，20%的女性在其一生中会发生尿路感染。大多数（70-90%）的尿路感染是革兰氏阴性菌产生脂多糖（LPS）的结果，而脂多糖是革兰氏阴性菌细胞壁的成分，负责导致疼痛和炎症的细胞反应。多种器官系统（胃肠道、呼吸系统、泌尿系统）的粘膜是抵抗革兰氏阴性菌感染的第一道天然防线；然而，调节尿路上皮细胞对LPS反应的细胞过程仍不清楚。尽管其他研究人员报道尿路上皮细胞不表达CD14 （LPS的主要同源细胞受体），但最近的一份报告描述了人膀胱粘膜中CD14的信息存在，我们实验室的初步调查证实了人尿路上皮细胞中存在信息和蛋白质。这些细胞也表达toll样受体4 (TLR4)，它似乎提供了响应LPS的跨膜信号通路。尿路上皮细胞有能力产生细胞因子来招募和激活白细胞。本研究的目的是验证CD14和TLR4在尿路上皮对LPS的反应中起关键作用的假设。利用CD14或TLR4基因破坏的人类尿路上皮细胞和小鼠的培养，我们将研究在存在或不存在功能性CD14和TLR4的情况下，体外和体内尿路上皮对LPS的反应。此外，我们将确定CD14的可溶性形式是否会增强尿路上皮细胞对LPS的反应。所提出的实验将更好地确定这些受体在存在（体内）和不存在（体外）其他先天免疫成分的情况下参与尿路上皮细胞对LPS的反应。我们预计这些研究将为进一步研究提供基础，以确定可能参与这一过程的其他蛋白质，并解剖将LPS结合尿路上皮细胞转化为细胞反应的信号通路。本研究的长期目标是确定策略，以尽量减少疼痛和有害的细胞效应与膀胱炎由于脂多糖产生革兰氏阴性菌。

DESCRIPTION (provided by applicant): Urinary tract infections result in approximately 9.6 million doctor visits annually, and 20% of women will develop a urinary tract infection during their lifetime. A majority (70-90%) of urinary tract infections are the result of gram-negative bacteria that produce lipopolysaccharide (LPS), and LPS is the component of the cell wall of gram-negative bacteria responsible for cellular response resulting in pain and inflammation. The mucosa of a variety of organ systems (GI, respiratory, urinary) functions as the first line of innate defense against infection by gram-negative bacteria; however, cellular processes that regulate the response of urothelial cells to LPS remain unknown. Although other investigators have reported that urothelial cells do not express CD14 (the primary cognate cellular receptor for LPS), a recent report described the presence of message for CD14 in human bladder mucosa, and preliminary investigations in our laboratory demonstrated the presence of message and protein in human urothelial cells. These cells also express Toll-Like Receptor 4 (TLR4), which appears to provide the transmembrane pathway for signaling in response to LPS. Urothelial cells have the capacity to produce cytokines which recruit and activate leukocytes. The goal of this research is to test the hypothesis that CD14 and TLR4 play a crucial role in the response of the urothelium to LPS. Using cultures of human urothelial cells and mice with genetic disruption of either CD14 or TLR4, we will investigate the response of the urothelium in vitro and in vivo to LPS in the presence or absence of functional CD14 and TLR4. Further, we will determine whether or not the soluble form of CD14 amplifies the response of urothelial cells to LPS. The proposed experiments will better define participation of these receptors in the response of urothelial cells to LPS in the presence (in vivo) and absence (in vitro) of other components of innate immunity. We anticipate that these studies will provide the foundation for continued investigations to identify additional proteins that may participate in this process and dissect signaling pathways which translate LPS binding by urothelial cells into cellular response. The long-range goal of this research is to identify strategies to minimize pain and harmful cellular effects associated with cystitis due to LPS-producing gram-negative bacteria.