Substance P in pathogenesis of cryptosporidiosis in AIDS

描述（由申请人提供）：隐孢子虫病由原生动物寄生虫小隐孢子虫引起，在正常宿主体内具有自限性，但可导致艾滋病患者危及生命的慢性腹泻。对于晚期艾滋病相关的隐孢子虫病，目前还没有安全有效的治疗方法。细小梭菌感染引起肠道生理变化，如氯阴离子分泌增加（CI）和上皮屏障破坏导致水样腹泻。P物质（SP）是一种神经肽，是一种疼痛递质，可引起人肠道外植体分泌C1离子。我们之前已经研究了SP在自然严重隐孢子虫病艾滋病患者和正常志愿者空肠活检中的表达。与患有轻度自限性隐孢子虫病的正常志愿者相比，艾滋病患者的SP表达较强。我们假设SP是艾滋病相关隐孢子虫病慢性肠道症状的关键介质。我们也假设SP在免疫缺陷宿主肠道组织中的表达会因隐孢子虫病感染而升高，单独感染HIV不会引起SP表达升高。为了验证这些假设，我们建议使用免疫缺陷的隐孢子虫病动物模型，即。灵长类动物与艾滋病（实验SIV感染后）和隐孢子虫病作为机会性自然发生的感染。动物模型的优点是，从动物模型中更容易获得大的组织样本，从艾滋病隐孢子虫病患者中获取组织样本，并且，旨在确定疾病发病机制的分子靶点和特异性拮抗剂的初步治疗试验的研究可以最好地使用动物来源的组织进行研究。本项目的目的是验证SP在免疫缺陷宿主中介导隐孢子虫病严重症状的假设。特异性目的1：确定与没有隐孢子虫病的免疫缺陷动物或患有亚临床实验隐孢子虫病的正常免疫能力猕猴相比，患有慢性自然感染隐孢子虫病的免疫缺陷动物的肠道SP是否上调。我们将比较免疫缺陷猕猴（艾滋病患者）感染和不感染天然细小梭菌的猕猴与感染和不感染亚临床实验细小梭菌的正常猕猴之间回肠SP mRNA和蛋白的表达水平。特异性目的2：验证SP是介导免疫缺陷宿主自然发生的慢性隐孢子虫病水样腹泻的肠道生理改变的关键因素。采用Ussing chamber技术比较SIV感染猕猴（伴艾滋病）回肠组织在SP受体拮抗剂存在和不存在的情况下，C1离子分泌和屏障完整性。这些研究将确定SP在小孢子虫引起的腹泻发病机制中的作用。SP参与疾病过程的证据将支持使用SP受体拮抗剂治疗与艾滋病相关的隐孢子虫病和其他肠道病原体相关的危及生命的疾病。

DESCRIPTION (provided by applicant): Cryptosporidiosis, caused by the protozoan parasite, Cryptosporidium parvum, is self-limited in normal hosts but can cause life threatening, chronic diarrhea in AIDS patients. No safe and effective treatment has been successfully developed for cryptospofidiosis associated with advanced AIDS. C. parvum infection causes intestinal physiologic changes like, increased chloride anion secretion (CI) and epithelial barrier disruption that leads to watery diarrhea. Substance P (SP), a neuropeptide, is a pain transmitter and can cause C1- ion secretion in human intestinal explants. We have previously studied SP expression in jejunal biopsies of AIDS patients with natural severe cryptosporidiosis and normal volunteers experimentally challenged with C. parvum (mild disease). SP expression was stronger in AIDS patients compared to normal volunteers with mild self-limited cryptosporidiosis. We hypothesize that SP is a key mediator of chronic intestinal symptoms in AIDS associated cryptosporidiosis. We also hypothesize that SP expression will be elevated in intestinal tissues of immunodeficient hosts because of cryptosporidiosis infection, HIV infection alone will not cause increased SP expression. To verify these hypotheses, we propose to use an immunodeficient animal model of cryptosporidiosis, ie. primates with AIDS (after experimental SIV infection) and cryptosporidiosis as an opportunistic naturally occurring infection. Advantage of an animal model is that, it is easier to procure large tissue samples from an animal model to that from AIDS patients with cryptosporidiosis, and, studies aimed at defining molecular targets responsible for disease pathogenesis and initial therapeutic testing of specific antagonists can best be studied using animal derived tissues. The goal of this project is to test the hypothesis that SP mediates severe symptoms of cryptosporidiosis in immunodeficient hosts. Specific aim 1: To determine if intestinal SP is upregulated in immunodeficient animals with chronic naturally infected cryptosporidiosis as compared to immunodeficient animals without cryptosporidiosis or normal immunocompetent macaques with subclinical experimental cryptosporidiosis. Ileal expression of SP mRNA and protein levels will be compared between immunodeficient macaques (with AIDS) with and without naturally occurring C. parvum infection and in normal macaques with and without subclinical experimental C. parvum infection. Specific aim 2: To test the hypothesis that SP is a key factor that mediates intestinal physiological alterations that lead to watery diarrhea in naturally occurring chronic cryptosporidiosis associated with immunodeficient hosts. C1- ion secretion and barrier integrity will be compared between ileal tissues from SIV infected macaques (with AIDS) with and without naturally occurring C. parvum infection in the presence and absence of SP receptor antagonist by the Ussing chamber technique. These studies will determine the role of SP in the pathogenesis of C. parvum induced diarrhea. Evidence implicating SP in the disease process would support the use of SP receptor antagonists as a therapy for the life threatening illness associated with AIDS related cryptosporidiosis and perhaps other intestinal pathogens.