Substance P in pathogenesis of cryptosporidiosis in AIDS

描述(申请人提供)：隐孢子虫病，由原生动物寄生虫微小隐孢子虫引起，在正常宿主中是自限性的，但可在艾滋病患者中引起危及生命的慢性腹泻。对于与晚期艾滋病相关的隐孢子虫病，目前还没有成功的安全有效的治疗方法。微小弧菌感染引起肠道生理变化，如氯离子分泌增加(CI)和上皮屏障破坏，导致水样腹泻。P物质(SP)是一种神经肽，是一种痛觉递质，可引起人体肠道外植体中的氯离子分泌。我们之前已经研究了患有自然严重隐孢子虫病的艾滋病患者和实验感染微小隐孢子虫(轻度疾病)的正常志愿者空肠活检组织中SP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表达更强。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。我们还假设，由于隐孢子虫病感染，免疫缺陷宿主的肠道组织中SP的表达会增加，HIV感染本身并不会导致SP的表达增加。为了验证这些假设，我们建议使用一种免疫缺陷的隐孢子虫病动物模型，即。患有艾滋病(在实验性SIV感染后)和隐孢子虫病的灵长类动物被认为是一种机会性的自然感染。动物模型的优点是，从动物模型获取大量组织样本更容易从患有隐孢子虫病的艾滋病患者身上获得组织样本，旨在确定疾病发病机制的分子靶点和特定拮抗剂的初步治疗测试的研究最好可以使用动物来源的组织进行研究。该项目的目标是验证SP在免疫缺陷宿主中介导严重隐孢子虫病症状的假设。具体目的1：与非隐孢子虫病免疫缺陷动物和免疫功能正常的亚临床实验性隐孢子虫病猕猴相比，慢性自然感染隐孢子虫病免疫缺陷动物的肠道SP表达是否上调。我们将比较免疫缺陷猕猴(艾滋病)自然感染和不感染细小弧菌的免疫缺陷猕猴以及有和不存在亚临床实验性细小弧菌感染的正常猕猴的回肠SP基因和蛋白的表达水平。具体目的2：验证SP是介导肠道生理改变的关键因素的假设，这些改变导致与免疫缺陷宿主相关的自然发生的慢性隐孢子虫病的水样腹泻。用Ussing小室技术比较了SIV感染的猕猴(患有艾滋病)和未自然感染微小弧菌的猕猴回肠组织在SP受体拮抗剂存在和不存在的情况下的Cl-离子分泌和屏障完整性。这些研究将确定SP在微小弧菌引起的腹泻发病机制中的作用。有关SP参与疾病过程的证据将支持使用SP受体拮抗剂作为与艾滋病相关的隐孢子虫病以及可能与其他肠道病原体相关的危及生命的疾病的治疗方法。

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.