Mucosal Defense Mechanisms in Substance Abuse

沙门氏菌和其他肠道侵袭性微生物的肠道感染是艾滋病和其他免疫抑制疾病患者发病率和死亡率的主要因素。人的肠粘膜有很大的表面积，与外界环境直接接触，必须与共生的肠道微生物共存，但不包括管腔病原体。此外，小肠中的Peyer‘s Patches(PP)通过跨过黏膜屏障将管腔抗原运送到粘膜下的抗原提呈细胞，在粘膜免疫中发挥关键作用，但PP也可能是肠道侵袭性微生物感染的入口，从而导致肠炎和败血症。在之前的资助期间，我们获得的证据表明，猪回肠黏膜下层的一种新的抑制性阿片受体在肠道神经回路中表达，炎性介质通过该回路激发上皮阴离子分泌。在这些实验过程中，我们有了一个令人兴奋的新发现，即高致死性霍乱沙门氏菌对猪空肠PP和非PP上皮细胞内的摄取可能受到宿主肠道神经系统的调节。这项申请旨在通过解决以下假设来继续这一新的研究路线：(1)细菌对PP和非PP肠道上皮细胞的吸收会因肠道神经活动和关键的肠道神经抑制分子(包括阿片和大麻)的变化而改变；以及(2)沙门氏菌吸收的神经调节不是这种微生物所特有的。这些假说将通过检测药物和其他治疗方法对霍乱沙门氏菌内化到分离的空肠PP和来自猪的非PP黏膜的影响来检验，具体目的如下：(1)表征支配猪空肠段PP的肠神经神经元的投射、化学编码和受体特征；(2)从功能上鉴定调节沙门氏菌内化到PP的肠道神经递质类别；(3)确定去甲肾上腺素、阿片和大麻素在改变PP和非PP粘膜对沙门氏菌摄取的作用；以及(4)研究去甲肾上腺素和其他神经抑制药物对PP和非PP肠粘膜细菌特异性和非特异性粘膜摄取过程的作用。拟议的实验结果应能加深我们对滥用药物在艾滋病相关肠道感染中的有害影响的了解，并可能确定能够提高口服疫苗效力的新药物治疗方法，该疫苗旨在预防人类免疫缺陷病毒和其他肠道病原体的感染。

Enteric infections by Salmonella and other enteroinvasive microorganisms are a major contributing factor to morbidity and mortality in individuals with AIDS and other immunosuppressed conditions. The large surface area of the human intestinal mucosa is in direct contact with the external environment, and must coexist with commensal enteric microflora but exclude luminal pathogens. In addition, Peyer's patches (PP) in the small intestine play a key role in mucosal immunity by delivering luminal antigens across the mucosal barrier to submucosal antigen-presenting cells, but PP may also serve as entryways for infection by enteroinvasive microorganisms which can produce enteritis and septicemia. In the previous funding period, we obtained evidence that a novel inhibitory opioid receptor in the porcine ileal submucosa is expressed in an enteric neural circuit through which inflammatory mediators evoke epithelial anion secretion. In the course of these experiments, we made the exciting new discovery that the intracellular uptake of the highly lethal Salmonella serovar choleraesuis into PP and non-PP epithelia of porcine jejunum may be modulated by the enteric nervous system of the host. This application seeks to continue this novel line of investigation by addressing the hypotheses that (1) bacterial uptake into PP and non-PP intestinal epithelia is altered by changes in enteric neural activity and key enteric neuroinhibitory molecules, including opioids and cannabinoids; and (2) that neuroregulation of Salmonella uptake is not specific for this microorganism. These hypotheses will be tested by examining the effects of drugs and other treatments on the internalization of S. choleraesuis into isolated jejunal PP and non-PP mucosa from pigs through the following Specific Aims: (1) to characterize the projections, chemical coding and receptor signatures of enteric neurons innervating PP in porcine jejunal segments; (2) to functionally identify classes of enteric neurotransmitters modulating Salmonella internalization into PP; (3) to define the actions of norepinephrine, opioids and cannabinoids in altering PP and non-PP mucosal uptake of Salmonella; and (4) to examine the actions of norepinephrine and other neuroinhibitory drugs on bacteria-specific and non-specific mucosal uptake processes in PP and non-PP intestinal mucosae. The results of the proposed experiments should enhance our understanding of the deleterious effects of abused drugs in AIDS- related enteric infections and may identify new pharmacological treatments capable of enhancing the efficacy of oral vaccines designed to prevent infections by the human immunodeficiency virus and other enteropathogens.