Mucosal Defense Mechanisms in Substance Abuse

沙门氏菌和其他肠道侵入性微生物引起的肠道感染是艾滋病和其他免疫抑制疾病患者发病和死亡的主要因素。人类肠道粘膜的大表面积与外界环境直接接触，必须与共生的肠道菌群共存，但不包括肠道病原体。此外，小肠中的Peyer's patches （PP）在粘膜免疫中发挥关键作用，通过粘膜屏障将腔内抗原传递到粘膜下抗原呈递细胞，但PP也可能作为肠侵入性微生物感染的入口通道，可产生肠炎和败血症。在之前的资助期内，我们获得了证据，证明猪回肠粘膜下层的一种新型抑制性阿片受体在肠神经回路中表达，炎症介质通过肠神经回路引起上皮阴离子分泌。在这些实验过程中，我们有了令人兴奋的新发现，高致死率的血清型霍乱沙门氏菌进入猪空肠PP和非PP上皮的细胞内摄取可能受到宿主肠神经系统的调节。本申请旨在通过解决以下假设来继续这一新颖的研究路线：(1)细菌对PP和非PP肠上皮的摄取通过肠内神经活动和关键肠内神经抑制分子（包括阿片类药物和大麻素）的变化而改变；(2)沙门氏菌摄取的神经调节并不是这种微生物所特有的。这些假设将通过研究药物和其他治疗对霍乱链球菌内化到离体猪空肠PP和非PP粘膜的影响来验证，具体目的如下：(1)表征支配猪空肠PP的肠神经元的投射、化学编码和受体特征；(2)功能性鉴定调节沙门氏菌内化到PP的肠内神经递质的种类；(3)确定去甲肾上腺素、阿片类药物和大麻素对沙门氏菌PP和非PP粘膜摄取的影响；(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。拟议实验的结果将增进我们对滥用药物对艾滋病相关肠道感染的有害影响的理解，并可能确定新的药理学治疗方法，以提高旨在预防人类免疫缺陷病毒和其他肠道病原体感染的口服疫苗的功效。

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.