The Role of Sphingolipids in Inflammatory Bowel Disease

描述（由申请人提供）：本提案的广泛长期目标是检查炎症性肠病（IBD）病理学中的鞘氨醇-1-磷酸（S1 P）来源，并将其合成酶鞘氨醇激酶1（SKI）与特异性抑制剂靶向，作为该疾病的潜在新型治疗方法。在具体目标1中，我们将确定结肠组织SK 1是IBD中SIP的来源。这将通过使用小鼠骨髓嵌合体模型结合溃疡性结肠炎小鼠模型来完成。我们将（i）通过照射野生型和鞘氨醇激酶-1敲除小鼠建立骨髓嵌合小鼠，并用宿主血液或来自相反品系的血液重建它们的血液。然后，我们将（ii）给予DSS以诱导骨髓嵌合小鼠的结肠炎，并检查疾病参数：如体重减轻、结肠缩短、脾肿大、疾病的组织学标志物、血液和组织SK 1活性以及鞘脂变化。接下来，我们将（iii）用流式细胞术检查细胞炎症反应和结肠组织中炎症的潜在机制，例如来自DSS施用的骨髓嵌合小鼠的TNFor和C 0X 2产生。这些研究与人类健康有明显的联系，应该牢固地建立IBD中SIP的来源，以允许潜在的靶向治疗。在具体目标2中，我们将开发新型SKI抑制剂作为治疗IBD的潜在新治疗方式。这一目的将检验抑制SKI将改善IBD严重程度的子假设。为此，我们将（1）评估我们新开发的SK 1抑制剂在FHC人结肠上皮细胞中抑制TNF α激活SK 1和下游信号传导的能力。我们将通过真实的时间-RTPCR和蛋白质印迹法检测这些下游效应。我们还将（ii）通过腹膜内或静脉内注射用SKI抑制剂治疗小鼠，并评估组织和血液SKI以及它们对DSS诱导的结肠炎的反应，如针对特定目标1所做的。这些研究推进了SKI抑制剂作为IBD的新型治疗方法。 公共卫生相关性：这项研究与人类健康和疾病直接相关。目前IBD的治疗涉及全身免疫抑制和使用类固醇，在患者中引起不良副作用，导致依从性问题。治疗该病需要更特异的靶点，副作用更小。这项研究的目标是找到一种新的治疗方法，抑制疾病的细胞机制，改善患者的健康。

DESCRIPTION (provided by applicant): The broad long-term objective of this proposal is to examine the source sphingosine-1-phosphate (S1P) in the pathology involved in inflammatory bowel disease (IBD), and target its synthetic enzyme, sphingosine kinase 1 (SKI) with specific inhibitors as a potential novel therapeutic approach to the disease. In Specific Aim 1, we will establish that colon tissue SK1 is the source of SI P in IBD. This will be done by using a mouse bone marrow chimera model in conjunction with a mouse model of ulcerative colitis. We will (i) establish bone marrow chimeric mice, by irradiating both wild-type and sphingosine kinase-1 knockout mice and reconstitute their blood with either host blood or blood from the opposite strain. We will then (ii) administer DSS to induce colitis in bone marrow chimeric mice, and examine parameters of disease: such as weight loss, colon shortening, splenomegaly, histologic markers of disease, blood and tissue SK1 activity, as well as sphingolipid changes. Next we will (Hi) examine the cellular inflammatory responses with flow cytometry and the underlying mechanisms of inflammation in colon tissue, such as TNFor and C0X2 production from DSS-administered, bone marrow chimeric mice. These studies have an obvious link to human health and should firmly establish the source of SI P in IBD to allow potential targeting of therapy. In Specific Aim 2 we will develop novel SKI inhibitors as potential new therapeutic modalities for the treatment of IBD. This aim will test the sub-hypothesis that inhibiting SKI will ameliorate the severity of IBD. To this end we will (1) evaluate our newly developed SK1 inhibitors as to their ability to inhibit TNFa activation of SK1 and downstream signaling in FHC human colon epithelial cells. We will examine these downstream effects by Real Time-RTPCR and western blotting. We will also (ii) treat mice with SKI inhibitors by either i.p. or i.v. injection and evaluate tissue and blood SKI as well as their response to DSS- induced colitis as done for Specific Aim 1. These studies advance SKI inhibitors as novel therapeutic approaches to IBD. PUBLIC HEALTH RELEVANCE: This research is directly related to human health and disease. Current therapies for IBD involve general immunosuppression and the use of steriods, causing undesirable side effects in patients leading to issues of compliance. More specific targets with less side effects are needed for treatment of this disease. This research is targeted at finding just that, a novel therapeutic that inhibits the cellular mechanism of disease and improving patient health.