Role of Na+/H+ exchanger in diabetic diarrhea

腹泻是来自美国的部署军事人员最常见的抱怨之一，并已 对服役人员的健康产生重大不利影响。据报道，76.8%的士兵在行动中 伊拉克自由和持久自由行动经历了腹泻。糖尿病正在成为一种日益严重的 对退伍军人的健康关注，四分之一的退伍军人得到退伍军人事务部的照顾 患有糖尿病。糖尿病常见的令人头疼的胃肠道并发症是腹泻。糖尿病腹泻 因其严重性和难治性而具有临床意义。糖尿病的总发病率 腹泻的比例可能高达22%。虽然2型糖尿病(T2 DM)患者腹泻的发生率较低，但 T2 DM患者腹泻的常见原因与药物有关，包括二甲双胍，它通常用于 2型糖尿病患者的血糖控制。与退伍军人事务部的临床相关性是，超过70%的 退伍军人管理局的患者超重或肥胖，T2 DM是一个主要的健康问题。然而，造成这种情况的根本原因 T2 DM的腹泻尚未被研究，因此有必要改进糖尿病腹泻的治疗。腹泻 是由电解质和液体的肠道运输改变引起的，但异常的电解质之间的联系 运输和糖尿病腹泻没有得到证实。肠道对Na+的主要吸收机制是 Na+/H+交换器3(NHE3)介导的电中性盐吸收。对NHE3的抑制与 同时伴有肠毒素和炎症性腹泻。我们最近对1型糖尿病(T1 DM)的研究表明 在T1 DM患者和小鼠中，NHE3的表达下调，这有助于识别特定的离子 转运蛋白首次被认为是糖尿病腹泻的原因。初步研究表明，NHE3 在人类糖尿病组织和T2 DM小鼠模型db/db小鼠中表达减少。此外，我们 有令人信服的证据表明，治疗T2 DM的广泛处方药二甲双胍抑制NHE3，这表明 NHE3功能障碍与二甲双胍引起的频繁腹泻有关。一磷酸腺苷激酶 AMPK是抗糖尿病药物二甲双胍的主要效应者，AMPK的激活导致NHE3抑制， 提示AMPK在二甲双胍对NHE3的调控中起关键作用。拟议研究的目标是 确定NHE3表达和活性降低在糖尿病腹泻，特别是T2 DM中的影响。这个 这项研究的中心假设是，PKCα升高对NHE3的抑制是NHE3的主要原因 对T2 DM的抑制和抗糖尿病药物如二甲双胍对AMPK的激活进一步抑制NHE3， 导致一些患者腹泻。旨在检验这一假设的拟议研究将建立一个 新的范式认为，糖尿病腹泻是由NHE3对钠和液体运输的异常调节引起的。我们 建议检验PKCα是T2 DM患者NHE3活性降低和液体吸收减少的主要原因这一假设 使用肠上皮细胞的小鼠和T2 DM的实验模型(目标1)。我们建议确定 AMPK在体内外对NHE3均有抑制作用。我们将通过以下方法确定NHE3抑制的潜在机制 AMPK通过研究导致NHE3抑制的信号通路。我们还计划确定AMPK是否 在人源化小鼠中，二甲双胍激活通过泛素化NHE3介导对NHE3的抑制(目标2)。 作为一项改善糖尿病腹泻治疗的临床前试验，我们将从生物学角度测试一种 出现磷脂、溶血磷脂酸和益生菌以减轻对NHE3的抑制(目标3)。

Diarrhea is one of the most frequent complaints in deployed military personnel from the United States and has significant adverse effects on the health of service members. It was reported that 76.8 % of soldiers in Operation Iraqi Freedom and Operation Enduring Freedom experienced diarrhea. Diabetes is becoming an increasing health concern for veterans, with one in four veterans receiving care from the Department of Veterans Affairs has diabetes. A common troublesome gastrointestinal complication of diabetes is diarrhea. Diabetic diarrhea attains clinical significance because of its severity and refractory nature. The overall incidence of diabetic diarrhea can reach as high as 22%. Although diarrhea is less frequent in type 2 diabetic mellitus (T2DM), the frequent cause of diarrhea in T2DM is associated with drugs, including metformin, which is commonly used for glycemic control in T2DM. Clinical Relevance to the Department of Veterans Affair is that with more than 70% of patients in VA facilities being overweight or obese, T2DM is a major health concern. Yet the underlying cause of diarrhea in T2DM has not been studied and there is a need to improve treatment for diabetic diarrhea. Diarrhea is caused by altered intestinal transport of electrolytes and fluid, but the link between the aberrant electrolyte transport and diabetic diarrhea is not established. The major Na+ absorptive mechanism in the intestine is electroneutral NaCl absorption mediated by the Na+/H+ exchanger 3 (NHE3). Inhibition of NHE3 is associated with both enterotoxigenic and inflammatory diarrhea. Our recent study of type 1 diabetic mellitus (T1DM) showed that NHE3 expression is downregulated in T1DM humans and mice, which helped to identify a specific ion transporter as a cause of diabetic diarrhea for the first time. Preliminary studies have demonstrated that NHE3 expression is decreased in human diabetic tissues and db/db mice, a mouse model for T2DM. Additionally, we have compelling evidence that metformin, a widely prescribed drug to treat T2DM, inhibits NHE3, suggesting NHE3 dysfunction is associated with frequent diarrhea caused by metformin. Adenosine monophosphate kinase (AMPK) is a major effector of anti-diabetic metformin, and activation of AMPK causes NHE3 inhibition, suggesting the critical role of AMPK in NHE3 regulation by metformin. The objective of the proposed study is determine the impact of decreased NHE3 expression and activity in diabetic diarrhea, in particular T2DM. The central hypothesis of this proposed study is that inhibition of NHE3 by elevated PKCα is a major cause of NHE3 inhibition in T2DM, and activation of AMPK by anti-diabetic drugs such as metformin further inhibits NHE3, contributing to diarrhea in some patients. The proposed studies designed to test this hypothesis will establish a new paradigm that diabetic diarrhea is caused by aberrant regulation of sodium and fluid transport by NHE3. We propose to test the hypothesis that PKCα is a major cause of reduced NHE3 activity and fluid absorption in T2DM mice using intestinal epithelial cells and experimental models of T2DM (Aim 1). We propose to determine that AMPK inhibits NHE3 in vitro and in vivo. We will determine the underlying mechanism of NHE3 inhibition by AMPK by investigating signal pathways responsible NHE3 inhibition. We also plan to determine whether AMPK activation by metformin mediates NHE3 inhibition via ubiquitination of NHE3 by using humanized mice (Aim 2). As a pre-clinical test to improve the treatment for diabetic diarrhea, we will test the efficacy of a biologically occurring phospholipid, lysophosphatidic acid, and probiotics in mitigating the inhibition of NHE3 (Aim 3).