Mechanisms of Oxalobacter-Induced Enteric Oxalate Excretion

描述(申请人提供)：高草酸尿症被认为是草酸钙结石病的主要危险因素，这种疾病在大约12%的美国人口中发生，除了减少草酸盐的饮食来源外，目前还没有可用的药物治疗方法。与高草酸尿和高草酸血症相关的更严重的并发症，如肾功能衰竭、结石和草酸组织沉积(草酸沉着症)也会发生在原发性高草酸尿症(PH)中，这是一种进行性遗传性代谢疾病，由肝脏丙氨酸-乙醛转氨酶(AGT)酶缺乏引起，只有通过肝肾联合移植才能治愈。显然，需要注意草酸相关疾病的替代治疗方案，我们建议研究促进草酸的肠道消除和降解，以此作为减少高草酸尿和高草酸血症的一种方式。从我们最近对大鼠/小鼠结肠草酸转运的研究中获得的几个关键信息为这里提出的研究提供了方向。首先，我们已经证明，除了降解草酸的饮食来源外，底物/草酸特有的微生物草酸杆菌通过诱导结肠草酸的分泌/排泄，可以显著降低尿中草酸的排泄。我们也有令人信服的结果表明，这种厌氧细菌通过产生一种促分泌剂与运输粘膜相互作用，激活了肠道草酸处理中的这些有益变化。其次，基于我们对野生型(WT)和基因敲除(KO)小鼠的研究，我们发现了两个关键的阴离子交换器，即PAT1(Slc26a6)和DRA(Slc26a3)，它们对肠道中草酸的移动有重要贡献，我们假设草酸转运的变化依赖于这些转运蛋白的表达和功能。因此，我们将研究草酸在PAT1和DRA单一KO小鼠以及PAT1/AGT双KO小鼠模型和被草酸杆菌定植的WT小鼠中的肠道和肾脏处理，并与相应的非定植小鼠进行比较。了解肠道草酸盐处理过程中细菌细胞调节的机制基础，对于未来确定哪些菌株和/或细菌产品将有效治疗原发性高草酸尿症和草酸钙结石病是至关重要的。公共卫生相关性：这些研究提出了草酸杆菌在大肠的定植导致肠道草酸排泄和降低尿草酸排泄的机制。此外，这些研究将使用靶向缺失slc26a3(DRA)和slc26a6(PAT1)的敲除小鼠模型来评估草酸杆菌是否调节两个已知的重要草酸转运体的功能。然而，与本申请特别重要和相关的是最近获得的PAT1/AGT双KO小鼠模型，该模型为我们提供了一个独特的机会来评估在有和没有草酸杆菌定植的原发性高草酸尿症1型(PH1)的情况下PAT1的功能/活性。了解PH1中草酸的处理以及PAT1和草酸杆菌在调节这种遗传疾病中草酸过量的身体负担方面的潜在作用和相互作用现在是可能的。

DESCRIPTION (provided by applicant): Hyperoxaluria is considered to be a major risk factor in calcium oxalate stone disease which occurs in about 12% of the American population and, apart from reducing dietary sources of oxalate, there is currently no pharmacological treatment available. More serious complications associated with hyperoxaluria and hyperoxalemia, such as renal failure, stones, and tissue deposits of oxalate (oxalosis) also occur in Primary Hyperoxaluria (PH), a progressive inherited metabolic disorder caused by a liver deficiency of the enzyme alanine-glyoxylate aminotransferase (AGT) which is cured only by a combined liver-kidney transplant. Clearly, alternative treatment options for the oxalate-associated diseases require attention and we are proposing to examine promoting intestinal elimination and degradation of oxalate as a way of reducing both hyperoxaluria and hyperoxalemia. Several key pieces of information from our recent studies of colonic oxalate transport in rats/mice have provided the direction for the studies proposed here. First, we have shown that, in addition to degrading dietary sources of oxalate, the substrate/oxalate-specific microorganism, Oxalobacter sp., which resides exclusively in the large intestine, can significantly lower urinary oxalate excretion by inducing colonic oxalate secretion/excretion. We also have compelling results indicating this anaerobic bacterium interacts with the transporting mucosa by producing a secretagogue that activates these beneficial changes in intestinal oxalate handling. Second, based upon our studies using wild type (WT) and knockout (KO) mice, we have identified two key anion exchangers, namely PAT1 (slc26a6) and DRA (slc26a3), that contribute significantly to the movements of oxalate in the intestine and we hypothesize that the changes in oxalate transport induced by Oxalobacter are dependent upon the expression and function of these transporters. Thus, we will examine intestinal and renal handling of oxalate in PAT1 and DRA single KO mice as well as in a PAT1/AGT double KO mouse model and WT mice colonized with Oxalobacter and compared to their appropriate non-colonized counterparts. An understanding of the mechanistic basis for bacterial cell modulation of intestinal oxalate handling is fundamental to future efforts in identifying which strains of bacteria and/or bacterial products will be effective in the treatment of Primary Hyperoxaluria and calcium oxalate stone disease. PUBLIC HEALTH RELEVANCE: The studies proposed address the mechanisms by which Oxalobacter colonization of the large intestine induces enteric oxalate elimination and lowers urinary oxalate excretion. In addition, these studies will evaluate whether Oxalobacter modulates the function of two known important oxalate transporters using knockout mouse models with targeted deletion of slc26a3 (DRA) and slc26a6 (PAT1). Especially important and relevant to the present application, however, is the recent availability of a PAT1/AGT double KO mouse model which affords us a unique opportunity to evaluate PAT1 function/activity in the setting of Primary Hyperoxaluria, type1 (PH1), with and without Oxalobacter colonization. Understanding oxalate handling in PH1 and the potential roles and interactions of PAT1 and Oxalobacter in modulating the excessive body burden of oxalate in this genetic disease are now possible.