Molecular Regulation of Intestinal Calcium Absorption

背景：钙(Ca~(2+))在多种生理过程中起着中心作用。因此，它在等离子体中保持在狭窄的范围内。这是通过肠道吸收、骨骼沉积/吸收以及肾滤过和随后的再吸收之间的动态相互作用实现的。第一步是肠道钙吸收，这是NSERC资助的这项研究计划的重点，通过两种途径之一进行。Ca~(2+)可通过主动的跨细胞途径(通过细胞)或被动的细胞旁途径(上皮细胞之间)被吸收。当饮食中有足够的钙离子时，被动的细胞旁吸收占主导地位。然而，当饮食中的钙离子含量较低时，跨细胞转运是很重要的。这是由一系列运输和承运人事件调节的(图1)。十二指肠被认为是活跃的跨细胞钙吸收的主要部位，尽管包括我的项目在内的新证据表明，大肠近端具有重要作用。肠道钙吸收的主要调节剂是维生素D，它增加了主动的跨肠道钙转运。肾脏直接对细胞外钙离子做出反应，我们已经在肠道中证明了类似的直接钙感应机制，但这一途径在整体钙稳态中的作用尚不清楚。 模式系统：我在这方面的研究计划集中在调节小鼠模型肠道钙吸收的基本分子机制上，因为它们：i)可以在整个生物体的背景下研究钙离子的动态平衡；ii)可以很容易地采用药理学和遗传学方法，包括关键因子的过度表达和击倒；以及iii)可以在Ussing小室中直接测量跨越肠上皮细胞的钙通量，在Ussing小室中，电压和浓度梯度可以得到严格控制，从而能够独立地询问旁细胞和跨细胞途径。 目标与方法：我未来5年的肠道钙吸收研究计划的主要目标有三个方面。1)探讨近端大肠对钙离子吸收的调节作用。我们假设这是一个重要的钙挽救部位，使钙损失最小化，或者当血浆钙水平较高时，有助于显著的细胞旁分泌。为此，我们将测量近端大肠标本对钙的吸收，并检测野生型和转基因动物的尿液、粪便和血浆中的钙水平。 2)探讨血清钙水平对细胞旁肠钙吸收的直接影响以及钙离子受体在这一过程中的作用。为此，我们将使用CASR的药理调节剂，并敲除肠道中的CASR。 3)使用类似的方法，我们将描绘出在紧密连接中有助于正钙平衡和生命早期适当矿化的骨骼的分子。 影响：这些研究将描述维持哺乳动物钙稳态的基本机制。

BACKGROUND: Calcium (Ca2+) is central to a myriad of physiological processes. Consequently it is maintained within narrow limits in plasma. This occurs via dynamic interplay between intestinal absorption, deposition/resorption from bone and by renal filtration and subsequent reabsorption. The first step is intestinal Ca2+ absorption, the focus of this NSERC funded research program, which occurs via one of two routes. Ca2+ can be absorbed by an active transcellular (through the cell) pathway, or by a passive paracellular (between epithelial cells) pathway. When there is sufficient Ca2+ in the diet, passive paracellular absorption predominates. However, when dietary Ca2+ is low, transcellular transport is important. This is mediated by a series of transport and carrier events (Fig 1). The duodenum is proposed to be the primary site of active transcellular Ca2+ absorption, although emerging evidence, including from my program, suggests a significant role for the proximal large bowel. The primary regulator of intestinal Ca2+ absorption is vitamin D, which increases active transintestinal Ca2+ transport. The kidney responds directly to extracellular Ca2+ and we have demonstrated a similar direct Ca2+ sensing mechanism in the intestine, but the role of this pathway in overall Ca2+ homeostasis is not clear. MODEL SYSTEMS: My research program in this area focuses on basic molecular mechanisms regulating intestinal Ca2+ absorption in mouse models because they: i) permit the study of Ca2+ homeostasis within the context of the whole organism; ii) are readily amenable to pharmacologic and genetic approaches, including over-expression and knockdown of key players; and iii) permit direct measurement of Ca2+ flux across intestinal epithelium in Ussing chambers where voltage and concentration gradients can be tightly controlled, enabling the interrogation of paracellular and transcellular pathways independently. OBJECTIVES & APPROACH: The main objectives of my research program in intestinal Ca2+ absorption over the next 5 years are three-fold. 1) To delineate the regulatory role of the proximal large bowel in Ca2+ absorption. We hypothesize this is an important site of Ca2+ salvaging minimizing Ca2+ loss or when plasma Ca2+ is high, contributing significant paracellular secretion. To this end, we will measure Ca2+ absorption across proximal large bowel preparations and examine urine, fecal and plasma Ca2+ levels in wild-type and genetically modified animals. 2) We will explore the direct effect of serum Ca2+ levels on paracellular intestinal Ca2+ absorption and the role of the CaSR in this process. To do so we will use pharmacological modulators of the CaSR and knockout the CaSR in the intestine. 3) Using similar approaches we will delineate molecules in the tight junction contributing to a positive Ca2+ balance and properly mineralized bone early in life. IMPACT: These studies will delineate fundamental mechanisms maintaining mammalian Ca2+ homeostasis.