Hormone Regulation of [Ca2+] in Pancreatic Acinar Cells

胰腺腺泡细胞中[Ca2]的激素调节

• 批准号: 7905573
• 负责人: Shmuel Muallem
• 金额: $ 7.68万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-05 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7905573
• 关键词: Acinar Cell; Address; Affect; Agonist; Alcohols; Amylases; Apoptosis; Bile Acids; Binding; Caerulein; Caspase; Cell Death; Cell membrane; Cell model; Cell physiology; Cells; Cellular Stress; Charge; Complex; Disease; Dyes; Electrostatics; Environment; Enzymes; Esters; Exocytosis; Functional disorder; Gene Expression; Genes; Goals; Hormones; In Vitro; Individual; Inflammation; Ions; Isotonic Exercise; Knockout Mice; Knowledge; Lactate Dehydrogenase; Liquid substance; Measures; Mediating; Modeling; Molecular; Mus; Neurotransmitters; Nodal; Pancreas; Pancreatitis; Patients; Physiological; Process; Protein Biosynthesis; Regulation; Reporter; Role; STIM1 gene; Second Messenger Systems; Signal Transduction; Stress; System; Time; Toxin; acute pancreatitis; cell injury; extracellular; hormone regulation; palmitoleic acid; public health relevance; receptor; response; scaffold; second messenger; sensor

DESCRIPTION (provided by applicant): The chief function of pancreatic acinar cells is the secretion of digestive enzymes and a small amount of isotonic fluid. Both functions are regulated by hormones and neurotransmitters that use Ca2+ as the second messenger. At the same time, aberrant Ca2+ signaling is the nodal point in all forms of pancreatitis. The hypothesis of this proposal is that specifically aberrant Ca2+ influx by store-operated Ca2+ channels (SOCs) is responsible for pancreatitis. Therefore, understanding regulation and function of Ca2+ influx channels is critical for understanding acinar cell function and dysfunction in the context of pancreatitis. Acinar cells have several Ca2+ influx channels, TRPC1, TRPC3, TRPC4, TRPC6 and Orai1, all of which are gated by the ER Ca2+ sensor STIM1. TRPCs are also gated by the scaffold Homer1 that inhibits their activity. Considering the involvement of SOCs in pancreatitis, the overall goal of this proposal is to understand gating mechanism of the pancreatic SOCs by STIM1 and Homer1 and their role in pancreatitis. This will be achieved in four aims: Aim 1: Decipher the molecular mechanism of TRPCs gating by STIM1 and Homer1. Homer1 interacts with a PPXF motif to keep the channels in a close state. STIM1 opens the TRPCs by interacting with two conserved residues that are only 4 aresidues upstream of the Homer1 binding motif. We propose to determine a) the gating mechanism of TRPCs by STIM1; b) how Homer1 and STIM1 function in tandem to gate the TRPCs. Aim 2: Determine molecular mechanism of Orai1 gating by STIM1. Agonist-stimulated pancreatic Ca2+ influx is mediated by a combination of TRPCs and Orai1 that are differentially gated by STIM1. We discovered that the STIM1(234-535) fragment is sufficient for full activation of Orai1. Hence we will: a) determine the minimal STIM1 domain that gates Orai1; b) how STIM1 opens Orai1; c) what is the relationship between the NATIVE TRPCs and NATIVE Orai1. Aim 3: The role of TRPCs, Orai1 and STIM1 in physiological and pathological acinar cells Ca2+ signaling and exocytosis will be determined by characterizing Ca2+ signaling and exocytosis in acinar cells from TRPC1-/-, TRPC3-/-, TRPC6-/-, Orai1-/- and STIM1-/- mice. Aim 4: The role of TRPC channels, Orai1, STIM1 will be studies in vitro-induced pancreatitis and measure cellular reporters of cell stress and cell damage. Then the KO mice will be used to evaluate the role of the Ca2+ influx channels in careluin and bile acid models of pancreatitis. PUBLIC HEALTH RELEVANCE: The pancreas secretes digestive enzymes and fluid in response to stimulation by hormones and neurotransmitters. The stimulants tell the pancreatic cells how to do so by changing the concentration of Ca2+ ions within the cells. Most of the Ca2+ enters the cells from the extracellular environment. When this process is aberrant, the digestive enzymes remain trapped within the cells and the cells dye and the patient develop the disease called pancreatitis. The aims of this proposal are to understand how Ca2+ ions enter the cells and how the aberrant Ca2+ entry causes the pancreatitis. This will be achieved by studying Ca2+ entry into the cells in model cell systems and in the cells of the pancreas of mice from which the genes that are responsible for the entry of Ca2+ into the cells have been deleted.

描述（由申请人提供）：胰腺腺泡细胞的主要功能是消化酶的分泌和少量的同量流体。这两个功能均由使用Ca2+作为第二使者的激素和神经递质调节。同时，异常的Ca2+信号传导是所有形式的胰腺炎中的淋巴结点。该提议的假设是，商店经营的Ca2+通道（SOC）特别异常的Ca2+涌入负责胰腺炎。因此，了解Ca2+涌入通道的调节和功能对于理解胰腺炎的腺泡细胞功能和功能障碍至关重要。腺泡细胞具有几个Ca2+涌入通道，TRPC1，TRPC3，TRPC4，TRPC6和ORAI1，所有这些通道都由ER Ca2+传感器STIM1封闭。 TRPC还通过抑制其活性的支架Homer1门控。考虑到SOC参与胰腺炎，该提案的总体目标是了解STIM1和HOMER1及其在胰腺炎中的作用的胰腺SOC的门控机制。这将在四个目的中实现：目标1：通过STIM1和HOMER1拆除TRPC的分子机制。 HOMER1与PPXF基序相互作用，以使通道保持近距离状态。 STIM1通过与Homer1结合基序上游的两个保守残基相互作用来打开TRPC。我们建议通过STIM1确定TRPC的门控机制； b）HOMER1和STIM1在串联中的功能如何在TRPC中发挥作用。 AIM 2：确定通过Stim1进行Orai1门控的分子机制。激动剂刺激的胰腺CA2+流入由TRPC和ORAI1的组合介导，而TRPC和ORAI1由STIM1差异化。我们发现Stim1（234-535）片段足以充分激活Orai1。因此，我们将：a）确定大门orai1的最小刺激域； b）STIM1如何打开Orai1； c）天然TRPC和本机Orai1之间的关系是什么？ AIM 3：TRPC，ORAI1和STIM1在生理和病理腺泡细胞中的作用CA2+信号传导和胞胞病的作用将通过表征Ca2+信号传导和胞吐作用来确定TRPC1 - / - ，TRPC3-/ - ，TRPC3-/ - ，TRPC6-，TRPC6-/ - ，ORAI1-/ - ， - / - ， - / - / - / - 和刺激。 AIM 4：TRPC通道，ORAI1，STIM1的作用将是在体外诱发的胰腺炎和测量细胞应激和细胞损伤的细胞记者的研究。然后，KO小鼠将用于评估Ca2+涌入通道在胰腺炎和胆汁酸模型中的作用。公共卫生相关性：胰腺分泌消化酶和液体，以响应激素和神经递质刺激。刺激物通过改变细胞内Ca2+离子的浓度来告诉胰腺细胞如何做到这一点。大多数CA2+从细胞外环境进入细胞。当此过程异常时，消化酶仍然被困在细胞中，细胞染料和患者发展为胰腺炎。该提案的目的是了解Ca2+离子如何进入细胞以及异常Ca2+进入如何导致胰腺炎。这将通过研究模型细胞系统中的Ca2+进入细胞和小鼠的细胞中的细胞来实现这一目标，从而删除了Ca2+进入细胞的基因。