Molecular Characterization of Folate Uptake in Pancreatic Acinar Cells

胰腺腺泡细胞叶酸摄取的分子表征

• 批准号: 7612291
• 负责人: Lisa Mee
• 金额: $ 3.57万
• 依托单位: UNIVERSITY OF CALIFORNIA-IRVINE
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-09-16 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7612291
• 关键词: Acids; Acinar Cell; Acinus organ component; Address; Affect; Alcoholism; Alcohols; Amino Acids; Anemia; Anions; Buffers; Calories; Carbon; Carrier Proteins; Cell physiology; Cells; Chronic; Clinical; Coenzymes; Condition; Coupled; DNA biosynthesis; DNA chemical synthesis; Development; Diet; Enzymes; Ethanol; Event; Folate; Folic Acid Deficiency; Genes; Genetic Transcription; Growth; Homeostasis; Homocysteine; Homocystine; Human; Immunohistochemistry; Impairment; In Situ Hybridization; Investigation; Kinetics; Light; Liquid substance; Liver; Malignant neoplasm of pancreas; Mammalian Cell; Mediating; Messenger RNA; Metabolic; Metabolism; Methylation; Molecular; Neural Tube Defects; Nutrient; Pancreas; Pancreatic Diseases; Pancreatitis; Pathogenesis; Pattern; Pharmaceutical Preparations; Phenytoin; Physiological; Physiology; Process; Proteins; Protons; RNA; RNA Precursors; Range; Rate; Rattus; Reaction; Regulation; Relative (related person); Research Personnel; Risk; SLC19A1 gene; Source; Specificity; Sulfasalazine; Supplementation; System; Temperature; Testing; Time; Tissue Sample; Tissues; Training; Vitamins; Water-Soluble Vitamin; Week; Work; analog; base; cancer risk; carcinogenesis; cell type; chronic alcohol ingestion; design; feeding; folate-binding protein; inhibitor/antagonist; insight; nutrition; small hairpin RNA; uptake

DESCRIPTION (provided by applicant): Folate, an essential vitamin, is critical for RNA/DNA synthesis, metabolism of certain amino acids (including homocysteine), and methylation. In the pancreas, folate deficiency causes a decrease in digestive enzyme secretion and increases risk of cancer. Pancreatic cells are unable to synthesize folate, obtaining it from exogenous sources. Since cellular and molecular mechanisms involved in folate uptake and its regulation in pancreatic acinar cells are unclear, we initiated preliminary studies using primary and cultured (AR42J) rat pancreatic acinar cells obtaining evidence suggesting involvement of a carrier-mediated mechanism for folate uptake. We showed that these cells express the major folate transporters, the reduced folate carrier (RFC) and the proton coupled folate transporter (PCFT). We propose to continue studies to delineate the specificity, requirements, and kinetics of the folate uptake process and to determine the relative functional contribution of RFC and PCFT toward total carrier-mediated folate uptake via an shRNA approach. We also propose to examine if the pancreatic folate uptake process is adaptively regulated by substrate availability (deficiency and over-supplementation) and whether or not it undergoes developmental regulation. Our studies will be extended to the human situation by examining the pattern of RFC and PCFT expression at both the RNA (in situ hybridization) and protein (immunohistochemistry) levels using normal human pancreatic tissue samples. Chronic alcoholism is a leading cause of pancreatitis and systemic folate deficiency. Although the effects of chronic alcohol ingestion on pancreatic physiology has been intensely studied, little is known about its effects on folate transport specifically. Thus, in preliminary studies we showed that chronic alcohol feeding to rats leads to a significant reduction in RFC and PCFT mRNA levels in the pancreas compared to their pair-fed controls. We propose to continue these studies and to examine the functional consequences of these effects on kinetics of folate uptake by pancreatic acinar cells, level of expression of the folate carrier proteins, and on intracellular folate content. We also propose to examine the effect of chronic alcohol consumption on transcription rates of the genes SLC19A1 and SLC46A1 (encoding RFC and PCFT respectively) in these cells. Results of these investigations should provide valuable information regarding the mechanisms of folate uptake and regulation in pancreatic acinar cells and on the effect of chronic alcohol consumption on these parameters. Such kowledge may shed light into the pathogenesis of certain pancreatic diseases and may ultimately assist in the designing of effective strategies to optimize pancreatic acinar folate homeostasis.

描述（由申请人提供）：叶酸是一种必需维生素，对于 RNA/DNA 合成、某些氨基酸（包括同型半胱氨酸）的代谢和甲基化至关重要。在胰腺中，叶酸缺乏会导致消化酶分泌减少并增加患癌症的风险。胰腺细胞无法合成叶酸，只能从外源获取叶酸。由于叶酸摄取及其在胰腺腺泡细胞中的调节所涉及的细胞和分子机制尚不清楚，我们利用原代和培养（AR42J）大鼠胰腺腺泡细胞进行了初步研究，获得了表明叶酸摄取涉及载体介导机制的证据。我们发现这些细胞表达主要的叶酸转运蛋白、还原叶酸转运蛋白（RFC）和质子偶联叶酸转运蛋白（PCFT）。我们建议继续研究，以描述叶酸摄取过程的特异性、要求和动力学，并确定 RFC 和 PCFT 通过 shRNA 方法对载体介导的叶酸摄取总量的相对功能贡献。我们还建议检查胰腺叶酸摄取过程是否受到底物可用性（缺乏和过度补充）的适应性调节，以及它是否经历发育调节。我们的研究将扩展到人类情况，使用正常人胰腺组织样本检查 RNA（原位杂交）和蛋白质（免疫组织化学）水平上的 RFC 和 PCFT 表达模式。慢性酒精中毒是胰腺炎和全身叶酸缺乏的主要原因。尽管人们已经深入研究了长期饮酒对胰腺生理学的影响，但对其对叶酸转运的具体影响知之甚少。因此，在初步研究中，我们表明，与配对喂养的对照组相比，长期饮酒的大鼠会导致胰腺中 RFC 和 PCFT mRNA 水平显着降低。我们建议继续这些研究，并检查这些影响对胰腺腺泡细胞叶酸摄取动力学、叶酸载体蛋白表达水平以及细胞内叶酸含量的功能影响。我们还建议检查长期饮酒对这些细胞中基因 SLC19A1 和 SLC46A1（分别编码 RFC 和 PCFT）转录率的影响。这些研究的结果应该提供有关胰腺腺泡细胞叶酸摄取和调节机制以及长期饮酒对这些参数的影响的有价值的信息。这些知识可能有助于揭示某些胰腺疾病的发病机制，并最终有助于设计优化胰腺腺泡叶酸稳态的有效策略。