Chymotrypsin in pancreatitis

摘要 这项资助的主要目的是使用基因工程小鼠模型来确定 人糜蛋白酶的遗传改变增加慢性胰腺炎的风险。以前的研究 证明了糜蛋白酶在人类和小鼠模型中对胰腺炎的保护作用。 在这个提议中，首要的假设是，保护性糜蛋白酶功能的丧失导致 胰腺炎的背景下，额外的遗传或环境风险因素，促进 胰内胰蛋白酶激活。为了证明这一概念，CTRB 1缺失的小鼠品系（Ctrb 1-del）将 与携带各种胰腺炎风险等位基因的小鼠杂交，或暴露于酒精饮食。 尽管CTRB 1缺失的小鼠在诱发疾病时表现出更高的胰腺炎反应， 实验通过雨蛙肽刺激;他们不自然发展胰腺炎。引入 胰凝乳蛋白酶缺乏症的其他危险因素预计会导致自发性 胰腺炎的发生和进展。或者，小鼠将表现出对雨蛙肽的加性易感性- 诱发胰腺炎。将研究以下具体目标。(1)胰凝乳蛋白酶缺乏 胰蛋白酶原突变导致胰腺炎在这个目标中，假设是， 胰凝乳蛋白酶和携带突变的小鼠阳离子胰蛋白酶原（亚型T7）将发展 自发性急性胰腺炎将进展为慢性胰腺炎。或者，小鼠将表现出 增加对雨蛙肽诱导的胰腺炎的易感性。通过组合调整基因剂量 杂合子和纯合子等位基因的突变应导致发病年龄和/或严重程度的变化。（二） 胰凝乳蛋白酶和胰蛋白酶抑制剂联合不足的胰腺炎。为此，Ctrb 1-del小鼠 将与SPINK 1缺失株（Spink 1-KO）或携带T7胰蛋白酶原的小鼠杂交 具有p.G199R突变的等位基因，其传递了降解特性（p.G199R突变的功能直向同源物）。 人中胰蛋白酶）。假设胰凝乳蛋白酶和胰蛋白酶联合缺乏的小鼠 抑制剂将发展为自发性胰腺炎或将表现出雨蛙肽严重性增加， 诱发胰腺炎。(3)胰凝乳蛋白酶缺乏的酒精性胰腺炎。在这一目标下， 胰凝乳蛋白酶缺陷的小鼠将被喂食Lieber-DeCarli酒精液体饮食。假设是， 胰凝乳蛋白酶缺乏的背景下，酒精会引起自发性胰腺炎， 胰蛋白酶原的选择性上调。这些目标的成功实现将提供决定性的 有证据表明，人类胰凝乳蛋白酶基因的改变促进了胰腺炎的发展， 与其他遗传和环境因素相互作用，增加胰内胰蛋白酶活性。

ABSTRACT The main objective of this grant is to use genetically engineered mouse models to determine how genetic changes in human chymotrypsin increase risk for chronic pancreatitis. Previous studies demonstrated the protective role of chymotrypsin against pancreatitis in humans and in mouse models. In this proposal, the overarching hypothesis is that loss of protective chymotrypsin function leads to pancreatitis in the context of additional genetic or environmental risk factors that facilitate intrapancreatic trypsin activation. To prove this notion, a CTRB1-deleted mouse strain (Ctrb1-del) will be crossed with mice carrying various pancreatitis risk alleles or will be exposed to an alcohol diet. Although CTRB1-deleted mice show heightened pancreatitis responses when disease is induced experimentally by cerulein stimulation; they do not develop pancreatitis naturally. Introduction of additional risk factors in the setting of chymotrypsin deficiency is predicted to result in the spontaneous onset and progression of pancreatitis. Alternatively, mice will exhibit additive susceptibility to cerulein- induced pancreatitis. The following specific aims will be studied. (1) Chymotrypsin deficiency synergizes with trypsinogen mutations to cause pancreatitis. In this aim, the hypothesis is that mice deficient in chymotrypsin and carrying mutations in mouse cationic trypsinogen (isoform T7) will develop spontaneous acute pancreatitis that will progress to chronic pancreatitis. Alternatively, mice will exhibit increased susceptibility to cerulein-induced pancreatitis. Adjusting gene dosage through combinations of heterozygous and homozygous alleles should result in variations in age of onset and/or severity. (2) Combined insufficiency of chymotrypsin and trypsin inhibitor in pancreatitis. In this aim, Ctrb1-del mice will be crossed either with a SPINK1-deleted strain (Spink1-KO) or with mice carrying a T7 trypsinogen allele with the p.G199R mutation, which conveys inhibitor-degrading properties (functional ortholog of human mesotrypsin). The hypothesis is that mice with combined deficiency in chymotrypsin and trypsin inhibitor will develop either spontaneous pancreatitis or will exhibit increased severity of cerulein- induced pancreatitis. (3) Alcoholic pancreatitis in the setting of chymotrypsin deficiency. In this aim, chymotrypsin-deficient mice will be fed the Lieber-DeCarli alcohol liquid diet. The hypothesis is that in the background of chymotrypsin-deficiency alcohol will cause spontaneous pancreatitis through selective upregulation of trypsinogen. Successful completion of these aims will offer conclusive evidence that alterations in chymotrypsin genes in humans promote the development of pancreatitis by interacting with other genetic and environmental factors that increase intrapancreatic trypsin activity.