Department of Molecular and Cell Biology, Boston University, Henry M Goldman School of Dental Medicine, Boston, Massachusetts 02118, USA.
Gut. 2010 Mar;59(3):365-72. doi: 10.1136/gut.2009.198903. Epub 2009 Nov 30.
Chronic pancreatitis is a progressive inflammatory disorder of the pancreas characterised by permanent destruction of acinar cells. Mutations in the chymotrypsinogen C (CTRC) gene have been linked to the development of chronic pancreatitis. The aim of the present study was to explore whether CTRC mutants induce endoplasmic reticulum (ER) stress in pancreatic acinar cells.
Dexamethasone-differentiated AR42J rat acinar cells and freshly isolated mouse acini were transfected with recombinant adenovirus carrying wild-type CTRC or the p.A73T pancreatitis-associated mutant. ER stress markers were assessed by reverse transcription-PCR and western blotting. Apoptosis was characterised by caspase-3/7 activity and the TUNEL assay.
Acinar cells transfected with the p.A73T mutant, but not those with wild-type CTRC, developed significant ER stress as judged by elevated mRNA and protein levels of the ER chaperone immunoglobulin-binding protein (BiP), increased splicing of the X-box binding protein-1 (XBP1) mRNA and marked induction of the transcription factor C/EBP-homologous protein (CHOP), a mediator of ER stress-associated apoptosis. Consistent with higher CHOP expression, AR42J cells expressing the p.A73T mutant became detached over time and showed considerably increased caspase-3/7 activity and TUNEL staining.
Pancreatitis-associated CTRC mutations can markedly increase the propensity of chymotrypsinogen C to elicit ER stress in pancreatic acinar cells. Thus, carriers of CTRC mutations may be at a higher risk of developing ER stress in the exocrine pancreas, which may contribute to parenchymal damage through acinar cell apoptosis.
慢性胰腺炎是一种胰腺的进行性炎症性疾病，其特征为腺泡细胞的永久性破坏。糜蛋白酶原 C（CTRC）基因的突变与慢性胰腺炎的发展有关。本研究旨在探讨 CTRC 突变体是否会在胰腺腺泡细胞中诱导内质网（ER）应激。
用携带野生型 CTRC 或 p.A73T 胰腺炎相关突变体的重组腺病毒转染地塞米松分化的 AR42J 大鼠腺泡细胞和新鲜分离的小鼠腺泡。通过逆转录-PCR 和 Western blot 评估 ER 应激标志物。通过 caspase-3/7 活性和 TUNEL 测定法来表征细胞凋亡。
与转染野生型 CTRC 的细胞相比，转染 p.A73T 突变体的腺泡细胞表现出明显的 ER 应激，其依据为内质网伴侣免疫球蛋白结合蛋白（BiP）的 mRNA 和蛋白水平升高、X 盒结合蛋白-1（XBP1）mRNA 的剪接增加以及转录因子 C/EBP 同源蛋白（CHOP）的显著诱导，CHOP 是 ER 应激相关细胞凋亡的介质。与更高的 CHOP 表达一致，表达 p.A73T 突变体的 AR42J 细胞随时间推移而脱落，并且 caspase-3/7 活性和 TUNEL 染色明显增加。
胰腺炎相关的 CTRC 突变可显著增加胰蛋白酶原 C 引发胰腺腺泡细胞 ER 应激的倾向。因此，携带 CTRC 突变的个体可能在外分泌胰腺中更容易发生 ER 应激，这可能通过腺泡细胞凋亡导致实质损伤。
