单层培养豚鼠胃主细胞中胃蛋白酶原基因表达的调控机制

Regulatory mechanism of pepsinogen gene expression in monolayer cultured guinea pig gastric chief cells.

作者信息

Okayama N, Itoh M, Joh T, Miyamoto T, Takeuchi T, Moriyama A, Kato T

机构信息

First Department of Internal Medicine, Nagoya City University Medical School, Nagoya, Aichi, Japan.

出版信息

Dig Dis Sci. 1998 Dec;43(12):2744-9. doi: 10.1023/a:1026623932350.

DOI:10.1023/a:1026623932350

PMID:9881509

Abstract

We have investigated the effects of dibutyryl cAMP, forskolin, carbamylcholine chloride (carbachol), ionomycin, and 12-O-tetradecanoylphorbol-13-acetate (TPA) on the expression of guinea pig pepsinogen mRNA in monolayer cultured gastric chief cells. After exposure of the cells to each of these compounds for 4 to 24 hr, and at 48 hr after primary culture, total cellular RNA was isolated using acid guanidium-phenol-chloroform and then was reverse transcribed to cDNA. Obtained cDNA was amplified by polymerase chain reaction (PCR) using primers detecting guinea pig pepsinogen mRNA and human beta-actin mRNA as an internal standard. The PCR products were separated and quantified using capillary electrophoresis. Dibutyryl cAMP and forskolin significantly increased pepsinogen mRNA, but carbachol, ionomycin, and TPA failed to increase that. These findings suggested that pepsinogen gene expression was up-regulated by intracellular cAMP, but not by intracellular calcium or protein kinase C in guinea pig chief cells.

摘要

我们研究了二丁酰环磷腺苷（dibutyryl cAMP）、福斯高林（forskolin）、氯化氨甲酰胆碱（carbachol）、离子霉素（ionomycin）和十四酰佛波醇乙酯（12-O-tetradecanoylphorbol-13-acetate，TPA）对单层培养的豚鼠胃主细胞中胃蛋白酶原mRNA表达的影响。将细胞分别暴露于这些化合物4至24小时后，以及原代培养48小时后，使用酸性胍盐-苯酚-氯仿法分离总细胞RNA，然后将其逆转录为cDNA。使用检测豚鼠胃蛋白酶原mRNA的引物和作为内参的人β-肌动蛋白mRNA，通过聚合酶链反应（PCR）扩增获得的cDNA。使用毛细管电泳分离并定量PCR产物。二丁酰环磷腺苷和福斯高林显著增加胃蛋白酶原mRNA，但氯化氨甲酰胆碱、离子霉素和TPA未能增加其表达。这些发现表明，豚鼠胃主细胞中胃蛋白酶原基因表达受细胞内cAMP上调，但不受细胞内钙或蛋白激酶C上调。