Pancreatic and biliary secretion are both altered in cystic fibrosis pigs

Abstract

The pancreas, liver, and gallbladder are commonly involved in cystic fibrosis (CF), and acidic, dehydrated, and protein-rich secretions are characteristic findings. Pancreatic function studies in humans have been done by sampling the jejunal fluid. However, it has been difficult to separately study the function of pancreatic and biliary systems in humans with CF, because jejunal fluid contains a mixture of bile and pancreatic fluids. In contrast, pancreatic and biliary ducts open separately into the porcine intestine; therefore, biliary and pancreatic fluid can be individually analyzed in CF pigs. We studied newborn wild-type (WT) and CF pigs and found that CFTR was localized to the pancreatic ducts. We collected bile and pancreatic fluid and analyzed pancreatic enzymes with activity assays and immunoblot. Pancreatic enzyme expression was significantly decreased in CF compared with WT pigs. The volume and pH of pancreatic fluid were significantly lower and protein concentration was >5-fold higher in CF pigs. Secretin stimulation increased pancreatic fluid volume and pH in WT, but not CF, pigs. Baseline bile volume did not differ between WT and CF pigs, but volume did not increase in response to secretin in CF pigs. Bile pH was lower and protein concentration was twofold higher in CF pigs. These results indicate that pancreatic and biliary secretions are altered in CF pigs. Abnormal pancreatic and biliary secretion in CF may have important implications in disease pathogenesis.

Fig. 1.

Pancreatic and biliary duct anatomy in pigs: biliary and pancreatic duct openings to the duodenum in a newborn pig. S, stomach; L, liver; GB, gallbladder; P, pancreas; Py, pylorus; D, duodenum. White arrow points to common bile duct; black arrow points to pancreatic duct.

Fig. 2.

Pancreatic enzyme expression in wild-type (WT) pigs and pigs with cystic fibrosis (CF). Pancreatic amylase (A), lipase (B), and trypsin (C) activities (n = 4 each) and pancreatic elastase (PE) and chymotrypsinogen (Chy) expression (D) (n = 3 each) were severely diminished in CF compared with WT pigs. *P < 0.01 vs. WT.

Fig. 3.

Immunocytochemical localization of CFTR in pancreas from newborn pigs shown as stacks of confocal and differential interference contrast microscopic images. Scale bars, 10 μm. A: WT pancreas. CFTR is green, β-catenin is red, and nuclei are blue (4′,6-diamidino-2-phenylindole). B: CFTR^−/− pancreas. Arrows point to intercalated ducts.

Fig. 4.

Pancreatic fluid composition in WT and CF pigs. A: pancreatic fluid volume increased after secretin in WT, but not CF, pigs (n = 4 WT and 5 CF). *P < 0.01. After secretin, pancreatic fluid volume was lower in CF than WT pigs. **P < 0.01. B: pancreatic fluid pH was lower in CF than WT pigs at baseline and after secretin (n = 4 WT and 5 CF). *P < 0.01. C: protein concentration was >5 fold higher in pancreatic secretion from CF than WT pigs (n = 3 WT and 2 CF).

Fig. 5.

Bile composition in WT and CF pigs. A: at baseline, bile volume was not significantly different between WT and CF pigs. Bile volume increased after secretin in WT, but not CF, pigs (n = 4 WT and 5 CF). *P < 0.05. B: bile pH did not differ significantly by genotype before and after secretin (n = 4 WT and 5 CF). C: volume of bile in CF pigs was not sufficient to measure amount of protein at baseline. Protein concentration after secretin was 2-fold higher in CF than WT pig bile (n = 3 WT and 2 CF).

Fig. 6.

Gallbladder bile pH. Gallbladder bile pH in newborn pigs at the time of euthanasia was measured using a needle-type fiber-optic pH meter. Gallbladder bile pH was lower in CF than WT pigs (n = 8 WT and 12 CF). *P < 0.0001 vs. WT.