Corticosteroids correct aberrant CFTR localization in the duct and regenerate acinar cells in autoimmune pancreatitis

Abstract

Background & aims: Corticosteroids are now widely accepted as a treatment for autoimmune pancreatitis (AIP). However, the molecular mechanism by which steroid treatment improves AIP remains largely unknown. The aim of this study was to elucidate cellular mechanisms by which corticosteroids improve both pancreatic exocrine function and histopathology in AIP.

Methods: Pancreatic exocrine function was evaluated by the secretin-stimulated function test and pancreatic biopsy specimens were processed for histologic analysis at the time of diagnosis and 3 months after initiation of steroid treatment. Expression and localization of proteins was assayed by immunohistochemistry. Analysis of immunoglobulin (Ig)G4-positive plasma cells was used to verify inflammation in AIP.

Results: The number of IgG4-positive plasma cells in pancreatic sections was decreased by steroid treatment, indicating reduced inflammation. Fluid, bicarbonate (HCO(3)(-)), and digestive enzyme secretions all were impaired in most patients with AIP. Corticosteroids improved both HCO(3)(-) and digestive enzyme secretion. A large fraction of the cystic fibrosis transmembrane conductance regulator (CFTR), which plays a central role in pancreatic duct HCO(3)(-) secretion, was mislocalized to the cytoplasm of duct cells before treatment. Corticosteroids corrected the localization of CFTR to the apical membrane, accounting for the improved HCO(3)(-) secretion. Steroid treatment resulted in regeneration of acinar cells, accounting for restored digestive enzyme secretion.

Conclusions: Corticosteroids reduce inflammation and restore both digestive enzyme and HCO(3)(-) secretion in patients with AIP by regenerating acinar cells and correcting CFTR localization in pancreatic duct cells. Mislocalization of CFTR may explain aberrant HCO(3)(-) secretion in other forms of pancreatitis.

Figure 1

(A) Pancreatic exocrine function in AIP was examined by the secretin test. Tests were performed at the time of diagnosis (0 months), 3 months, and 12 months after initiation of corticosteroids. Lower limits of normal are marked by dashed lines. Black lines indicate change of exocrine function in patients whose function was evaluated at 0 and 3 months after initiation of corticosteroids. Gray lines indicate changes of exocrine function in patients whose exocrine function was evaluated at 0, 3, and 12 months. (a) Total secreted volume for an hour (normal, ≥183 mL/h). (b) Maximum HCO₃⁻ concentration (mbc) (normal, ≥80 mEq/L). (c) Total amylase output for an hour (normal, ≥99,000 U/h). (B) Change in pancreatic exocrine functions caused by short-term treatment. (a) Secreted volume was not altered by steroids (n = 8, mean ± standard error of the mean; at 0 months, 123.5 ± 12.9 mL/h; at 3 months, 122.1 ± 12.7 mL/h). (b) MBC (n = 7; at 0 months, 53.3 ± 5.8 mEq/L; at 3 months, 72.5 ± 7.2 mEq/L), and (c) total amylase output (n = 8; at 0 months, 6138 ± 2107 U/h; at 3 months, 17,143 ± 2710 U/h) were improved significantly by treatment. (C) Change of pancreatic exocrine functions at 0, 3, and 12 months after initiation of corticosteroids (n = 3). (a) Secretory volume and (b) MBC were not altered by prolonging treatment up to 12 months. (c) Amylase output is improved by short-term treatment (at 0 months, 6683 ± 444 U/h; at 3 months, 17,200 ± 1968 U/h) and further by long-term treatment (at 12 months, 28,962 ± 4894 U/h).

Figure 2

Pancreatic sections were stained with H&E. (A) Normal subject. (B) AIP, surgically resected tissue. Masson's trichrome staining of (C) normal pancreas and (D) AIP. H&E staining of specimen (E) before and (F) 3 months after initiation of steroid treatment. (E and F) Sections were obtained from the same patient. Bars, 100 μm. Insets show images at a higher magnification.

Figure 3

Immunolocalization of AQP1 in the pancreas. (A) Control subject. (B) AIP, surgically resected. Biopsy specimen (C) before treatment and (D) after treatment. (C and D) Sections were obtained from the same patient. The pathologic changes were seen in all sections examined. Bars, 20 μm. Insets show images at a higher magnification.

Figure 4

Immunolocalization of CFTR in the pancreas. (A) Normal subject. (B) AIP, surgically resected. Biopsy specimen (C) before and (D) after treatment. (C and D) Sections are from the same patient. (E) Pancreatic sections obtained from normal subjects (n = 4), from AIP patients at 0 (n = 7) or 3 months (n = 7) treatment were scored at 0 (none), 1 (slight), 2 (moderate), and 3 (severe) for the degree of cytoplasmic staining of CFTR in pancreatic ducts. CFTR localization also was examined in the pancreas from the patients with (F) alcoholic, (G) obstructive, and (H) idiopathic chronic pancreatitis. Each panel represents 6 alcoholic, 2 obstructive, and 3 idiopathic cases of pancreatitis, respectively. Bars, 20 μm. Insets show images at a higher magnification.

Figure 5

Immunohistochemical staining for IgG4. (A) Marked IgG4-positive plasma cell infiltrates are present in tissue before treatment. (B) The number of IgG4-positive plasma cells was decreased significantly after treatment. (C) Pancreatic sections obtained from normal subjects (n = 4), and AIP patients at 0 (n = 7) or 3 months (n = 7) treatment were scored at 0 (none), 1 (slight), 2 (moderate), and 3 (severe) for extent of IgG4-positive plasma cell infiltration.

Figure 6

Immunoperoxidase labeling of CD133 (prominine-1) in the pancreas. (A) Normal subject. (B) Absence of acinar cells and massive fibrosis are evident before treatment. CD133 was detected in the apical membrane of residual pancreatic ducts. (C) CD133 and fibrosis after short-term steroid treatment. (D) CD133-positive cells are absent and acinar cell regeneration did not occur. Bars, 20 μm. Insets show images at a higher magnification.