Protective effect of caspase inhibitor on intestinal integrity in experimental severe acute pancreatitis.
OBJECTIVE
Endotoxin/bacterial translocation (E/BT) plays an important role in systemic complications in severe acute pancreatitis (SAP). The breakdown of intestinal integrity is considered to be implicated in E/BT. We recently demonstrated that accelerated apoptosis of intestinal mucosa may have a part in E/BT. On the other hand, caspase is believed to play a central role in apoptosis. The aim of this study was to investigate the efficacy of caspase inhibitor on intestinal integrity and E/BT in SAP.


METHODS
SAP was induced by retrograde injection of 3% sodium deoxycholate into the biliopancreatic ducts in rats. At the same time, polycaspase inhibitor (Z-VAD-fmk) was administered intraperitoneally. Caspase activation in the intestine was evaluated by immunohistochemical staining and Western blotting. Apoptosis of intestinal mucosa was detected by TdT-mediated dUTP-biotin nick end labeling staining and DNA fragmentation enzyme-linked immunosolvent assay. Intestinal permeability was assayed ex vivo by measuring the leaked amount of FITC-dextran. Blood endotoxin level, bacterial culture of the ascites and mesenteric lymph nodes, and 24-h mortality rate were evaluated.


RESULTS
Immunoreactivities for activated caspase-10, -9, and -3 were increased 2 h after induction of SAP. Apoptosis and permeability of ileum were significantly increased 6 h after induction of SAP. Caspase inhibitor significantly improved the increasing apoptosis and permeability. It did not prevent the bacterial translocation but improved the disorder of intestinal mucosa and elevation of blood endotoxin 18 h after induction of SAP. Moreover, caspase treatment significantly improved the 24-h mortality rate. Z-VAD-fmk indeed inhibited the caspase-3 activation in intestinal mucosa of SAP.


CONCLUSIONS
These results suggest that caspase activation has a key role in the accelerated apoptosis of intestinal epithelial cells in SAP and that breakdown of intestinal mucosa via accelerated apoptosis causes the increase in intestinal permeability following endotoxin translocation in SAP.
Yasuda
Takeo
T
Department of Gastroenterological Surgery, Kobe University Graduate School of Medical Sciences, Kobe, Japan. yasudatk@med.kobe-u.ac.jp
Takeyama
Yoshifumi
Y
Ueda
Takashi
T
Shinzeki
Makoto
M
Kishi
Shinji
S
Sawa
Hidehiro
H
Nakajima
Takahiro
T
Kuroda
Yoshikazu
Y
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology*; D000681:Amylases / Q000097:blood; D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D001201:Ascites / Q000188:drug therapy / Q000473:pathology; D018988:Bacterial Translocation / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004731:Endotoxins / Q000097:blood; D004789:Enzyme Activation / Q000187:drug effects; D007408:Intestinal Absorption / Q000187:drug effects; D007421:Intestine, Small / Q000201:enzymology / Q000473:pathology*; D008198:Lymph Nodes / Q000473:pathology; D008297:Male; D010179:Pancreas / Q000473:pathology; D019283:Pancreatitis, Acute Necrotizing / Q000188:drug therapy* / Q000473:pathology; D051381:Rats; D017208:Rats, Wistar; D012720:Severity of Illness Index; D015996:Survival Rate
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D004731:Endotoxins; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; D000681:Amylases; C505410:Casp3 protein, rat; D053148:Caspase 3