Development and Application of a Mouse Intestinal Loop Model To Study the In Vivo Action of Clostridium perfringens Enterotoxin

Abstract

Clostridium perfringens enterotoxin (CPE) is responsible for causing the gastrointestinal symptoms of C. perfringens type A food poisoning, the second most commonly identified bacterial food-borne illness in the United States. CPE is produced by sporulating C. perfringens cells in the small intestinal lumen, where it then causes epithelial cell damage and villous blunting that leads to diarrhea and cramping. Those effects are typically self-limiting; however, severe outbreaks of this food poisoning, particularly two occurring in psychiatric institutions, have involved deaths. Since animal models are currently limited for the study of the CPE action, a mouse ligated intestinal loop model was developed. With this model, significant lethality was observed after 2 h in loops receiving an inoculum of 100 or 200 g of CPE but not using a 50-g toxin inoculum. A correlation was noted between the overall intestinal histological damage and lethality in mice. Serum analysis revealed a dose-dependent increase in serum CPE and potassium levels. CPE binding to the liver and kidney was detected, along with elevated levels of potassium in the serum. These data suggest that CPE can be absorbed from the intestine into the circulation, followed by the binding of the toxin to internal organs to induce potassium leakage, which can cause death. Finally, CPE pore complexes similar to those formed in tissue culture cells were detected in the intestine and liver, suggesting that (i) CPE actions are similar in vivo and in vitro and (ii) CPE-induced potassium release into blood may result from CPE pore formation in internal organs such as the liver.

Clostridium perfringens Type A Enterotoxin Damages the Rabbit Colon

“…Therefore, damage from prolonged contact between the colon and CPE may have facilitated entry of the enterotoxin into the bloodstreams of these people, so it could then damage their internal organs and contribute to death. Similar enterotoxemia was experimentally demonstrated recently in mice intestinally challenged with CPE (29).…”

“…6A). Similar to the CH-1-like CPE complex formed in Caco-2 cells (13,28) or mouse intestine (29), the CPE large complex formed in colonic tissues migrated on SDS-containing 4% polyacrylamide gels with an apparent size of ϳ155 kDa. A similarly migrating large complex was also detected in CPE-treated rabbit small intestine (Fig.…”

“…Despite its true size of ϳ450 kDa, the CH-1 complex migrates anomalously in SDS-PAGE as a smeary ϳ155-kDa species on the SDS-containing 4% polyacrylamide gels necessary to analyze this large protein species (13). Previous studies had demonstrated that CPE binds and forms a similarly migrating CH-1-like large complex in mouse internal organs, including the small intestine (29). Therefore, the current study analyzed whether CPE also binds and forms a large CPE complex in the rabbit colon.…”

“…(A) CPE Western blot analyses of homogenates from colonic or small intestinal loops challenged for 6 h with 50 or 100 g/ml of CPE. In both the small intestine and colon, CPE bound and formed a large complex that migrates as a 155-kDa CPE species, similar to the CH-1-like CPE complex formed in mouse small intestine or in Caco-2 cells (25,29). (B) Claudin-3 (upper panel) or claudin-4 (lower panel) Western blot analyses of homogenates from small intestines and colons from four control rabbits.…”

Toxin Plasmids of Clostridium perfringens

“…It is thought that this lethality results when the medication reduces intestinal motility and interferes with CPEinduced diarrhea, thus prolonging contact between CPE and the intestinal mucosa. Based upon animal model studies (184), this longer presence of CPE in the intestines could facilitate absorption of the toxin into the circulation to cause a lethal enterotoxemia.…”