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Cryptosporidium parvum infection of intestinal epithelium: morphologic and functional studies in an in vitro model
R B Adams1, R L Guerrant, S Zu
1Department of Internal Medicine, University of Virginia Health Sciences Center, Charlottesville 22908.
Abstract:
A monolayer of mature polarized colonic epithelial cells (T84) able to generate and maintain a barrier to macromolecular flow was used to study pathophysiologic events that occur on microvillus cell exposure to Cryptosporidium parvum. By 24-48 h, several life cycle forms were seen in parasitophorous vacuoles near the apical cell surface, along with a time- and oocyst dose-dependent reduction in epithelial barrier function. As few as 10(5) organisms constituted a successful infecting dose, and heat inactivation of organisms markedly reduced the monolayer barrier alteration. Horseradish peroxidase flux studies demonstrated a substantial increase in macromolecular permeability of the monolayer, and lactate dehydrogenase determinations indicated modest injury of the T84 epithelial cells on exposure to oocysts. Thus, disruption of the epithelial cell barrier, not just opening of transcellular channels for ion flow as reported previously, is responsible for the effects of C. parvum oocysts on intestinal epithelium.
Insights
Cryptosporidium parvum disrupts the intestinal epithelial barrier by damaging T84 cells. This pathogen causes increased macromolecular permeability, impacting gut barrier function.
Area of Science:
- Cell Biology
- Parasitology
- Gastroenterology
Background:
- The intestinal epithelium forms a critical barrier against pathogens.
- Cryptosporidium parvum is an opportunistic parasite causing significant diarrheal disease.
- Understanding the mechanisms of C. parvum-induced intestinal damage is crucial for developing effective treatments.
Purpose of the Study:
- To investigate the pathophysiological effects of Cryptosporidium parvum oocysts on a mature colonic epithelial cell monolayer.
- To elucidate the specific mechanisms by which C. parvum disrupts intestinal barrier function.
Main Methods:
- Utilized T84 colonic epithelial cell monolayers as a model system.
- Assessed epithelial barrier function by measuring macromolecular flux (horseradish peroxidase).
- Determined cell injury using lactate dehydrogenase release assays.
- Observed parasite development within epithelial cells using microscopy.
Main Results:
- Exposure to C. parvum oocysts led to a time- and dose-dependent reduction in epithelial barrier function.
- Increased macromolecular permeability was observed, indicating barrier disruption.
- Modest T84 cell injury was detected, evidenced by lactate dehydrogenase release.
- Heat inactivation of oocysts significantly reduced the observed barrier alterations.
Conclusions:
- Cryptosporidium parvum directly disrupts the integrity of the colonic epithelial barrier.
- The primary mechanism involves direct damage and disruption of the epithelial cell barrier, not solely ion channel opening.
- These findings highlight the importance of epithelial barrier integrity in intestinal health and C. parvum pathogenesis.