Background: Small intestinal epithelial cells (IEC) show apoptosis in physiological turnover of cells and in certain inflammatory diseases.
Aims: To investigate the role of caspases in the progression of IEC apoptosis in vivo.
Methods: IEC were separated along the villus-crypt axis from the jejunum of normal and Nippostrongylus brasiliensis infected rats at 4°C. Caspases were examined by a fluorometric assay method, histochemistry, and immunoblotting.
Results: Villus cell rich IEC from normal rats exhibited a high level of caspase-3-like activity whereas activities of caspase-1, -8, and -9 were negligible. Immunoblotting analysis of villus cell rich IEC revealed partial cleavage of procaspase-3 into a 17 kDa molecule as well as cleavage of a caspase-3 substrate, poly(ADP-ribose) polymerase (PARP), whereas in crypt cell rich IEC, caspase-3 cleavage was less significant. Caspase-3 activity was also observed histochemically in villus epithelium on frozen sections of the normal small intestine. IEC prepared at 4°C did not reveal nuclear degradation whereas subsequent incubation in a suspension at 37°C induced intense nuclear degradation within one hour in accordance with increases in active caspase-3. This apoptosis was partially suppressed by the caspase inhibitor Z-VAD-fmk. Nematode infected animals showed villus atrophy together with significant increases in levels of caspase-3 in IEC but not of caspase-1, -8, or -9.
Conclusion: Caspase-3 may have an important role in the physiological replacement of IEC as well as in progression of IEC apoptosis induced by nematode infection.
- small intestine
- IEC, intestinal epithelial cells
- PARP, poly(ADP-ribose) polymerase
- DMEM, Dulbecco's modified Eagle's medium
- FCS, fetal calf serum
- BrdU, bromodeoxyuridine
- PCNA, proliferating cell nuclear antigen
- ALP, alkaline phosphatase
- VCU, villus-crypt units
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