| Autor: |
Singh, B. N., Lucas, J. J., Hayes, G. R., Kumar, Ish, Beach, D. H., Frajblat, Marcel, Gilbert, R. O., Sommer, U., Costello, C. E. |
| Zdroj: |
Infection and Immunity; July 2004, Vol. 72 Issue: 7 p4151-4158, 8p |
| Abstrakt: |
ABSTRACTTritrichomonas foetusis a serious veterinary pathogen, causing bovine trichomoniasis, a sexually transmitted disease leading to infertility and abortion. T. foetusinfects the mucosal surfaces of the reproductive tract. Infection with T. foetusleads to apoptotic cell death of bovine vaginal epithelial cells (BVECs) in culture. An affinity-purified cysteine protease (CP) fraction yielding on sodium dodecyl sulfate-polyacrylamide gel electrophoresis a single band with an apparent molecular mass of 30 kDa (CP30) also induces BVEC apoptosis. Treatment of CP30 with the protease inhibitors TLCK (Nα-p-tosyl-l-lysine chloromethyl ketone) and E-64 [l-trans-epoxysuccinyl-leucylamide-(4-guanido)-butane] greatly reduces induction of BVEC apoptosis. Matrix-assisted laser desorption ionization-time-of-flight MALDI-TOF mass spectrometry analysis of CP30 reveals a single peak with a molecular mass of 23.7 kDa. Mass spectral peptide sequence analysis of proteolytically digested CP30 reveals homologies to a previously reported cDNA clone, CP8 (D. J. Mallinson, J. Livingstone, K. M. Appleton, S. J. Lees, G. H. Coombs, and M. J. North, Microbiology 141:3077-3085, 1995). Induction of apoptosis is highly species specific, since the related human parasite Trichomonas vaginalisand associated purified CPs did not induce BVEC death. Fluorescence microscopy along with the Cell Death Detection ELISAPLUSassay and flow cytometry analyses were used to detect apoptotic nuclear condensation, DNA fragmentation, and changes in plasma membrane asymmetry in host cells undergoing apoptosis in response to T. foetusinfection or incubation with CP30. Additionally, the activation of caspase-3 and inhibition of cell death by caspase inhibitors indicates that caspases are involved in BVEC apoptosis. These results imply that apoptosis is involved in the pathogenesis of T. foetusinfection in vivo, which may have important implications for therapeutic interference with host cell death that could alter the course of the pathology in vivo. |
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