الفهرس 
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Abstract
The aim of the present study is to investigate the in vitro effect of iron and spermine on the growth kinetics, cytoadherence and cytotoxicity of T. vaginalis and to examine the in vitro effect of iron on MLC of metronidazole on T. vaginalis isolates.
In order to achieve this aim, vaginal washouts and urine samples were collected from 100 female patients attending the Early Cancer Detection Unit, and the out-patients clinics, Gynecology and Obstetrics Hospital, Faculty of Medicine, Ain Shams University. They were examined for the presence of T. vaginalis trophozoites by wet mount microscopic examination and in vitro culture on TYM media. Two different T. vaginalis isolates were maintained to conduct the study; maintenance was done by subculturing of T. vaginalis every 24-48 hours in new culture medium.
Growth kinetics was assessed by counting parasites grown on culture media with different conditions every 24 hours for 7 days. Parasites were grown on iron-rich culture medium supplemented with 250 µM FAS, iron-depleted culture medium supplemented with 100 µM 2,2-DP, TYM medium supplemented with 20 mM DAB for polyamine metabolism inhibition and polyamine metabolism restoration was done by harvesting DAB-treated parasites after 18-hrs growth, washed with sterile PBS and transferred to TYM-serum medium supplemented with 50 µM spermine at 37 ºC.
The cytotoxicity and cytoadherence levels of trichomonads, grown on culture media with different conditions, over Vero cell were assessed.
Aerobic and anaerobic metronidazole susceptibility assays were done to evaluate the effect of iron on MLC of metronidazole on T. vaginalis isolates. Susceptibility assays were done on T. vaginalis grown on normal and iron-rich media.
Concerning the effect of iron on the growth kinetics pattern of T. vaginalis isolates, trichomonads grown on iron rich media showed significant increase in parasites count which was obvious for both isolates. As regards generation time, isolate 1 showed no significant difference between trichomonads grown on iron-rich media and control untreated trichomonads, while isolate 2 showed significant increase in generation time of iron treated parasites in comparison to control. Regarding the effect of iron on cytoadherence and toxicity to Vero cells, both isolates showed different results. Isolate 1 revealed significant lower levels of cytotoxicity and increased level of adherence to Vero cells when compared to untreated organisms handled identically. While, isolate 2 showed significant higher levels of cytotoxicity with increased level of adherence to Vero cells compared to untreated parasites.
Regarding trichomonads grown on iron-depleted media, there were opposite results of both isolates. Isolate 1 showed a significant decrease in parasites count, while isolate 2, showed significant increase of parasites count. As regards generation time, both isolates showed no significant difference between trichomonads grown on iron-depleted media and control untreated trichomonads. Concerning the effect on cytoadherence and toxicity to Vero cells, both isolates showed different results. Isolate 1 showed significant higher levels of cytotoxicity and decreased level of adherence to Vero cells when compared to untreated parasites. While isolate 2 revealed lower levels of cytotoxicity and increased adherence percentage to Vero cells.
Regarding the effect of polyamine on the growth kinetics pattern of T. vaginalis isolates, trichomonads grown on polyamine depleted media, DAB treated, showed significant decrease in parasites count which was obvious for both isolates. As regards trichomonads grown on spermine restored media, isolate 2, showed a reversion of growth inhibition caused by DAB after addition of exogenous spermine. While isolate 1, cannot reverse the growth inhibition caused by DAB after addition of spermine. Both isolates showed a significant increase in generation time of both DAB treated and spermine restored trichomonads in comparison to untreated parasites.
Concerning the effect of polyamine on cytoadherence and toxicity to Vero cells, trichomonads grown on polyamine depleted media, showed different results with both isolates. Isolate 1 revealed enhanced adherence with significant higher levels of cytotoxicity to Vero cells when compared to untreated organisms handled identically. While, isolate 2 showed increased level of adherence and significant decrease level of cytotoxicity to Vero cells compared to untreated parasites. Regarding trichomonads grown on spermine restored media; there was no reversion of the enhanced adherence caused by DAB treatment by addition of spermine. Also both isolates showed significant lower levels of cytotoxicity to Vero cells when compared to untreated parasites.
Addition of iron decreased the MLC of metronidazole on T. vaginalis isolates. The aerobic MLC of metronidazole on FAS treated trichomonads of both isolates was 0.38 μg /ml while that on untreated T. vaginalis was 12.5 μg /ml. As regards anaerobic assay, the MLC of metronidazole on FAS treated parasites of isolate 1 was 0.097μg /ml, while that on untreated T. vaginalis was 3.12 μg /ml. Concerning isolate 2, MLC of metronidazole on FAS treated trichomonads was 0.19 μg /ml, while that on untreated T. vaginalis was 3.12 μg /ml.