Induction of Apoptotic-like Cell Death in Trichomonas vaginalis by Metronidazole / 热带医学杂志

Objective Apoptosis or programmed cell death(PCD) has been studied extensively in multicellular organisms,however,very little is known about the molecular mechanisms by which apoptosis occurs in unicellular protozoan parasites.The aim of this study is to characterize the apoptosis or PCD of Trichomonas vaginalis induced by metronidazole (MTZ).Methods T. Vaginalis strain cultures were treated with various concentrations of MTZ and the number of viable cells were determined at different time intervals.The genomic DNA of MTZ treated T. Vaginalis was extracted and DNA fragmentation was analyzed.TUNEL assay was carried out to detect the endonuclease activity in T. Vaginalis after MTZ treatment.Flow cytometric analysis was used to analyse the phosphatidylserine (PS) exposure of T. Vaginalis.Results Metronidazole (MTZ) induced an apoptotic-like cell death in T. Vaginalis.This apoptotic-like cell death was demonstrated by cell shrinkage,phosphatidylserine exposure,and nuclear chromatin condensation.However, no oligonucleosmal DNA laddering was detected.Conclusion The regulatory pathway of apoptotic cell death in T. Vaginalis may be different from multicellular organisms.The determination of protozoan apoptotic pathways leading to cell death might ultimately allow the identification of new therapeutic targets.

Molecular Cloning and Characterization of a RRas Homologue Gene from Trichomonas vaginalis / 中国寄生虫学与寄生虫病杂志

Objective To clone and characterize a RRas-like gene from Trichomonas vaginalis for studying cellular signal transduction pathways in the organism. Methods A cDNA clone, which showed homology with RRas proteins of human being, was isolated and sequenced from a cDNA expression library of T. vaginalis. The genomic DNA corresponding to the cDNA sequence was amplified using PCR technique and sequenced. Sequence analysis was per-formed using BLASTP, RPS-BLAST and ClustalW programs. Phylogenetic tree was constructed and bootstrapped with 1 050 replicates using the software MEGA3. Results The cDNA sequence showed a length of 705 bp with an open reading frame of 615 bp. The deduced amino acid sequence from the open reading frame possesses 205 residuals. Sequencing of the PCR product of genomic DNA revealed that the genomic DNA sequence encompassing the putative 5′-ATG and 3′-stop codons was identical to the cDNA sequence. Sequence analysis demonstrated that this gene was most homologous to the RRas members of Homo sapiens and Mus musculus (both having 51% identity and 70% similarity), and the amino acid sequence contains highly conserved GTP-binding domains and a fully conserved effector domain of human RRas member. Phylogenetic analysis showed that TvRRas clustered with RAS oncoprotein branch and RRAS branch of human. Conclusion The encoding protein probably belongs to a RRas family of T. vaginalis.