Novel Insights into the Molecular Events Linking to Cell Death Induced by Tetracycline in the Amitochondriate Protozoan Trichomonas vaginalis

Abstract

f Trichomonas vaginalis colonizes the human urogenital tract and causes trichomoniasis, the most common nonviral sexually transmitted disease. Currently, 5-nitroimidazoles are the only recommended drugs for treating trichomoniasis. However, increased resistance of the parasite to 5-nitroimidazoles has emerged as a highly problematic public health issue. Hence, it is essential to identify alternative chemotherapeutic agents against refractory trichomoniasis. Tetracycline (TET) is a broad-spectrum antibiotic with activity against several protozoan parasites, but the mode of action of TET in parasites remains poorly understood. The in vitro effect of TET on the growth of T. vaginalis was examined, and the mode of cell death was verified by various apoptosis-related assays. Next-generation sequencing-based RNA sequencing (RNA-seq) was employed to elucidate the transcriptome of T. vaginalis in response to TET. We show that TET has a cytotoxic effect on both metronidazole (MTZ)-sensitive and -resistant T. vaginalis isolates, inducing some features resembling apoptosis. RNA-seq data reveal that TET significantly alters the transcriptome via activation of specific pathways, such as aminoacyl-tRNA synthetases and carbohydrate metabolism. Functional analyses demonstrate that TET disrupts the hydrogenosomal membrane potential and antioxidant system, which concomitantly elicits a metabolic shift toward glycolysis, suggesting that the hydrogenosomal function is impaired and triggers cell death. Collectively, we provide in vitro evidence that TET is a potential alternative therapeutic choice for treating MTZ-resistant T. vaginalis. The in-depth transcriptomic signatures in T. vaginalis upon TET treatment presented here will shed light on the signaling pathways linking to cell death in amitochondriate organisms.T richomoniasis is the most common nonviral sexually transmitted infection (STI) and is caused by the parasitic protozoan Trichomonas vaginalis, with more than 275 million cases reported annually worldwide (1). Infected women develop vaginitis, urethritis, and cervicitis, potentially leading to serious health outcomes, such as infertility, preterm delivery, low-birth-weight infants, susceptibility to herpesvirus and human papillomavirus infection, and cervical cancer (2). Trichomoniasis has also been considered a cofactor of human immunodeficiency virus transmission and lethal prostate cancer (3, 4). Currently, metronidazole (MTZ) and other 5-nitroimidazoles are the only recommended drugs for the treatment of trichomoniasis. However, it is estimated that approximately 5 to 10% of all clinical cases of trichomoniasis display resistance to the above-mentioned drugs (5, 6). The only option for treating MTZ-refractory trichomoniasis is to increase the dose of MTZ. However, the teratogenic effect of MTZ on animal models is well documented (7-9), and up to 12% of patients suffer from nausea (10). Hence, it is essential to identify alternative chemotherapeutic agents to combat MTZ-resistant T. vaginalis.Tetracyclines (TETs) ar...

Cell death offers exceptional cellular and molecular windows for pharmacological interventions in protozoan parasites

“…Comparative bioinformatics examinations in Trichomonas vagina and Giardia intestinalis have shown the existence of the nuclease, NM23 gene family homologs, which has been implicated in DNA fragmentation. The in-depth transcriptomic signatures in Tetracycline (TET) treated-T. vaginalis showed the involvement of hydrogenosome in programmed cell death [25]. In response to beta-Lapachone treatment and starvation, Giardia lamblia presented some autophagic-like (myelinic figures in large vacuoles and LC-3 staining) and apoptotic-like (Cell shrinkage, chromatin condensation, membrane blebbing and vacuolization) programmed cell death characteristics suggesting the possible role of mitosomes, a relic organelle in these processes [26].…”

DNA analysis of cattle parasitic protozoan Tritrichomonas foetus after photodynamic therapy

“…degradation of cytoplasmatic components, hydrogenosome shape changes, plasmatic membrane projections (blebs), flagellar internalization, release of lysosomal enzyme, cell shape changes, cytoplasm vacuolization, photosensitizing (PS) externalization, and caspase activation [4,25,[28][29][30][31][32][33][34][35]. In the present study, T. foetus treated with PDT associated with photosensitizer tetrasulfonated aluminium phthalocyanine (AlPcS 4 ) showed "ladder-patterned" DNA fragments, in conventional electrophoresis assay and longer tails in Comet Assay.…”

Cell Death after Photodynamic Therapy Treatment in Unicellular Protozoan Parasite Tritrichomonas foetus

“…Recent studies showed that alterations in lysosomes and hydrogenosomes were also observed in T. foetus after proteossome inhibitors treatment [24]. Other studies suggest that tetracycline disrupts hydrogenosomal function since it reduced the hydrogenosomal energy metabolism efficiency in T. vaginalis [25].…”

“…However, taxol and nocodazole induced the formation of multinucleated cells, abnormal distribution of the nuclear contents and, sometimes, nuclear fragmentation [22]. In recent studies utilizing tetracycline, it was observed DNA fragmentation in T. vaginalis [25].…”

“…During treatment with taxol cell size was increased, and giant, multinucleate and abnormal cells were observed, as well as the presence of zoids and membrane blebbing formation [22]. In T. vaginalis tetracycline-treatment the cell exhibited a round shape, a not disrupted plasma membrane, and a rough cell surface [25]. However, the parasites treated with mepoxomicin and bortezomib showed the appearance of wrinkled or rounded cells with externalized flagella, membrane blebbing, cell lysis, intense cytosolic and nuclear vacuolization, cytoplasmic disintegration and abnormal Golgi reduction [24].…”