Glucose-restriction increasesTrichomonas vaginaliscellular damage towards HeLa cells and proteolytic activity of cysteine proteinases (CPs), such as TvCP2

Abstract

Trichomonas vaginalis induces cellular damage to the host cells (cytotoxicity) through the proteolytic activity of multiple proteinases of the cysteine type (CPs). Some CPs are modulated by environmental factors such as iron, zinc, polyamines, etc. Thus, the goal of this study was to assess the effect of glucose on T. vaginalis cytotoxicity, proteolytic activity and the particular role of TvCP2 (TVAG_057000) during cellular damage. Cytotoxicity assays showed that glucose-restriction (GR) promotes the highest HeLa cell monolayers destruction (~95%) by trichomonads compared to those grown under high glucose (~44%) condition. Zymography and Western blot using different primary antibodies showed that GR increased the proteolytic activity, amount and secretion of certain CPs, including TvCP2. We further characterized the effect of glucose on TvCP2. TvCP2 increases in GR, localized in vesicles close to the plasma membrane and on the surface of T. vaginalis. Furthermore, pretreatment of GR-trichomonads with an anti-TvCP2r polyclonal antibody specifically reduced the levels of cytotoxicity and apoptosis induction to HeLa cells in a concentration-dependent manner. In conclusion, our data show that GR, as a nutritional stress condition, promotes trichomonal cytotoxicity to the host cells, increases trichomonad proteolytic activity and amount of CPs, such as TvCP2 involved in cellular damage.

The effect of iron on Trichomonas vaginalis TvCP2: a cysteine proteinase found in vaginal secretions of trichomoniasis patients

“…For parasites grown under high-iron (HI) or IR condition (250 or 0 μ m iron, respectively), the culture medium was supplemented with 250 μ m ferrous ammonium sulphate or 150 μ m 2-2′-dipyridyl (an iron-chelating agent; Sigma-Aldrich, Co.), respectively (Gorrell, 1985; Alvarez-Sánchez et al ., 2007). To obtain confluent HeLa cell monolayers, cells were grown in Dulbecco's modified Eagle medium (DMEM) (Gibco Lab, Grand Island, NY, USA) supplemented with 10% HIBS at 37 °C for 48 h in a 5% CO 2 atmosphere (Alvarez-Sánchez et al ., 2000; Miranda-Ozuna et al ., 2019).…”

“…The expression of some CP genes is differentially modulated by iron concentrations (Arroyo et al ., 2015). Moreover, CPs are considered to be virulence factors (Hernández et al ., 2014), participating in haemolysis (Dailey et al ., 1990; Fiori et al ., 1993, 1996, 1999; Lubick and Burgess, 2004; Cárdenas-Guerra et al ., 2013, 2015), complement resistance (Alderete et al ., 1995), cytoadherence (Arroyo and Alderete, 1989, 1995; Mendoza-López et al ., 2000; Rendón-Gandarilla et al ., 2013), cytotoxicity (Alvarez-Sánchez et al ., 2000, 2007; Ramón-Luing et al ., 2011; Carvajal-Gámez et al ., 2014; Arroyo et al ., 2015) and apoptosis induction in HeLa and human vaginal epithelial cells (Sommer et al ., 2005; Kummer et al ., 2008; Miranda-Ozuna et al ., 2019).…”

“…The tvcp2 (TVAG_057000) gene (945-bp) was identified as one of the cDNA clones in an expression library together with three other genes that encode similar trichomonad cathepsin L-like CPs (Mallinson et al ., 1994), which were also found in in vitro secretion products (SPs) involved in the induction of apoptosis in vaginal epithelial cells (Sommer et al ., 2005). TvCP2 is downregulated by glucose and is involved in cellular damage (Miranda-Ozuna et al ., 2019). Moreover, several immunogenic proteinases from T. vaginalis (immunoproteome) were identified in an immunoproteomic analysis, combined with mass spectrometry (MS) and the use of patient sera in 2-DE-Western-blotting(2-DE-WB) assays (Ramón-Luing et al ., 2010).…”

Mucosal microbial parasites/symbionts in health and disease: an integrative overview

“…These different aspects associated with T. vaginalis infections illustrates the intricate associations of the parasite with bacterial ( Mycoplasma ) and viral (TVV) endosymbionts, the bacterial members of the UGT microbiota and how these interactions can influence the parasite pathobiology including increasing human infecting virus transmission rates. These considerations will be important to complement more traditional investigations focusing on the study of specific aspects of host–parasite interactions, such as the potential role of environmental glucose concentration variation (Miranda-Ozuna et al ., 2019) and cell surface and secreted factors such as exosomes (Mercer and Johnson, 2018), in modulating the virulence of the parasite. These examples illustrate dramatically the importance to investigate host–MMPS–microbiota–virus interactions in an integrative manner to develop more refined diagnostics and novel prophylactic and therapeutic strategies to eventually promote reproductive and sexual health more efficiently.…”

“…Several papers associated with this Special Issue are derived from talks that were delivered at the EMBO Conference ‘Anaerobic protists: Integrating parasitology with mucosal microbiota and immunology’ () (Labruyere et al ., 2017; Leitsch, 2017; Dessi et al ., 2019; Miranda-Ozuna et al ., 2019; Stensvold, 2019). These are complemented by articles providing broader perspectives on the study of the MMPS (Chabra et al , 2019; Liu et al , 2019; Midlej et al , 2019; Bartley et al ., 2018; Deere et al ., 2018; van Gestel et al ., 2018; Vargas Rigo et al ., 2018; Chihi et al ., 2019; Rush et al ., 2019).…”