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Nitric oxide maintains cell survival of Trichomonas vaginalis upon iron depletion.

Cheng WH, Huang KY, Huang PJ, Hsu JH, Fang YK, Chiu CH, Tang P - Parasit Vectors (2015)

Bottom Line: The free radical signaling molecules reactive oxygen species (ROS) and reactive nitrogen species (RNS) have been proven to participate in iron deficiency in eukaryotes.We found that the inhibition of proteasome activity shortened the survival of iron-deficient cells compared with untreated iron-deficient cells.Surprisingly, the addition of arginine restored both NO level and the survival of proteasome-inhibited cells, suggesting that proteasome-derived NO is crucial for cell survival under iron-limited conditions.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kweishan, Taoyuan, Taiwan. m9701203@stmail.cgu.edu.tw.

ABSTRACT

Background: Iron plays a pivotal role in the pathogenesis of Trichomonas vaginalis, the causative agent of highly prevalent human trichomoniasis. T. vaginalis resides in the vaginal region, where the iron concentration is constantly changing. Hence, T. vaginalis must adapt to variations in iron availability to establish and maintain an infection. The free radical signaling molecules reactive oxygen species (ROS) and reactive nitrogen species (RNS) have been proven to participate in iron deficiency in eukaryotes. However, little is known about the roles of these molecules in iron-deficient T. vaginalis.

Methods: T. vaginalis cultured in iron-rich and -deficient conditions were collected for all experiments in this study. Next generation RNA sequencing was conducted to investigate the impact of iron on transcriptome of T. vaginalis. The cell viabilities were monitored after the trophozoites treated with the inhibitors of nitric oxide (NO) synthase (L-NG-monomethyl arginine, L-NMMA) and proteasome (MG132). Hydrogenosomal membrane potential was measured using JC-1 staining.

Results: We demonstrated that NO rather than ROS accumulates in iron-deficient T. vaginalis. The level of NO was blocked by MG132 and L-NMMA, indicating that NO production is through a proteasome and arginine dependent pathway. We found that the inhibition of proteasome activity shortened the survival of iron-deficient cells compared with untreated iron-deficient cells. Surprisingly, the addition of arginine restored both NO level and the survival of proteasome-inhibited cells, suggesting that proteasome-derived NO is crucial for cell survival under iron-limited conditions. Additionally, NO maintains the hydrogenosomal membrane potential, a determinant for cell survival, emphasizing the cytoprotective effect of NO on iron-deficient T. vaginalis. Collectively, we determined that NO produced by the proteasome prolonged the survival of iron-deficient T. vaginalis via maintenance of the hydrogenosomal functions.

Conclusion: The findings in this study provide a novel role of NO in adaptation to iron-deficient stress in T. vaginalis and shed light on a potential therapeutic strategy for trichomoniasis.

No MeSH data available.


Related in: MedlinePlus

NO accumulated in iron-deficient T. vaginalis.a Antioxidant capacity of iron-rich (IR) and -deficient (ID) cells representing the reducing power (Cu++ to Cu+) of each lysate. ROS (CM-DCF DA) b and NO (CM-DAF FM) c were examined. IR, iron rich (80 μM FAC); ID, iron deficient (180 μM DIP). The data are presented as the mean ± SD of three independent experiments. *** p < 0.001, compared with the IR group
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Fig2: NO accumulated in iron-deficient T. vaginalis.a Antioxidant capacity of iron-rich (IR) and -deficient (ID) cells representing the reducing power (Cu++ to Cu+) of each lysate. ROS (CM-DCF DA) b and NO (CM-DAF FM) c were examined. IR, iron rich (80 μM FAC); ID, iron deficient (180 μM DIP). The data are presented as the mean ± SD of three independent experiments. *** p < 0.001, compared with the IR group

Mentions: Previous studies have indicated that iron-deficient cells exhibit up- and down-regulation of thiol- and iron-dependent antioxidant defense systems, respectively [15, 21]. We confirmed the expression patterns of the iron-dependent antioxidants SOD (TVAG_039980, TVAG_120340) and rubrerythrin (TVAG_064490, TVAG_275660) and the thiol-dependent thioredoxin peroxidase (TVAG_114310, TVAG_455310) in cells cultured under iron-rich and -deficient conditions by using quantitative RT-PCR. The results revealed a trend similar to previous studies (Additional file 4). Furthermore, we examined the cellular reducing power in iron-rich and -deficient cells by measuring the amount of copper reduction (Cu++ to Cu+), which is used as a general indicator for antioxidant capacity. As shown in Fig. 2a, there is a significant increase in copper reduction in the iron-deficient group compared with iron-rich group. Together with quantitative RT-PCR analysis of multiple antioxidant genes, these data confirm that the antioxidative response of iron-deficient cells is stronger than that of iron-rich cells.Fig. 2


Nitric oxide maintains cell survival of Trichomonas vaginalis upon iron depletion.

Cheng WH, Huang KY, Huang PJ, Hsu JH, Fang YK, Chiu CH, Tang P - Parasit Vectors (2015)

NO accumulated in iron-deficient T. vaginalis.a Antioxidant capacity of iron-rich (IR) and -deficient (ID) cells representing the reducing power (Cu++ to Cu+) of each lysate. ROS (CM-DCF DA) b and NO (CM-DAF FM) c were examined. IR, iron rich (80 μM FAC); ID, iron deficient (180 μM DIP). The data are presented as the mean ± SD of three independent experiments. *** p < 0.001, compared with the IR group
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4513698&req=5

Fig2: NO accumulated in iron-deficient T. vaginalis.a Antioxidant capacity of iron-rich (IR) and -deficient (ID) cells representing the reducing power (Cu++ to Cu+) of each lysate. ROS (CM-DCF DA) b and NO (CM-DAF FM) c were examined. IR, iron rich (80 μM FAC); ID, iron deficient (180 μM DIP). The data are presented as the mean ± SD of three independent experiments. *** p < 0.001, compared with the IR group
Mentions: Previous studies have indicated that iron-deficient cells exhibit up- and down-regulation of thiol- and iron-dependent antioxidant defense systems, respectively [15, 21]. We confirmed the expression patterns of the iron-dependent antioxidants SOD (TVAG_039980, TVAG_120340) and rubrerythrin (TVAG_064490, TVAG_275660) and the thiol-dependent thioredoxin peroxidase (TVAG_114310, TVAG_455310) in cells cultured under iron-rich and -deficient conditions by using quantitative RT-PCR. The results revealed a trend similar to previous studies (Additional file 4). Furthermore, we examined the cellular reducing power in iron-rich and -deficient cells by measuring the amount of copper reduction (Cu++ to Cu+), which is used as a general indicator for antioxidant capacity. As shown in Fig. 2a, there is a significant increase in copper reduction in the iron-deficient group compared with iron-rich group. Together with quantitative RT-PCR analysis of multiple antioxidant genes, these data confirm that the antioxidative response of iron-deficient cells is stronger than that of iron-rich cells.Fig. 2

Bottom Line: The free radical signaling molecules reactive oxygen species (ROS) and reactive nitrogen species (RNS) have been proven to participate in iron deficiency in eukaryotes.We found that the inhibition of proteasome activity shortened the survival of iron-deficient cells compared with untreated iron-deficient cells.Surprisingly, the addition of arginine restored both NO level and the survival of proteasome-inhibited cells, suggesting that proteasome-derived NO is crucial for cell survival under iron-limited conditions.

View Article: PubMed Central - PubMed

Affiliation: Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kweishan, Taoyuan, Taiwan. m9701203@stmail.cgu.edu.tw.

ABSTRACT

Background: Iron plays a pivotal role in the pathogenesis of Trichomonas vaginalis, the causative agent of highly prevalent human trichomoniasis. T. vaginalis resides in the vaginal region, where the iron concentration is constantly changing. Hence, T. vaginalis must adapt to variations in iron availability to establish and maintain an infection. The free radical signaling molecules reactive oxygen species (ROS) and reactive nitrogen species (RNS) have been proven to participate in iron deficiency in eukaryotes. However, little is known about the roles of these molecules in iron-deficient T. vaginalis.

Methods: T. vaginalis cultured in iron-rich and -deficient conditions were collected for all experiments in this study. Next generation RNA sequencing was conducted to investigate the impact of iron on transcriptome of T. vaginalis. The cell viabilities were monitored after the trophozoites treated with the inhibitors of nitric oxide (NO) synthase (L-NG-monomethyl arginine, L-NMMA) and proteasome (MG132). Hydrogenosomal membrane potential was measured using JC-1 staining.

Results: We demonstrated that NO rather than ROS accumulates in iron-deficient T. vaginalis. The level of NO was blocked by MG132 and L-NMMA, indicating that NO production is through a proteasome and arginine dependent pathway. We found that the inhibition of proteasome activity shortened the survival of iron-deficient cells compared with untreated iron-deficient cells. Surprisingly, the addition of arginine restored both NO level and the survival of proteasome-inhibited cells, suggesting that proteasome-derived NO is crucial for cell survival under iron-limited conditions. Additionally, NO maintains the hydrogenosomal membrane potential, a determinant for cell survival, emphasizing the cytoprotective effect of NO on iron-deficient T. vaginalis. Collectively, we determined that NO produced by the proteasome prolonged the survival of iron-deficient T. vaginalis via maintenance of the hydrogenosomal functions.

Conclusion: The findings in this study provide a novel role of NO in adaptation to iron-deficient stress in T. vaginalis and shed light on a potential therapeutic strategy for trichomoniasis.

No MeSH data available.


Related in: MedlinePlus