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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

T. vaginalis extends the survival time when cultured under iron-deficient conditions. The growth of cells cultured in iron-rich (IR, 80 μM FAC, a) and -deficient (ID, 180 μM DIP, b) media. The number of viable cells was monitored every three hours using the trypan blue exclusion assay. The initial cell density is 1 × 106cells/ ml. The data are presented as the mean ± SD of three independent experiments
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Fig1: T. vaginalis extends the survival time when cultured under iron-deficient conditions. The growth of cells cultured in iron-rich (IR, 80 μM FAC, a) and -deficient (ID, 180 μM DIP, b) media. The number of viable cells was monitored every three hours using the trypan blue exclusion assay. The initial cell density is 1 × 106cells/ ml. The data are presented as the mean ± SD of three independent experiments

Mentions: It is known that iron is essential for cell proliferation in T. vaginalis. Iron restriction causes an increase in doubling time and a decrease in the maximum cell density [27]. Until now, no report indicates the viability of T. vaginalis upon iron deficiency. To monitor the cell viability under iron-deficient condition, we cultured the cells with a higher initial cell density (106 cells/ml) in the medium containing 180 μM DIP [24, 28]. The iron-rich cells reached the maximum cell density of approximately 3.5 × 106 (cells/ml) at 6 h after inoculation, followed by a rapid decline (Fig. 1a). The viable cells were reduced to 1 × 105 (cells/ml) at approximately 50 h in iron-rich cultivation. In contrast, we found that the maximum cell density of iron-deficient cells was 2-fold (~1.5 × 106 cells/ml) less than that of iron-rich cells (Fig. 1b). Interestingly, our result showed that the survival of iron-deficient cells was further extended to 66 h with a cell density of 1 × 105 viable cells per ml. This phenomenon reflects that although iron deficiency affects cell proliferation, T. vaginalis is capable of adapting to an iron-deficient environment and survives for a longer period. However, the underlying mechanism responsible for cell survival during iron deficiency is still unclear, which might be an important issue in elucidating how T. vaginalis establishes and maintains infection in the vaginal region.Fig. 1


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)

T. vaginalis extends the survival time when cultured under iron-deficient conditions. The growth of cells cultured in iron-rich (IR, 80 μM FAC, a) and -deficient (ID, 180 μM DIP, b) media. The number of viable cells was monitored every three hours using the trypan blue exclusion assay. The initial cell density is 1 × 106cells/ ml. The data are presented as the mean ± SD of three independent experiments
© Copyright Policy - open-access
Related In: Results  -  Collection

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

Fig1: T. vaginalis extends the survival time when cultured under iron-deficient conditions. The growth of cells cultured in iron-rich (IR, 80 μM FAC, a) and -deficient (ID, 180 μM DIP, b) media. The number of viable cells was monitored every three hours using the trypan blue exclusion assay. The initial cell density is 1 × 106cells/ ml. The data are presented as the mean ± SD of three independent experiments
Mentions: It is known that iron is essential for cell proliferation in T. vaginalis. Iron restriction causes an increase in doubling time and a decrease in the maximum cell density [27]. Until now, no report indicates the viability of T. vaginalis upon iron deficiency. To monitor the cell viability under iron-deficient condition, we cultured the cells with a higher initial cell density (106 cells/ml) in the medium containing 180 μM DIP [24, 28]. The iron-rich cells reached the maximum cell density of approximately 3.5 × 106 (cells/ml) at 6 h after inoculation, followed by a rapid decline (Fig. 1a). The viable cells were reduced to 1 × 105 (cells/ml) at approximately 50 h in iron-rich cultivation. In contrast, we found that the maximum cell density of iron-deficient cells was 2-fold (~1.5 × 106 cells/ml) less than that of iron-rich cells (Fig. 1b). Interestingly, our result showed that the survival of iron-deficient cells was further extended to 66 h with a cell density of 1 × 105 viable cells per ml. This phenomenon reflects that although iron deficiency affects cell proliferation, T. vaginalis is capable of adapting to an iron-deficient environment and survives for a longer period. However, the underlying mechanism responsible for cell survival during iron deficiency is still unclear, which might be an important issue in elucidating how T. vaginalis establishes and maintains infection in the vaginal region.Fig. 1

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