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Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line
BACKGROUND: Autophagy is an important part of pathogenesis of IBD. Thiopurines such as azathioprine (AZA) are approved drugs for clinical practices in IBD patients. Besides, as an escape strategy, Toxoplasma gondii can use the mTORC1 complex to inactivate autophagy. METHODS: In this study, we invest...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10029279/ https://www.ncbi.nlm.nih.gov/pubmed/36941573 http://dx.doi.org/10.1186/s12866-023-02819-8 |
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author | Nemati, Sara Mohammad Rahimi, Hanieh Meyfour, Anna Pazoki, Hossein Asadzadeh Aghdaei, Hamid Shahrokh, Shabnam Mirjalali, Hamed |
author_facet | Nemati, Sara Mohammad Rahimi, Hanieh Meyfour, Anna Pazoki, Hossein Asadzadeh Aghdaei, Hamid Shahrokh, Shabnam Mirjalali, Hamed |
author_sort | Nemati, Sara |
collection | PubMed |
description | BACKGROUND: Autophagy is an important part of pathogenesis of IBD. Thiopurines such as azathioprine (AZA) are approved drugs for clinical practices in IBD patients. Besides, as an escape strategy, Toxoplasma gondii can use the mTORC1 complex to inactivate autophagy. METHODS: In this study, we investigated whether T. gondii tachyzoites may modulate autophagy and interfere the effects of azathioprine in IBD treatment. PMA-activated human monocyte cell line (THP-1) was infected with fresh T. gondii RH tachyzoites. After 5 h of infection, the cells were treated with AZA for 6 h. The expression of atg5, atg7, atg12, lc3b, and β-actin (BACT) genes was evaluated using quantitative real-time PCR. To analyze the phosphorylation of ribosomal protein S6 (rpS6), western blot using specific primary antibodies was performed. RESULTS: The results of real-time PCR revealed that AZA, T. gondii tachyzoites, and a combination of AZA and T. gondii tachyzoites upregulated atg5 gene for 4.297-fold (P-value = 0.014), 2.49-fold (P-value = 0.006), and 4.76-fold (P-value = 0.001), respectively. The atg7 gene showed significant upregulation (2.272-fold; P-value = 0.014) and (1.51-fold; P-value = 0.020) in AZA and AZA / T. gondii, respectively. The expression of atg12 gene was significantly downregulated in AZA and T. gondii tachyzoites for (8.85-fold; P-value = 0.004) and (2.005-fold; P-value = 0.038), respectively, but upregulated in T. gondii/AZA (1.52-fold; P-value = 0.037). In addition, the lc3b gene was only significantly changed in AZA / T. gondii (3.028-fold; P-value = 0.001). Western blot analysis showed that T. gondii tachyzoites significantly phosphorylated rpS6, and tachyzoites did not interfere the effects of AZA to phosphorylate the rpS6. CONCLUSION: Taken together, although AZA and T. gondii similarly affects the expression levels of atg5, atg7, and atg12, but T. gondii does not seem to modulate the effects of AZA via mTORC functions. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12866-023-02819-8. |
format | Online Article Text |
id | pubmed-10029279 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-100292792023-03-22 Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line Nemati, Sara Mohammad Rahimi, Hanieh Meyfour, Anna Pazoki, Hossein Asadzadeh Aghdaei, Hamid Shahrokh, Shabnam Mirjalali, Hamed BMC Microbiol Research BACKGROUND: Autophagy is an important part of pathogenesis of IBD. Thiopurines such as azathioprine (AZA) are approved drugs for clinical practices in IBD patients. Besides, as an escape strategy, Toxoplasma gondii can use the mTORC1 complex to inactivate autophagy. METHODS: In this study, we investigated whether T. gondii tachyzoites may modulate autophagy and interfere the effects of azathioprine in IBD treatment. PMA-activated human monocyte cell line (THP-1) was infected with fresh T. gondii RH tachyzoites. After 5 h of infection, the cells were treated with AZA for 6 h. The expression of atg5, atg7, atg12, lc3b, and β-actin (BACT) genes was evaluated using quantitative real-time PCR. To analyze the phosphorylation of ribosomal protein S6 (rpS6), western blot using specific primary antibodies was performed. RESULTS: The results of real-time PCR revealed that AZA, T. gondii tachyzoites, and a combination of AZA and T. gondii tachyzoites upregulated atg5 gene for 4.297-fold (P-value = 0.014), 2.49-fold (P-value = 0.006), and 4.76-fold (P-value = 0.001), respectively. The atg7 gene showed significant upregulation (2.272-fold; P-value = 0.014) and (1.51-fold; P-value = 0.020) in AZA and AZA / T. gondii, respectively. The expression of atg12 gene was significantly downregulated in AZA and T. gondii tachyzoites for (8.85-fold; P-value = 0.004) and (2.005-fold; P-value = 0.038), respectively, but upregulated in T. gondii/AZA (1.52-fold; P-value = 0.037). In addition, the lc3b gene was only significantly changed in AZA / T. gondii (3.028-fold; P-value = 0.001). Western blot analysis showed that T. gondii tachyzoites significantly phosphorylated rpS6, and tachyzoites did not interfere the effects of AZA to phosphorylate the rpS6. CONCLUSION: Taken together, although AZA and T. gondii similarly affects the expression levels of atg5, atg7, and atg12, but T. gondii does not seem to modulate the effects of AZA via mTORC functions. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1186/s12866-023-02819-8. BioMed Central 2023-03-21 /pmc/articles/PMC10029279/ /pubmed/36941573 http://dx.doi.org/10.1186/s12866-023-02819-8 Text en © The Author(s) 2023 https://creativecommons.org/licenses/by/4.0/Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/ (https://creativecommons.org/licenses/by/4.0/) . The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/ (https://creativecommons.org/publicdomain/zero/1.0/) ) applies to the data made available in this article, unless otherwise stated in a credit line to the data. |
spellingShingle | Research Nemati, Sara Mohammad Rahimi, Hanieh Meyfour, Anna Pazoki, Hossein Asadzadeh Aghdaei, Hamid Shahrokh, Shabnam Mirjalali, Hamed Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title | Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title_full | Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title_fullStr | Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title_full_unstemmed | Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title_short | Evaluation of the mTORC activity in the presence of Toxoplasma gondii and azathioprine in human monocyte cell line |
title_sort | evaluation of the mtorc activity in the presence of toxoplasma gondii and azathioprine in human monocyte cell line |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10029279/ https://www.ncbi.nlm.nih.gov/pubmed/36941573 http://dx.doi.org/10.1186/s12866-023-02819-8 |
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