Evaluation of Helicobacter pylori-derived outer membrane vesicles on the expression of inflammatory cytokines
-
Maryam Mohammadbeigi
,
Amir Peymani
,
Shahin Bolori
,
Simin Sotoudeh
,
Rasoul Samimi
,
Anahita Bakhtiari
,
Saina Shegefti
,
Mehdi Bakht
,
Milad Badri
,
Amir Javadi
,
Farhad Nikkhahi
Abstract
Background and Objectives: Helicobacter pylori infection has been increasingly linked to extra-gastric diseases. Outer membrane vesicles are a key virulence factor of H. pylori. This study investigates the influence of H. pylori-derived outer membrane vesicles on inflammatory marker expression in human hepatoma cells (HepG2).
Materials and Methods: Outer membrane vesicles were isolated through ultracentrifugation and characterized using dynamic light scattering technique (DLS) and a Field Emission Scanning Electron Microscope (FE-SEM). Protein concentrations were measured via the Bradford assay. HepG2 cells treated with outer membrane vesicles were analyzed for IL-6, TNF-α, TLR-4, TGF-β, and PPAR-γ mRNA expression by RT-qPCR. Cell viability was assessed through an MTT assay. The prevalence of H. pylori virulence-associated genes (babA2, sabA, and oipA) was determined by PCR.
Results: The results showed a high prevalence of sabA (91.7%), babA2 (75%), and oipA (66.7%). FE-SEM and DLS analyses confirmed the presence of bleb-shaped nanovesicles ranging in size from 50 to 450 nm. H. pylori-derived outer membrane vesicles significantly upregulated the expression of pro-inflammatory markers (TLR-4, PPAR-γ, TNF-α, and IL-6), while downregulating TGF-β expression.
Conclusion: These findings underscore the potential role of nanoparticles in driving inflammatory responses and influencing host cell signaling, which may play a key role in liver-related pathologies.
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2. Lormohammadi L, Nikkhahi F, Bolori S, Karami AA, Hajian S, Rad N, et al. High level of resistance to metronidazole and clarithromycin among Helicobacter pylori clinical isolates in Qazvin province, Iran. Gene Rep 2022; 26: 101494.
3. Reshetnyak VI, Burmistrov AI, Maev IV. Helicobacter pylori: Commensal, symbiont or pathogen? World J Gastroenterol 2021; 27: 545-560.
4. Kim J, Wang TC. Helicobacter pylori and gastric cancer. Gastrointest Endosc Clin N Am 2021; 31: 451-465.
5. Huang Y, Wang Q-l, Cheng D-d, Xu W-t, Lu N-h. Adhesion and invasion of gastric mucosa epithelial cells by Helicobacter pylori. Front Cell Infect Microbiol 2016; 6: 159.
6. Pellicano R, Ianiro G, Fagoonee S, Settanni CR, Gasbarrini A. Extragastric diseases and Helicobacter pylori. Helicobacter 2020; 25 Suppl 1: e12741.
7. Mohammadi M, Attar A, Mohammadbeigi M, Peymani A, Bolori S, Fardsanei F. The possible role of Helicobacter pylori in liver diseases. Arch Microbiol 2023; 205: 281.
8. Tamura A, Fujioka T, Nasu M. Relation of Helicobacter pylori infection to plasma vitamin B12, folic acid, and homocysteine levels in patients who underwent diagnostic coronary arteriography. Am J Gastroenterol 2002; 97: 861-866.
9. He J, Liu Y, Ouyang Q, Li R, Li J, Chen W, et al. Helicobacter pylori and unignorable extragastric diseases: Mechanism and implications. Front Microbiol 2022; 13: 972777.
10. Rabelo-Gonçalves EMA, Roesler B, Zeitune J (2014). Helicobacter pylori and Liver–Detection of Bacteria in liver Tissue from patients with Hepatocellular carcinoma using Laser Capture Microdissection Technique (LCM). Trends in Helicobacter pylori Infection.
11. Mekonnen HD, Fisseha H, Getinet T, Tekle F, Galle PR. Helicobacter pylori infection as a risk factor for hepatocellular carcinoma: a case-control study in ethiopia. Int J Hepatol 2018; 2018: 1941728.
12. Xu G, Ma S, Dong L, Mendez-Sanchez N, Li H, Qi X. Relationship of Helicobacter pylori infection with nonalcoholic Fatty Liver Disease: A meta-analysis. Can J Gastroenterol Hepatol 2023; 2023: 5521239.
13. Chen X, Peng R, Peng D, Xiao J, Liu D, Li R. An update: is there a relationship between H. pylori infection and nonalcoholic fatty liver disease? why is this subject of interest? Front Cell Infect Microbiol 2023; 13: 1282956.
14. Turner L, Bitto NJ, Steer DL, Lo C, D’Costa K, Ramm G, et al. Helicobacter pylori outer membrane vesicle size determines their mechanisms of host cell entry and protein content. Front Immunol 2018; 9: 1466.
15. Chew Y, Chung H-Y, Lin P-Y, Wu D-C, Huang S-K, Kao M-C. Outer membrane vesicle production by Helicobacter pylori represents an approach for the delivery of virulence factors caga, vaca and urea into human gastric adenocarcinoma (Ags) cells. Int J Mol Sci 2021; 22: 3942.
16. Mommersteeg MC, Yu J, Peppelenbosch MP, Fuhler GM. Genetic host factors in Helicobacter pylori-induced carcinogenesis: Emerging new paradigms. Biochim Biophys Acta Rev Cancer 2018; 1869: 42-52.
17. Taubenheim C (2019). The genomic content of outer membrane vesicles (OMVs) in freshwater cyanobacteria.
18. Jarzab M, Posselt G, Meisner-Kober N, Wessler S. Helicobacter pylori-derived outer membrane vesicles (OMVs): role in bacterial pathogenesis? Microorganisms 2020; 8: 1328.
19. Gong J, Tu W, Liu J, Tian D. Hepatocytes: A key role in liver inflammation. Front Immunol 2023; 13: 1083780.
20. Schulze RJ, Schott MB, Casey CA, Tuma PL, McNiven MA. The cell biology of the hepatocyte: A membrane trafficking machine. J Cell Biol 2019; 218: 2096-2112.
21. Melo J, Pinto V, Fernandes T, Malheiro AR, Osório H, Figueiredo C, et al. Isolation method and characterization of outer membranes vesicles of Helicobacter pylori grown in a chemically defined medium. Front Microbiol 2021; 12: 654193.
22. Zhang T, Liu C-F, Zhang T-N, Wen R, Song W-L. Overexpression of peroxisome proliferator-activated receptor γ coactivator 1-α protects cardiomyocytes from lipopolysaccharide-induced mitochondrial damage and apoptosis. Inflammation 2020; 43: 1806-1820.
23. Amano A, Takeuchi H, Furuta N. Outer membrane vesicles function as offensive weapons in host–parasite interactions. Microbes Infect 2010; 12: 791-798.
24. Ait-Ahmed Y, Lafdil F. Novel insights into the impact of liver inflammatory responses on primary liver cancer development. Liver Res 2023; 7: 26-34.
25. Chmiela M, Kupcinskas J. Pathogenesis of Helicobacter pylori infection. Helicobacter 2019; 24 Suppl 1(Suppl Suppl 1): e12638.
26. Liu Y, Li D, Liu Y, Shuai P. Association between Helicobacter pylori infection and non-alcoholic fatty liver disease, hepatic adipose deposition and stiffness in Southwest China. Front Med (Lausanne) 2021; 8: 764472.
27. Wang S, Gao J, Wang Z. Outer membrane vesicles for vaccination and targeted drug delivery. Wiley Interdiscip Rev Nanomed Nanobiotechnol 2019; 11(2): e1523.
28. Yue Y, Xu J, Li Y, Cheng K, Feng Q, Ma X, et al. Antigen-bearing outer membrane vesicles as tumour vaccines produced in situ by ingested genetically engineered bacteria. Nat Biomed Eng 2022; 6: 898-909.
29. Nilsson H-O, Mulchandani R, Tranberg K-G, Stenram U, Wadström T. Helicobacter species identified in liver from patients with cholangiocarcinoma and hepatocellular carcinoma. Gastroenterology 2001; 120: 323-324.
30. Zoaiter M, Nasser R, Hage-Sleiman R, Abdel-Sater F, Badran B, Zeaiter Z. Helicobacter pylori outer membrane vesicles induce expression and secretion of oncostatin M in AGS gastric cancer cells. Braz J Microbiol 2021; 52: 1057-1066.
31. Furuyama N, Sircili MP. Outer membrane vesicles (OMVs) produced by gram‐negative bacteria: structure, functions, biogenesis, and vaccine application. Biomed Res Int 2021; 2021: 1490732.
32. Zahmatkesh ME, Jahanbakhsh M, Hoseini N, Shegefti S, Peymani A, Dabin H, et al. Effects of exosomes derived from Helicobacter pylori outer membrane vesicle-infected hepatocytes on hepatic stellate cell activation and liver fibrosis induction. Front Cell Infect Microbiol 2022; 12: 857570.
33. Bolori S, Shegefti S, Baghaei K, Yadegar A, Moon K-M, Foster LJ, et al. The Effects of Helicobacter pylori-Derived outer Membrane Vesicles on hepatic stellate cell activation and liver Fibrosis In Vitro. Biomed Res Int 2023; 2023: 4848643.
34. Shegefti S, Bolori S, Nabavi-Rad A, Dabiri H, Yadegar A, Baghaei K. Helicobacter pylori-derived outer membrane vesicles suppress liver autophagy: A novel mechanism for H. pylori-mediated hepatic disorder. Microb Pathog 2023; 183: 106319.
35. Chen X, Peng R, Peng D, Xiao J, Liu D, Li R. An update: is there a relationship between H. pylori infection and nonalcoholic fatty liver disease? why is this subject of interest? Front Cell Infect Microbiol 2023; 13: 1282956.
36. Sun Y, Gu J, Liu R, Zhou H, Lu L, Dai X, et al. IL‐2/IL‐6 ratio correlates with liver function and recovery in acute liver injury patients. APMIS 2019; 127: 468-474.
37. Yildirim Z, Bozkurt B, Ozol D, Armutcu F, Akgedik R, Karamanli H, et al. Increased exhaled 8‐isoprostane and interleukin‐6 in patients with Helicobacter pylori infection. Helicobacter 2016; 21: 389-394.
38. Tsai C-C, Kuo T-Y, Hong Z-W, Yeh Y-C, Shih K-S, Du S-Y, et al. Helicobacter pylori neutrophil-activating protein induces release of histamine and interleukin-6 through G protein-mediated MAPKs and PI3K/Akt pathways in HMC-1 cells. Virulence 2015; 6: 755-765.
39. Polyzos SA, Kountouras J, Papatheodorou A, Patsiaoura K, Katsiki E, Zafeiriadou E, et al. Helicobacter pylori infection in patients with nonalcoholic fatty liver disease. Metabolism 2013; 62: 121-126.
40. Ponzetto A, Figura N. Interleukin 6 predicts mortality in patients with End-Stage Liver Disease. Clin Gastroenterol Hepatol 2018; 16: 783.
41. Abdel-Razik A, Mousa N, Shabana W, Refaey M, Elhelaly R, Elzehery R, et al. Helicobacter pylori and non‐alcoholic fatty liver disease: A new enigma? Helicobacter 2018; 23(6): e12537.
42. Noori M, Mahboobi R, Nabavi-Rad A, Jamshidizadeh S, Fakharian F, Yadegar A, et al. Helicobacter pylori infection contributes to the expression of Alzheimer's disease-associated risk factors and neuroinflammation [Heliyon 9(9) (September 2023) e19607]. Heliyon 2023; 9(9): e19944.
43. Fabregat I, Moreno-Càceres J, Sánchez A, Dooley S, Dewidar B, Giannelli G, et al. TGF-β signalling and liver disease. FEBS J 2016; 283: 2219-2232.
44. Dooley S, ten Dijke P. TGF-β in progression of liver disease. Cell Tissue Res 2012; 347: 245-256.
45. Goo M-J, Ki M-R, Lee H-R, Yang H-J, Yuan D-W, Hong I-H, et al. Helicobacter pylori promotes hepatic fibrosis in the animal model. Lab Invest 2009; 89: 1291-1303.
46. Konturek PC, Kania J, Kukharsky V, Raithel M, Ocker M, Rembiasz K, et al. Implication of peroxisome proliferator-activated receptor γ and proinflammatory cytokines in gastric carcinogenesis: link to Helicobacter pylori-infection. J Pharmacol Sci 2004; 96: 134-143.
2. Lormohammadi L, Nikkhahi F, Bolori S, Karami AA, Hajian S, Rad N, et al. High level of resistance to metronidazole and clarithromycin among Helicobacter pylori clinical isolates in Qazvin province, Iran. Gene Rep 2022; 26: 101494.
3. Reshetnyak VI, Burmistrov AI, Maev IV. Helicobacter pylori: Commensal, symbiont or pathogen? World J Gastroenterol 2021; 27: 545-560.
4. Kim J, Wang TC. Helicobacter pylori and gastric cancer. Gastrointest Endosc Clin N Am 2021; 31: 451-465.
5. Huang Y, Wang Q-l, Cheng D-d, Xu W-t, Lu N-h. Adhesion and invasion of gastric mucosa epithelial cells by Helicobacter pylori. Front Cell Infect Microbiol 2016; 6: 159.
6. Pellicano R, Ianiro G, Fagoonee S, Settanni CR, Gasbarrini A. Extragastric diseases and Helicobacter pylori. Helicobacter 2020; 25 Suppl 1: e12741.
7. Mohammadi M, Attar A, Mohammadbeigi M, Peymani A, Bolori S, Fardsanei F. The possible role of Helicobacter pylori in liver diseases. Arch Microbiol 2023; 205: 281.
8. Tamura A, Fujioka T, Nasu M. Relation of Helicobacter pylori infection to plasma vitamin B12, folic acid, and homocysteine levels in patients who underwent diagnostic coronary arteriography. Am J Gastroenterol 2002; 97: 861-866.
9. He J, Liu Y, Ouyang Q, Li R, Li J, Chen W, et al. Helicobacter pylori and unignorable extragastric diseases: Mechanism and implications. Front Microbiol 2022; 13: 972777.
10. Rabelo-Gonçalves EMA, Roesler B, Zeitune J (2014). Helicobacter pylori and Liver–Detection of Bacteria in liver Tissue from patients with Hepatocellular carcinoma using Laser Capture Microdissection Technique (LCM). Trends in Helicobacter pylori Infection.
11. Mekonnen HD, Fisseha H, Getinet T, Tekle F, Galle PR. Helicobacter pylori infection as a risk factor for hepatocellular carcinoma: a case-control study in ethiopia. Int J Hepatol 2018; 2018: 1941728.
12. Xu G, Ma S, Dong L, Mendez-Sanchez N, Li H, Qi X. Relationship of Helicobacter pylori infection with nonalcoholic Fatty Liver Disease: A meta-analysis. Can J Gastroenterol Hepatol 2023; 2023: 5521239.
13. Chen X, Peng R, Peng D, Xiao J, Liu D, Li R. An update: is there a relationship between H. pylori infection and nonalcoholic fatty liver disease? why is this subject of interest? Front Cell Infect Microbiol 2023; 13: 1282956.
14. Turner L, Bitto NJ, Steer DL, Lo C, D’Costa K, Ramm G, et al. Helicobacter pylori outer membrane vesicle size determines their mechanisms of host cell entry and protein content. Front Immunol 2018; 9: 1466.
15. Chew Y, Chung H-Y, Lin P-Y, Wu D-C, Huang S-K, Kao M-C. Outer membrane vesicle production by Helicobacter pylori represents an approach for the delivery of virulence factors caga, vaca and urea into human gastric adenocarcinoma (Ags) cells. Int J Mol Sci 2021; 22: 3942.
16. Mommersteeg MC, Yu J, Peppelenbosch MP, Fuhler GM. Genetic host factors in Helicobacter pylori-induced carcinogenesis: Emerging new paradigms. Biochim Biophys Acta Rev Cancer 2018; 1869: 42-52.
17. Taubenheim C (2019). The genomic content of outer membrane vesicles (OMVs) in freshwater cyanobacteria.
18. Jarzab M, Posselt G, Meisner-Kober N, Wessler S. Helicobacter pylori-derived outer membrane vesicles (OMVs): role in bacterial pathogenesis? Microorganisms 2020; 8: 1328.
19. Gong J, Tu W, Liu J, Tian D. Hepatocytes: A key role in liver inflammation. Front Immunol 2023; 13: 1083780.
20. Schulze RJ, Schott MB, Casey CA, Tuma PL, McNiven MA. The cell biology of the hepatocyte: A membrane trafficking machine. J Cell Biol 2019; 218: 2096-2112.
21. Melo J, Pinto V, Fernandes T, Malheiro AR, Osório H, Figueiredo C, et al. Isolation method and characterization of outer membranes vesicles of Helicobacter pylori grown in a chemically defined medium. Front Microbiol 2021; 12: 654193.
22. Zhang T, Liu C-F, Zhang T-N, Wen R, Song W-L. Overexpression of peroxisome proliferator-activated receptor γ coactivator 1-α protects cardiomyocytes from lipopolysaccharide-induced mitochondrial damage and apoptosis. Inflammation 2020; 43: 1806-1820.
23. Amano A, Takeuchi H, Furuta N. Outer membrane vesicles function as offensive weapons in host–parasite interactions. Microbes Infect 2010; 12: 791-798.
24. Ait-Ahmed Y, Lafdil F. Novel insights into the impact of liver inflammatory responses on primary liver cancer development. Liver Res 2023; 7: 26-34.
25. Chmiela M, Kupcinskas J. Pathogenesis of Helicobacter pylori infection. Helicobacter 2019; 24 Suppl 1(Suppl Suppl 1): e12638.
26. Liu Y, Li D, Liu Y, Shuai P. Association between Helicobacter pylori infection and non-alcoholic fatty liver disease, hepatic adipose deposition and stiffness in Southwest China. Front Med (Lausanne) 2021; 8: 764472.
27. Wang S, Gao J, Wang Z. Outer membrane vesicles for vaccination and targeted drug delivery. Wiley Interdiscip Rev Nanomed Nanobiotechnol 2019; 11(2): e1523.
28. Yue Y, Xu J, Li Y, Cheng K, Feng Q, Ma X, et al. Antigen-bearing outer membrane vesicles as tumour vaccines produced in situ by ingested genetically engineered bacteria. Nat Biomed Eng 2022; 6: 898-909.
29. Nilsson H-O, Mulchandani R, Tranberg K-G, Stenram U, Wadström T. Helicobacter species identified in liver from patients with cholangiocarcinoma and hepatocellular carcinoma. Gastroenterology 2001; 120: 323-324.
30. Zoaiter M, Nasser R, Hage-Sleiman R, Abdel-Sater F, Badran B, Zeaiter Z. Helicobacter pylori outer membrane vesicles induce expression and secretion of oncostatin M in AGS gastric cancer cells. Braz J Microbiol 2021; 52: 1057-1066.
31. Furuyama N, Sircili MP. Outer membrane vesicles (OMVs) produced by gram‐negative bacteria: structure, functions, biogenesis, and vaccine application. Biomed Res Int 2021; 2021: 1490732.
32. Zahmatkesh ME, Jahanbakhsh M, Hoseini N, Shegefti S, Peymani A, Dabin H, et al. Effects of exosomes derived from Helicobacter pylori outer membrane vesicle-infected hepatocytes on hepatic stellate cell activation and liver fibrosis induction. Front Cell Infect Microbiol 2022; 12: 857570.
33. Bolori S, Shegefti S, Baghaei K, Yadegar A, Moon K-M, Foster LJ, et al. The Effects of Helicobacter pylori-Derived outer Membrane Vesicles on hepatic stellate cell activation and liver Fibrosis In Vitro. Biomed Res Int 2023; 2023: 4848643.
34. Shegefti S, Bolori S, Nabavi-Rad A, Dabiri H, Yadegar A, Baghaei K. Helicobacter pylori-derived outer membrane vesicles suppress liver autophagy: A novel mechanism for H. pylori-mediated hepatic disorder. Microb Pathog 2023; 183: 106319.
35. Chen X, Peng R, Peng D, Xiao J, Liu D, Li R. An update: is there a relationship between H. pylori infection and nonalcoholic fatty liver disease? why is this subject of interest? Front Cell Infect Microbiol 2023; 13: 1282956.
36. Sun Y, Gu J, Liu R, Zhou H, Lu L, Dai X, et al. IL‐2/IL‐6 ratio correlates with liver function and recovery in acute liver injury patients. APMIS 2019; 127: 468-474.
37. Yildirim Z, Bozkurt B, Ozol D, Armutcu F, Akgedik R, Karamanli H, et al. Increased exhaled 8‐isoprostane and interleukin‐6 in patients with Helicobacter pylori infection. Helicobacter 2016; 21: 389-394.
38. Tsai C-C, Kuo T-Y, Hong Z-W, Yeh Y-C, Shih K-S, Du S-Y, et al. Helicobacter pylori neutrophil-activating protein induces release of histamine and interleukin-6 through G protein-mediated MAPKs and PI3K/Akt pathways in HMC-1 cells. Virulence 2015; 6: 755-765.
39. Polyzos SA, Kountouras J, Papatheodorou A, Patsiaoura K, Katsiki E, Zafeiriadou E, et al. Helicobacter pylori infection in patients with nonalcoholic fatty liver disease. Metabolism 2013; 62: 121-126.
40. Ponzetto A, Figura N. Interleukin 6 predicts mortality in patients with End-Stage Liver Disease. Clin Gastroenterol Hepatol 2018; 16: 783.
41. Abdel-Razik A, Mousa N, Shabana W, Refaey M, Elhelaly R, Elzehery R, et al. Helicobacter pylori and non‐alcoholic fatty liver disease: A new enigma? Helicobacter 2018; 23(6): e12537.
42. Noori M, Mahboobi R, Nabavi-Rad A, Jamshidizadeh S, Fakharian F, Yadegar A, et al. Helicobacter pylori infection contributes to the expression of Alzheimer's disease-associated risk factors and neuroinflammation [Heliyon 9(9) (September 2023) e19607]. Heliyon 2023; 9(9): e19944.
43. Fabregat I, Moreno-Càceres J, Sánchez A, Dooley S, Dewidar B, Giannelli G, et al. TGF-β signalling and liver disease. FEBS J 2016; 283: 2219-2232.
44. Dooley S, ten Dijke P. TGF-β in progression of liver disease. Cell Tissue Res 2012; 347: 245-256.
45. Goo M-J, Ki M-R, Lee H-R, Yang H-J, Yuan D-W, Hong I-H, et al. Helicobacter pylori promotes hepatic fibrosis in the animal model. Lab Invest 2009; 89: 1291-1303.
46. Konturek PC, Kania J, Kukharsky V, Raithel M, Ocker M, Rembiasz K, et al. Implication of peroxisome proliferator-activated receptor γ and proinflammatory cytokines in gastric carcinogenesis: link to Helicobacter pylori-infection. J Pharmacol Sci 2004; 96: 134-143.
| Files | ||
| Issue | Vol 17 No 6 (2025) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijm.v17i6.20365 | |
| Keywords | ||
| Helicobacter pylori Gene expression Gastric cancer Real-time polymerase chain reaction Virulence factor | ||
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How to Cite
1.
Mohammadbeigi M, Peymani A, Bolori S, Sotoudeh S, Samimi R, Bakhtiari A, Shegefti S, Bakht M, Badri M, Javadi A, Nikkhahi F. Evaluation of Helicobacter pylori-derived outer membrane vesicles on the expression of inflammatory cytokines. Iran J Microbiol. 2025;17(6):966-976.
