Evaluation of the Cytokine Genes Expression in Vaccinated BALB/c Mice with pEGFP-C2-leoA DNA Vaccine

Zahra  Ahmadzadeh Chaleshtori; Ali Asghar Rastegari; Hashem Nayeri; Abbas Doosti

doi:10.18502/ssu.v30i12.11966

Zahra Ahmadzadeh Chaleshtori Department of Biochemistry, Falavarjan Branch, Islamic Azad University, Isfahan, Iran.
Ali Asghar Rastegari Department of Biochemistry, Falavarjan Branch, Islamic Azad University, Isfahan, Iran.
Hashem Nayeri Department of Biochemistry, Falavarjan Branch, Islamic Azad University, Isfahan, Iran.
Abbas Doosti Biotechnology Research Center, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran.

DOI: https://doi.org/10.18502/ssu.v30i12.11966

Keywords: Gastric cancer, Helicobacter pylori, cytokine, vaccine, leoA, BALB/c.

Abstract

Introduction: Helicobacter pylori (H. pylori) causes successive changes in the stomach wall, which starts with inflammation of the stomach mucosa and in some cases leads to stomach cancer. The leoA gene, by encoding GTPase, plays a vital role in the pathogenicity of this bacterium in the gastric mucosa. The secretory vesicles produced by the leoA gene release poison and stimulate the immune system in the host's body. The aim of the present study was to evaluate the gene vaccine pEGFP-C2-leoA on the expression changes of cytokines such as IL6, IL4, and interferon-gamma in inflammation caused by Helicobacter pylori infection in a mouse model.

Methods: In this interventional experimental study, recombinant plasmid (pEGFP-C2-leoA) was produced, propagated, and extracted through transformation into susceptible bacterial cells. Then, suitable concentrations of 1% chitosan nanoparticles solution were prepared for injection into the quadriceps muscle of BALB/c mice. Finally, the gene expression and changes of the mentioned cytokines were measured by Real-Time RT-PCR method, the obtained results were statistically analyzed by SPSS version 16 software; One way ANOVA test and subsequent LSD test, as well as independent t-test, were used to check the existence of correlation and the significance level of the data.

Results: After the injection of the vaccine into the quadriceps muscle of mice during the treatment period, cytokines IFNᵧ (<0.038) and IL6 (<0.049) showed a significant increase in expression. On the other hand, cytokine IL4 and the leoA gene also showed a significant decrease in expression (>0.042).

Conclusion: Based on the results, the leoA gene cloned in the expression vector pEGFP-C2 has the ability to express and produce the specific protein product of this gene in eukaryotic cells, and according to the results obtained in the animal model and the immune characteristic obtained in this research it is shown that the final construct pEGFP-C2-leoA has the necessary potential to investigate immunogenicity in a human model as a gene vaccine.