PPD in HBsAg vaccine formulation suppressed IFN-γ and IL-4 cytokine responses and induced long-lived humoral immune responses: Results from 220-day monitoring of specific IgG responses

Document Type : Original Article


1 Department of Biotechnology, Faculty of Advanced Sciences & Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran

2 Department of Microbiology, Faculty of Advanced Sciences & Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran

3 Department of FMD vaccine, Razi Vaccine and Serum Research Institute, Agricultural Research, Education and Extension Organization (AREEO), Karaj, Iran

4 Department of Biology, Faculty of basic science, Shahrekord Branch, Islamic Azad University, Shahrekord, Iran

5 Advanced Therapy Medicinal Product (ATMP) Department, Breast Cancer Research Center, Motamed Cancer Institute, Academic Center for Education, Culture and Research (ACECR), Tehran, Iran

6 Recombinant Vaccine Research Center, Tehran University of Medical Sciences, Tehran, Iran

7 Immunotherapy Group, The Institute of Pharmaceutical Sciences (TIPS), Tehran University of Medical Sciences, Tehran, Iran

8 Department of Immunology, Pasteur Institute of Iran, Tehran, Iran


Objective(s): HBsAg vaccine is unable to induce Th1 immune responses. Here, immune responses and long-lived IgG responses of HBsAg-Alum, HBsAg-MF59, as well as HBsAg-MF59 were compared when formulated with PPD. 
Materials and Methods: BALB/c mice were injected with the vaccines subcutaneously three times with a two-week interval. Then, specific IgG, long-lived IgG responses up to 220 days, and also IgG1 and IgG2a isotypes were assessed using ELISA. Furthermore, IFN-γ and IL-4 were assessed on spleen cell culture supernatant by ELISA. 
Results: IFN-γ cytokine response between MF59- and Alum-adjuvanted vaccines did not show a significant difference. HBsAg-Alum vaccine revealed an increase in IL-4 cytokine, as compared with those immunized with HBsAg-MF59 at borderline (P=0.0553). In addition, HBsAg-MF59+PPD 10 µg showed a significant decrease in IL-4 and IFN-γ cytokines, as compared with HBsAg-MF59. Furthermore, the HBsAg-MF59+PPD10 µg group showed a significant increase in the IL-2/IL-4 ratio, as compared with HBsAg-MF59 (P=0.0339). Specific IgG antibody showed a significant increase in HBsAg-MF59, as compared with HBsAg-Alum. Furthermore, HBsAg-MF59 plus PPD showed a significant increase in IgG responses, as compared with HBsAg-MF59 and HBsAg-Alum groups. Long-lived IgG responses up to 220 days after the final shot showed a significant increase in HBsAgMF59 versus HBsAg-Alum group and PPD in the HBsAg-MF59 vaccine formulation, resulting in a significant increase in IgG responses versus HBsAg-MF59 group. In addition, PPD in the HBsAg-MF59 vaccine formulation suppressed IgG1 response versus HBsAg-Alum. However, HBsAg-MF59 showed a significant increase in IgG2α versus the HBsAg-Alum group (P=0.0190). Immunization with HBsAg-MF59+PPD (10 µg) showed a significant increase versus the HBsAg-MF59 group (P=0.0040). Results from the IgG2a/IgG1 ratio in HBsAg-MF59+PPD1µg and HBsAg-MF59+PPD10 µg groups showed a significant increase as compared with HBsAg-MF59 groups (P<0.0345). 
Conclusion: PPD in the HBsAg vaccine formulation suppressed IL-4 and IFN-γ responses, but increased the IL-2/IL-4 ratio, IgG2a, and IgG2a/IgG1 responses, which may show Th1 polarization. Furthermore, PPD leads to more potent long-lived IgG responses in the HBsAg vaccine, highlighting its potential as a component of a complex adjuvant.


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