Evaluation of the Efficacy of Saponin Extract from Glycine Max (L.) Merr. as an Adjuvant on the Hepatitis B Vaccine and Hepatitis B Surface Antigen in Balb/C Mice

Abstract

The most prevalent and dangerous type of liver infection worldwide is hepatitis B. Hepatitis B virus infects and damages the liver. A chronic hepatitis B infection affects roughly 300 million people, whereas two billion people, or one in three, have already contracted the virus. Hepatitis B is preventable and treatable, but it nevertheless claims the lives of up to a million people annually. Hepatitis B vaccination is the primary means of preventing infections and complications caused by the Hepatitis B Virus (HBV). Subunit vaccines such as the recombinant hepatitis B vaccines have been shown not to activate many facets of the immune response, as compared to whole-organism-based vaccines. Immuno-stimulatory adjuvants that improve immune responses caused by low immunogenic antigens are critical for improving immunogenicity in subunit vaccines. Saponin-based adjuvants extracted from Quillaja saponaria and Glycine max (L.) Merr. can stimulate cell-mediated and enhance antibody production. These saponins are now being applied in human vaccines, with several clinical trials proving safety and efficacy. Using mice models (N=51), this study assessed the adjuvant activities of saponins extracted from Glycine max (L.) Merr. on BALB/c mice vaccinated with either Revac BTM vaccine, a commercial hepatitis B vaccine or purified hepatitis B surface antigen. Saponins were extracted from soybean meal and characterized using the Fourier Transform Infrared Analysis and Ultra-violet/Visible Spectrophotometry. Eight-weeks-old female BALB/c mice were divided into 17 groups and vaccinated in triplicate, with 50µl of either Revac BTM vaccine, or Hepatitis B surface Antigen (HBsAg) supplemented with 100%, 50% or 25% concentrations of the saponin extract. A booster shot was administered two weeks after the first vaccination. A negative control involved the administration of phosphate buffer saline. Enzyme-linked Immunosorbent Assay was used to compare the humoral immune response using serum collected on day 14 and day 30. Spleen tissues were harvested on day 30 and used to analyze the cellular immune response by determining the mRNA expression levels of the Interleukin-6 (IL-6) and Tumor Necrosis Factor-alpha (TNF-α) genes. Hematological analysis was performed on the whole blood collected from the mice through a cardiac puncture. The findings from this study demonstrated that soybean saponin extracts did not have a statistically significant effect on the immune response of BALB/c mice. The immune response was slightly higher for Revac-BTM vaccine (0.758 + 0.012), and HBsAg (0.799 + 0.013) than vaccination with the Revac-BTM vaccine without the saponin extract. A novel finding was that the HBV vaccine suppressed the mRNA expression on IL-6 gene (Fold change in transcription (FCT) = 0.603, while promoting the expression of the TNF-α gene (FCT = 28.84). This study demonstrated that the Glycine max (L). Merr. saponin extract did not have any effect on most of the hematological parameters. However, the neutrophil and platelet counts were reduced (p value= 0.027 and 0.592 respectively). Glycine max (L). Merr. saponin extract showed minimal adjuvant activity for HBV vaccine, unique expression profile of IL-6 and TNF-α genes, and high safety profile in BALB/c mice. The findings of this study demonstrated that the saponin extract from Glycine max (L). Merr. did not elicit significant immune response in BALB/c mice and hence cannot be used in combination with either the Revac-BTM or the HBsAg.