Screening Of CpG-ODN And The Effect Of CpG-ODN As The Immune Adjuvant Of The Hepatitis B Vaccine On The Pregnant Mice

Objective: Design and synthesize the oligodeoxynucleotides containing unm-ethylated CpG dinucleotides (CpG-ODN) to screen out the optimal CpG-ODN that could markedly enhance the immune function of cord blood mononuclear cells in health newborns. Moreover, observe the action of CpG-1826 (5' -TCCATGACGTTC CTGACGTT-3' ) as the immune adjuvant of the hepatitis B vaccine on the pregnant mice and their neonatal mice.Methods: Using the tetrazolium salt assay (MTT) to detect the immunostimul-atory effects of seventeen CpG-ODNs on cord blood mononuclear cells in normal term infants, primarily single out the CpG-ODNs that had better immunological function. To further validate immunostimulatory properties of the CpG-ODNs, the expression of TLR9 in cord blood mononuclear cells induced by these CpG-ODNs was tested by RT-PCR. It is well-known that CpG-1826 have potently property of enhancing immunity of mouse. Thereby, we employ different concentrations of CpG-1826 as adjuvant of the hepatitis B vaccine to immune the pregnant mice. Serum hepatitis B surface antibody (HBsAb) levels in pregnant mice and their neonatal mice were detected by ELISA assays. At the same time, the growth and development index plus the survival quantity of the neonatal mice were inspected.Results: We screened out CpG-1: 5' -TCGCGTCGTCGTCGTCGTCGTCGT-3' andCpG-6:5' -GACGTTCGAACGTCGACGTA-3' , which could evidently activate immunity of cord blood mononuclear cells in neonate. The absorbency of CpG-1 and CpG-6 measured by MTT assay were significantly higher than negative control group. The TLR9/GAPDH absorbency ratio of CpG-1 and CpG-6 measured by RT-PCR assay were dramatically higher than the positive control groups CpG-2006 and the negative control group. When the pregnant mice were injected CpG-1826 (20 g) with hepatitis B vaccine, the serum quantity of HBsAb in pregnant mice and neonatal mice increased distinctly than the CpG-1826 (10 g) group , theCpG-1826 (40 wg) group, the hepatitis B vaccine group or negative control group. But there were no obvious difference in the growth and development index plus the survival quantity of neonatal mice in the five groups.Conclusion: CpG-ODN 5' -TCGCGTCGTCGTCGTCGTCGTCGT-3' and5 ' -GACGTTCGAACGTCGACGTA-3 ' both have capacity of reinforcement immunity of cord blood mononuclear cell in newborn. CpG-1826 (20 g) not only could markedly increase the level of HBsAb in pregnant mice and their neonatal mice, but also have no any influence on the survival quantity , the growth and development of neonatal mice, while it was injected into pregnant mice with the hepatitis B vaccine. Therefore, CpG-1826 is an ideal immune adjuvant.