Frequencies of the expression of main protein antigens from Helicobacter pylori isolates and production of specific serum antibodies in infected patients

AIM: To investigate the frequencies of the expression of main protein antigens of Helicobacter pylori (H py/ori) isolates, such as UreB, VacA, CagA1, HpaA, NapA, FlaA and FlaB and the production of specific antibodies in sera from H pylori-infected patients, and to understand the correlations among the different clinical types of chronic gastritis and peptic ulcer and the infection and virulence of H pylori. METHODS: H pylori strains in biopsy specimens from 157 patients with chronic gastritis and peptic ulcer were isolated and serum samples from the patients were also collected. The target recombinant proteins rUreB, rVacA, rCagAl, rHpaA, rNapA, rFlaA and rFlaB expressed by the prokaryotic expression systems constructed in our previous studies were collected through Ni-NTA affinity chromatography. Rabbit antisera against rUreB, rVacA, rCagAl, rHpaA, rNapA, rFlaA and rFlaB were prepared by using routine subcutaneous immunization. By using ultrasonic lysates of the isolates as coated antigens, and the self-prepared rabbit antisera as the first antibodies and commercial HRP-labeling sheep anti-rabbit IgG as the second antibody, expression frequencies of the seven antigens in the isolates were detected by ELISA. Another ELISA was established to detect antibodies against the seven antigens in sera of the patients by using the corresponding recombinant proteins as coated antigens, and the sera as the first antibody and HRP-labeling sheep anti-human IgG as the second antibody respectively. Correlations among the different clinical types of chronic gastritis and peptic ulcer and the infection and virulence of H pylori were statistically analysed. RESULTS: In the 125 isolates of H pylori, the positive rates of UreB, VacA, CagAl, HpaA, NapA, FlaA and FlaB were 100%, 65.6%, 92.8%, 100%, 93.6%, 100% and 99.2% respectively. In the 125 serum samples from the H pylori infected patients, the positive rates of antibodies against recombinant UreB, VacA, CagA1, HpaA, NapA, FIaA and FlaB were 100%, 42.4%, 89.6%, 81.6%, 93.6%, 98.4% and 92.8% respectively. H pylori strains were isolated from 79.6% (125/157) of the biopsy specimens, but no close correlations among the H pylori infection frequencies and different types of chronic gastritis and peptic ulcer could be found (P>0.05, x2 = 0.01-0.87). The VacA positive rate (82.40%) in the strains isolated from the specimens of patients with peptic ulcer and the anti-VacA positive rate (54.3%) in the sera from the patients were significantly higher than those (51.5%, 32.3%) from the patients with chronic gastritis (P<0.01, x2= 13.19; P<0.05, x2= 6.13). When analysis was performed in the different types of chronic gastritis, the VacA in the strains isolated from the specimems of patients with active gastritis showed a higher expression frequency (90.0%) than those from superficial (47.9%) and atrophic gastritis (30.0%) (P<0.05, x2 = 5.93; P<0.01,x2 = 7.50). While analysis was carried out in the strains isolated from the specimens with superficial (93.8%) and active gastritis (100%), NapA showed a higher expression frequency compared to that from atrophic gastritis (60.0%) (P<0.01, x2 = 8.88; P<0.05, X2=5.00). CONCLUSION: The types of chronic gastritis and peptic ulcer and their severity are not associated with H pylori infection frequency but closely related to the infection frequency of different virulent H pylori strains. The optimal antigens for developing vaccine and diagnostic kit are UreB, FlaA, HpaA, FlaB, NapA and CagAl, but not VacA.

Assessment of 188Re marked anti MHC class II antibody by peripheral blood mononuclear cells stimulated by donor alloantigen

Background Previous studies showed that anti MHC-II monoclone antibody （MAb） only had partial inhibiting effect of alloreactive mixed lymphocyte reaction （MLR） in vitro and it was unsteady and non-persistent. The aim of this research was to determine whether radioactive isotope 188Re marked MHC-II antibody could benefit the allograft acceptance in transplantation as compared to normal MHC-II antibody. Methods 188Re was incorporated to 2E9/13F（ab＇）2 which is against swine MHC class II antigen （MAb-~88Re）. Porcine peripheral blood mononuclear （PBMC） cells were examined for proliferation and cytokine mRNA expression after stimulation with MHC-II MAb or MAb-188Re. Results The proliferative response of recipient PBMCs in mixed lymphocyte reaction （MLR） to donor alloantigen showed that the stimulation index of MAb-188Re group was significantly lower than the MHC-II MAb group and control （P 〈0.05）. mRNA expression of interleukin 2, interferon y and tumor necrosis factor a （type 1 cytokines） was lower in MAb-188Re group than the MHC-II MAb group, while interleukin 10 （type 2 cytokines） was higher in MAb-~aSRe group in the first 24 hours. Conclusion MAb-188Re could help the graft acceptance by inhibiting T cell proliferation, lowering the expression of type 1 cytokines and elevating the type 2 cytokines produced by PBMC.