Objective: To study the immunological protection of Helicobacter pylori (H. pylori) vaccine with chitosan as adjuvant and it's mechanism.
Methods: One-grade female BALB/c mice were randomly divided into nine groups and immunized by (1) PBS alone, (2) chitosan solution alone, (3) chitosan particles alone, (4) H. pylori antigen alone, (5) H. pylori antigen plus chitosan solution, (6) H. pylori antigen plus chitosan particles, (7) H. pylori antigen plus CT, (8) H. pylori antigen plus chitosan solution and cholera toxin (CT), (9) H. pylori antigen plus chitosan particles and CT orally respectively once a week for four weeks. At 4 weeks after the last immunization, these mice were challenged by alive H. pylori (1 x 10(9)/ml) twice at two days intervals. At 4 weeks after the last challenge, these mice were all killed and Gastric mucosa were embedded in paraffin, sectioned and assayed with Giemsa stain. The other gastric mucosa were used to quantitatively culture H. pylori. An ELISA was used to detect anti-H. pylori IgA in saliva and gastric mucosa and a quantitative ELISA was used to detect IL-2, IL-4, IL-10 content in gastric mucosa, and SP immunohistochemical method was used to detect secretory immunoglobulin A (sIgA) in gastric mucosa.
Results: (1) In the groups with chitosan as adjuvant, 60% mice could achieve immunological protection, which was according to that with CT as adjuvant (58.33%), and was significantly higher than H. pylori antigen alone and other groups without H. pylori antigen (P < 0.01 or P < 0.05). While the rates of protection in the groups with chitosan plus CT as adjuvant were 84.62%, 85.71% and the H. pylori colonization score in it was significantly lower than that in the groups with CT as adjuvant and without adjuvants (P < 0.01 or P < 0.05). (2) the labeling index for sIgA-positive lumen of glands and special anti-H. pylori IgA levels in gastric mucosa in the groups with chitosan as an adjuvant had no difference with those in the group with CT as an adjuvant (P > 0.05) and were significantly higher than those in non-adjuvant groups, while those in the groups with chitosan plus CT were significantly higher than those in the group with CT as an adjuvant (P < 0.01 or P < 0.05). (3) Before challenge, the content of IL-2 in gastric mucosa were no different among different groups (P > 0.05). After challenge the content of IL-2 were significantly higher in the groups with adjuvant than those in the control group (P < 0.01 or P < 0.05), Moreover, those in the groups with antigen after challenge were significantly higher than those before challenge (P < 0.05). (4) Before challenge, the content of IL-10 in gastric mucosa were significantly higher in the groups with chitosan as adjuvant than those in the control group and the group without adjuvant (P < 0.01 or P < 0.05). After challenge, the content of IL-10 were no different among different groups (P > 0.05). Moreover, those in the groups with adjuvant after challenge were significantly lower than those before challenge (P < 0.01). (5) Before challenge, the content of IL-4 in gastric mucosa were significantly higher in the groups with chitosan as adjuvant than those in the control group and the group without adjuvant (P < 0.05), After challenge, the content of IL-4 were significantly higher in the groups with chitosan particles as an adjuvant than those in the group with CT as an adjuvant (P < 0.05), and those in the group with chitosan solution as an adjuvant were significantly higher than those in control group, non-adjuvant group and the groups with CT (P < 0.01 or P < 0.05), Moreover, those in the groups with adjuvant after challenge were significantly lower than those before challenge (P < 0.01 or P < 0.05).
Conclusions: (1) H. pylori vaccine with chitosan as adjuvant could protect against H. pylori infection, this suggested that chitosan could be a mucosa adjuvant of H. pylori vaccine, and it could effectively elicit special humoral immune response of systemic and local mucosa, which might be one of its protective mechanism. (2) H. pylori vaccine with chitosan as adjuvant may induce both Th1 and Th2 type immune response, and after challenge it could reverse the inhibition of Th2 induced by H. pylori infection and recover the Th1/Th2 imbalance. which might contribute to the immune protection against H. pylori.