Dynamics of Radiation attenuated sporozoites induced Liver-stage specific CD8+ Memory T cells following Infectious Sporozoite Challenge that ensue long-lived protection against Plasmodium berghei infection

Abstract

Developing effective anti-malarial vaccine has been a challenge. Radiation attenuated sporozoite (RAS) vaccine induces sterile protection against Plasmodium liver-stage (LS) infection, but the protection lasts 6-9 months in rodents. Interestingly, we have shown that the intermittent infectious sporozoite challenge within few weeks post last boost immunization enhances the sterile protection up to 18 months. While the loss of protection within 6 months was correlated with a sharp decline of CD8+ memory T cells in liver, the intermittent challenge appeared to rescue the loss of T cell responses. Here, an interval of 6 months or longer between the last boost immunization and the challenge is associated with the loss of protection. In this thesis work, we wanted to understand how the delayed challenge might impair the liver-stage specific CD8+ memory T cell responses. The C57BL/6 mice were vaccinated with multiple doses of Plasmodium berghei RAS (Pb-RAS). Two weeks later, they were challenged with 10K infectious P. berghei sporozoites. (1) We observed that CD8+ memory T cells were maintained in RAS vaccinated mice irrespective of their status of challenge, although the frequencies of the memory T cells were higher in the challenged group. The expression of anti-apoptotic factor Bcl-2 was reflected in the status of CD8+ memory T cells in liver. (2) Upon evaluating the effector function of T cells, we found that RAS-mice w/o challenge showed significantly declined cytotoxic (CD107a+) CD8+ T cell response in liver after six months of RAS vaccination, while it was maintained in challenged mice. Although the IFN-+ CD8+ T cells were maintained in both the groups, the overall frequencies of T cells were significantly higher in the challenged mice. More importantly, challenged mice showed CD8+ T cells expressing higher IFN- compared to RAS vaccinated mice (w/o challenged). (3) We further measured the homing potential of the CD8+ T cells in terms of chemokine receptor expression of CXCR3 and CXCR6. We did not find much difference in the expression of CXCR3 molecules on the CD8+ T cells from both groups of mice even after six months, but we observed the higher expression of CXCR6 on the effector T cells characterized from spleen and liver draining LNs of challenged mice compared to unchallenged RAS immune mice. Moreover, we observed that the retention of CXCR6 expression on the effector T cells in the livers of challenged mice is relatively longer compared to the unchallenged RAS immune mice. It is known that CXCR6 expression on CD8+ T cells is critical for their survival in liver. Thus, our results are the suggestive of the fact that infectious sporozoite challenge helps retain the LS specific CD8 memory T cells in the liver of RAS immunized mice. In summary, our results demonstrate that intermittent infectious sporozoite challenge generates qualitatively better CD8+ T cells in terms of their effector function as well as retention in the liver that might ensure the longer-lived sterile protection against liver-stage Plasmodium infection. While addressing our hypothesis, we also demonstrated that induction of sterile protection primarily requires frequent immunizations with RAS vaccine and the parasite liver-load during RAS vaccination and/or challenge could influence the longevity of the protection.