Generation of T-Cell Response through Vaccination by Mimicking Natural Course of Viral Infection

Abstract

SUMMARY: The goal of vaccination is to generate and maintain a heterogeneous pool of long lived memory T cells, which can provide long term protection. But often, we notice that vaccines, particularly subunit vaccines, fail to achieve the desired goal in case of many diseases such as HIV, tuberculosis and malaria. One of the possible reasons can be the inability of vaccine regimen to mimic natural course of infection. Hence, we had proposed a strategy in which we injected TLR ligands and Ag in a manner analogous to the course of natural infection. We believe that sensitization of host with increasing concentration of antigen along with TLR ligands will mimic a condition in host which is analogous to replicating pathogen as in natural infection and increasing the overall antigen exposure to the immune system. Creating such conditions can lead to generation of memory type CD4+ T cells and CD8+ T cells. The immunization strategy includes use of different combination of Ag and TLR ligands to compare natural course of infection and vaccine regimen. First two groups mimic course of natural infection. In first group, the immune system will encounter TLR ligands without antigen on day 1 and increasing doses of antigen and TLR ligands on subsequent two days. In second group, very little amount of antigen is administered along with TLR ligands on day 1. After successive days increasing concentration of antigen (OVA) and TLR ligands was injected to the same mice to further mimic the course. In the third group, which mimicked the vaccine regimen all the components (TLR ligands and antigen) were administered at one shot. Mice were sacrificed so as to measure acute phase, early memory phase and late memory stage of immune response. Cells from draining lymph nodes (Inguinal LNs and poplitael LNs) were harvested and stimulated in-vitro to reactivate antigen specific cells. Cells were surface stained with flourochrome conjugated antibodies for various cell surface markers and acquired on BD FACS Calibur and BD FACS Aria-III. Intarcellular staining was also performed for pro-inflammatory cytokines – IFN-γ, IL-2 and TNF-α. Results for CD 4+ T memory generation by Mimicking Natural Course of Infection compared to Vaccine Regimen (Day 12 and 30) when immunized with different combination of TLR ligands and OVA Ag shows that during acute phase there are considerable no. of central and effector memory cells in case of vaccine regimen, although 46 less than the natural course. This may be because during vaccination, high antigen dose is given in one shot which would generate more no. of effector cells. So, although vaccination generates memory it is not true memory and it diminishes during later phase of infection. Later phase (Day 66) results showed highest no. of memory cells in groups mimicking natural infection which may be heterogeneous in nature. Thus, we can say that we were able to follow natural course in some way which gave rise to long lived heterogeneous population of memory T cells. Results for CD4+ T memory generation when immunized with PAD4 along with TLR ligand indicated that there was increase in number of CD4+ central and effector memory response when immunized mice were compared with unimmunized ones. Though we cannot comment on memory response but still we were able to create conditions which would favour CD4-memory generation. Results for T cell memory generation by mimicking natural course of infection compared to vaccine regimen when immunized with different combination of TLR ligands and OVA Plasmid, on day 35, CI 2 (Course of Infection group 2) shows higher number of CD4+ and CD8+ T central memory and effector memory response as well as antigen specific response in terms of IFN-γ response compare to vaccine group. While IL-2 response shows a relative similarity in CI 1 and CI 2 group in case of CD4+ T cell response, CD8+ T cell responses, CI 2 group shows increase in IL-2 response as well as TNF-α. On Day 65, CI 2 group shows higher number of central memory responses, effector memory responses and IFN-γ producing cells as well as TNF-α producing cells for both CD4+ and CD8+ T cell responses when compared to vaccine regimen group. From the above findings one might infer that there is generation of qualitatively better T cell response in both CI 1 and CI 2 groups compared to vaccine. Moreover, this provides us insights that qualitatively better memory T cells following natural course of infection compared to vaccine regimen.