Overexpression Of IL-7 Extends The Humoral Immune Response Induced By Rabies Vaccination

Rabies is one of the oldest human infectious diseases,which targets the central nervous system in almost all species of mammals.The number of rabies cases in China ranks second in the word and is the top five types of infectious diseases nationwide.More than 95% human rabies cases are related to animal bites,indicating that it is possible to eliminate human rabies by controlling rabies in domestic animals,especially dogs.The mass vaccination of domestic dogs(>70%)has nearly eliminated cases of human rabies in developed and some developing countries.However,the traditional animal inactivated vaccines induce protective antibody responses only after multiple shots and generally require repeated booster doses to provide long-lasting protection in pre-exposure settings,which increases the operational costs and restricts their wide use in developing countries.Therefore,it is still an urgent need to develop a single-dose and long-lasting RABV vaccine that can induce robust antibodies,especially VNA,to protect animals from rabies.Our previous studies with rRABVs expressing cytokines/chemokines,show that these immune factors help to induce rapid and enhanced humoral immune responses by recruiting and maturating DCs,which activate B cells and result in the enhanced production of antibodies,especially VNA.However,the duration and maintenance of the VNA is relatively short.And little information is available regarding the mechanisms underlying immunological memory,which can broaden humoral responses following rabies vaccination.In this study,a recombinant rabies virus(rRABV)that expressed murine interleukin-7(IL-7)which plays a central role in the proliferation and maintenance of both T-and B-lymphocytes in the immune responses,referred to here as r LBNSE-IL7,was constructed by reverse genetic system,and its effectiveness was evaluated in a mouse model.No significant difference was found in virus growth kinetics in vitro between the r RABV and parental virus.IL-7 was highly expressed in a dose-dependent manner in the r LBNSE-IL7 infected BSR cells but not rLBNSE or r LBNSE-IL7(-),as measured by ELISA.The rRABV encoding the IL-7 or IL7(-)gene without any side effects was demonstrated to be stable for at least ten generations in BSR cells,which confirmed by RT-PCR and sequencing.To evaluate any adverse effects that may have been caused by the expression of IL-7 on animals,we examined the presence of IL-7 genes could affect the pathogenicity in immunocompetent and immunocompromised mice by the intracerebral route.The result demonstrates that IL-7 expression in the brain rather than the increased size of the RABV genome attenuates RABV pathogenicity in vivo.To investigate the immunogenicity of rLBNSE-IL7,mice were immunized with rRABVs by the intramuscular route to analyze T cell-dependent B cell mechanisms induced by rRABVs.rLBNSE-IL7 induced higher rates of Tfh cells and GC B cells from draining lymph nodes,and increased the percentages of long-lived Bmem in the draining LNs and PCs in the bone marrow for up to 360 days post immunization than the parent virus rLBNSE.As a result of the expansion of long-lived PCs by r LBNSE-IL7,it also generated prolonged and higher virus-neutralizing antibodies(VNA)and IgG,resulting in better protection against a lethal challenge than rLBNSE.Moreover,consistent with the increased numbers of Bmem and PCs after boost with rLBNSE,rLBNSE-IL7-immunized mice promptly produced a more potent secondary anti-RABV neutralising antibody response than rLBNSE-immunized mice.In our present study,the expression of IL-7 may facilitate GC responses with the assistance of increased Tfh cells.The GC-derived B cells subsequently further differentiate into Bmem and PCs after an extensive selection step,following an improvement of primary and secondary antibody responses.In summary,our data indicate that the rRABV-expressing IL-7 designed for this study demonstrated changeless viral growth kinetics after the insertion of foreign gene and an ability to attenuate the pathogenicity,and high potency of inducing immunological memory for long-lasting primary and secondary antibody responses by Tfh cells,GC B cells,long-lived Bmem and PCs.This rRABV therefore has the potential to be used as a single-dose,long-lasting avirulent vaccine for animals.