Critical Role of Perforin-dependent CD8+ T Cell Immunity for Rapid Protective Vaccination in a Murine Model for Human Smallpox

Kremer, Melanie; Suezer, Yasemin; Volz, Asisa; Frenz, Theresa; Majzoub, Monir; Hanschmann, Kay-Martin; Lehmann, Michael H.; Kalinke, Ulrich; Sutter, Gerd

Cytotoxic effector molecule perforin is essential for rapid protection T cells seemed to play a dominant role in rapid protection mediated by MVA. CD4+ and CD8+ T cells exert different effector functions to control infections. CD4+ T cells primarily activate other immune cells like B cells and macrophages through expression of cytokines [57] [58] [59] and they are also recognized as being crucially involved in the activation of antigen-specific CD8 + T cells [60] . In contrast, CD8+ T cells can directly kill infected cells which is mediated by the release of cytotoxic granules containing notably perforin and granzymes. Furthermore, studies showed that cell-mediated cytotoxicity and especially perforin is important for recovery from ECTV infection [61, 62] . To analyze the role of perforin mediated cellular cytotoxicity in rapid protection, we vaccinated perforin deficient mice (Prf 2/2 ) i.n. with 10 8 PFU MVA two days before a lethal challenge infection with ECTV. In contrast to wt mice, Prf 2/2 mice were not protected, developed severe disease and all mice succumbed to ECTV infection until day 18 post infection ( Figure 7A ). Correspondingly, we detected high levels of virus in lung and liver of MVA vaccinated Prf 2/2 mice at the time point of death, while vaccinated wt mice had cleared the virus at the end of the experiment (21 dpi) ( Figure 7B ). Nevertheless, livers of MVA immunized Prf 2/2 mice contained reduced virus loads in comparison to mock vaccinated Prf 2/2 controls. This observation appeared to correlate with the somewhat prolonged course of disease in vaccinated animals (not statistically significant) and might be the consequence of innate or humoral adaptive immune responses including the possible contribution of type I and/or type II interferons. Histopathologic examination revealed multiple randomly located foci of necrosis and inflammation in the liver of vaccinated and ECTV challenged Prf 2/2 mice ( Figure 7C , upper panel). These lesions were characterized by hepatocytic necrosis and infiltration by macrophages and lymphocytes. However, MVA immunized wt mice had no necrotic and inflammatory lesions in liver tissues ( Figure 7C , lower panel). Thus, the availability of the cytotoxic effector protein perforin was essential to maintain the protective capacity of MVA immunization suggesting the induction of T cell mediated cytotoxicity as key mechanism of protective immunity.