For in vivo effector priming, 4 106 CFSE (Invitrogen) labeled CD4+ T cells were resuspended in 100 L of PBS (Dibco) and injected intravenously into B6 mice
June 14, 2021
For in vivo effector priming, 4 106 CFSE (Invitrogen) labeled CD4+ T cells were resuspended in 100 L of PBS (Dibco) and injected intravenously into B6 mice. antibody response to PI-LPS, similar antibody profiles were observed but IgG titers were significantly higher after vaccination with PI-WCV. Furthermore, higher frequency of antigen-specific CD4+ T cells was detected in mice immunized with PI-WCV. PI-WCVCstimulated DCs displayed significantly higher levels of CCR7 and migratory ability to secondary lymphoid organs. Challenge-protection studies in wild-type and CCR7-deficient mice confirmed that CCR7 is critical for PI-WCVCinduced cellular immunity. Conclusions PI-WVC stimulates protective immunity to in mice through stimulation of migratory behavior in DCs for protective cellular immunity. Additionally, the humoral immune response to LPS is an important component of protective immunity. is a facultative, gram-negative intracellular bacteria and etiological agent of the aerosol-transmitted zoonotic disease, Q fever . The high infectivity of and its hardiness in severe environments has raised concerns that it could be used as a biowarfare agent [2, 3]. Therefore, the development of a safe and efficacious vaccine is warranted. lipopolysaccharide (LPS) undergoes a phase variation where virulent phase I (PI, Nine Mile strain, NMI) converts to avirulent phase II (PII, NMII) under specific conditions due to an irreversible mutation [4C7]. This phase variation in is similar to that observed in enterobacteria like that transition from smooth to rough LPS variants. NMI organisms have smooth LPS with complete O-antigen, while phase II have a rough-type LPS, missing the branched-chain sugars virenose and di-hydrohydroxystreptose . Interestingly, LPS was the first virulence factor to be defined, where PI are able Auristatin E to cause infection in guinea pigs and mice but PII are not . Vaccines from inactivated PI and PII called whole-cell vaccines (PI-WCV and PII-WCV), have been developed and tested in animal models and humans [10C13]. PI-WCV confers protection against NMI challenge Auristatin E in guinea pig and mouse models, whereas PII-WCV does not [14, 15]. A commercial form of PI-WCV (Q-Vax; Commonwealth Serum Laboratories), has shown an extraordinary ability to prevent human Q fever and has been licensed for use in Australia . Unfortunately, immunization with this DUSP2 vaccine can result in severe local or systemic adverse reactions, especially when administered to previously sensitized individuals, and therefore is not licensed for use in the United States [17, 18]. Early mouse studies suggested that PI-WCV induces both humoral and cell-mediated immune responses and adoptive transfer of either sera or T cells conferred protection against infection in immune-competent Auristatin E mice [14, 19C23]. Furthermore, evaluation of protective responses in mice suggests that antibodies play an important role in preventing the development of clinical disease, whereas T-cell mediated immunity is required for clearance of . The importance of T cells to vaccine-induced immunity is further highlighted by the lack of protection after passive transfer of immune sera to athymic or severe combined immune deficiency (SCID) mice, indicating that T cells are required for antibody-mediated protection [14, 24]. Although PI-WVC is protective in mice, PII-WVC is not and the underlying cause for why PI-WCV and PII-WCV differ so dramatically in their ability to confer protective immunity is largely unknown. Conventional thought is that PI and PII share highly similar antigen contents with the exception of their unique LPS. Indeed, previous studies have highlighted the importance of antibodies against PI-LPS in mediating protection in mice, where vaccination with PI-LPS confers similar levels of protection in mice to PI-WCV . There is a great interest in deciphering the underlying mechanistic differences in PI-WCV and PII-WCVCbased immunity as the PII organism is classified as biosafety level 2 and therefore it would be much more economical to create a vaccine from the exempt BL2 strain than from the select agent virulent Nine Mile phase I (RSA 493) and Nine Mile phase II (RSA 439) were grown in embryonated chicken eggs, purified by gradient centrifugation, and inactivated by.