Adult-Like Anti-Mycobacterial T Cell and In Vivo Dendritic Cell Responses Following Neonatal Immunization with Ag85B-ESAT-6 in the IC31® Adjuvant

BACKGROUND: With the exception of some live vaccines, e.g. BCG, subunit vaccines formulated with "classical" adjuvants do not induce similar responses in neonates as in adults. The usual neonatal profile is characterized by lower levels of TH1-associated biomarkers. This has hampered the development of new neonatal vaccines for diseases that require early protection. Tuberculosis is one of the major targets for neonatal immunization. In this study, we assessed the immunogenicity of a novel candidate vaccine comprising a mycobacterial fusion protein, Ag85B-ESAT-6, in a neonatal murine immunization model. METHODS/FINDINGS: The Ag85B-ESAT-6 fusion protein was formulated either with a classical alum based adjuvant or with the novel IC31 adjuvant. Following neonatal or adult immunization, 3 parameters were studied in vivo: (1) CD4(+) T cell responses, (2) vaccine targeting/activation of dendritic cells (DC) and (3) protection in a surrogate mycobacterial challenge model. Conversely to Alum, IC31 induced in both age groups strong Th1 and Th17 responses, characterized by multifunctional T cells expressing IL-2 and TNF-alpha with or without IFN-gamma. In the draining lymph nodes, a similarly small number of DC contained the adjuvant and/or the antigen following neonatal or adult immunization. Expression of CD40, CD80, CD86 and IL-12p40 production was focused on the minute adjuvant-bearing DC population. Again, DC targeting/activation was similar in adults and neonates. These DC/T cell responses resulted in an equivalent reduction of bacterial growth following infection with M. bovis BCG, whereas no protection was observed when Alum was used as adjuvant. CONCLUSION: Neonatal immunization with the IC31-adjuvanted Ag85B-ESAT-6 subunit vaccine elicited adult-like multifunctional protective anti-mycobacterial T cell responses through the induction of an adult pattern of in vivo DC activation

Multikomponenten-Darstellung staubbildender Sternhüllen

In den kühlen, ausgedehnten Hüllen weitentwickelter Sterne auf dem asymptotischen Riesenast (AGB) bilden sich kleine Festkörper. Dieser Staub führt durch den auf ihn wirkenden Strahlungsdruck zu einem massiven Materiestrom bzw. Wind. Eine realistische Beschreibung dieses aus vielen chemischen Komponenten bestehenden Materiestroms erfordert die konsistente Behandlung der Hydrodynamik, der Staubbildung, des Strahlungstransports und der Chemie der Gasphase aller am Wind beteiligten Komponenten. Das langfristige Ziel, das dieser Arbeit zu Grunde liegt, ist ein Mehrkomponenten-Hydrodynamik-Modell, das den Materiestrom sowohl der einzelnen unterschiedlichen Staub bildenden Moleküle, als auch das der daraus gebildeten Staubteilchen mit den unterschiedlichen Größen adäquat beschreibt. In dieser Arbeit wird ein Teilaspekt behandelt, der die Wechselwirkung zwischen der Gasphase und dem Staubanteil im betrachteten Materiestrom untersucht. An Hand eines stationären, sphärisch symmetrischen Modells wird in einem ersten Schritt das Verhalten des Gleichungsystems betrachtet. Die Wechselwirkungen der Staub- mit der Gasphase werden durch Austauschterme dargestellt, die sich aus der detaillierten Darstellung der hydrodynamischen Gleichungen ergeben. Es werden die speziellen Anforderungen und die daraus resultierenden Schwierigkeiten für die numerische Behandlung des erweiterten Gleichungssystems aufgezeigt.The outer regions of the cool, extended shells of evolved stars on the asymptotic giant branch (AGB) are the source of small grains. Driven by radiation, the newly formed dust leads to a massive outflow, respectively wind from the star. In order to model this outflow in a realistic way, a consistent description of hydrodynamics, dust formation, radiative transfer, and chemistry of the gas phase with all involved components is needed. In the long term, the basic goal consists in a multicomponent model of hydrodynamics including all dust-forming molecules as well as their successor dust particles with individual size and composition. The part of the project exploring the interac- tions between gas and dust in the outflow is subject matter of this work. Based on a stationary model in spherical symmetry, the behaviour of the underlying system of equations is studied. Interactions between the gas and the dust phase are introduced by coupling terms derived from the detailed elaboration of the hydrodynamic equations. The specific conditions and the resulting difficulties for the numerical treatment of the extended system of equations are discussed

Fetal calf serum-free generation of functionally active murine dendritic cells suitable for in vivo therapeutic approaches

Standard protocols to generate mouse dendritic cells (DC) generally use culture medium supplemented with fetal calf serum; however, reinjection in vivo of DC cultured in fetal calf serum results in priming to xenogeneic proteins that clearly limits the use of such DC. We therefore established a fetal calf serum-free culture system for the generation of murine DC from bone marrow precursors. DC can be generated fetal calf serum-free using RPMI supplemented with 1.5% syngeneic mouse serum. Although the yield of DC grown under fetal calf serum-free conditions was somewhat lower than that of the standard culture, large numbers of DC could be generated without the exposure to xenogeneic proteins. The yield of fetal calf serum-free cultured DC was further enhanced by addition of the proinflammatory cytokines TNF-alpha and IL-1beta with the combination resulting in up to 10% more DC. Phenotypically, CD11c DC cultured fetal calf serum-free homogenously coexpressed the DC-specific molecule DEC-205 as well as the costimulatory molecules CD40, CD80, and CD86. In contrast, only a subpopulation of the CD11c DC cultured in fetal calf serum-containing medium coexpressed these molecules. Functionally, fetal calf serum-free DC showed strong stimulatory capacity for naive allogeneic CD4 and CD8 T cells. Importantly, fetal calf serum-free DC showed spontaneous in vivo migratory activity. Moreover, 5 x 105 subcutaneously injected TNBS-conjugated fetal calf serum-free DC were able to mediate contact sensitivity. Furthermore, the intravenous or subcutaneous injection of a single dose of 5 x 105 OVA-pulsed fetal calf serum-free DC resulted in the induction of an OVA-specific immune response in naive TCR transgenic animals. Thus DC cultured under fetal calf serum-free conditions are suitable instruments for in vivo therapeutic approaches, especially in autoimmune models. Keywords: DC vaccines/dendritic cell development/fetal calf serum-free culture conditions for DC/in vivo therapeutic DC approaches