Apoptosis is essential for neutrophil functional shutdown and determines tissue damage in experimental pneumococcal meningitis.

During acute bacterial infections such as meningitis, neutrophils enter the tissue where they combat the infection before they undergo apoptosis and are taken up by macrophages. Neutrophils show pro-inflammatory activity and may contribute to tissue damage. In pneumococcal meningitis, neuronal damage despite adequate chemotherapy is a frequent clinical finding. This damage may be due to excessive neutrophil activity. We here show that transgenic expression of Bcl-2 in haematopoietic cells blocks the resolution of inflammation following antibiotic therapy in a mouse model of pneumococcal meningitis. The persistence of neutrophil brain infiltrates was accompanied by high levels of IL-1beta and G-CSF as well as reduced levels of anti-inflammatory TGF-beta. Significantly, Bcl-2-transgenic mice developed more severe disease that was dependent on neutrophils, characterized by pronounced vasogenic edema, vasculitis, brain haemorrhages and higher clinical scores. In vitro analysis of neutrophils demonstrated that apoptosis inhibition completely preserves neutrophil effector function and prevents internalization by macrophages. The inhibitor of cyclin-dependent kinases, roscovitine induced apoptosis in neutrophils in vitro and in vivo. In wild type mice treated with antibiotics, roscovitine significantly improved the resolution of the inflammation after pneumococcal infection and accelerated recovery. These results indicate that apoptosis is essential to turn off activated neutrophils and show that inflammatory activity and disease severity in a pyogenic infection can be modulated by targeting the apoptotic pathway in neutrophils.

Phagocytosis-induced apoptosis of macrophages is linked to uptake, killing and degradation of bacteria.

Phagocytosis and intracellular destruction of pathogens by phagocytes is a crucial defense mechanism of the innate immune response during infection. It has been reported a number of times that the interaction with pyogenic, extracellular bacteria leads to the apoptotic death of phagocytes. The signaling events that cause this form of cell death are largely unknown. In this study, we demonstrate a link between uptake, killing and degradation of Escherichia coli bacteria and induction of apoptosis in macrophages. Treatment of murine RAW 264.7 macrophages with bafilomycin A(1), a phagosome acidification inhibitor, reduced killing and degradation of phagocytosed bacteria and significantly decreased macrophage apoptosis. The stable overexpression of constitutively active or dominant-negative mutants of the small GTPase Rab5a increased bacterial phagocytosis and consecutively apoptosis. In these cells, relative killing and degradation were not affected, linking the increased apoptosis to enhanced uptake and suggesting that the apoptosis-inducing signal derives from the higher incidence of degradation events or an accumulation of phagosomes of a late maturation stage. These results thus provide a link between bacterial phagocytosis and degradation and the induction of apoptosis in macrophages. We propose that this form of apoptosis is the physiological conclusion of an innate immune response against pyogenic bacteria.

BimS-induced apoptosis requires mitochondrial localization but not interaction with anti-apoptotic Bcl-2 proteins.

Release of apoptogenic proteins such as cytochrome c from mitochondria is regulated by pro- and anti-apoptotic Bcl-2 family proteins, with pro-apoptotic BH3-only proteins activating Bax and Bak. Current models assume that apoptosis induction occurs via the binding and inactivation of anti-apoptotic Bcl-2 proteins by BH3-only proteins or by direct binding to Bax. Here, we analyze apoptosis induction by the BH3-only protein Bim(S). Regulated expression of Bim(S) in epithelial cells was followed by its rapid mitochondrial translocation and mitochondrial membrane insertion in the absence of detectable binding to anti-apoptotic Bcl-2 proteins. This caused mitochondrial recruitment and activation of Bax and apoptosis. Mutational analysis of Bim(S) showed that mitochondrial targeting, but not binding to Bcl-2 or Mcl-1, was required for apoptosis induction. In yeast, Bim(S) enhanced the killing activity of Bax in the absence of anti-apoptotic Bcl-2 proteins. Thus, cell death induction by a BH3-only protein can occur through a process that is independent of anti-apoptotic Bcl-2 proteins but requires mitochondrial targeting.

The IgLON family in epithelial ovarian cancer: expression profiles and clinicopathologic correlates.

PURPOSE: The IgLON family of cell adhesion molecules, comprising OPCML, HNT, LSAMP, and NEGR1, has recently been linked to cancer, through two of its members being proposed as tumor suppressors. We examined the expression profile of the family in human sporadic epithelial ovarian cancer and the normal ovary. EXPERIMENTAL DESIGN: We determined the expression level of each IgLON in a panel comprising 57 tumor and 11 normal ovarian samples by quantitative real-time reverse transcription-PCR. The results were statistically tested for associations with clinicopathologic variables. RESULTS: OPCML, LSAMP and NEGR1 exhibited reduced expression in the tumor samples relative to the normal samples, whereas HNT expression was elevated. Statistically significant changes were specific to histologic type. The expression levels of individual IgLONs were correlated, the most significant finding being a positive correlation between LSAMP and NEGR1. LSAMP expression was also negatively correlated with overall survival and was found to be a negative predictor of outcome. CONCLUSIONS: The expression of the IgLON family is altered in sporadic epithelial ovarian tumors in comparison to the normal ovary. In our small but representative cohort of patients, we have found significant correlations and associations in expression and clinicopathology that suggest a wider role of the family in ovarian cancer.

Protective T cell immunity against malaria liver stage after vaccination with live sporozoites under chloroquine treatment.

In this study we present the first systematic analysis of the immunity induced by normal Plasmodium yoelii sporozoites in mice. Immunization with sporozoites, which was conducted under chloroquine treatment to minimize the influence of blood stage parasites, induced a strong protection against a subsequent sporozoite and, to a lesser extent, against infected RBC challenges. The protection induced by this immunization protocol proved to be very effective. Induction of this protective immunity depended on the presence of liver stage parasites, as primaquine treatment concurrent with sporozoite immunization abrogated protection. Protection was not found to be mediated by the Abs elicited against pre-erythrocytic and blood stage parasites, as demonstrated by inhibition assays of sporozoite penetration or development in vitro and in vivo assays of sporozoite infectivity or blood stage parasite development. CD4(+) and CD8(+) T cells were, however, responsible for the protection through the induction of IFN-gamma and NO.