Long-term elevation of complement factors in cerebrospinal fluid of patients with Borna disease virus 1 (BoDV-1) encephalitis.

BACKGROUND: Borna disease virus 1 (BoDV-1) causes rare but severe zoonotic infections in humans, presenting as severe encephalitis. The case-fatality risk is very high and no effective countermeasures have been established so far. An immunopathology is presumed, while data on immune responses in humans are limited. Evidence of a role of the complement system in various neurological disorders and central nervous viral infections is increasing and specific inhibitors are available as therapeutic options. METHODS: In this study, we investigated factors of the complement system in the cerebrospinal fluid (CSF) of patients with BoDV-1 infections (n = 17) in comparison to non-inflammatory control CSF samples (n = 11), using a bead-based multiplex assay. In addition, immunohistochemistry was performed using post-mortem brain tissue samples. RESULTS: We found an intrathecal elevation of complement factors of all complement pathways and an active cascade during human BoDV-1 infections. The increase of certain complement factors such as C1q was persistent and C3 complement deposits were detected in post-mortem brain sections. Intrathecal complement levels were negatively correlated with survival. CONCLUSION: Further investigations are warranted to clarify, whether targeting the complement cascade by specific inhibitors might be beneficial for patients suffering from severe BoDV-1 encephalitis.

Enzyme-triggered antigen release enhances cross-presentation by dendritic cells.

Nanoparticles hold great potential as vaccine carriers due to their highly versatile structure and the possibility to influence intracellular trafficking and antigen presentation by their design. In this study, we developed a nanoparticulate system with a new enzyme-triggered antigen release mechanism. For this novel approach, nanoparticle and model antigen ovalbumin were linked with a substrate of the early endosomal protease cathepsin S. This construct enabled the transfer of antigens delivered to bone marrow-derived dendritic cells from the endo-lysosomal compartments in the cytosol. Consecutively, our particles enhanced cross-presentation on dendritic cells and subsequently promoted a stronger activation of CD8+ T cells. Our findings suggest that enzyme-triggered antigen release allows the endosomal escape of the antigen, leading to increased MHC-I presentation. Since T cell immunity is central for the control of viral infections and cancer, this release mechanism offers a promising approach for the development of both prophylactic and therapeutic vaccines.

A Polymorphism of Bactericidal/Permeability-Increasing Protein Affects Its Neutralization Efficiency towards Lipopolysaccharide.

Gram-negative sepsis driven by lipopolysaccharide (LPS) has detrimental outcomes, especially in neonates. The neutrophil-derived bactericidal/permeability-increasing protein (BPI) potently neutralizes LPS. Interestingly, polymorphism of the BPI gene at position 645 (rs4358188) corresponds to a favorable survival rate of these patients in the presence of at least one allele 645 A as opposed to 645 G. When we exploited the existing X-ray crystal structure, the corresponding amino acid at position 216 was revealed as surface exposed and proximal to the lipid-binding pocket in the N-terminal domain of BPI. Our further analysis predicted a shift in surface electrostatics by a positively charged lysine (BPI216K) exchanging a negatively charged glutamic acid (BPI216E). To investigate differences in interaction with LPS, we expressed both BPI variants recombinantly. The amino acid exchange neither affected affinity towards LPS nor altered bactericidal activity. However, when stimulating human peripheral blood mononuclear cells, BPI216K exhibited a superior LPS-neutralizing capacity (IC50 12.0 ± 2.5 pM) as compared to BPI216E (IC50 152.9 ± 113.4 pM, p = 0.0081) in respect to IL-6 secretion. In conclusion, we provide a functional correlate to a favorable outcome of sepsis in the presence of BPI216K.

Bactericidal/permeability-increasing protein instructs dendritic cells to elicit Th22 cell response.

Neutrophil-derived bactericidal/permeability-increasing protein (BPI) is known for its bactericidal activity against gram-negative bacteria and neutralization of lipopolysaccharide. Here, we define BPI as a potent activator of murine dendritic cells (DCs). As shown in GM-CSF-cultured, bone-marrow-derived cells (BMDCs), BPI induces a distinct stimulation profile including IL-2, IL-6, and tumor necrosis factor expression. Conventional DCs also respond to BPI, while M-CSF-cultivated or peritoneal lavage macrophages do not. Subsequent to BPI stimulation of BMDCs, CD4+ T cells predominantly secrete IL-22 and, when naive, preferentially differentiate into T helper 22 (Th22) cells. Congruent with the tissue-protective properties of IL-22 and along with impaired IL-22 induction, disease severity is significantly increased during dextran sodium sulfate-induced colitis in BPI-deficient mice. Importantly, physiological diversification of intestinal microbiota fosters BPI-dependent IL-22 induction in CD4+ T cells derived from mesenteric lymph nodes. In conclusion, BPI is a potent activator of DCs and consecutive Th22 cell differentiation with substantial relevance in intestinal homeostasis.

Scorpionfish BPI is highly active against multiple drug-resistant Pseudomonas aeruginosa isolates from people with cystic fibrosis.

Chronic pulmonary infection is a hallmark of cystic fibrosis (CF) and requires continuous antibiotic treatment. In this context, Pseudomonas aeruginosa (Pa) is of special concern since colonizing strains frequently acquire multiple drug resistance (MDR). Bactericidal/permeability-increasing protein (BPI) is a neutrophil-derived, endogenous protein with high bactericidal potency against Gram-negative bacteria. However, a significant range of people with CF (PwCF) produce anti-neutrophil cytoplasmic antibodies against BPI (BPI-ANCA), thereby neutralizing its bactericidal function. In accordance with literature, we describe that 51.0% of a total of 39 PwCF expressed BPI-ANCA. Importantly, an orthologous protein to human BPI (huBPI) derived from the scorpionfish Sebastes schlegelii (scoBPI) completely escaped recognition by these autoantibodies. Moreover, scoBPI exhibited high anti-inflammatory potency towards Pa LPS and was bactericidal against MDR Pa derived from PwCF at nanomolar concentrations. In conclusion, our results highlight the potential of highly active orthologous proteins of huBPI in treatment of MDR Pa infections, especially in the presence of BPI-ANCA.

Cystic fibrosis is a genetic disorder that makes people produce unusually thick and sticky mucus that clogs their lungs and airways. This inevitably leads to recurring bacterial infections, particularly those caused by the Gram-negative bacterium Pseudomonas aeruginosa. Antibiotics are needed to treat these infections. However, over time most bacteria build modes of resistance to these drugs and, once multiple drug-resistant bacteria colonize the lung, very limited treatment options are left. Therefore, new therapeutic approaches are desperately needed. Notably, humans themselves express a highly potent antimicrobial protein called BPI (short for Bactericidal/permeability­increasing protein) that attacks Gram-negative bacteria, including multiple drug-resistant strains of P. aeruginosa. Unfortunately, many people with cystic fibrosis also generate antibodies that bind to BPI and interfere with its antimicrobial function. Faced with this conundrum, Holzinger et al. set out to find BPIs made by other animals which might not be recognized by human antibodies and also display a high potential to attack Gram-negative bacteria. Based on specific selection criteria, Holzinger et al. focused their attention on BPI made by scorpionfish, a type of venomous fish that live near coral reefs. Compared to other BPI proteins they investigated, the one produced by scorpionfish appeared to be the most capable of binding to P. aeruginosa via a prominent surface molecule exclusively found on Gram-negative bacteria. Furthermore, when Holzinger et al. tested whether the antibodies present in people with cystic fibrosis could recognize scorpionfish BPI, they found that the BPI completely evaded detection. The scorpionfish BPI was also able to pre-eminently attack P. aeruginosa. In fact, it was even able to potently kill drug-resistant strains of the bacteria that had been isolated from people with cystic fibrosis. This study suggests that scorpionfish BPI could serve as an alternative to antibiotics in people with cystic fibrosis that have otherwise untreatable bacterial infections. Drug-resistant bacteria which cause life threatening conditions are on the rise across the globe, and scorpionfish BPI could be a potential candidate to treat affected patients. In the future, animal experiments will be needed to explore how highly potent non-human BPIs function in whole living organisms.

Low Intestinal IL22 Associates With Increased Transplant-Related Mortality After Allogeneic Stem Cell Transplantation.

The role of IL-22 in adult patients undergoing allogeneic stem cell transplantation (SCT) is of major interest since animal studies showed a protective and regenerative effect of IL-22 in graft versus host disease (GvHD). However, no clinical data exist on the tissue expression. Here we demonstrate that patients not suffering from transplant-related mortality (TRM) show significantly upregulated IL22 expression during histological and clinical GI-GvHD (p = 0.048 and p = 0.022, respectively). In contrast, in GvHD patients suffering from TRM, IL22 was significantly lower (p = 0.007). Accordingly, lower IL22 was associated with a higher probability of TRM in survival analysis (p = 0.005). In a multivariable competing risk Cox regression analysis, low IL22 was identified as an independent risk factor for TRM (p = 0.007, hazard ratio 2.72, 95% CI 1.32 to 5.61). The expression of IL22 seemed to be microbiota dependent as broad-spectrum antibiotics significantly diminished IL22 expression (p = 0.019). Furthermore, IL22 expression significantly correlated with G-protein coupled receptor (GPR)43 (r = 0.263, p = 0.015) and GPR41 expression (r = 0.284, p = 0.009). In conclusion, our findings reveal an essential role of IL-22 for the prognosis of patients undergoing allogeneic SCT.

Lipopolysaccharide Binding Protein and Bactericidal/Permeability-Increasing Protein as Biomarkers for Invasive Pulmonary Aspergillosis.

Early diagnosis of invasive pulmonary aspergillosis (IPA) is crucial to prevent lethal disease in immunocompromized hosts. So far, lipopolysaccharide binding protein (LBP) and bactericidal/permeability-increasing protein (BPI) levels have not been evaluated as biomarkers for IPA. IL-8, previously introduced as a biomarker for IPA, was also included in this study. Bronchoalveolar lavage fluid (BALF) of IPA patients and control patients with non-infectious lung disease was collected according to clinical indications. Measurements in BALF displayed significantly higher levels of LBP (p < 0.0001), BPI (p = 0.0002) and IL-8 (p < 0.0001) in IPA compared to control patients. Receiver operating characteristic curve analysis revealed higher AUC for LBP (0.98, 95% CI 0.95-1.00) than BPI (0.84, 95% CI 0.70-0.97; p = 0.0301). Although not significantly different, AUC of IL-8 (0.93, 95% CI 0.85-1.00) also tended to be higher than AUC for BPI (p = 0.0624). When the subgroup of non-hematological patients was analyzed, test performance of LBP (AUC 0.99, 95% CI 0.97-1.00), BPI (AUC 0.97, 95% CI 0.91-1.00) and IL-8 (AUC 0.96, 95% CI: 0.90-1.00) converged. In conclusion, LBP and-to a lesser extend-BPI displayed high AUCs that were comparable to those of IL-8 for diagnosis of IPA in BALF. Further investigations are worthwhile, especially in non-hematological patients in whom sensitive biomarkers for IPA are lacking.

Bactericidal/Permeability-Increasing Protein Is an Enhancer of Bacterial Lipoprotein Recognition.

Adequate perception of immunologically important pathogen-associated molecular patterns like lipopolysaccharide and bacterial lipoproteins is essential for efficient innate and adaptive immune responses. In the context of Gram-negative infection, bactericidal/permeability-increasing protein (BPI) neutralizes endotoxic activity of lipopolysaccharides, and thus prohibits hyperactivation. So far, no immunological function of BPI has been described in Gram-positive infections. Here, we show a significant elevation of BPI in Gram-positive meningitis and, surprisingly, a positive correlation between BPI and pro-inflammatory markers like TNFα. To clarify the underlying mechanisms, we identify BPI ligands of Gram-positive origin, specifically bacterial lipopeptides and lipoteichoic acids, and determine essential structural motifs for this interaction. Importantly, the interaction of BPI with these newly defined ligands significantly enhances the immune response in peripheral blood mononuclear cells (PBMCs) mediated by Gram-positive bacteria, and thereby ensures their sensitive perception. In conclusion, we define BPI as an immune enhancing pattern recognition molecule in Gram-positive infections.

CpG and LPS can interfere negatively with prion clearance in macrophage and microglial cells.

Cells of the innate immune system play important roles in the progression of prion disease after peripheral infection. It has been found in vivo and in vitro that the expression of the cellular prion protein (PrP(c)) is up-regulated on stimulation of immune cells, also indicating the functional importance of PrP(c) in the immune system. The aim of our study was to investigate the impact of cytosine-phosphate-guanosine- and lipopolysaccharide-induced PrP(c) up-regulation on the uptake and processing of the pathological prion protein (PrP(Sc)) in phagocytic innate immune cells. For this purpose, we challenged the macrophage cell line J774, the microglial cell line BV-2 and primary bone marrow-derived macrophages in a resting or stimulated state with various prion strains, and monitored the uptake and clearance of PrP(Sc). Interestingly, stimulation led either to a transient increase in the level of PrP(Sc) relative to unstimulated cells or to a decelerated degradation of PrP(Sc). These features were dependent on cell type and prion strain. Our data indicate that the stimulation of innate immune cells may be able to support transient prion propagation, possibly explained by an increased PrP(c) cell surface expression in stimulated cells. We suggest that stimulation of innate immune cells can lead to an imbalance between the propagation and degradation of PrP(Sc).