Stephanie Verschuere

β-Glucan microparticles are good candidates for mucosal antigen delivery in oral vaccination

Continuously improving the developmental process and the efficacy of oral vaccines is essential in the fight against intestinal pathogens. A promising strategy for vaccination applying safe, biodegradable and non-replicating antigen delivery systems has gained increased interest for eliciting cellular and humoral immune responses. The current study evaluates the potential of β-glucan particles (GP) as an oral antigen delivery system and their adjuvant characteristics. GP are efficiently internalized by human intestinal epithelial cell lines (Caco-2 and HT-29 cells), without exerting negative effects on cell viability. GP triggered the expression of pro-inflammatory cytokines IL-23p19, IL-8 and the β-glucan receptors dectin-1 and TLR2 by activated Caco-2 cells, and CCL20 in HT-29 cells. In contrast, the expression level of TGF-β, an important mediator of oral tolerance, was significantly downregulated in HT-29 cells. Additionally, adoptive transfer experiments showed proliferating ovalbumin (OVA)-specific CD4(+) T cells mainly in the spleens of GP-OVA-fed mice. Furthermore, we detected a significantly increased IL-17 and a trend towards increased IFN-γ production in the spleen of GP-OVA-fed mice upon antigen restimulation. Oral administration of GP-OVA induced increased OVA-specific IgA, secretory-IgA (S-IgA) and secretory component (SC) production in intestinal fluids. Our data show that GP vehicles are able to deliver OVA via an oral route allowing efficient antigen presentation alongside adaptive immune activation, resulting in a Th17-biased response and the production of OVA-specific IgA, secretory-IgA and secretory component antibodies.

Biosupported bimetallic Pd-Au nanocatalysts for dechlorination of environmental contaminants

Biologically produced monometallic palladium nanoparticles (bio-Pd) have been shown to catalyze the dehalogenation of environmental contaminants, but fail to efficiently catalyze the degradation of other important recalcitrant halogenated compounds. This study represents the first report of biologically produced bimetallic Pd/Au nanoparticle catalysts. The obtained catalysts were tested for the dechlorination of diclofenac and trichlorethylene. When aqueous bivalent Pd(II) and trivalent Au(III) ions were both added to concentrations of 50 mg L(-1) and reduced simultaneously by Shewanella oneidensis in the presence of H(2), the resulting cell-associated bimetallic nanoparticles (bio-Pd/Au) were able to dehalogenate 78% of the initially added diclofenac after 24 h; in comparison, no dehalogenation was observed using monometallic bio-Pd or bio-Au. Other catalyst-synthesis strategies did not show improved dehalogenation of TCE and diclofenac compared with bio-Pd. Synchrotron-based X-ray diffraction, (scanning) transmission electron microscopy and energy dispersive X-ray spectroscopy indicated that the simultaneous reduction of Pd and Au supported on cells of S. oneidensis resulted in the formation of a unique bimetallic crystalline structure. This study demonstrates that the catalytic activity and functionality of possibly environmentally more benign biosupported Pd-catalysts can be improved by coprecipitation with Au.

Chronic cigarette smoke exposure induces microbial and inflammatory shifts and mucin changes in the murine gut

Inflammatory bowel diseases (IBD) are complex multifactorial diseases characterized by an inappropriate host response to an altered commensal microbiome and dysfunctional mucus barrier. Cigarette smoking is the best known environmental risk factor in IBD. Here, we studied the influence of chronic smoke exposure on the gut microbiome, mucus layer composition and immune factors in conventional mice. We compared smoke-exposed with air-exposed mice (n = 12) after a smoke exposure of 24 weeks. Both Illumina sequencing (n = 6) and denaturing gradient gel electrophoresis (n = 12) showed that bacterial activity and community structure were significantly altered in the colon due to smoke exposure. Interestingly, an increase of Lachnospiraceae sp. activity in the colon was observed. Also, the mRNA expression of Muc2 and Muc3 increased in the ileum, whereas Muc4 increased in the distal colon of smoke-exposed mice (n = 6). Furthermore, we observed increased Cxcl2 and decreased Ifn-γ in the ileum, and increased Il-6 and decreased Tgf-β in the proximal colon. Tight junction gene expression remained unchanged. We infer that the modulating role of chronic smoke exposure as a latently present risk factor in the gut may be driven by the altered epithelial mucus profiles and changes in microbiome composition and immune factors.

Cigarette smoking alters epithelial apoptosis and immune composition in murine GALT

Smokers have a twofold increased risk to develop Crohns disease (CD). However, little is known about the mechanisms through which smoking affects CD pathogenesis. Especially Crohns ileitis is negatively influenced by smoking. Interestingly, the ileum and, more in particular, the Peyers patches in the terminal ileum are also the sites where the first CD lesions are found. Several chemokines are implicated in the pathogenesis, among which is the CCL20-CCR6 pathway. Here, we studied the gut-associated lymphoid tissue in C57BL/6 wild-type mice and in CCR6-deficient mice after exposure to air or cigarette smoke for 24 weeks. Apoptotic index of the follicle-associated epithelium overlying the Peyers patches was evaluated. We found that chronic smoke exposure induced apoptosis in the follicle-associated epithelium. Furthermore, immune cell numbers and differentiation along with chemokine expression were determined in Peyers patches. Important changes in immune cell composition were observed: total dendritic cells, CD4+ T cells (including regulatory T cells) and CD8+ T cells increased significantly after smoke exposure. The CD11b+ dendritic cell subset almost doubled. Interestingly, these changes were accompanied by an upregulated mRNA expression of the chemokines CCL9 and CCL20. However, no differences in the increase of dendritic cells were observed between wild-type and CCR6-deficient mice. Our results show that cigarette smoke exposure increases apoptosis in the follicle-associated epithelium and is associated with immune cell accumulation in Peyers patches.

The effect of smoking on intestinal inflammation: What can be learned from animal models?

Epidemiological evidence demonstrates that smoking is the most important environmental risk factor in Crohns disease while it positively interferes with the disease course of ulcerative colitis. However, the underlying mechanisms through which smoking exerts this divergent effect and affects pathogenesis of inflammatory bowel disease are largely unknown. Animal smoke models are good models to investigate the impact of cigarette smoke on intestinal physiology and inflammation. They enable one to explore the interaction of smoke components and the gut on cellular and molecular level, clarifying how smoking interferes with normal gut function and with disease course in inflammatory conditions. This review describes the currently used animal models for studying the impact of cigarette smoke on the intestinal tract. We first discuss the different methods for simulation of smoking. Furthermore, we focus on the effect of smoke exposure on normal gut physiology and immunology, on experimental (entero)colitis, and on inflammation-induced neoplasia. Based on this current knowledge, a hypothesis is formulated about the mechanisms through which cigarette smoke interferes with the gut in normal and pathological conditions.

Biogenic nanopalladium based remediation of chlorinated hydrocarbons in marine environments.

Biogenic catalysts have been studied over the last 10 years in freshwater and soil environments, but neither their formation nor their application has been explored in marine ecosystems. The objective of this study was to develop a biogenic nanopalladium-based remediation method for reducing chlorinated hydrocarbons from marine environments by employing indigenous marine bacteria. Thirty facultative aerobic marine strains were isolated from two contaminated sites, the Lagoon of Mar Chica, Morocco, and Priolo Gargallo Syracuse, Italy. Eight strains showed concurrent palladium precipitation and biohydrogen production. X-ray diffraction and thin section transmission electron microscopy analysis indicated the presence of metallic Pd nanoparticles of various sizes (5-20 nm) formed either in the cytoplasm, in the periplasmic space, or extracellularly. These biogenic catalysts were used to dechlorinate trichloroethylene in simulated marine environments. Complete dehalogenation of 20 mg L(-1) trichloroethylene was achieved within 1 h using 50 mg L(-1) biogenic nanopalladium. These biogenic nanoparticles are promising developments for future marine bioremediation applications.

Cigarette smoke and the terminal ileum: increased autophagy in murine follicle-associated epithelium and Peyer’s patches

Cigarette smoke (CS) exposure is associated with increased autophagy in several cell types, such as bronchial epithelial cells. Smoking is also an environmental risk factor in Crohn’s disease, in which impairment of the autophagy-mediated anti-bacterial pathway has been implicated. So far, it is unknown whether CS induces autophagy in the gut. Here, we examined the effect of chronic CS exposure on autophagy in the follicle-associated epithelium (FAE) of murine Peyer’s patches. Transmission electron microscopy revealed that the proportion of cell area occupied by autophagic vesicles significantly increased in the FAE after CS exposure. An increased number of autophagic vesicles was observed in the FAE, whereas the vesicle size remained unaltered. Besides enterocytes, also M-cells contain more autophagic vesicles upon CS exposure. In addition, the mRNA level of the autophagy-related protein Atg7 in the underlying Peyer’s patches is increased after CS exposure, which indicates that the autophagy-inducing effect of CS is not limited to the FAE. In conclusion, our results demonstrate that CS exposure induces autophagy in murine FAE and in the underlying immune cells of Peyer’s patches, suggesting that CS exposure increases the risk for Crohn’s disease by causing epithelial oxidative damage, which needs to be repaired by autophagy.

Spray-dried polyelectrolyte microparticles in oral antigen delivery: stability, biocompatibility, and cellular uptake.

During the past decade, extensive research has undeniably improved the formulation and delivery of oral vaccines. Nevertheless, several factors, such as the harsh gastrointestinal environment together with tolerance induction to exogenous antigens, have thus far impeded the optimal effectiveness and clinical application of oral delivery systems. The current study encompasses an initial evaluation of the stability, biocompatibility, and cellular uptake of two promising candidate systems for oral antigen delivery, that is, calcium carbonate- (CP) and mannitol-templated (MP) porous microspheres. Both spray-dried formulations were efficiently internalized by human intestinal epithelial cells (Caco-2 and HT-29) and degraded into phagolysosomal intracellular compartments. In addition, cellular particle uptake and processing significantly up-regulated the expression of (HLA) class-II and costimulatory molecules on intestinal epithelial cells. Even though the high surface-area-to-volume ratio of the microspheres was expected to favor protease access, antigen release was remarkably limited in simulated intestinal fluid and was even absent under gastric conditions. Finally, neither CP nor MP exerted cytotoxicity upon prolonged in vitro incubation with high antigen concentration. Altogether, these data support the potential of CP and MP for oral antigen delivery and motivate the further development of these promising carrier systems in in vivo studies.

Agreement of Gleason Score on Prostate Biopsy and Radical Prostatectomy Specimen: Is There Improvement With Increased Number of Biopsy Cylinders and the 2005 Revised Gleason Scoring?

INTRODUCTION The objectives of this study were to assess the agreement of GS on biopsy compared with RP specimens and to assess whether an increased number of biopsy cylinders and the 2005 International Society of Urological Pathology (ISUP) GS modification improved this agreement. MATERIALS AND METHODS Pathological data of biopsy and RP specimens were analyzed in 328 consecutive patients, before (group 1; n = 135) and after (group 2; n = 193) implementation of the 2005 ISUP modification. Additionally, patients had more biopsy cylinders taken in group 2 (mean 10 vs. 6.9). The agreement of GS between biopsy and RP specimens was evaluated using the kappa coefficient. GS was pooled into 3 grades: low- (GS ≤ 6), intermediate- (GS = 7), and high-grade (GS ≥ 8) prostate cancer. RESULTS Kappa coefficient for GS in group 1 and 2 was 0.261 and 0.341, respectively. For tumor grade, this was 0.308 and 0.359 for group 1 and 2, respectively. For RP specimens, there was more agreement between biopsy and RP GS in group 2 compared with group 1 (53.9% vs. 37.8%). Upgrading was almost exclusively (89.5%) seen in patients with biopsy GS ≤ 6 and was lower in group 2 (25.4% vs. 48.1%) because of classification of more intermediate- and high-grade tumors using the 2005 ISUP modification. Taking > 6 biopsy cylinders was associated with better GS and tumor grade agreement. CONCLUSION Extended biopsy template and the 2005 ISUP modification resulted in an improved agreement between biopsy GS and RP GS and a shift toward more aggressive tumors.

Contemporary minimally-invasive extended pelvic lymph node dissection for prostate cancer before curative radiotherapy: Prospective standardized analysis of complications

To prospectively assess contemporary complication rates of minimally‐invasive staging pelvic lymph node dissection before curative radiotherapy for prostate cancer using a standardized classification.