Osteopontin Deficiency Accelerates Spontaneous Colitis in Mice with Disrupted Gut Microbiota and Macrophage Phagocytic Activity

Background

Osteopontin (OPN) is a multifunctional protein expressed in a variety of tissues and cells. Recent studies revealed increased OPN expression in the inflamed intestinal tissues of patients with inflammatory bowel disease (IBD). The role of OPN in the pathophysiology of IBD, however, remains unclear.

Aims

To investigate the role of OPN in the development of intestinal inflammation using a murine model of IBD, interleukin-10 knock out (IL-10 KO) mice.

Methods

We compared the development of colitis between IL-10 KO and OPN/IL-10 double KO (DKO) mice. OPN expression in the colonic tissues of IL-10 KO mice was examined by fluorescence in situ hybridization (FISH) analysis. Enteric microbiota were compared between IL-10 KO and OPN/IL-10 DKO mice by terminal restriction fragment length polymorphism analysis. The effect of OPN on macrophage phagocytic function was evaluated by phagocytosis assay.

Results

OPN/IL-10 DKO mice had an accelerated onset of colitis compared to IL-10 KO mice. FISH analysis revealed enhanced OPN synthesis in the colonic epithelial cells of IL-10 KO mice. OPN/IL-10 DKO mice had a distinctly different enteric bacterial profile with a significantly lower abundance of Clostridium subcluster XIVa and a greater abundance of Clostridium cluster XVIII compared to IL-10 KO mice. Intracellular OPN deletion in macrophages impaired phagocytosis of fluorescence particle-conjugated Escherichia coli in vitro. Exogenous OPN enhanced phagocytosis by OPN-deleted macrophages when administered at doses of 1 to 100 ng/ml, but not 1000 ng/ml.

Conclusions

OPN deficiency accelerated the spontaneous development of colitis in mice with disrupted gut microbiota and macrophage phagocytic activity.     

Baseline Assessment of Mesophotic Reefs of the Vitória-Trindade Seamount Chain Based on Water Quality,Microbial Diversity,Benthic Cover and Fish Biomass Data

Seamounts are considered important sources of biodiversity and minerals. However, their biodiversity and health status are not well understood; therefore, potential conservation problems are unknown. The mesophotic reefs of the Vitória-Trindade Seamount Chain (VTC) were investigated via benthic community and fish surveys, metagenomic and water chemistry analyses, and water microbial abundance estimations. The VTC is a mosaic of reef systems and includes fleshy algae dominated rhodolith beds, crustose coralline algae (CCA) reefs, and turf algae dominated rocky reefs of varying health levels. Macro-carnivores and larger fish presented higher biomass at the CCA reefs (4.4 kg per frame) than in the rhodolith beds and rocky reefs (0.0 to 0.1 kg per frame). A larger number of metagenomic sequences identified as primary producers (e.g., Chlorophyta and Streptophyta) were found at the CCA reefs. However, the rocky reefs contained more diseased corals (>90%) than the CCA reefs (~40%) and rhodolith beds (~10%). Metagenomic analyses indicated a heterotrophic and fast-growing microbiome in rocky reef corals that may possibly lead to unhealthy conditions possibly enhanced by environmental features (e.g. light stress and high loads of labile dissolved organic carbon). VTC mounts represent important hotspots of biodiversity that deserve further conservation actions.     

A novel factor FLOURY ENDOSPERM2 is involved in regulation of rice grain size and starch quality

Rice (Oryza sativa) endosperm accumulates a massive amount of storage starch and storage proteins during seed development. However, little is known about the regulatory system involved in the production of storage substances. The rice flo2 mutation resulted in reduced grain size and starch quality. Map-based cloning identified FLOURY ENDOSPERM2 (FLO2), a member of a novel gene family conserved in plants, as the gene responsible for the rice flo2 mutation. FLO2 harbors a tetratricopeptide repeat motif, considered to mediate a protein-protein interactions. FLO2 was abundantly expressed in developing seeds coincident with production of storage starch and protein, as well as in leaves, while abundant expression of its homologs was observed only in leaves. The flo2 mutation decreased expression of genes involved in production of storage starch and storage proteins in the endosperm. Differences between cultivars in their responsiveness of FLO2 expression during high-temperature stress indicated that FLO2 may be involved in heat tolerance during seed development. Overexpression of FLO2 enlarged the size of grains significantly. These results suggest that FLO2 plays a pivotal regulatory role in rice grain size and starch quality by affecting storage substance accumulation in the endosperm.     

Alterations in hepatitis B virus nucleotide sequences in a chronic virus carrier from immunotolerant to immunoactive phase

Factors involved in transition from the immunotolerant to immunoactive phase in chronic hepatitis B virus (HBV) infection remain unclear. We investigated viral mutations occurring during transition and elucidated their virological and immunological significance. Full-length HBV DNA sequences were serially determined in a chronic HBV carrier from the immunotolerant to immunoactive phase. Viral replicative competence was examined by transfection analysis. HBV-specific CD8⁺ T cell response was evaluated by coculture of CD8⁺ T cells with autologous dendritic cells followed by interferon-γ Elispot assay. Eleven point mutations and two deletions appeared around the onset of the immunoactive phase. Viral replicative competence declined significantly after the onset of active hepatitis. Examination of the CD8⁺ T cell response against two putative T-cell epitopes, which contained substituted amino acids from the immunotolerant to immunoactive phase, showed that mutant HBV epitopes gave a lesser T cell response than wild-type HBV ones. In summary, point mutations and deletions may occur prior to or concurrent with the onset of the immunoactive phase during chronic HBV infection. These mutations may result in a significant decrease in both viral replicative competence and HBV-specific CD8⁺ T cell response, suggesting a possible adaptation for the maintenance of viral persistence.     

Highly efficient rescue of dengue virus using a co-culture system with mosquito/mammalian cells

The production rate of dengue viruses (DENVs), especially low-passage virus isolates, is low, and, therefore, the isolates are generally used only after several passages. However, in vitro passages could induce mutation(s). In this study, we established a system for the characterization of low-passage viral isolates using an infectious cDNA clone. We used R05-624, a plaque derived from type 2 (DENV-2) Thai strain, for the construction of the cDNA clone, named pmMW/R05-624. We found that transfection of both of mammalian Vero cells and mosquito C6/36 cells with viral RNA derived from the cDNA clone produced a significant amount of progeny virus: 3.2 × 10⁶ focus-forming units (FFU) production per ml of cultured fluid only 3 days after transfection with 2 μg RNA. Conversely, no detectable level of viruses was produced by conventional methods using a single cell line, Vero or C6/36. When this system was applied for the characterization of eight low-passage clinical viral isolates by placing their 5′-half or 3′-half in the above cDNA clone, we found that all the isolates, except for L04-225, produced similar levels of progeny virus. Among a total of eight cDNA clones reconstructed with the NS4A-3′NCR region derived from L04-225, one clone carried an insertion and produced a low level of progeny virus. Thus, our system to efficiently rescue clinical samples or low-passage viral isolates could be useful for assessing the virological and molecular characteristics of DENV that could be related to disease pathogenesis.     

Extended gene map reveals tripartite motif, C-type lectin, and Ig superfamily type genes within a subregion of the chicken MHC-B affecting infectious disease

MHC haplotypes have a remarkable influence on whether tumors form following infection of chickens with oncogenic Marek's disease herpesvirus. Although resistance to tumor formation has been mapped to a subregion of the chicken MHC-B region, the gene or genes responsible have not been identified. A full gene map of the subregion has been lacking. We have expanded the MHC-B region gene map beyond the 92-kb core previously reported for another haplotype revealing the presence of 46 genes within 242 kb in the Red Jungle Fowl haplotype. Even though MHC-B is structured differently, many of the newly revealed genes are related to loci typical of the MHC in other species. Other MHC-B loci are homologs of genes found within MHC paralogous regions (regions thought to be derived from ancient duplications of a primordial immune defense complex where genes have undergone differential silencing over evolutionary time) on other chromosomes. Still others are similar to genes that define the NK complex in mammals. Many of the newly mapped genes display allelic variability and fall within the MHC-B subregion previously shown to affect the formation of Marek's disease tumors and hence are candidates for genes conferring resistance.     

Removal and recovery of phosphorous from swine wastewater by demonstration crystallization reactor and struvite accumulation device

A demonstration crystallization reactor and struvite accumulation device for the removal and recovery of phosphorous was constructed and their performance was evaluated using actual swine wastewater for 3.5 years. The wastewater pH was increased by aeration, and the concentrations of total P and soluble PO(4)-P were reduced by a struvite crystallization reaction induced under a high pH condition. A 30% MgCl(2) addition was effective in enhancing the struvite crystallization reaction. The concentrations of suspended solids, total Zn and total Cu, were also decreased by the settling function of the reactor. On removing the efficiencies of these components, no noticeable seasonal fluctuation in performance was observed during the 3.5-year operation. In terms of maximum yield, 171g struvite was obtained from 1m(3) swine wastewater by the demonstration accumulation device for struvite recovery. The recovered struvite needed only air-drying before use since it was approximately 95% pure even without washing.     

Neurotoxic prion protein (PrP) fragment 106-126 requires the N-terminal half of the hydrophobic region of PrP in the PrP-deficient neuronal cell line

The cytotoxicity of aged PrP(106-126) was examined using an immortalized prion protein (PrP) gene-deficient neuronal cell line. The N-terminal half of the hydrophobic region (HR) but not the octapeptide repeat (OR) of PrP was required for aged PrP(106-126) neurotoxicity, suggesting that neurotoxic signals of aged PrP(106-126) are mediated by this region.