Mannose-6-phosphate regulates destruction of lipid-linked oligosaccharides

Mannose-6-phosphate (M6P) is an essential precursor for mannosyl glycoconjugates, including lipid-linked oligosaccharides (LLO; glucose(3)mannose(9)GlcNAc(2)-P-P-dolichol) used for protein N-glycosylation. In permeabilized mammalian cells, M6P also causes specific LLO cleavage. However, the context and purpose of this paradoxical reaction are unknown. In this study, we used intact mouse embryonic fibroblasts to show that endoplasmic reticulum (ER) stress elevates M6P concentrations, leading to cleavage of the LLO pyrophosphate linkage with recovery of its lipid and lumenal glycan components. We demonstrate that this M6P originates from glycogen, with glycogenolysis activated by the kinase domain of the stress sensor IRE1-α. The apparent futility of M6P causing destruction of its LLO product was resolved by experiments with another stress sensor, PKR-like ER kinase (PERK), which attenuates translation. PERK's reduction of N-glycoprotein synthesis (which consumes LLOs) stabilized steady-state LLO levels despite continuous LLO destruction. However, infection with herpes simplex virus 1, an N-glycoprotein-bearing pathogen that impairs PERK signaling, not only caused LLO destruction but depleted LLO levels as well. In conclusion, the common metabolite M6P is also part of a novel mammalian stress-signaling pathway, responding to viral stress by depleting host LLOs required for N-glycosylation of virus-associated polypeptides. Apparently conserved throughout evolution, LLO destruction may be a response to a variety of environmental stresses.     

Involvement of mitochondrial dysfunction in human islet amyloid polypeptide-induced apoptosis in INS-1E pancreatic beta cells: An effect attenuated by phycocyanin

Misfolded human islet amyloid polypeptide (hIAPP) in pancreatic islets is associated with the loss of insulin-secreting beta cells in type 2 diabetes. Insulin secretion impairment and cell apoptosis can be due to mitochondrial dysfunction in pancreatic beta cells. Currently, there is little information about the effect of hIAPP on mitochondrial function. In this study, we used INS-1E rat insulinoma beta cells as a model to investigate the role of mitochondria in hIAPP-induced apoptosis and the protective effects of phycocyanin (PC). We demonstrated that hIAPP induced apoptosis in INS-1E cells was associated with the disruption of mitochondrial function, as evidenced by ATP depletion, mitochondrial mass reduction, mitochondrial fragmentation and loss of mitochondrial membrane potential (ΔΨ(m)). Further molecular analysis showed that hIAPP induced changes in the expression of Bcl-2 family members, release of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria into cytosol, activation of caspases and cleavage of poly (ADP-ribose) polymerase. Interestingly, the hIAPP-induced mitochondrial dysfunction in INS-1E cells was effectively restored by co-treatment of PC. Moreover, there was crosstalk between the extrinsic and intrinsic apoptotic pathways as demonstrated by cleavage of Bid by caspase-8 in the apoptotic process triggered by hIAPP. Taken together, we demonstrated for the first time the involvement of mitochondrial dysfunction in hIAPP-induced INS-1E cell apoptosis. Attenuation of mitochondrial dysfunction provides a mechanism for the protective effects of PC.     

Population density, water supply, and the risk of dengue fever in Vietnam: cohort study and spatial analysis

Background

Aedes aegypti, the major vector of dengue viruses, often breeds in water storage containers used by households without tap water supply, and occurs in high numbers even in dense urban areas. We analysed the interaction between human population density and lack of tap water as a cause of dengue fever outbreaks with the aim of identifying geographic areas at highest risk.

Methods and Findings

We conducted an individual-level cohort study in a population of 75,000 geo-referenced households in Vietnam over the course of two epidemics, on the basis of dengue hospital admissions (n = 3,013). We applied space-time scan statistics and mathematical models to confirm the findings. We identified a surprisingly narrow range of critical human population densities between around 3,000 to 7,000 people/km² prone to dengue outbreaks. In the study area, this population density was typical of villages and some peri-urban areas. Scan statistics showed that areas with a high population density or adequate water supply did not experience severe outbreaks. The risk of dengue was higher in rural than in urban areas, largely explained by lack of piped water supply, and in human population densities more often falling within the critical range. Mathematical modeling suggests that simple assumptions regarding area-level vector/host ratios may explain the occurrence of outbreaks.

Conclusions

Rural areas may contribute at least as much to the dissemination of dengue fever as cities. Improving water supply and vector control in areas with a human population density critical for dengue transmission could increase the efficiency of control efforts. Please see later in the article for the Editors'' Summary     

Isolation and identification of mannose‐binding proteins and estimation of their abundance in sera from hepatocellular carcinoma patients

The interaction of glycan‐binding proteins (GBPs) and glycans plays a significant biological role that ranges from cell–cell recognition to cell trafficking, and glycoprotein targeting. The anomalies of GBPs related to the types and/or quantities were not clearly known in cancer incidence. It is imperative to identify and annotate the GBPs related with the canceration. Here the mannose‐binding proteins (MBPs) from the clinical sera were isolated and identified by the mannose‐magnetic particle conjugates and the high‐accuracy MS analysis. Seventy‐five MBPs from normal donors’ sera and 79 MBPs from hepatocellular carcinoma patients’ sera were identified and annotated. By using the stringent criteria of exponentially modified protein abundance index (emPAI) quantification, 12 MBPs were estimated to be significantly upregulated (emPAI ratio > 4) and nine MBPs were estimated to be significantly downregulated (emPAI ratio < 0.25) in the hepatocellular carcinoma sera. Real‐time quantitative PCR, Western blotting, and protein microarrays were also used to confirm the altered MBPs expression level and the specific binding between the isolated MBPs and mannose. The sequence recognition motifs and structure preference of the isolated MBPs were characterized. The functional enrichment analysis revealed that over 57% of the isolated MBPs were binding protein and the upregulated MBPs were involved in cell death, tumor progression, and macromolecular complex remodeling.     

A new species of Devriesia causing sooty blotch and flyspeck on Rubber Trees in China

Sooty blotch and flyspeck is a fungal disease complex that colonizes a wide range of plants. In a survey of host plants for sooty blotch and flyspeck fungi, we obtained an isolate associated with discrete flyspeck symptom on stems of the Rubber Trees, Ficus elastica, from Haikou, Hainan, China. A pure culture was obtained, and the nuclear ITS region of rDNA was amplified by PCR. Molecular phylogeny inferred from ITS sequence data suggested that it was close to Devriesia strelitziae, reported from South Africa. Morphologically, it was characterized by longer conidia, more conidial septa, and having chlamydospores. Therefore, we describe our isolate as a novel species of Devriesia.     

Ratio of Metastatic to Examined Lymph Nodes,a Helpful Staging System and Independent Prognostic Factor of Esophagogastric Junction Cancer

Background

The incidence of the esophagogastric junction cancer is growing rapidly. The purpose of this study is to clarify the outcome of the ratio between metastatic and examined lymph nodes in esophagogastric junction cancer patients with or without 7 examined lymph nodes.

Methods

A total of 3,481 patients who underwent operation are identified from the Surveillance, Epidemiology, and End Results database. Different lymph nodes resected groups are analyzed to test the lymph nodes ratio factor.

Results

There are 2522 patients with 7 or more lymph nodes resected and 959 patients with less than 7 lymph nodes resected. Lymph nodes ratio and lymph node involvement are independent prognostic factors. But the lymph nodes ratio categories have a better prognostic value than the lymph node involvement categories. Compared with lymph node involvement categories, lymph nodes ratio categories represent patients with more homogeneous overall survival rate.

Conclusions

This study defines that the lymph nodes ratio is an independent prognostic factor for esophagogastric junction cancer. The lymph nodes ratio can prevent stage migration and may be a helpful system to predict the prognosis of esophagogastric junction cancer patients.     

Epigenome-wide association study reveals longitudinally stable DNA methylation differences in CD4+ T cells from children with IgE-mediated food allergy

Food allergy is mediated by a combination of genetic and environmental risk factors, potentially mediated by epigenetic mechanisms. CD4+ T-cells are key drivers of the allergic response, and may therefore harbor epigenetic variation in association with the disease phenotype. Here we retrospectively examined genome-wide DNA methylation profiles (~450 000 CpGs) from CD4+ T-cells on a birth cohort of 12 children with IgE-mediated food allergy diagnosed at 12-months, and 12 non-allergic controls. DNA samples were available at two time points, birth and 12-months. Case:control comparisons of CD4+ methylation profiles identified 179 differentially methylated probes (DMP) at 12-months and 136 DMP at birth (FDR-adjusted P value < 0.05, delta β > 0.1). Approximately 30% of DMPs were coincident with previously annotated SNPs. A total of 96 allergy-associated non-SNP DMPs were present at birth when individuals were initially disease-free, potentially implicating these loci in the causal pathway. Pathway analysis of differentially methylated genes identified several MAP kinase signaling molecules. Mass spectrometry was used to validate 15 CpG sites at 3 candidate genes. Combined analysis of differential methylation with gene expression profiles revealed gene expression differences at some but not all allergy associated differentially methylated genes. Thus, dysregulation of DNA methylation at MAPK signaling-associated genes during early CD4+ T-cell development may contribute to suboptimal T-lymphocyte responses in early childhood associated with the development of food allergy.     

Autophagy regulates the therapeutic potential of mesenchymal stem cells in experimental autoimmune encephalomyelitis

Mesenchymal stem cell (MSC)-based therapy is a promising approach to treat various inflammatory disorders including multiple sclerosis. However, the fate of MSCs in the inflammatory microenvironment is largely unknown. Experimental autoimmune encephalomyelitis (EAE) is a well-studied animal model of multiple sclerosis. We demonstrated that autophagy occurred in MSCs during their application for EAE treatment. Inflammatory cytokines, e.g., interferon gamma and tumor necrosis factor, induced autophagy in MSCs synergistically by inducing expression of BECN1/Beclin 1. Inhibition of autophagy by knockdown of Becn1 significantly improved the therapeutic effects of MSCs on EAE, which was mainly attributable to enhanced suppression upon activation and expansion of CD4⁺ T cells. Mechanistically, inhibition of autophagy increased reactive oxygen species generation and mitogen-activated protein kinase 1/3 activation in MSCs, which were essential for PTGS2 (prostaglandin-endoperoxide synthase 2 [prostaglandin G/H synthase and cyclooxygenase]) and downstream prostaglandin E2 expression to exert immunoregulatory function. Furthermore, pharmacological treatment of MSCs to inhibit autophagy increased their immunosuppressive effects on T cell-mediated EAE. Our findings indicate that inflammatory microenvironment-induced autophagy downregulates the immunosuppressive function of MSCs. Therefore, modulation of autophagy in MSCs would provide a novel strategy to improve MSC-based immunotherapy.