Dairy foods and dairy protein consumption is inversely related to markers of adiposity in obese men and women

A number of intervention studies have reported that the prevalence of obesity may be in part inversely related to dairy food consumption while others report no association. We sought to examine relationships between energy, protein and calcium consumption from dairy foods (milk, yoghurt, cheese, dairy spreads, ice-cream) and adiposity including body mass index (BMI), waist (WC) and hip circumference (HC), and direct measures of body composition using dual energy X-ray absorptiometry (% body fat and abdominal fat) in an opportunistic sample of 720 overweight/obese Australian men and women. Mean (SD) age, weight and BMI of the population were 51 ± 10 year, 94 ± 18 kg and 32.4 ± 5.7 kg/m2, respectively. Reduced fat milk was the most commonly consumed dairy product (235 ± 200 g/day), followed by whole milk (63 ± 128 g/day) and yoghurt (53 ± 66 g/day). Overall dairy food consumption (g/day) was inversely associated with BMI, % body fat and WC (all p < 0.05). Dairy protein and dairy calcium (g/day) were both inversely associated with all adiposity measures (all p < 0.05). Yoghurt consumption (g/day) was inversely associated with % body fat, abdominal fat, WC and HC (all p < 0.05), while reduced fat milk consumption was inversely associated with BMI, WC, HC and % body fat (all p < 0.05). Within a sample of obese adults, consumption of dairy products, dairy protein, and calcium was associated with more favourable body composition

CD8+ T Cells Mediate the Athero-Protective Effect of Immunization with an ApoB-100 Peptide

Immunization of hypercholesterolemic mice with selected apoB-100 peptide antigens reduces atherosclerosis but the precise immune mediators of athero-protection remain unclear. In this study we show that immunization of apoE (-/-) mice with p210, a 20 amino acid apoB-100 related peptide, reduced aortic atherosclerosis compared with PBS or adjuvant/carrier controls. Immunization with p210 activated CD8+ T cells, reduced dendritic cells (DC) at the site of immunization and within the plaque with an associated reduction in plaque macrophage immunoreactivity. Adoptive transfer of CD8+ T cells from p210 immunized mice recapitulated the athero-protective effect of p210 immunization in naïve, non-immunized mice. CD8+ T cells from p210 immunized mice developed a preferentially higher cytolytic response against p210-loaded dendritic cells in vitro. Although p210 immunization profoundly modulated DCs and cellular immune responses, it did not alter the efficacy of subsequent T cell dependent or independent immune response to other irrelevant antigens. Our data define, for the first time, a role for CD8+ T cells in mediating the athero-protective effects of apoB-100 related peptide immunization in apoE (-/-) mice

Rad21-Cohesin Haploinsufficiency Impedes DNA Repair and Enhances Gastrointestinal Radiosensitivity in Mice

Approximately half of cancer-affected patients receive radiotherapy (RT). The doses delivered have been determined upon empirical experience based upon average radiation responses. Ideally higher curative radiation doses might be employed in patients with genuinely normal radiation responses and importantly radiation hypersensitive patients would be spared the consequences of excessive tissue damage if they were indentified before treatment. Rad21 is an integral subunit of the cohesin complex, which regulates chromosome segregation and DNA damage responses in eukaryotes. We show here, by targeted inactivation of this key cohesin component in mice, that Rad21 is a DNA-damage response gene that markedly affects animal and cell survival. Biallelic deletion of Rad21 results in early embryonic death. Rad21 heterozygous mutant cells are defective in homologous recombination (HR)-mediated gene targeting and sister chromatid exchanges. Rad21+/− animals exhibited sensitivity considerably greater than control littermates when challenged with whole body irradiation (WBI). Importantly, Rad21+/− animals are significantly more sensitive to WBI than Atm heterozygous mutant mice. Since supralethal WBI of mammals most typically leads to death via damage to the gastrointestinal tract (GIT) or the haematopoietic system, we determined the functional status of these organs in the irradiated animals. We found evidence for GIT hypersensitivity of the Rad21 mutants and impaired bone marrow stem cell clonogenic regeneration. These data indicate that Rad21 gene dosage is critical for the ionising radiation (IR) response. Rad21 mutant mice thus represent a new mammalian model for understanding the molecular basis of irradiation effects on normal tissues and have important implications in the understanding of acute radiation toxicity in normal tissues

EndoS2 is a unique and conserved enzyme of serotype M49 group A Streptococcus that hydrolyses N-linked glycans on IgG and α1-acid glycoprotein

Many bacteria have evolved ways to interact with glycosylation functions of the immune system of their hosts. Streptococcus pyogenes [GAS (group A Streptococcus)] secretes the enzyme EndoS that cleaves glycans on human IgG and impairs the effector functions of the antibody. The ndoS gene, encoding EndoS, has, until now, been thought to be conserved throughout the serotypes. However, in the present study, we identify EndoS2, an endoglycosidase in serotype M49 GAS strains. We characterized EndoS2 and the corresponding ndoS2 gene using sequencing, bioinformatics, phylogenetic analysis, recombinant expression and LC–MS analysis of glycosidic activity. This revealed that EndoS2 is present exclusively, and highly conserved, in serotype M49 of GAS and is only 37% identical with EndoS. EndoS2 showed endo-β-N-acetylglucosaminidase activity on all N-linked glycans of IgG and on biantennary and sialylated glycans of AGP (α1-acid glycoprotein). The enzyme was found to act only on native IgG and AGP and to be specific for free biantennary glycans with or without terminal sialylation. GAS M49 expression of EndoS2 was monitored in relation to carbohydrates present in the culture medium and was linked to the presence of sucrose. We conclude that EndoS2 is a unique endoglycosidase in serotype M49 and differs from EndoS of other GAS strains by targeting both IgG and AGP. EndoS2 expands the repertoire of GAS effectors that modify key glycosylated molecules of host defence

Deciphering Protein <i>O</i>‑Glycosylation: Solid-Phase Chemoenzymatic Cleavage and Enrichment

Glycosylation plays a critical role in the biosynthetic-secretory pathway in the endoplasmic reticulum (ER) and Golgi apparatus. Over 50% of mammalian cellular proteins are typically glycosylated; this modification is involved in a wide range of biological functions such as barrier formation against intestinal microbes and serves as signaling molecules for selectins and galectins in the innate immune system. <i>N</i>-linked glycosylation analysis has been greatly facilitated owing to a range of specific enzymes available for their release. However, system-wide analysis on <i>O</i>-linked glycosylation remains a challenge due to the lack of equivalent enzymes and the inherent structural heterogeneity of <i>O</i>-glycans. Although <i>O</i>-glycosidase can catalyze the removal of core 1 and core 3 <i>O</i>-linked disaccharides from glycoproteins, analysis of other types of <i>O-</i>glycans remains difficult, particularly when residing on glycopeptides. Here, we describe a novel chemoenzymatic approach driven by a newly available <i>O-</i>protease and solid phase platform. This method enables the assignment of <i>O</i>-glycosylated peptides, <i>N</i>-glycan profile, sialyl <i>O</i>-glycopeptides linkage, and mapping of heterogeneous <i>O</i>-glycosylation. For the first time, we can analyze intact <i>O</i>-glycopeptides generated by <i>O</i>-protease and enriched using a solid-phase platform. We establish the method on standard glycoproteins, confirming known <i>O-</i>glycosites with high accuracy and confidence, and reveal up to 8-fold more glycosites than previously reported with concomitant increased heterogeneity. This technique is further applied for analysis of Zika virus recombinant glycoproteins, revealing their dominant <i>O</i>-glycosites and setting a basis set of <i>O</i>-glycosylation tracts in these important viral antigens. Our approach can serve as a benchmark for the investigation of protein <i>O</i>-glycosylation in diseases and other biomedical contexts. This method should become an indispensable tool for investigations where <i>O</i>-glycosylation is central

Deciphering Protein <i>O</i>‑Glycosylation: Solid-Phase Chemoenzymatic Cleavage and Enrichment

Glycosylation plays a critical role in the biosynthetic-secretory pathway in the endoplasmic reticulum (ER) and Golgi apparatus. Over 50% of mammalian cellular proteins are typically glycosylated; this modification is involved in a wide range of biological functions such as barrier formation against intestinal microbes and serves as signaling molecules for selectins and galectins in the innate immune system. <i>N</i>-linked glycosylation analysis has been greatly facilitated owing to a range of specific enzymes available for their release. However, system-wide analysis on <i>O</i>-linked glycosylation remains a challenge due to the lack of equivalent enzymes and the inherent structural heterogeneity of <i>O</i>-glycans. Although <i>O</i>-glycosidase can catalyze the removal of core 1 and core 3 <i>O</i>-linked disaccharides from glycoproteins, analysis of other types of <i>O-</i>glycans remains difficult, particularly when residing on glycopeptides. Here, we describe a novel chemoenzymatic approach driven by a newly available <i>O-</i>protease and solid phase platform. This method enables the assignment of <i>O</i>-glycosylated peptides, <i>N</i>-glycan profile, sialyl <i>O</i>-glycopeptides linkage, and mapping of heterogeneous <i>O</i>-glycosylation. For the first time, we can analyze intact <i>O</i>-glycopeptides generated by <i>O</i>-protease and enriched using a solid-phase platform. We establish the method on standard glycoproteins, confirming known <i>O-</i>glycosites with high accuracy and confidence, and reveal up to 8-fold more glycosites than previously reported with concomitant increased heterogeneity. This technique is further applied for analysis of Zika virus recombinant glycoproteins, revealing their dominant <i>O</i>-glycosites and setting a basis set of <i>O</i>-glycosylation tracts in these important viral antigens. Our approach can serve as a benchmark for the investigation of protein <i>O</i>-glycosylation in diseases and other biomedical contexts. This method should become an indispensable tool for investigations where <i>O</i>-glycosylation is central

Deciphering Protein <i>O</i>‑Glycosylation: Solid-Phase Chemoenzymatic Cleavage and Enrichment

Glycosylation plays a critical role in the biosynthetic-secretory pathway in the endoplasmic reticulum (ER) and Golgi apparatus. Over 50% of mammalian cellular proteins are typically glycosylated; this modification is involved in a wide range of biological functions such as barrier formation against intestinal microbes and serves as signaling molecules for selectins and galectins in the innate immune system. <i>N</i>-linked glycosylation analysis has been greatly facilitated owing to a range of specific enzymes available for their release. However, system-wide analysis on <i>O</i>-linked glycosylation remains a challenge due to the lack of equivalent enzymes and the inherent structural heterogeneity of <i>O</i>-glycans. Although <i>O</i>-glycosidase can catalyze the removal of core 1 and core 3 <i>O</i>-linked disaccharides from glycoproteins, analysis of other types of <i>O-</i>glycans remains difficult, particularly when residing on glycopeptides. Here, we describe a novel chemoenzymatic approach driven by a newly available <i>O-</i>protease and solid phase platform. This method enables the assignment of <i>O</i>-glycosylated peptides, <i>N</i>-glycan profile, sialyl <i>O</i>-glycopeptides linkage, and mapping of heterogeneous <i>O</i>-glycosylation. For the first time, we can analyze intact <i>O</i>-glycopeptides generated by <i>O</i>-protease and enriched using a solid-phase platform. We establish the method on standard glycoproteins, confirming known <i>O-</i>glycosites with high accuracy and confidence, and reveal up to 8-fold more glycosites than previously reported with concomitant increased heterogeneity. This technique is further applied for analysis of Zika virus recombinant glycoproteins, revealing their dominant <i>O</i>-glycosites and setting a basis set of <i>O</i>-glycosylation tracts in these important viral antigens. Our approach can serve as a benchmark for the investigation of protein <i>O</i>-glycosylation in diseases and other biomedical contexts. This method should become an indispensable tool for investigations where <i>O</i>-glycosylation is central

Deciphering Protein <i>O</i>‑Glycosylation: Solid-Phase Chemoenzymatic Cleavage and Enrichment

Glycosylation plays a critical role in the biosynthetic-secretory pathway in the endoplasmic reticulum (ER) and Golgi apparatus. Over 50% of mammalian cellular proteins are typically glycosylated; this modification is involved in a wide range of biological functions such as barrier formation against intestinal microbes and serves as signaling molecules for selectins and galectins in the innate immune system. <i>N</i>-linked glycosylation analysis has been greatly facilitated owing to a range of specific enzymes available for their release. However, system-wide analysis on <i>O</i>-linked glycosylation remains a challenge due to the lack of equivalent enzymes and the inherent structural heterogeneity of <i>O</i>-glycans. Although <i>O</i>-glycosidase can catalyze the removal of core 1 and core 3 <i>O</i>-linked disaccharides from glycoproteins, analysis of other types of <i>O-</i>glycans remains difficult, particularly when residing on glycopeptides. Here, we describe a novel chemoenzymatic approach driven by a newly available <i>O-</i>protease and solid phase platform. This method enables the assignment of <i>O</i>-glycosylated peptides, <i>N</i>-glycan profile, sialyl <i>O</i>-glycopeptides linkage, and mapping of heterogeneous <i>O</i>-glycosylation. For the first time, we can analyze intact <i>O</i>-glycopeptides generated by <i>O</i>-protease and enriched using a solid-phase platform. We establish the method on standard glycoproteins, confirming known <i>O-</i>glycosites with high accuracy and confidence, and reveal up to 8-fold more glycosites than previously reported with concomitant increased heterogeneity. This technique is further applied for analysis of Zika virus recombinant glycoproteins, revealing their dominant <i>O</i>-glycosites and setting a basis set of <i>O</i>-glycosylation tracts in these important viral antigens. Our approach can serve as a benchmark for the investigation of protein <i>O</i>-glycosylation in diseases and other biomedical contexts. This method should become an indispensable tool for investigations where <i>O</i>-glycosylation is central