Association between Polymorphisms in Glutathione Peroxidase and Selenoprotein P Genes, Glutathione Peroxidase Activity, HRT Use and Breast Cancer Risk.

Breast cancer (BC) is one of the most common cancers in women. Evidence suggests that genetic variation in antioxidant enzymes could influence BC risk, but to date the relationship between selenoproteins and BC risk remains unclear. In this report, a study population including 975 Danish cases and 975 controls matched for age and hormone replacement therapy (HRT) use was genotyped for five functional single nucleotide polymorphisms (SNPs) in SEPP1, GPX1, GPX4 and the antioxidant enzyme SOD2 genes. The influence of genetic polymorphisms on breast cancer risk was assessed using conditional logistic regression. Additionally pre-diagnosis erythrocyte GPx (eGPx) activity was measured in a sub-group of the population. A 60% reduction in risk of developing overall BC and ductal BC was observed in women who were homozygous Thr carriers for SEPP1 rs3877899. Additionally, Leu carriers for GPX1 Pro198Leu polymorphism (rs1050450) were at ∼2 fold increased risk of developing a non-ductal BC. Pre-diagnosis eGPx activity was found to depend on genotype for rs713041 (GPX4), rs3877899 (SEPP1), and rs1050450 (GPX1) and on HRT use. Moreover, depending on genotype and HRT use, eGPx activity was significantly lower in women who developed BC later in life compared with controls. Furthermore, GPx1 protein levels increased in human breast adenocarcinoma MCF7 cells exposed to β-estradiol and sodium selenite.In conclusion, our data provide evidence that SNPs in SEPP1 and GPX1 modulate risk of BC and that eGPx activity is modified by SNPs in SEPP1, GPX4 and GPX1 and by estrogens. Our data thus suggest a role of selenoproteins in BC development

Luminal-Applied Flagellin Is Internalized by Polarized Intestinal Epithelial Cells and Elicits Immune Responses via the TLR5 Dependent Mechanism

Bacteria release flagellin that elicits innate responses via Toll-like receptor 5 (TLR5). Here, we investigated the fate of apically administrated full length flagellin from virulent and avirulent bacteria, along with truncated recombinant flagellin proteins in intestinal epithelial cells and cellular responses. Flagellin was internalized by intestinal epithelial cell (IEC) monolayers of IEC-18. Additionally, apically applied flagellin was internalized by polarized human Caco-2BBe and T-84 cells in a TLR5 dependent mechanism. More, flagellin exposure did not affect the integrity of intestinal monolayers. With immunofluorescent staining, internalized flagellin was detected in both early endosomes as well as lysosomes. We found that apical exposure of polarized Caco-2BBe and T-84 to flagellin from purified Salmonella, Escherichia coli O83:H1 (isolate from Crohn’s lesion) or avirulent E. coli K12 induced comparable levels of basolateral IL-8 secretion. A recombinant protein representing the conserved amino (N) and carboxyl (C) domains (D) of the flagellin protein (ND1/2ECHCD2/1) induced IL-8 secretion from IEC similar to levels elicited by full-length flagellins. However, a recombinant flagellin protein containing only the D3 hypervariable region elicited no IL-8 secretion in both cell lines compared to un-stimulated controls. Silencing or blocking TLR5 in Caco-2BBe cells resulted in a lack of flagellin internalization and decreased IL-8 secretion. Furthermore, apical exposure to flagellin stimulated transepithelial migration of neutrophils and dendritic cells. The novel findings in this study show that luminal-applied flagellin is internalized by normal IEC via TLR5 and co-localizes to endosomal and lysosomal compartments where it is likely degraded as flagellin was not detected on the basolateral side of IEC cultures

The modular systems biology approach to investigate the control of apoptosis in Alzheimer's disease neurodegeneration

Apoptosis is a programmed cell death that plays a critical role during the development of the nervous system and in many chronic neurodegenerative diseases, including Alzheimer's disease (AD). This pathology, characterized by a progressive degeneration of cholinergic function resulting in a remarkable cognitive decline, is the most common form of dementia with high social and economic impact. Current therapies of AD are only symptomatic, therefore the need to elucidate the mechanisms underlying the onset and progression of the disease is surely needed in order to develop effective pharmacological therapies. Because of its pivotal role in neuronal cell death, apoptosis has been considered one of the most appealing therapeutic targets, however, due to the complexity of the molecular mechanisms involving the various triggering events and the many signaling cascades leading to cell death, a comprehensive understanding of this process is still lacking. Modular systems biology is a very effective strategy in organizing information about complex biological processes and deriving modular and mathematical models that greatly simplify the identification of key steps of a given process. This review aims at describing the main steps underlying the strategy of modular systems biology and briefly summarizes how this approach has been successfully applied for cell cycle studies. Moreover, after giving an overview of the many molecular mechanisms underlying apoptosis in AD, we present both a modular and a molecular model of neuronal apoptosis that suggest new insights on neuroprotection for this disease

TRANSIENT RESPONSES OF MYOSTATIN SIGNALING MARKERS TO ACUTE BOUTS OF RESISTANCE TRAINING

Mason C. McIntosh, Casey L. Sexton, Joshua L. Godwin, Bradley A. Ruple, Shelby C. Osburn, John M. Michel, Daniel L. Plotkin, Christopher B. Mobley, Michael D. Roberts. Auburn University, Auburn, AL. BACKGROUND: The myostatin (MSTN) gene has been heavily researched for its role in repressing skeletal muscle hypertrophy and the anabolic mTORC1 signaling pathway. Preliminary data from our laboratory has shown a significant hypermethylation of cytosine phosphate guanine sites within the MSTN gene as well as a significant downregulation of MSTN mRNA following acute bouts of resistance exercise to failure (i.e., training at 30% one repetition maximum versus 80% one repetition maximum). This study sought to determine the changes of MSTN-related and mTORC1 signaling proteins following two bouts of resistance exercise to failure. We hypothesized there to be significant time-effects of assayed markers in response to each bout of resistance training. METHODS: Eleven previously-trained college-aged men (age: 23 ± 4 years, 11.4 ± 6.4 percent fat, 4 ± 3 years training experience) participated in this study. Each participant performed two resistance training sessions (spaced one week apart) involving either: i) 30FAIL training; 4 sets of back squats and 4 sets of leg extensors to failure at 30% of one-repetition maximum (1RM), or: ii) 80FAIL training; 4 sets of both exercises at 80% of 1RM. Vastus lateralis muscle biopsies were obtained prior to each training session (PRE), 3 hours following training (3hPOST), and 6 hours following training (6hPOST). Western blots were performed on biopsied muscle to determine the relative expression of phosphorylated (p)-mTOR (Ser2448), p-p70S6K (Thr389), p-AKT (Ser473), p-rpS6 (Ser235/236), follistatin (FST), and MSTN. RESULTS: There were no significant bout*time interactions for any of the assayed markers. There were significant differences observed for p-p70S6K (p=0.001; PRE to 3hPOST) and FST (p=0.021; PRE to 6hPOST). There were no significant main time effects observed for p-mTOR, p-AKT, or p-rps6. CONCLUSIONS: The two modes of resistance training elicited similar effects on p-p70S6K and FST protein expression. These results suggest, regardless of training load, mTORC1 markers and MTSN-related protein expression respond similarly in previously-trained college-aged men

Testosterone and trenbolone enanthate increase mature myostatin protein expression despite increasing skeletal muscle hypertrophy and satellite cell number in rodent muscle

Dalbo, VJ ORCiD: 0000-0002-5944-7558The androgen-induced alterations in adult rodent skeletal muscle fibre cross-sectional area (fCSA), satellite cell content and myostatin (Mstn) were examined in 10-month-old Fisher 344 rats (n = 41) assigned to Sham surgery, orchiectomy (ORX), ORX + testosterone (TEST; 7.0 mg week−1) or ORX + trenbolone (TREN; 1.0 mg week−1). After 29 days, animals were euthanised and the levator ani/bulbocavernosus (LABC) muscle complex was harvested for analyses. LABC muscle fCSA was 102% and 94% higher in ORX + TEST and ORX + TREN compared to ORX (p <.001). ORX + TEST and ORX + TREN increased satellite cell numbers by 181% and 178% compared to ORX, respectively (p <.01), with no differences between conditions for myonuclear number per muscle fibre (p =.948). Mstn protein was increased 159% and 169% in the ORX + TEST and ORX + TREN compared to ORX (p <.01). pan-SMAD2/3 protein was ~30–50% greater in ORX compared to SHAM (p =.006), ORX + TEST (p =.037) and ORX + TREN (p =.043), although there were no between-treatment effects regarding phosphorylated SMAD2/3. Mstn, ActrIIb and Mighty mRNAs were lower in ORX, ORX + TEST and ORX + TREN compared to SHAM (p <.05). Testosterone and trenbolone administration increased muscle fCSA and satellite cell number without increasing myonuclei number, and increased Mstn protein levels. Several genes and signalling proteins related to myostatin signalling were differentially regulated by ORX or androgen therapy. © 2016 Blackwell Verlag GmbHAssociated Grant:This study was also supported in part by work supported from the Office of Research and Development, Rehabilitation Research and Development (RR&D) Service, Department of Veterans Affairs Merit Award to S.E.B., Veteran’s Health Administration Advanced Geriatrics Fellowship to D.T.B., and RR&D CDA-2 to J.F.Y

Testosterone and trenbolone enanthate increase mature myostatin protein expression despite increasing skeletal muscle hypertrophy and satellite cell number in rodent muscle

The androgen-induced alterations in adult rodent skeletal muscle fibre cross-sectional area (fCSA), satellite cell content and myostatin (Mstn) were examined in 10-month-old Fisher 344 rats (n = 41) assigned to Sham surgery, orchiectomy (ORX), ORX + testosterone (TEST; 7.0 mg week−1) or ORX + trenbolone (TREN; 1.0 mg week−1). After 29 days, animals were euthanised and the levator ani/bulbocavernosus (LABC) muscle complex was harvested for analyses. LABC muscle fCSA was 102% and 94% higher in ORX + TEST and ORX + TREN compared to ORX (p <.001). ORX + TEST and ORX + TREN increased satellite cell numbers by 181% and 178% compared to ORX, respectively (p <.01), with no differences between conditions for myonuclear number per muscle fibre (p =.948). Mstn protein was increased 159% and 169% in the ORX + TEST and ORX + TREN compared to ORX (p <.01). pan-SMAD2/3 protein was ~30–50% greater in ORX compared to SHAM (p =.006), ORX + TEST (p =.037) and ORX + TREN (p =.043), although there were no between-treatment effects regarding phosphorylated SMAD2/3. Mstn, ActrIIb and Mighty mRNAs were lower in ORX, ORX + TEST and ORX + TREN compared to SHAM (p <.05). Testosterone and trenbolone administration increased muscle fCSA and satellite cell number without increasing myonuclei number, and increased Mstn protein levels. Several genes and signalling proteins related to myostatin signalling were differentially regulated by ORX or androgen therapy. © 2016 Blackwell Verlag Gmb

Mucosal immunization with a urease B DNA vaccine induces innate and cellular immune responses against Helicobacter pylori.

BACKGROUND: Helicobacter pylori is recognized as a major risk factor for recurrent gastroduodenal inflammatory diseases and gastric adenocarcinoma. The high prevalence of H. pylori infection worldwide, the risks of side-effects from antibiotic therapy, and increasing resistance to antibiotics are the main primers for the development of improved H. pylori vaccines. The antigenic potential of its urease enzyme, a critical virulence factor required for colonization of the gastric mucosa, has been demonstrated in animal and human studies. An important but controversial issue in H. pylori vaccine studies is the type of immune response required to control infection. A new approach in H. pylori vaccinology is the administration of DNA vaccines, which has included heat-shock protein and catalase DNA vaccines. MATERIALS AND METHODS: The H. pylori urease subunit B construct or vector alone was administered to mice via the intranasal route. Spleens and stomachs were examined on day 0 and weeks 3, 6, and 12 after immunization. Proliferation of spleen cells was assessed using the carboxyfluorescein diacetate succinimidyl ester-based flow cytometry assay and cytokine secretion from cultured spleen cells was detected by ELISA, after stimulation with the urease subunit B recombinant antigen. Total RNA was isolated from stomach and spleen tissue and the expression of beta-defensin and cytokine genes was monitored by reverse transcription followed by polymerase chain reaction (RT-PCR). Immunized mice were challenged with H. pylori and bacterial DNA quantified by TaqMan PCR. RESULTS: The urease B subunit DNA vaccine increased INF-gamma secretion and splenocyte proliferation without inducing adverse effects in the spleen. Increase in gastric beta-defensin 1 and marked induction in local IL-10 : IFN-gamma ratio up to 12 weeks post-immunization suggest a potential role for local innate immune responses in protection at the site of infection. Although significant bacterial reduction in the stomachs of urease B subunit DNA-immunized mice was observed, intermediate reduction was also noted in the vector group. Increased defensin expression and adjuvant effects of the cytosine preceding guanosine motifs may contribute to this phenomenon. Our data confirm that cytosine preceding guanosine motifs, even without coadministration with antigen, can reduce extracellular bacterial load. CONCLUSIONS: In this study, a DNA construct encoding the urease B subunit was assessed for its immune profile and its ability to reduce bacterial colonization in the murine stomach. Our studies suggest that local innate immune responses may play a greater role than previously supposed in limiting H. pylori colonization in the gastric mucosa