Enhanced mitochondrial superoxide scavenging does not Improve muscle insulin action in the high fat-fed mouse

Improving mitochondrial oxidant scavenging may be a viable strategy for the treatment of insulin resistance and diabetes. Mice overexpressing the mitochondrial matrix isoform of superoxide dismutase (sod2(tg) mice) and/or transgenically expressing catalase within the mitochondrial matrix (mcat(tg) mice) have increased scavenging of O2(˙-) and H2O2, respectively. Furthermore, muscle insulin action is partially preserved in high fat (HF)-fed mcat(tg) mice. The goal of the current study was to test the hypothesis that increased O2(˙-) scavenging alone or in combination with increased H2O2 scavenging (mtAO mice) enhances in vivo muscle insulin action in the HF-fed mouse. Insulin action was examined in conscious, unrestrained and unstressed wild type (WT), sod2(tg), mcat(tg) and mtAO mice using hyperinsulinemic-euglycemic clamps (insulin clamps) combined with radioactive glucose tracers following sixteen weeks of normal chow or HF (60% calories from fat) feeding. Glucose infusion rates, whole body glucose disappearance, and muscle glucose uptake during the insulin clamp were similar in chow- and HF-fed WT and sod2(tg) mice. Consistent with our previous work, HF-fed mcat(tg) mice had improved muscle insulin action, however, an additive effect was not seen in mtAO mice. Insulin-stimulated Akt phosphorylation in muscle from clamped mice was consistent with glucose flux measurements. These results demonstrate that increased O2(˙-) scavenging does not improve muscle insulin action in the HF-fed mouse alone or when coupled to increased H2O2 scavenging

Tumor-associated endothelial cells display GSTP1 and RARβ2 promoter methylation in human prostate cancer

BACKGROUND: A functional blood supply is essential for tumor growth and proliferation. However, the mechanism of blood vessel recruitment to the tumor is still poorly understood. Ideally, a thorough molecular assessment of blood vessel cells would be critical in our comprehension of this process. Yet, to date, there is little known about the molecular makeup of the endothelial cells of tumor-associated blood vessels, due in part to the difficulty of isolating a pure population of endothelial cells from the heterogeneous tissue environment. METHODS: Here we describe the use of a recently developed technique, Expression Microdissection, to isolate endothelial cells from the tumor microenvironment. The methylation status of the dissected samples was evaluated for GSTP1 and RARβ2 promoters via the QMS-PCR method. RESULTS: Comparing GSTP1 and RARβ2 promoter methylation data, we show that 100% and 88% methylation is detected, respectively, in the tumor areas, both in epithelium and endothelium. Little to no methylation is observed in non-tumor tissue areas. CONCLUSION: We applied an accurate microdissection technique to isolate endothelial cells from tissues, enabling DNA analysis such as promoter methylation status. The observations suggest that epigenetic alterations may play a role in determining the phenotype of tumor-associated vasculature

Cost-Effectiveness of Adding Cetuximab to Platinum-Based Chemotherapy for First-Line Treatment of Recurrent or Metastatic Head and Neck Cancer

To assess the cost effectiveness of adding cetuximab to platinum-based chemotherapy in first-line treatment of patients with recurrent or metastatic head and neck squamous cell carcinoma (HNSCC) from the perspective of the Canadian public healthcare system.We developed a Markov state transition model to project the lifetime clinical and economic consequences of recurrent or metastatic HNSCC. Transition probabilities were derived from a phase III trial of cetuximab in patients with recurrent or metastatic HNSCC. Cost estimates were obtained from London Health Sciences Centre and the Ontario Case Costing Initiative, and expressed in 2011 CAD. A three year time horizon was used. Future costs and health benefits were discounted at 5%.In the base case, cetuximab plus platinum-based chemotherapy compared to platinum-based chemotherapy alone led to an increase of 0.093 QALY and an increase in cost of $36,000 per person, resulting in an incremental cost effectiveness ratio (ICER) of$386,000 per QALY gained. The cost effectiveness ratio was most sensitive to the cost per mg of cetuximab and the absolute risk of progression among patients receiving cetuximab.The addition of cetuximab to standard platinum-based chemotherapy in first-line treatment of patients with recurrent or metastatic HNSCC has an ICER that exceeds $100,000 per QALY gained. Cetuximab can only be economically attractive in this patient population if the cost of cetuximab is substantially reduced or if future research can identify predictive markers to select patients most likely to benefit from the addition of cetuximab to chemotherapy

Both telomeric and non-telomeric DNA damage are determinants of mammalian cellular senescence

<p>Abstract</p> <p>Background</p> <p>Cellular senescence is a state reached by normal mammalian cells after a finite number of cell divisions and is characterized by morphological and physiological changes including terminal cell-cycle arrest. The limits on cell division imposed by senescence may play an important role in both organismal aging and in preventing tumorigenesis. Cellular senescence and organismal aging are both accompanied by increased DNA damage, seen as the formation of γ-H2AX foci (γ-foci), which may be found on uncapped telomeres or at non-telomeric sites of DNA damage. However, the relative importance of telomere- and non-telomere-associated DNA damage to inducing senescence has never been demonstrated. Here we present a new approach to determine accurately the chromosomal location of γ-foci and quantify the number of telomeric versus non-telomeric γ-foci associated with senescence in both human and mouse cells. This approach enables researchers to obtain accurate values and to avoid various possible misestimates inherent in earlier methods.</p> <p>Results</p> <p>Using combined immunofluorescence and telomere fluorescence <it>in situ </it>hybridization on metaphase chromosomes, we show that human cellular senescence is not solely determined by telomeric DNA damage. In addition, mouse cellular senescence is not solely determined by non-telomeric DNA damage. By comparing cells from different generations of telomerase-null mice with human cells, we show that cells from late generation telomerase-null mice, which have substantially short telomeres, contain mostly telomeric γ-foci. Most notably, we report that, as human and mouse cells approach senescence, all cells exhibit similar numbers of total γ-foci per cell, irrespective of chromosomal locations.</p> <p>Conclusion</p> <p>Our results suggest that the chromosome location of senescence-related γ-foci is determined by the telomere length rather than species differences <it>per se</it>. In addition, our data indicate that both telomeric and non-telomeric DNA damage responses play equivalent roles in signaling the initiation of cellular senescence and organismal aging. These data have important implications in the study of mechanisms to induce or delay cellular senescence in different species.</p

Protective Unfolded Protein Response in Human Pancreatic Beta Cells Transplanted into Mice

Background: There is great interest about the possible contribution of ER stress to the apoptosis of pancreatic beta cells in the diabetic state and with islet transplantation. Methods and Findings: Expression of genes involved in ER stress were examined in beta cell enriched tissue obtained with laser capture microdissection (LCM) from frozen sections of pancreases obtained from non-diabetic subjects at surgery and from human islets transplanted into ICR-SCID mice for 4 wk. Because mice have higher glucose levels than humans, the transplanted beta cells were exposed to mild hyperglycemia and the abnormal environment of the transplant site. RNA was extracted from the LCM specimens, amplified and then subjected to microarray analysis. The transplanted beta cells showed an unfolded protein response (UPR). There was activation of many genes of the IRE-1 pathway that provide protection against the deleterious effects of ER stress, increased expression of ER chaperones and ERAD (ER-associated protein degradation) proteins. The other two arms of ER stress, PERK and ATF-6, had many down regulated genes. Downregulation of EIF2A could protect by inhibiting protein synthesis. Two genes known to contribute to apoptosis, CHOP and JNK, were downregulated. Conclusions: Human beta cells in a transplant site had UPR changes in gene expression that protect against the proapoptotic effects of unfolded proteins

Pleiotropic Roles of a Ribosomal Protein in Dictyostelium discoideum

The cell cycle phase at starvation influences post-starvation differentiation and morphogenesis in Dictyostelium discoideum. We found that when expressed in Saccharomyces cerevisiae, a D. discoideum cDNA that encodes the ribosomal protein S4 (DdS4) rescues mutations in the cell cycle genes cdc24, cdc42 and bem1. The products of these genes affect morphogenesis in yeast via a coordinated moulding of the cytoskeleton during bud site selection. D. discoideum cells that over- or under-expressed DdS4 did not show detectable changes in protein synthesis but displayed similar developmental aberrations whose intensity was graded with the extent of over- or under-expression. This suggested that DdS4 might influence morphogenesis via a stoichiometric effect – specifically, by taking part in a multimeric complex similar to the one involving Cdc24p, Cdc42p and Bem1p in yeast. In support of the hypothesis, the S. cerevisiae proteins Cdc24p, Cdc42p and Bem1p as well as their D. discoideum cognates could be co-precipitated with antibodies to DdS4. Computational analysis and mutational studies explained these findings: a C-terminal domain of DdS4 is the functional equivalent of an SH3 domain in the yeast scaffold protein Bem1p that is central to constructing the bud site selection complex. Thus in addition to being part of the ribosome, DdS4 has a second function, also as part of a multi-protein complex. We speculate that the existence of the second role can act as a safeguard against perturbations to ribosome function caused by spontaneous variations in DdS4 levels

Association of cetuximab with adverse pulmonary events in cancer patients: a comprehensive review

Compounds derived from biologic sources, or biologicals, are increasingly utilized as therapeutic agents in malignancy. Development of anti-cancer targeted therapies from biologics is increasingly being utilized. Cetuximab, a chimeric monoclonal antibody, is one such anti-cancer targeted therapeutic that has shown efficacy in quelling the rate of patient decline in colorectal, head/neck, and non-small cell lung cancer. However, due to the relatively recent addition of biologic compounds to the therapeutic arsenal, information related to adverse reactions is less well known than those seen in traditional chemotherapeutics. Dermatologic reactions have been demonstrated as the most frequent side effect cited during cetuximab therapy for malignancy; however, other effects may lead to greater morbidity. In general, pulmonary complications of therapeutics can lead to significant morbidity and mortality. The purpose of this review is to compile the various pulmonary side effects seen in patients treated with cetuximab for various malignancies, and to compare the incidence of these adverse reactions to standard therapies

Gene Expression Profiles of Beta-Cell Enriched Tissue Obtained by Laser Capture Microdissection from Subjects with Type 2 Diabetes

Background: Changes in gene expression in pancreatic beta-cells from type 2 diabetes (T2D) should provide insights into their abnormal insulin secretion and turnover. Methodology/Principal Findings: Frozen sections were obtained from cadaver pancreases of 10 control and 10 T2D human subjects. Beta-cell enriched samples were obtained by laser capture microdissection (LCM). RNA was extracted, amplified and subjected to microarray analysis. Further analysis was performed with DNA-Chip Analyzer (dChip) and Gene Set Enrichment Analysis (GSEA) software. There were changes in expression of genes linked to glucotoxicity. Evidence of oxidative stress was provided by upregulation of several metallothionein genes. There were few changes in the major genes associated with cell cycle, apoptosis or endoplasmic reticulum stress. There was differential expression of genes associated with pancreatic regeneration, most notably upregulation of members of the regenerating islet gene (REG) family and metalloproteinase 7 (MMP7). Some of the genes found in GWAS studies to be related to T2D were also found to be differentially expressed. IGF2BP2, TSPAN8, and HNF1B (TCF2) were upregulated while JAZF1 and SLC30A8 were downregulated. Conclusions/Significance: This study made possible by LCM has identified many novel changes in gene expression tha