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Research Papers: Gerotarget (Focus on Aging):
Transcriptome profiling reveals divergent expression shifts in brown and white adipose tissue from long-lived GHRKO mice
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Michael B. Stout1,*, William R. Swindell2,*, Xu Zhi3,4, Kyle Rohde4, Edward O. List5, Darlene E. Berryman5, John J. Kopchick5, Adam Gesing6, Yimin Fang7 and Michal M. Masternak4,8
1 Robert and Arlene Kogod Center on Aging, Mayo Clinic, Rochester, MN, USA
2 Department of Dermatology, University of Michigan, Ann Arbor, MI, USA
3 Center for Reproductive Medicine, Department of Obstetrics and Gynecology, Peking University Third Hospital, Beijing, China
4 College of Medicine, Burnett School of Biomedical Sciences, University of Central Florida, Orlando, FL, USA
5 Edison Biotechnology Institute and Heritage College of Osteopathic Medicine, Ohio University, Athens, OH, USA
6 Department of Oncological Endocrinology, Medical University of Lodz, Lodz, Poland
7 Geriatrics Research Laboratory, Department of Internal Medicine, Southern Illinois University School of Medicine, Springfield, IL, USA
8 Department of Head and Neck Surgery, The Greater Poland Cancer Centre, Poznan, Poland
* These authors have contributed equally to this work
Michal M. Masternak, email:
Michael B. Stout, email:
Keywords: brown adipose tissue, growth hormone, inflammation, metabolism, white adipose tissue, Gerotarget section
Received: August 09, 2015 Accepted: August 29, 2015 Published: September 21, 2015
Mice lacking the growth hormone receptor (GHRKO) exhibit improved lifespan and healthspan due to loss of growth hormone signaling. Both the distribution and activity of brown and white adipose tissue (BAT and WAT) are altered in GHRKO mice, but the contribution of each tissue to age-related phenotypes has remained unclear. We therefore used whole-genome microarrays to evaluate transcriptional differences in BAT and WAT depots between GHRKO and normal littermates at six months of age. Our findings reveal a unique BAT transcriptome as well as distinctive responses of BAT to Ghr ablation. BAT from GHRKO mice exhibited elevated expression of genes associated with mitochondria and metabolism, along with reduced expression of genes expressed by monocyte-derived cells (dendritic cells [DC] and macrophages). Largely the opposite was observed in WAT, with increased expression of DC-expressed genes and reduced expression of genes associated with metabolism, cellular respiration and the mitochondrial inner envelope. These findings demonstrate divergent response patterns of BAT and WAT to loss of GH signaling in GHRKO mice. These patterns suggest both BAT and WAT contribute in different ways to phenotypes in GHRKO mice, with Ghr ablation blunting inflammation in BAT as well as cellular metabolism and mitochondrial biogenesis in WAT.
Growth hormone (GH) is a pivotal modulator of postnatal growth, metabolism, and adiposity in mammals. More recent studies have also determined that the magnitude of GH action in mammalian species is a major determinant of healthspan and in some cases longevity [1, 2]. Transgenic GH-overexpressing mice develop phenotypes associated with premature aging and have a shortened lifespan , while human acromegaly patients display greater disease burden and compressed life expectancy . In contrast, Ames dwarf, Snell dwarf, and GH receptor-knockout (GHRKO) mice are protected from a variety of age-related diseases and live between 30-60% longer than their respective normal littermates [5-7]. The effects of genetic GH-deficiency and GH-resistance on human lifespan remains unclear, yet available evidence indicates these mutations confer protection from several age-related diseases including atherosclerosis, diabetes, and cancer [8-10]. Multiple interrelated mechanisms involving metabolic adjustments, reduced inflammation, and enhanced stress resistance contribute to the healthspan- and lifespan-extending effects of curtailed GH action [1, 2]. Interestingly, many of these mechanisms are closely linked with adipose tissue homeostasis, which is often perturbed with advancing age [11, 12].
Both white (WAT) and brown (BAT) adipose tissues undergo significant age-related functional changes in mice and humans [11, 12]. WAT, particularly subcutaneous depots, begin to display declines in preadipocyte replication and adipogenic potential in mid-to-late life [11, 13]. The aging process also promotes declines in BAT mass and activity [14-17], which may be a result of progenitor exhaustion, mitochondrial dysfunction, and/or perturbed endocrine control of brown adipocyte formation and function [18-20]. These declines are believed to be significant contributors to the age-related exacerbation of visceral adiposity, ectopic lipid deposition, and accompanying metabolic disorders [11, 12]. Long-lived GH-related mutant mice are protected from lipid redistribution and metabolic disorders with advancing age. We and others previously reported that Ames dwarf, Snell dwarf, and GHRKO mice preferentially deposit lipid in subcutaneous WAT depots, which is likely in response to the preservation of preadipocyte differentiation capacity [21-25]. Surprisingly, visceral WAT in GHRKO mice has a protective role by enhancing systemic insulin sensitivity and metabolic homeostasis , which lies in sharp contrast to other models [27, 28]. In alignment with these observations, BAT mass [29, 30] and thermogenesis-related gene expression also appear to be enhanced in GHRKO mice .
Although it is generally accepted that improvements in adipose tissue homeostasis in Ames dwarf, Snell dwarf, and GHRKO mice are mechanistically linked to reductions in inflammation and increased adiponectin expression [21, 22, 26, 31-33], it remains unclear if depot-specific changes occur and what role BAT modifications may play in this process. To address these questions, we evaluated transcriptional differences in WAT and BAT depots between GHRKO and normal littermates at six months of age. Our findings show that BAT and WAT have distinctive global gene expression profiles and that both types of adipose tissue respond differently to Ghr ablation. These response patterns involve modulation of inflammatory and metabolic pathways, with expression shifts in GHRKO BAT suggestive of decreased inflammation and heightened cellular metabolism. Our findings thus identify novel mechanisms by which BAT may mediate favorable effects of Ghr ablation on aging, and illustrate how such mechanisms may contrast sharply with those involving WAT.
BAT expression profiles are distinct from WAT and show divergent responses to Ghr ablation
We used in situ oligonucleotide whole-transcript arrays to profile gene expression in 48 adipose tissue samples from GHRKO (KO) and normal (N) littermate mice. An initial cluster analysis revealed no outliers among the 48 samples, but it was immediately clear that BAT samples were distinct from WAT depots (SubQ, EPI and PERI), regardless of genotype (Figure 1A). This same pattern was discernable when all samples (BAT + WAT) were plotted with respect to the first two principal component axes (Figure 1B). BAT expression profiles were thus distinct from all WAT depots analyzed. Among WAT depots, differences in gene expression profiles were less pronounced (Figures 1A and 1B), and in general, genotype (KO or N) had a stronger impact on WAT than anatomical location. However, cluster and principle components analyses of WAT samples alone (excluding BAT) revealed some distinction between EPI and SubQ, with PERI samples showing an intermediate “EPI/SubQ” expression profile (Supplemental Figure 1). The EPI/SubQ distinction was stronger in normal mice and attenuated in GHRKO animals (Supplemental Figure 1). For all WAT depots, however, fold-changes (KO/N) followed a moderate, yet significant, negative genome-wide correlation with those calculated for BAT (Figure 1C-1E). Global effects of Ghr ablation on gene expression in BAT thus tended to be opposite of those in WAT.
Figure 1: BAT shows distinct expression patterns and divergent expression shifts in GHRKO as compared to normal mice. A. The 48 samples were clustered based upon expression of 18,124 genes with detectable expression in at least 33% of samples (16 of 48 samples). The Euclidean distance between expression profiles was calculated and hierarchical clustering was performed using average linkage. B. Principal components plot. Samples were plotted with respect to the first two principal components extracted from the normalized expression matrix (18,124 genes × 48 samples). Parts C. - E. compare estimated fold-changes (KO/N) in BAT with those from C. EPI, D. PERI and E. SubQ. Each point represents an individual gene expressed in both depots being compared. Yellow circles encompass the 90% of genes closest to the bivariate mean (Mahalanobis distance), and the dashed red line represents the robust linear model fit. The Spearman correlation coefficient and associated p-value are listed in the lower right corner of C. - E..
Differential expression analysis was next performed to identify genes with significantly altered expression in GHRKO mice with respect to each adipose depot (Supplemental Table 1). The largest number of DEGs was found in SubQ WAT (153 increased; 193 decreased), followed by PERI WAT (129 increased; 190 decreased), BAT (78 increased; 202 decreased), and EPI WAT (48 increased; 144 decreased) (Figures 2A and 2B). Consistent with the above trends (Figure 1), many DEGs were shared among WAT depots, but few of these also overlapped with those identified in BAT (Figures 2A and 2B). For instance, aside from Ghr, only six genes were significantly decreased in all adipose tissue depots analyzed (Ifi27l2a, Hcar1, Hcar2, Fgf10, Sh3pxd2a, Slc25a10; Figure 2D). Only a few genes, moreover, were significantly elevated in BAT as well as one or more WAT depots (e.g., Orm3, Mmrn1, Tc2n, Gm10419; Figure 2C). More genes, however, could be identified with BAT-specific or WAT-specific changes in gene expression (Figures 2A and 2B). For instance, 74 DEGs were increased only in BAT (Figure 2A), while 178 DEGs were decreased only in BAT (Figure 2B). Analysis of DEGs again indicated that Ghr ablation elicits unique gene expression shifts in BAT as compared to WAT, consistent with trends observed at the global level of genome-wide expression (Figure 1A).
Figure 2: Shared or partially shared BAT/WAT expression changes in GHRKO mice as compared to normal controls. We identified 346, 319, 280 and 192 DEGs with respect to subcutaneous (SubQ), renal (PERI), brown (BAT) and epididymal fat (EPI) (FC > 1.50 or FC < 0.67 with FDR < 0.05). Venn diagrams show the numbers of shared and depot-specific DEGs either A. increased or B. decreased in GHRKO mice as compared to normal controls. Heatmaps show genes most consistently C. increased or D. decreased across all four depots. Genes were primarily sorted according to the number of depots in which they were differentially expressed (FC > 1.50 or FC < 0.67 with FDR < 0.05) and then secondarily sorted according to estimated fold-change (KO/N). The 25 top-ranked genes most consistently altered across fat depots are shown in C. and D..
Ghr ablation modulates BAT metabolic and inflammatory gene expression profiles
We identified a number of genes divergently altered by Ghr ablation in BAT and WAT depots. These included genes increased in BAT from GHRKO mice, despite being uniformly decreased in WAT depots (e.g., Gys2, Acss2, Me1, Abhd1, Echdc1; Figure 3A). Gene ontology (GO) biological process (BP) terms enriched among such genes were disproportionately related to metabolism, including organic acid metabolic process, oxidation-reduction process, cholesterol biosynthesis, and positive regulation of lipid & acetyl-CoA metabolic processes (Supplemental Figure 2A). Conversely, we also identified genes with reduced expression in BAT from GHRKO mice and increased expression in at least one WAT depot (e.g., Endod1, Kirrel, Cd74, Col12a1, Apol6, H2-Eb1, Trp53i11, Sema5a, Pkp2; Figure 3B). Such genes were disproportionately associated with immune and inflammation-related GO BP terms such as regulation of immune system process, cell motility and activation, innate immune response, inflammatory response, and wound healing (Supplemental Figure 2B).
Figure 3: Genes with divergent gene expression changes in BAT versus WAT (GHRKO versus normal mice). The 17,831 genes expressed in BAT and each WAT depot were filtered to identify those showing the most divergent gene expression changes in BAT as compared to WAT. A. DEGs increased in BAT and decreased/unaltered in WAT. GHRKO-increased genes (BAT) were filtered based upon the difference in fold-change estimated for BAT and the largest fold-change estimated for any WAT depot (log2 scale). B. DEGs decreased in BAT and increased/unaltered in WAT. GHRKO-decreased genes (BAT) were ranked based upon the difference in fold-change estimated for BAT and the largest fold-change estimated for any WAT (log2 scale). The 40 top-ranked genes with divergent expression shifts in BAT and WAT are shown in A. and B.
Ghr ablation curtails WAT expression of mitochondria-associated genes
In contrast to the robust increase in metabolism-related gene expression in BAT from GHRKO mice, WAT depots all displayed reduced expression of genes associated with mitochondria (Figure 4). DEGs with decreased expression in WAT, for instance, were significantly enriched with genes associated with mitochondrion, mitochondrial envelope, and mitochondrial inner membrane (Figure 4B-4D). This was not the case, however, for DEGs with decreased expression in BAT, which were instead associated with extracellular proteins (Figure 4A). Cluster analysis of 1,450 genes associated with the mitochondrion GO CC term (GO:0005739) revealed a distinct group of genes with decreased expression in all WAT depots, even though their expression was unaltered or elevated in BAT (Figure 5A). Examples of such mitochondrion genes included Cpt2, Tst, Slc25a1 and Cox8b, each of which was decreased in all WAT depots without corresponding changes in BAT (Figure 5B). Many other mitochondrion genes were decreased in at least two WAT depots and not similarly altered in BAT (Figure 5B).
Figure 4: Genes with GHRKO-decreased expression in WAT (but not BAT) are associated with mitochondria and the mitochondrion inner envelope. GHRKO-decreased DEGs were identified with respect to each fat depot and analyzed to identify significantly overrepresented GO cellular component (CC) terms. The top-ranked eight most enriched GO CC terms are listed for each depot. The number of GHRKO-decreased DEGs associated with each GO CC term is listed (left margin, within parentheses) along with exemplar GHRKO-decreased DEGs (right margin).
Figure 5: BAT and WAT show divergent mitochondrial gene expression changes in GHRKO as compared to normal controls. A. Cluster analysis of 1,450 BAT/WAT-expressed genes associated with the “mitochondrion” GO BP term (GO:0005739). Complete linkage hierarchical clustering was performed based upon the Euclidean distance between fold-change estimates (BAT, EPI, PERI and SubQ). B. Mitochondrion genes (GO:0005739) most consistently decreased in WAT from GHRKO mice. The 1,450 genes were primarily sorted to identify those most consistently differentially expressed in WAT depots (FC < 0.67 with FDR < 0.05) and secondarily sorted based upon fold-change (i.e., largest fold-change estimated for any WAT depot). The 40 top-ranked genes based upon these criteria are shown.
Anti-inflammatory gene expression changes in BAT from GHRKO mice: evidence for reduced dendritic cell and macrophage infiltration
Genes specifically decreased in BAT from GHRKO mice were associated with immune and inflammatory GO BP terms (Supplemental Figure 2B). One possible explanation for this pattern is that Ghr ablation leads to reduced immune cell infiltration of BAT in GHRKO mice, resulting in decreased expression of genes characteristically expressed by particular immune cell subsets. To address this possibility, we utilized the Immunological Genome Project (IGP) database  to determine whether genes with decreased expression in BAT included “signature genes” expressed primarily by cell types from the innate or adaptive immune systems (Figure 6).
GHRKO-decreased DEGs in BAT were significantly enriched with genes specifically expressed by DCs and macrophages (Figures 6A and 6E). For instance, GHRKO-decreased DEGs in BAT included H2-Ea-ps, C3ar1, Cxcl9, C7, C1qb and Ifi205; each of these genes was expressed at significantly higher levels in lung-derived DCs relative to all other cell types in the IGP database (i.e., IGP population DC.103-11b+.PolyIC.Lu; Figure 6E and Supplemental Figure 3A). Overall, GHRKO-decreased DEGs in BAT were significantly more likely to be specifically expressed by this DC population, as compared to all other BAT-expressed genes (P = 3.01 × 10-28; Wilcoxon rank sum test; Supplemental Figure 3B).
We repeated the same analysis in WAT but observed much weaker enrichment of immune cell-specific genes among GHRKO-decreased DEGs (Figures 6B-6E). GHRKO-decreased DEGs, for instance, showed no enrichment for genes specifically expressed by lung-derived DCs (P ≥ 0.727; Supplemental Figure 3C-3E). To the contrary, GHRKO-increased DEGs were modestly enriched for genes specifically expressed by DCs (PERI and SubQ; Figures 6C and 6D), suggesting possibly increased inflammatory status of these WAT depots. Interestingly, DEGs with elevated expression in WAT (PERI and SubQ) tended to be expressed by non-immune cell types, including stromal cells, fibroblastic reticular cells, skin fibroblasts, and blood endothelial cells (Figures 6C and 6D). For example, GHRKO-increased DEGs in SubQ were expressed at significantly elevated levels in blood endothelial cells as compared to all other IGP cell types (e.g., Akr1c14, Palmd, Shroom4, Prex2, Sorbs2; P = 8.08 × 10-14). These observations suggest increased DC infiltration of WAT in GHRKO mice, accompanied by heightened vascularity and possibly altered adipocyte differentiation status (PERI and SubQ).
Figure 6: Genes with decreased expression in BAT are enriched with genes specifically expressed by dendritic cells and macrophages (Immunological Genome Project). A. - D.GRHKO-increased and -decreased DEGs were screened with respect to 222 cell populations included within the Immunological Genome Project expression database. Enrichment statistics were calculated for each cell population and with respect to GHRKO-increased DEGs (horizontal axis) and GHRKO-decreased DEGs (vertical axis). For each cell population, enrichment statistics represent -log10-transformed p-values derived from the test of whether DEGs are disproportionately included among “signature genes” with a cell type-specific expression pattern (Wilcoxon rank sum test). Positive statistics indicate that DEGs are enriched with genes specifically expressed by a given cell type, while negative statistics indicate that DEGs are enriched with genes showing a non-specific expression pattern. In A. - D., each point represents an individual cell type, with different symbols for the main cell type categories included within the Immunological Genome Project (see legend). E. Cell populations for which signature genes most strongly overlap with GHRKO-decreased DEGs identified in BAT (DC = dendritic cell; st = stromal cell; MF = macrophage; Fi = fibroblast). Further information on each cell type is available online (http://www.immgen.org/). For each cell population, enrichment statistics (-log10-transformed p-values) are shown with respect to BAT, EPI, PERI and SubQ fat, respectively.
Curtailed GH action elicits beneficial effects on age-related diseases through a variety of mechanisms [1, 2]. Many of the enhancements observed in long-lived GH-related mutants are often associated with the preservation of adipose tissue homeostasis with advancing age [11, 12]. Our previous reports have established that subcutaneous WAT lipid storage capacity remains intact in GHRKO mice into late-life, which likely prevents age-related lipid redistribution and metabolic dysfunction [22-25]. Visceral WAT in GHRKO mice has also been reported to enhance metabolic homeostasis through alterations in lipid metabolism and adipokine production , which is certainly contrary to traditional dogma [27, 28]. Although fewer studies have evaluated the effects of Ghr ablation on BAT activity, GHRKO mice have been reported to have greater BAT mass [29, 30] and increased expression of genes responsible for thermogenic activity . These studies suggest GH may differentially modulate adipose tissue phenotypes in a depot-specific manner. Given that transcriptional alterations are likely associated with the previously described adipose tissue functional changes in GHRKO mice, we evaluated transcriptome profiles in WAT and BAT to determine if the beneficial effects associated with Ghr ablation is more closely aligned with specific depots.
Our analysis revealed distinct BAT and WAT expression profiles and divergent responses of each adipose type to Ghr ablation. Consistent with this, previous reports have also shown that WAT and BAT possess intrinsically different transcriptomes [39, 40], likely due to differences in progenitor lineage , resident immune cells , and metabolic phenotypes . Our findings extend this work and demonstrate that BAT and WAT respond differently to the altered in vivo endocrine status of GHRKO mice. This differential response involved genes associated with cellular respiration and metabolism, with GHRKO BAT showing increased expression of mitochondrial genes and GHRKO WAT featuring decreased expression of such genes. These expression shifts mirror quintessential differences in BAT versus WAT physiology. Whitening of adipose tissue during the early postnatal period in sheep, for instance, is characterized by decreased mitochondrial abundance and expression of genes involved in mitochondrial function . This may underlie physiological adipose shifts in GHRKO mice, evidenced in previous reports by enhanced BAT metabolism  and reduced WAT metabolism [22-24]. Such shifts may be connected to GHRKO aging and longevity in two ways. First, heightened BAT metabolism could improve energy dissipation, in some ways mimicking systemic consequences of low-calorie diets [44, 45], while leading to improvements in glucose tolerance and insulin sensitivity . Secondly, dampening metabolic processes in WAT may favorably impact oxidative stress levels , but perhaps more importantly, could reduce WAT-derived pro-inflammatory cytokines (e.g., IL-6 and TNF-α) .
GHRKO mice appear resistant to some favorable effects of caloric restriction (CR). For instance, GHRKO mice do not exhibit increased lifespan when provided a CR diet [49, 50]. Additionally, key expression markers of mitochondrial biogenesis are not further enhanced by CR in GHRKO mice . These important observations suggest CR and GH signaling repression improve longevity through mechanisms that at least partially overlap. In this context, it is interesting to note that an earlier proteomic study of rats has also identified divergent responses of BAT and WAT to CR . In contrast to our findings, however, CR increased biomarkers of mitochondrial activity in WAT, but did not alter or decreased the same biomarkers in BAT . These findings are contrasting with our own observations. This discrepancy may be due to our focus on the transcriptome, rather than the proteome, or by the fact that we studied mice, rather than rats. Alternatively, our findings and those of earlier work  may reflect a key difference between two well-studied aging interventions (CR and loss of GH signaling), wherein these two interventions have divergent, but opposite, effects on BAT and WAT metabolism. In some respects, this is not surprising, since CR and loss of GH action have opposing effects on systemic adiposity, with CR tending to reduce total adipose mass and curtailed GH signaling tending to increase it [22, 23, 49]. In future work, it may be valuable to compare CR responses of BAT and WAT in GHRKO and normal mice. For instance, assuming that CR and Ghr ablation have additive effects, we predict that BAT metabolism would be highest in ad lib-fed GHRKO mice, lowest in CR-fed normal mice, and intermediate in both ad lib-fed normal and CR-fed GHRKO mice.
Immune cells reside in adipose tissue and interactions involving these cell types play a significant role in modulating systemic metabolism and inflammatory status [42, 53, 54]. Genes with decreased expression in BAT were disproportionately associated with immunity and inflammation response (Supplemental Figure 2), suggesting attenuated BAT immune cell infiltration in GHRKO mice. Given the close associations between inflammation and mitochondrial dysfunction , we used data from the Immunological Genome Project  to computationally discern the identity of immune cell types contributing to this pattern [56-58]. This showed that genes with decreased expression in GHRKO BAT were often expressed by DC and macrophage populations. Both of these monocyte-derived cell types are commonly associated with adipose tissue and tend to become more numerous in adipose as a result of obesity . This inflammatory process has been best characterized in WAT (rather than BAT), although prior studies have identified increased BAT macrophage numbers in obese mouse genotypes . DCs are also commonly associated with adipose tissue, particularly in the obese state, and their presence appears to facilitate development of Th17 responses and obesity-associated insulin resistance [61, 62]. In GHRKO mice, reduced BAT infiltration by macrophages and/or DCs may dampen systemic inflammation with aging and contribute to insulin sensitivity. These immune cell types, moreover, are reported to have deleterious effects on adipose metabolism . Their reduced prominence in GHRKO BAT could contribute to the apparent enhancement of BAT mitochondrial activity discussed above. Interestingly, the same analysis in WAT revealed that immune cell populations were affected to a far lesser extent by Ghr ablation (Figure 6). In fact, in WAT we detected increased expression of genes expressed by non-immune cell types, particularly cells of stromal and fibroblastic origin. This may reflect a pro-proliferative and stem cell-like phenotype of GHRKO WAT, which may account for the greater progenitor differentiation capacity previously observed in young  and old adult GHRKO mice [24, 25]
Our analyses identified strong differences between BAT and WAT expression profiles, although we could also discern more subtle differences among the three WAT depots. At a genome-wide level, for instance, we could identify distinctive expression signatures for EPI and SubQ, even though each of these overlapped with and could not clearly be distinguished from PERI (Supplemental Figure 1). This result resonates with other studies that have documented transcriptional differences between visceral and subcutaneous WAT depots in rodents and humans [64-66]. It is interesting to note that EPI versus SubQ expression differences were relatively greater in controls and weaker in GHRKO mice (Supplemental Figure 1). The fact that the EPI/SubQ WAT distinction was weakened in GHRKO mice may reflect a unique aspect of WAT physiology in response to Ghr ablation. Previously, we showed that removal of visceral fat from normal mice improves insulin sensitivity and glucose homeostasis; however, these same benefits were absent when the same depots were removed from GHRKO animals . Therefore, in GHRKO mice, visceral WAT physiology may more closely resemble SubQ and not generate the same deleterious factors that shorten lifespan in normal mice. Physiological and functional studies will thus be needed in future work to better understand whether changes in adipose tissue are mechanistically linked to the improved healthspan and lifespan of GHRKO mice.
In summary, our study demonstrates that BAT and WAT depots respond to Ghr ablation in divergent manners. We found that BAT and WAT respond by modulating metabolic and inflammatory pathways in opposing fashions, with expression shifts in BAT suggestive of decreased inflammation and heightened cellular metabolism in GHRKO mice. Our findings point towards new mechanisms by which BAT may mediate favorable effects of Ghr ablation on aging, healthspan and lifespan, and identify several ways in which the physiological balance of BAT and WAT activity is perturbed with systemic loss of GH sensitivity.
MATERIALS AND METHODS
Animals and tissue collection
GHRKO and normal (N) littermate mice (GHR+/-; N = 6/group; heterogeneous background) were maintained at Southern Illinois University . All mice were housed 4-5 animals/cage at 22 ± 0.5°C on a 12:12-hour light-dark cycle. Mice had ad libitum access to standard laboratory chow and water. At six-months of age, mice were anesthetized with isoflurane and euthanized by cervical dislocation prior to dissection. WAT depots including inguinal (SubQ), epididymal (EPI), and perirenal (PERI) along with BAT were immediately excised, flash-frozen, and stored at −80°C for future analyses. All procedures were approved by the Institutional Animal Care and Use Committee of Southern Illinois University.
RNA was extracted from ~50 mg of frozen adipose tissues using RNeasy mini kits (Qiagen, Valencia, CA) according to the manufacturer’s protocol including on column DNase treatment to eliminate any DNA contamination. Samples were homogenized with zirconium oxide beads using a Bullet Blender Homogenizer (Next Advance, Averill Park, NY). Once homogenized adipose tissues were centrifuged at 12,000 x g for 10 min at 4°C and the upper lipid layer was excluded by transferring the supernatant to a new tube. Pure RNA was then extracted and eluted into 30 μl of nuclease-free water. RNA concentrations and purities were determined using an Epoch plate reader (BioTek, Winooski, VT). RNA samples with an A260/A280 ratio < 1.9 were re-extracted. Extracted RNA was stored at −80°C in preparation for microarray analyses.
RNA quality was assessed by the Agilent Bioanalyzer Nano chip (Agilent Technologies, Santa Clara, CA). Labeled single-stranded cDNA (sscDNA) was generated using 400ng of total RNA according to Whole-transcript (WT) Sense Target Labeling Assay protocol using Ambion’s WT Expression kit and GeneChip WT Terminal Labeling kit. Labeled sscDNA (5.5 µg) was then hybridized onto GeneChip Mouse Gene 1.0 ST Array (Affymetrix, Santa Clara, CA), which analyzes 28,853 gene transcripts using ~760,000 probe sets (on average 27 probes per gene). Staining and washing of the arrays were conducted using a Fluidics 450 station, and scanned using Affymetrix GeneChip Command Console Software (AGCC) and GeneChip® Scanner 3000 7G. All these procedures were conducted according to the manufacturer’s instructions.
Bioinformatics and statistics
Normalized gene expression values were calculated for the 48 arrays using the robust multichip average (RMA) algorithm (R package: oligo; R function: rma) . This generated log2-normalized expression values for 35,556 “transcript clusters”, with each cluster incorporating probes targeting one of 20,862 mouse genes (i.e., the “core” option was used with the oligo package rma function). For each sample, transcript clusters with expression scores beneath the 20th percentile were considered undetected. Based on this threshold, 28,445 clusters were detected (present) and 7,111 clusters were undetected (absent) in each sample. For subsequent analyses, a representative transcript cluster was chosen for each mouse gene, with the representative chosen as the cluster with highest average expression across all 48 arrays included in the analysis. This limited analyses to 20,862 clusters (one representative for each mouse gene). For differential expression testing, we further limited analyses to only those genes for which the associated transcript cluster was detected with respect to at least 3 of the 12 arrays analyzed for a given fat depot. This criterion yielded between 18,081 - 18,285 genes for each fat depot. These fat-expressed genes were subsequently analyzed for differential expression (GHR-/- vs. control) using linear models with empirical Bayes moderated t-statistics (R package: limma; R functions: lmFit and eBayes) . To correct for multiple hypothesis testing among the 18,081 - 18,285 genes analyzed with respect to a given depot, raw p-values associated with moderated t-statistics were adjusted using the Benjamini-Hochberg method to control the false discovery rate (R function: p.adjust) . Differentially expressed genes (DEGs) were identified with respect to each fat depot, respectively, based upon a fold-change (FC) threshold of 1.50 (or 1/1.50 = 0.67) and an FDR-adjusted p-value of 0.05.
AGCC, Affymetrix GeneChip Command Console; BAT, brown adipose tissue; BP, biological process; CC, cellular component; CR, calorie restriction; DC, dendritic cells; DEG, differentially expressed genes; EPI, epididymal; FC, fold-change; FDR, false discovery rate; Fi, fibroblast; GH, growth hormone; GHRKO, growth hormone receptor knockout; GO, gene ontology; IGP, Immunological Genome Project; KO, knockout; MF, macrophage; N, normal littermates; PERI, perirenal; RMA, robust multichip average; sscDNA, single-stranded cDNA; st, stromal cell; SubQ, inguinal; WAT, white adipose tissue; WT, whole-transcript.
The authors would like to thank Dr. Andrzej Bartke for providing the tissues for these analyses.
This work was supported by the National Institutes of Health (AG032290 and AG031736 to M.M.M.) and a Robert and Arlene Kogod Career Development Award in Aging Research (M.B.S.). W.R.S. is funded in part by the American Skin Association Carson Family Research Scholar Award in Psoriasis. X.Y. is funded in part by National Natural Science Foundation of China (81401201) and the Beijing Natural Science Foundation (7152154). A.G. is funded by Polish National Science Centre (DEC-2012/04/M/NZ4/00198; grant No. 507/1- 107- 05/507-10-050 of the Medical University of Lodz, Poland).
CONFLICTs OF INTEREST
The authors declare no conflicts of interest.
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13. Stout M, Tchkonia T, Kirkland J. The Aging Adipose Organ: Lipid Redistribution, Inflammation, and Cellular Senescence. In: Fantuzzi G, Braunschweig C, eds. Adipose Tissue and Adipokines in Health and Disease. Nutrition and Health: Humana Press; 2014:69-80.
14. Heaton JM. The distribution of brown adipose tissue in the human. J Anat. 1972;112:35-39.
15. Pfannenberg C, Werner MK, Ripkens S, Stef I, Deckert A, Schmadl M, et al. Impact of age on the relationships of brown adipose tissue with sex and adiposity in humans. Diabetes. 2010;59:1789-1793.
16. McDonald RB, Horwitz BA. Brown adipose tissue thermogenesis during aging and senescence. J Bioenerg Biomembr. 1999;31:507-516.
17. Rogers NH, Landa A, Park S, Smith RG. Aging leads to a programmed loss of brown adipocytes in murine subcutaneous white adipose tissue. Aging Cell. 2012;11:1074-1083.
18. Florez-Duquet M, McDonald RB. Cold-induced thermoregulation and biological aging. Physiol Rev. 1998;78:339-358.
19. Valle A, Guevara R, Garcia-Palmer FJ, Roca P, Oliver J. Caloric restriction retards the age-related decline in mitochondrial function of brown adipose tissue. Rejuvenation Res. 2008;11:597-604.
20. Nedergaard J, Cannon B. The changed metabolic world with human brown adipose tissue: therapeutic visions. Cell Metab. 2010;11:268-272.
21. Wang Z, Al-Regaiey KA, Masternak MM, Bartke A. Adipocytokines and lipid levels in Ames dwarf and calorie-restricted mice. J Gerontol A Biol Sci Med Sci. 2006;61:323-331.
22. Berryman DE, List EO, Coschigano KT, Behar K, Kim JK, Kopchick JJ. Comparing adiposity profiles in three mouse models with altered GH signaling. Growth hormone & IGF research : official journal of the Growth Hormone Research Society and the International IGF Research Society. 2004;14:309-318.
23. Berryman DE, List EO, Palmer AJ, Chung MY, Wright-Piekarski J, Lubbers E, et al. Two-year body composition analyses of long-lived GHR null mice. J Gerontol A Biol Sci Med Sci. 2010;65:31-40.
24. Stout MB, Tchkonia T, Pirtskhalava T, Palmer AK, List EO, Berryman DE, et al. Growth hormone action predicts age-related white adipose tissue dysfunction and senescent cell burden in mice. Aging (Albany NY). 2014;6:575-586.
25. Stout MB, Tchkonia T, Kirkland JL. Growth hormone in adipose dysfunction and senescence. Oncotarget. 2015;6:10667-10668.
26. Masternak MM, Bartke A, Wang F, Spong A, Gesing A, Fang Y, et al. Metabolic effects of intra-abdominal fat in GHRKO mice. Aging cell. 2012;11:73-81.
27. Muzumdar R, Allison DB, Huffman DM, Ma X, Atzmon G, Einstein FH, et al. Visceral adipose tissue modulates mammalian longevity. Aging Cell. 2008;7:438-440.
28. Tran TT, Yamamoto Y, Gesta S, Kahn CR. Beneficial effects of subcutaneous fat transplantation on metabolism. Cell Metab. 2008;7:410-420.
29. Li Y, Knapp JR, Kopchick JJ. Enlargement of interscapular brown adipose tissue in growth hormone antagonist transgenic and in growth hormone receptor gene-disrupted dwarf mice. Exp Biol Med (Maywood). 2003;228:207-215.
30. Egecioglu E, Bjursell M, Ljungberg A, Dickson SL, Kopchick JJ, Bergstrom G, et al. Growth hormone receptor deficiency results in blunted ghrelin feeding response, obesity, and hypolipidemia in mice. Am J Physiol Endocrinol Metab. 2006;290:E317-325.
31. Al-Regaiey KA, Masternak MM, Bonkowski M, Sun L, Bartke A. Long-lived growth hormone receptor knockout mice: interaction of reduced insulin-like growth factor i/insulin signaling and caloric restriction. Endocrinology. 2005;146:851-860.
32. Lubbers ER, List EO, Jara A, Sackman-Sala L, Cordoba-Chacon J, Gahete MD, et al. Adiponectin in mice with altered GH action: links to insulin sensitivity and longevity? J Endocrinol. 2013;216:363-374.
33. Sackmann-Sala L, Berryman DE, Lubbers ER, Zhang H, Vesel CB, Troike KM, et al. Age-related and depot-specific changes in white adipose tissue of growth hormone receptor-null mice. J Gerontol A Biol Sci Med Sci. 2014;69:34-43.
34. Panici JA, Wang F, Bonkowski MS, Spong A, Bartke A, Pawlikowska L, et al. Is altered expression of hepatic insulin-related genes in growth hormone receptor knockout mice due to GH resistance or a difference in biological life spans? J Gerontol A Biol Sci Med Sci. 2009;64:1126-1133.
35. Carvalho BS, Irizarry RA. A framework for oligonucleotide microarray preprocessing. Bioinformatics. 2010;26:2363-2367.
36. Smyth GK. Linear models and empirical bayes methods for assessing differential expression in microarray experiments. Stat Appl Genet Mol Biol. 2004;3:Article3.
37. Benjamini Y, Hochberg Y. Controlling the False Discovery Rate - a Practical and Powerful Approach to Multiple Testing. J Roy Stat Soc B Met. 1995;57:289-300.
38. Kim CC, Lanier LL. Beyond the transcriptome: completion of act one of the Immunological Genome Project. Curr Opin Immunol. 2013;25:593-597.
39. Unami A, Shinohara Y, Kajimoto K, Baba Y. Comparison of gene expression profiles between white and brown adipose tissues of rat by microarray analysis. Biochem Pharmacol. 2004;67:555-564.
40. Rosell M, Kaforou M, Frontini A, Okolo A, Chan YW, Nikolopoulou E, et al. Brown and white adipose tissues: intrinsic differences in gene expression and response to cold exposure in mice. Am J Physiol Endocrinol Metab. 2014;306:E945-964.
41. Timmons JA, Wennmalm K, Larsson O, Walden TB, Lassmann T, Petrovic N, et al. Myogenic gene expression signature establishes that brown and white adipocytes originate from distinct cell lineages. Proc Natl Acad Sci U S A. 2007;104:4401-4406.
42. Ferrante AW, Jr. The immune cells in adipose tissue. Diabetes Obes Metab. 2013;15 Suppl 3:34-38.
43. Basse AL, Dixen K, Yadav R, Tygesen MP, Qvortrup K, Kristiansen K, et al. Global gene expression profiling of brown to white adipose tissue transformation in sheep reveals novel transcriptional components linked to adipose remodeling. BMC Genomics. 2015;16:215.
44. Bloor ID, Symonds ME. Sexual dimorphism in white and brown adipose tissue with obesity and inflammation. Horm Behav. 2014;66:95-103.
45. Gao AW, Houtkooper RH. Mitochondrial fission: firing up mitochondria in brown adipose tissue. Embo J. 2014;33:401-402.
46. Poher AL, Altirriba J, Veyrat-Durebex C, Rohner-Jeanrenaud F. Brown adipose tissue activity as a target for the treatment of obesity/insulin resistance. Front Physiol. 2015;6:4.
47. Paglialunga S, Ludzki A, Root-McCaig J, Holloway GP. In adipose tissue, increased mitochondrial emission of reactive oxygen species is important for short-term high-fat diet-induced insulin resistance in mice. Diabetologia. 2015;58:1071-1080.
48. Forsythe LK, Wallace JM, Livingstone MB. Obesity and inflammation: the effects of weight loss. Nutr Res Rev. 2008;21:117-133.
49. Bonkowski MS, Rocha JS, Masternak MM, Al Regaiey KA, Bartke A. Targeted disruption of growth hormone receptor interferes with the beneficial actions of calorie restriction. Proc Natl Acad Sci U S A. 2006;103:7901-7905.
50. Arum O, Bonkowski MS, Rocha JS, Bartke A. The growth hormone receptor gene-disrupted mouse fails to respond to an intermittent fasting diet. Aging Cell. 2009;8:756-760.
51. Gesing A, Masternak MM, Wang F, Joseph AM, Leeuwenburgh C, Westbrook R, et al. Expression of key regulators of mitochondrial biogenesis in growth hormone receptor knockout (GHRKO) mice is enhanced but is not further improved by other potential life-extending interventions. J Gerontol A Biol Sci Med Sci. 2011;66:1062-1076.
52. Okita N, Hayashida Y, Kojima Y, Fukushima M, Yuguchi K, Mikami K, et al. Differential responses of white adipose tissue and brown adipose tissue to caloric restriction in rats. Mech Ageing Dev. 2012;133:255-266.
53. Grant RW, Dixit VD. Adipose tissue as an immunological organ. Obesity (Silver Spring). 2015;23:512-518.
54. Brestoff JR, Artis D. Immune regulation of metabolic homeostasis in health and disease. Cell. 2015;161:146-160.
55. Currais A. Ageing and inflammation - A central role for mitochondria in brain health and disease. Ageing Res Rev. 2015;21:30-42.
56. Swindell WR, Johnston A, Gudjonsson JE. Transcriptional profiles of leukocyte populations provide a tool for interpreting gene expression patterns associated with high fat diet in mice. PLoS One. 2010;5:e11861.
57. Swindell WR, Johnston A, Sun L, Xing X, Fisher GJ, Bulyk ML, et al. Meta-profiles of gene expression during aging: limited similarities between mouse and human and an unexpectedly decreased inflammatory signature. PLoS One. 2012;7:e33204.
58. Swindell WR, Johnston A, Voorhees JJ, Elder JT, Gudjonsson JE. Dissecting the psoriasis transcriptome: inflammatory- and cytokine-driven gene expression in lesions from 163 patients. BMC Genomics. 2013;14:527.
59. Lee BC, Lee J. Cellular and molecular players in adipose tissue inflammation in the development of obesity-induced insulin resistance. Biochim Biophys Acta. 2014;1842:446-462.
60. Herrero L, Shapiro H, Nayer A, Lee J, Shoelson SE. Inflammation and adipose tissue macrophages in lipodystrophic mice. Proc Natl Acad Sci U S A. 2010;107:240-245.
61. Bertola A, Ciucci T, Rousseau D, Bourlier V, Duffaut C, Bonnafous S, et al. Identification of adipose tissue dendritic cells correlated with obesity-associated insulin-resistance and inducing Th17 responses in mice and patients. Diabetes. 2012;61:2238-2247.
62. Chen Y, Tian J, Tian X, Tang X, Rui K, Tong J, et al. Adipose tissue dendritic cells enhances inflammation by prompting the generation of Th17 cells. PLoS One. 2014;9:e92450.
63. Olarescu NC, Berryman DE, Householder LA, Lubbers ER, List EO, Benencia F, et al. GH action influences adipogenesis of mouse adipose tissue-derived mesenchymal stem cells. J Endocrinol. 2015;226:13-23.
64. Gesta S, Bluher M, Yamamoto Y, Norris AW, Berndt J, Kralisch S, et al. Evidence for a role of developmental genes in the origin of obesity and body fat distribution. Proc Natl Acad Sci U S A. 2006;103:6676-6681.
65. Yamamoto Y, Gesta S, Lee KY, Tran TT, Saadatirad P, Kahn CR. Adipose depots possess unique developmental gene signatures. Obesity (Silver Spring). 2010;18:872-878.
66. Vohl MC, Sladek R, Robitaille J, Gurd S, Marceau P, Richard D, et al. A survey of genes differentially expressed in subcutaneous and visceral adipose tissue in men. Obes Res. 2004;12:1217-1222.
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domain: medical


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(LifeSiteNews) — According to a prominent Jesuit priest and former chaplain of the U.S. House of Representatives, supporting a woman’s right to “choose” to kill her unborn child is a “Catholic value.”
Defying 2,000 years of Church teaching, and the Catholic biblical view on the sanctity of human life, Fr. Pat Conroy SJ told the Washington Post in an interview that “a good Catholic” support a woman’s right to “choose” to commit an abortion.
“It’s an American value that each one of us can choose where our life is going,” stated Conroy on the topic of abortion. “That happens to be a Catholic value, too. That we should all use our gifts and our talents and our intelligence as best we can to make the best choices we have the freedom to make.”
“A good Catholic in our system could be saying: Given women in our system have this constitutional right [to abort their child], our task as fellow Christians, or as Catholics, is to make it possible for her to optimize her ability to make the choice,” he added.
Despite Conroy’s position, the Catholic Church authoritatively and definitively holds and has always held that abortion is an “abominable crime” and that “life must be protected with the utmost care from the moment of conception.”
In fact, even those who merely support abortion in the political realm and have never been directly involved in procuring, performing, or obtaining an abortion, are in a state of grave sin and receive the highest juridical penalty under the Church’s Canon law system, namely excommunication.
Justifying his rejection of the authentic teaching of the Catholic faith, Conroy twisted the writings of St. Thomas Aquinas to the interviewer in an attempt to rationalize his support for the murder of innocent human life.
“Thomas Aquinas says if your conscience says to do something the church says is a sin, you are bound to follow your conscience. That’s Thomas Aquinas!” mispresented Conroy.
In reality, Aquinas held that the Catholic Church is the supreme authority on issues of faith and morals, and while culpability is certainly lessened if someone sins without realizing they are sinning, there is still a moral obligation to seek to form one’s conscience in accordance with authoritative Church teaching.
“To say that one is bound to follow an erring conscience is not to say that one is morally justified in doing any- and everything one is inclined to do. If every action was morally right just because one wanted to do it, it would be logically impossible to commit any morally wrong act; and this would empty ‘morally right and wrong’ of any meaning,” explained Dr. Joseph Magee, whose PhD is in Thomistic Philosophy. “Moreover, everyone knows that certain actions by individuals just are morally wrong, and the individuals who did them are guilty of wrongdoing, despite the fact that the perpetrator thought it was a good thing … In such cases, even an ‘erring conscience’ does not justify or excuse the moral evil.”
“But short of having the subjective conviction (after exercising due diligence to form one’s conscience well) that one has to act contrary to Church authority, or that one may not morally obey Church authority, that is, if one is not sure one is right, one has (Aquinas would say) an erring opinion (not an erring conscience) (see ST II-II, q.5, a.3), and in that case, one is morally bound not to act contrary to a trustworthy authority (e.g., the Church),” added the Aquinas expert.
“Such a person would have a doubtful conscience, and one morally cannot act with such doubts, any more than a hunter may shoot at a rustling bush if it might be hiding another hunter and not a deer.”

domain: news


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Casino Blackjack Online Vs Blackjack Tournaments Help The Differences
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If the supplier’s open card is Ace, a participant can get an insurance coverage throughout the vary of half of of the guess amount. If the player just isn’t happy with the primary 2 playing cards, he/she will receive extra card with a hand sign. It is a technique to express oneself if the participant doesn’t want additional card, and the supplier should stay if the sum of the playing cards is 17 or higher. Quality There is always a seat at these exclusive, branded Bet365 baize tables, in games the place 21 is the successful number. With the highest high quality video streaming, Playtech has definitely come to the “high quality” get together with the software program platforms for these real-time video games. This is a fully useful blackjack simulator with a number of consumer modifiable options (i.e. variety of decks in use, recreation pace, initial bankroll, etc). The simulator will recommend the participant’s appropriate course of action for every determination primarily based on the card depend.
If there is a tie, the player’s wager will remain as it is and the hand shall be over. It refers to when the participant’s and supplier’s sum of cards are the identical.
What does 8 mean in blackjack?
If any player cannot go, they pick up the top card from the discard pile. The wild cards are as follows: eights make the next player miss a go (two eights makes two miss a go etc), king reverses direction of play, but both of these must be played on the right suit or rank.
At the time all coup have been skilled throughout the online game, a winner would be announced and even he/she would be awarded often the prize cash. Similarly to every Blackjack sport, you may need competed, you’ll might want to select your private transfer properly in a event scenario. Our specialists at CasinoTop10 most suggest sticking to typically the Blackjack method sheet as a way to choose whether or not to Hit, Endure, Split, Two instances Down or Surrender. On online Play on-line blackjack Tournaments, you’ll need to choose a move in a given time limit clause.
Your Guide to Rules, Strategy – nj.com
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Unlike inside different events the place particles usually have zero value, Live life Tournaments normally have worth. In advance of playing through the event, game lovers must get hold of chips that they’ll swap for earnings at the highly finish of your Live Money Blackjack Competition. While Blackjack Tournaments are primarily based on the online game of playing establishment Blackjack, there are a selection notable distinctions between the frequent model from the sport and tournaments. For the newcomers, slot machines are among few decisions left, however even these are totally occupied.
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To play Casino Poker Blackjack Slots you merely pick the sort of recreation you like finest, examine whether you could have sufficient digital cash, and start having enjoyable. If you play your playing cards proper you’ll be able to go away the game with way more money than you began with, and if you want, can invest it to enter more expensive and demanding but additionally far more fun leagues. Black-jack is certainly an gratifying betting home online video games that allows recreation lovers to not ever sole execute only for enjoyable however may also return out of your most-favored on line and smartphone on line casino. Players want to get a general card worth of 21 and even to come back nearer to it, with out going over, than the vendor, towards whom all playing is completed. For this operate aces rely as 1 or 11, face playing cards as 10, and the remaining at their catalog value. A hand counting 21 on the preliminary two cards is known as a pure or maybe a blackjack. If the vendor doesn’t have a Blackjack, the participant loses solely the insurance wager.
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In blackjack, the house edge comes from the dealer having a hidden card about which the player has to make assumptions. In addition, if the player busts, the dealer wins, even if they then bust in the same round. Learning blackjack odds and strategy makes this less of an issue.
Through sport, details from every individual round ordinarily advance to a different round. This procedure repeats alone till on-line players finally get to the last rounded and a champ is printed. Once players sit down, they don’t depart shortly and a few maintain bundles of 50,000-won and 10,000-received notes of their arms.
The Way To Play Competitors Blackjack
Any participant, including the dealer, whose hand worth exceeds 21 mechanically loses the sport, and this is referred to as a ‘Bust’. Still, the aspect gathered like that is certainly minuscular so it isn’longer properly price your effort inside slightest. The variety of the wheel is indicated on the desk format and the player can guess there. For winning bet, payouts are made accordingly and the vendor takes all the dropping bets.
The numbers on the wheel are indicated on the desk structure and the participant could choose the wager he/she desires. If the ball lands on exact quantity the player wagered, the participant will get payouts accordingly. If there are lots of gamers, every participant will use different colored play chips to stop confusion. Roulette is among the most popular on line casino games that even beginners can easily enjoy themselves without any previous experience. The dealer spins the ball and the player guess on the variety of the pocket where the ball will land. If the preliminary two playing cards drawn by the vendor or participant has an ace and a ten worth card, the hand totals 21 and is known as a “Blackjack”. When a participant has a Blackjack, the player shall be paid three to 2(1.5times) and can’t lose.

domain: game


Before we dig into the Support Debugging Tool, we need to install it. We will do that here and turn on the Administrator mode so that we don't have to revisit this with each new recipe.
To install and set up the Support Debugging Tool, perform the following steps:
DEBU_1200.zipfile from Microsoft via a Microsoft partner.
DEBU_1200.zipfile, select Extract All, and follow the prompts to unzip the file.
CNKfile from the new
DEBU_1100folder to the location where Dynamics GP is installed. By default this is
C:\Program Files (x86)\Microsoft Dynamics\GP 2013\or C:\Program Files\Microsoft Dynamics\GP 2013\on a 32-bit machine. ...


Toto Wolff addresses speculations about Jérôme d’Ambrosio potentially leaving Mercedes, citing personal reasons behind the Belgian’s contemplation.
Wolff addresses questions regarding Jérôme d’Ambrosio at Mercedes: he cites “personal reasons” for the Belgian’s contemplation of his future.
Is Jérôme d’Ambrosio poised to leave Mercedes for Ferrari? This rumor has been swirling in the paddock for several days, and Toto Wolff, Mercedes’ team principal, has commented on his close associate’s situation.
Wolff reminded that d’Ambrosio remains with the German squad for now. However, the Silver Arrows’ director hinted at “personal reasons” prompting the Belgian to consider his future.
“It’s up to him to discuss,” Wolff stated, while ensuring that a “good atmosphere” still prevails within the team. “We’ll see where it leads us,” he added, according to remarks reported by Crash.net.
This uncertain situation could be due to d’Ambrosio’s contract ending at the year’s close. Some sources suggest the 38-year-old former driver wishes to return to Europe.
It’s worth noting d’Ambrosio’s significant role at Mercedes in recent years. Joining in 2022, he notably filled in for Wolff as team principal during the Japanese and Qatar Grands Prix in 2023. His contributions have also extended to the development of young drivers, a crucial area for the Silver Arrows squad.
A move to Ferrari, therefore, cannot be ruled out, especially since d’Ambrosio is no stranger to the Scuderia’s circle. Before joining Mercedes, he worked alongside Susie Wolff, Toto’s wife, at the Venturi Formula E team.
His F1 driving career was relatively short-lived. Starting in 2011 with Virgin Racing, it concluded at the 2012 Italian Grand Prix, where he replaced Romain Grosjean, who was serving a suspension.
Whether d’Ambrosio’s journey with Mercedes will end soon remains to be seen. The answer will likely come from the driver himself in the coming weeks. Meanwhile, Toto Wolff remains circumspect, keeping the speculation alive.
d’Ambrosio Mercedes Ferrari Future. d’Ambrosio Mercedes Ferrari Future


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Yes Bingo, a relatively small online bingo site have recently made changes to their bonus situation in an attempt to throw themselves amongst the larger fish in the pond. Ever seeking to boost the numbers of players at their site, the online bingo site has recently included a bonus money wheel into the mix at their domain.
Towards the later part of 2012, bonus wheels were becoming vastly popular at online bingo sites and many of us correctly predicted that this phenomenon would continue on into 2013. Yes Bingo, evidently certainly thought so too. Their bonus wheel isn’t rewarding new players with a welcome bonus, but is instead designed for already existing players at their online bingo site.
Already existing members of Yes Bingo will be able to pocket up to £20 per day by spinning the bingo bonus wheel.
If that wasn’t enough to spice things up at the online bingo site however, they have also made a few other changes in an attempt to liven up the atmosphere and draw in the masses. Players at the bingo domain will also be offered the chance to pocket up to £25 daily via guaranteed bingo jackpot games which will costs a mere five pence per ticket.
The bingo site has also taken steps towards offering a free bingo scheme, where up to a £25 could be offered for the victor, as well as extending the value of their progressive jackpot 75 and 90 ball bingo games, although players are already bagging up to half a million pounds between them per month with this offer.
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text:
Quick Plastic Forming (QPF) is a recent adaptation of Superplastic Forming (SPF) that allows economical fabrication of complex components using superplastic material. QPF requires refined, equiaxed grains and high-angle grain boundaries in the microstructure to enhance sheet deformation by GBS at the high strain rates involved. This study evaluates the effects of laboratory rolling conditions on continuously cast AA5083 in the hot-band condition in anticipation of QPF. Orientation imaging microscopy, scanning electron microscopy and X-ray analysis were used to analyze roles of the geometric dynamic recrystallization and particle stimulated nucleation of recrystallization in the microstructure evolution during rolling. A refined microstructure was developed during the rolling procedures but mechanical property data indicated low to moderate ductility and failure by excessive cavity formation. Factors influencing the development of microstructure and mechanical properties during laboratory rolling were investigated. / US Navy (USN) author.
|Thompson, Matthew F.
|McNelley, Terry, Swaminathan, Srinivasan, Naval Postgraduate School (U.S.)
|Monterey, California. Naval Postgraduate School
|Naval Postgraduate School
|xvi, 41 p. : ill. (some col.) ;, application/pdf
|Approved for public release, distribution unlimited
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domain: literature


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Sense…I have none when it comes to spelling
Ventura is a nut and a 9/11 troofer.
He’s even wrong about Limbaugh receiving government health care in Hawaii.
My friend …you calling someone a nut is like…Pot…Kettle…Black. But it’s cool I somehow knew you’d be the first to comment.
Are you a troofer George?
The problem with populists like this is that they form their ideas before they learn the facts and then never let anything change thier minds.
No wonder Jesse only served for a single term.
He may be a nutjob and may have sucked as a Governor, but let his comments stand on their own merit.
Is he right about the Dems trampling on the Constitution? YES
Is he right about the Repubs trampling on the Constitution? YES
Fact is both parties whittle away at Constitutional restraints on the Federal Government at every opportunity - they just do it from opposite sides. The reality is that they both suck, but they’re really still the only game in town.
Is he right about using Hawaii as an example for why we need Federal healthcare? NO (apparently he fails his own Constitutional argument).
I can’t speak to whether HI really has state run healthcare or not or whether it’s any good if they do, but it doesn’t matter. The Constitution specifically limits the power of the Federal Government (Healthcare is not one of the enumerated powers of the Fed). On the other hand there are no such restrictions on the individual states. If HI or any other state wants to establish state run and funded healthcare, there is no U.S. Constitutional constraint preventing them from doing so, as state’s rights are not limited by the U.S. Constitution except for those specific powers which are reserved for the Fed. If some states have state run HC and some don’t, people can choose what state they want to live in. The population will vote with their feet. The same reason we see such mass exodus of businesses, jobs and people from states with oppressive policies.
It’s amazing how many of our state and federal politicians do not understand the fundamental differences in how the Constitution addresses Federal vs. state powers, yet it’s conceptually so simple: If the U.S. Constitution doesn’t say that the Fed can do something, it’s not supposed to be allowed to; Unless the U.S. Constitution specifically limits powers of the states, they can do whatever they choose in accordance with their own Constitution and laws.
The national defense of our country comes first… economics are close 2nd.
Health care is not a constitutional right…however the quickest fix is simply treat it as a commodity and tear down state regulation barriers that prevent US citizens from seeking coverage from whomever they may choose…including getting medication from outside sources other than US approved companies.
Jesse is indeed right about both parties… they both suck…all parties should be abolished and one simply votes for the individuals based upon their convictions they put forth. What is so hard about simply advertising that; My name John Doe, I believe in guns, state rights,… I am against abortion, taxes, and 90% of government run programs…blah, blah, blah,…vote for me in November (its not that difficult).
Institute term limits and that no lawyer can run for office and we would do 1000% better than what we have now.
A troofer is no better than a broken clock.
Just because it’s right twice a day doesn’t mean you can tell time with it.
That’s completely true. Rush Limbaugh should just listen to himself sometime. Oh, you were talking about Jesse V, not Rush!??
I kinda think they do know the difference–they just ignore the provisions for their own benefit. The Federal Government doesn’t want to give any authority or rights to the states if they can benefit from it themselves.
Which is exactly how “The War of Northern Aggression” got started. :twisted:

domain: law


text:
‘Reservoir’ by Michael Walker. Buy now.
‘Dogs of Teotihuacán’ by Michelle Loren. Buy now.
Feature Shoot has opened a photography store! After a decade of showcasing thousands of emerging and established photographers, we’ve launched Superfine Prints, a growing collection of imagery from some of the most popular and pioneering artists featured on our platform. We’ve always cared about preserving the value of physical prints, and we’re thrilled to make this step from your screens into your homes.
Superfine Prints is carefully curated by Alison Zavos, the Founder and Editor-in-Chief of Feature Shoot. The images in the store reflect the high standard you’ve come to associate with Feature Shoot, and the growing collection includes photographs we’d love to see on our own walls. All of the photographs available at Superfine Prints are exclusive and limited edition, and our affordable prices are perfect for any collector.
As with our website, we strive to represent a diverse, international group of artists who transcend geographical, social, and political boundaries through the language of photography. So far, Superfine Prints is home to photographers hailing from around the world, including the United States, Canada, Mexico, Brazil, Australia, Hong Kong, Tokyo, England, France, Spain, Ireland, Germany, Turkey, Poland, Ukraine, the Netherlands, Denmark, Norway, Sweden, and Greece. We use high quality, acid-free, pH-neutral archival paper for all our prints, ensuring products that are both beautiful and longlasting.
Superfine Prints is brand-new, so be sure to visit our website to see all the photos available, and follow along on Instagram for updates. We are offering free shipping for the first 100 orders, so act fast and use the code ‘FREESHIP’ at checkout. As our store continues to grow, please keep in mind that all photographers who participate in The Print Swap going forward will have the opportunity to join the Superfine Prints roster and sell their work. Tag your images #theprintswap to be considered for our global photo-sharing project, and visit The Print Swap website to learn how it works.
‘First Snow’ by Nigel Agar. Buy now.
‘Horses do make the landscape more beautiful’ by Kellie Kvern. Buy now.
‘The Birds’ by Brittany Kelley. Buy now.
‘Subterranean Homesick Alien’ by Andrea Lambe. Buy now.
‘Once Today’ by Christine Ruf. Buy now.
‘Golden Gate Bridge, 2017’ by Gwen Heeney. Buy now.
‘Umbrellas at the Opera’ by Chelsea London. Buy now.
‘Square.’ by Andreas Godwin. Buy now.
‘Wyeth and Idaho’ by Brad Curran. Buy now.
‘Fallen elevated water tank, 2017’ by Alfredo Esparza. Buy now.
‘Ella as Dorothy’ by Cathy Ronalds. Buy now.
‘Tian Tan Buddha – Pailou’ by Grzegorz Kurdziel. Buy now.
‘Nesodden’ by Stein Jarle Nilsen. Buy now.

domain: geography


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I thought Up a Rutted Road was progressing just fine. One editor with a major publishing house had looked at it and liked it. "But our middle-grade list is already too full," she wrote in the rejection letter.
Another editor was excited about the proposal. "I want to see the full manuscript when it is finished," she said. Unfortunately, she went on an extended medical leave before she could see the completed project.
Then I sent it to an agent with whom I had chatted at a conference. "It lacks a sense of place," he said. How dare he attack my opus, my firstborn! (One tends to mix metaphors when one is upset.)
Initial reaction aside, I began to ponder what the agent had said. Could he be right? I read it through and realized that he was. And so began the laborious revision, still in process.
Up a Rutted Road tells the story of Camie, who spends her tenth summer in the southeastern mountains of Kentucky on the hardscrabble farm of her Uncle Glen and Aunt Charlene. Before summer's end, she must confront death and her own feelings of guilt.
"It lacks a sense of place," the agent had said. How could I correct that? I had to become more familiar with certain elements in the story so that I could accurately show them to the reader.
Elsie Blue, for example. I knew from the beginning that she was a blue-tick coon hound. I knew her by yowl, but where were her spots? What exactly was her coloring? I only needed to do a quick search of the web to find her! Now, I keep her picture before me whenever she's in a scene.
I also went in search of Uncle Glen and Aunt Charlene's "old-timey" house, its siding weathered to an ashen gray. Once I found it, I drew the interior layout. Finally, I had a house I could accurately describe.
The next step was to interview a close friend and fellow storyteller who grew up in southeastern Kentucky. He told me about the coming together of the highlanders in the gloaming, to meet on a neighbor's porch and sing or play various folk instruments-- fiddles, home-made banjos, Jew's harps, mountain dulcimers, etc.--sending the music bouncing around from one mountainside to another and down in the hollows. That luscious story will find its way into the pages of Up a Rutted Road.
Finally, I reached deep down into my own fear, returned to my childhood, and looked out of the window of my father's car to where a mountain road appeared to drop away, leaving nothing but air between the car and the fast-flowing creek far below. I gave my churning stomach and my dizzy head to Camie for a moment in Chapter One, as she looks over just such a cliff.
Writers love that first passionate awen that produces the initial draft of a manuscript. Beginning writers resent and reject the various revisions that are necessary to arrive at a polished work fit for submission. They have not yet learned the value of an agent, an editor, or a critiquer who dares to say, "You need..." or "It lacks..." I have come to the point where I invite such comments and actually enjoy the edits and revisions, because the story is worth it.

domain: library


text:
Christopher Walken is to make his debut on a UK TV series, starring in The Offenders, a new comedy series from Stephen Merchant on the BBC.
Here's our first look at Trainspotting star Kelly Macdonald, set to make her debut in hit BBC police drama Line of Duty.
John Barrowman's Captain Jack Harkness will be back for the Doctor Who Christmas special, it's been confirmed.
Chris Chibnall says the 'complexity' of shooting in the era of coronavirus means each 'Doctor Who' episode will take longer to make.
Jodie Whittaker has described her crisis that she was 'doing Doctor Who wrong', after she took over from Peter Capaldi.
'Revolution of the Daleks' will force the TARDIS team to struggle without the Doctor, who is marooned in space prison.
The films, collectively titled Small Axe, will air on BBC One and BBC iPlayer in November.
David Tennant has been voted the best ever Doctor in the veteran BBC sci-fi series Doctor Who.
The TV drama about a fictional police anti-corruption squad averaged nearly 13 million viewers for its fifth series.
The 'Fleabag' creator will provide the voice for Andrew Scott's osprey dæmon in 'His Dark Materials'.
The BBC/HBO adaptation of Philip Pullman's classic fantasy trilogy will be back on TV in the autumn.
A producer of the BBC romance series has spoken out against the 'deeply disrespectful' sex scene compilation.
With The A Word about to start its third series, writer Peter Bowker explains how the drama is able to keep reinventing itself.
Stephen Graham has been talked about as potential dream casting for the BBC crime drama for years.
Doctor Who fans remaining indoors during the coronavirus outbreak have arranged a 'mass viewing' of one of the show's classic episodes.
Mark Kermode and Simon Mayo will continue to say "hello to Jason Isaacs", despite the coronavirus pandemic.
New streaming service Britbox has added warnings about racially insensitive content to some older episodes of Doctor Who.
The comment was made in a series of tweets after the actor discussed the media's treatment of Meghan Markle on 'Question Time'.
The BBC sci-fi show has seen a flag in ratings but BBC boss Piers Wenger insisted it still has a long way to go.

domain: entertainment


text:
Southeast Asian Sites Hacked Before Global Anonymous Protest
The Anonymous hacker group stepped up cyber attacks across Southeast Asia, targeting websites in the Philippines and Singapore before a global protest today against censorship and government corruption.
Anonymous Philippines said it infiltrated 115 government websites before a demonstration outside congress in Quezon City as part of a global “Million Mask March,” coinciding with Guy Fawkes Day in the U.K. The mask of Fawkes, who tried to blow up the English Parliament in the 17th century, has become a symbol of the movement. Seventy riot police were on hand at the start of the protest, outnumbering demonstrators more than 3-to-1.
Anonymous Philippines has called on the public to join the “revolution” today. “The government, in many ways, has failed its citizens,” the group said on its Facebook page. “Fairness, justice and freedom are more than just words.”
In Singapore, a website owned by the city’s biggest newspaper publisher was temporarily shut yesterday after being hacked on Nov. 1. A video uploaded on the YouTube website last week showed a person in a Guy Fawkes mask threatening to bring down Singapore’s infrastructure to protest Internet regulations.
“I do not see this as a people’s power but more of a few people going after those with power in the system through clandestine means,” said Bridget Welsh, an associate professor of political science at the Singapore Management University. “Cyber issues reflect a major arena of political contestation that will only intensify in the years ahead.”
The threats in the Philippines have prompted President Benigno Aquino’s government to say that it will take action against those who hack websites to protest corruption. The Singapore police said government websites are down for “maintenance” and warned against “misreporting” of “rumors.”
Anonymous has stepped up its criticism in the Philippines as Aquino’s government remains embroiled in a scandal over the misuse of public funds. Prosecutors have recommended corruption-related charges against a group of lawmakers and a businesswoman related to the alleged improper use of funds that lawmakers doled out at their discretion on infrastructure and development projects.
“There is sufficient democratic space, so there’s no need to resort to illegal acts,” Philippines’ Communications Secretary Sonny Coloma told reporters in Manila yesterday.
Singapore Press Holdings Ltd. (SPH) said it was investigating after some users had difficulty accessing the Straits Times online yesterday. The website was infiltrated last week by a hacker who said the newspaper distorted Anonymous’s position by saying the group is at war with Singapore instead of with its government. Singapore Press confirmed that infiltration.
Singapore from June 1 required websites that regularly publish news on the city state to be licensed and pay a S$50,000 ($40,200) bond, to be forfeited on the publication of “prohibited content” that “undermines racial or religious harmony.” The new laws has prompted criticism from Anonymous.
Singapore government agencies were put on alert for possible attacks, the Straits Times reported on Nov. 1. The Messiah, a hacker with Anonymous, claimed responsibility for infiltrating the website of the Ang Mo Kio Town Council last week, the municipal branch of Prime Minister Lee Hsien Loong’s constituency.
The city’s Infocomm Development Authority said in a statement yesterday that maintenance on Nov. 2 uncovered a fault involving “a routing issue and a hardware failure” that affected some government websites and was later fixed. Government agencies have been on “heightened vigilance” because of the threats, according to the statement.
Guy Fawkes Night is held annually in the U.K on Nov. 5 to mark the anniversary of the discovery of a plot organized by conspirators including Fawkes to blow up the Houses of Parliament in London in 1605.
To contact the reporter on this story: Andrea Tan in Singapore at email@example.com

domain: politics


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We can all fall on hard times now and again. A simple change in our situations can have a large impact on our finances and our ability to pay back the debts we owe.
If you find yourself struggling and unable to pay back your debts, there are some practical dos and don’ts to help you get back on track and find a sensible solution to your problem.
1. Don’t panic
So, you’ve missed a payment, and you know you will continue to struggle to pay back what you owe. Don’t panic – if you immediate take action, bailiffs are not going to turn up on your doorstep and you’re not going to be sent to prison!
2. Don’t borrow more
Borrowing more money is not the answer to this problem – it will only make your position in coming months more difficult as the amount you need to repay will increase. You are in a financial hole, take control and stop digging by adding to your debt.
3. Don’t ignore the problem
As much as you want it to, your debt isn’t going to disappear on its own. Burying your head in the sand will only make the problem worse, and then the bailiffs really will be getting in touch! Plus, if you have late payment charges, your debt problem is only getting worse. It’s time to open the letters and pick up the phone, and find a solution to your problem.
4. Do prioritise
Some bills are more important than others.
Your priority debts and bills are payments such as; mortgage/rent, gas and electric, car repayments, tax debts, child support and maintenance and parking fines.
Non-priority debts are payments such as; credit card and store card debts, payday loans and unsecured bank loans.
You also need to consider your essential expenses such as food and transport to work, ensure you cover your everyday living and priority debts before paying off your consumer debts.
5. Do contact a debt adviser
Of course, we would say this! – But if you feel that a debt is slipping further and further out of your control, it is time to ask for help. You are never alone and a debt adviser can find you a solution. We can talk to creditors on your behalf and make your life much easier.
If you have fallen into debt and are struggling to get out, please contact us today! We can help you find a solution. Click here to contact us or call Clive on 01656 661426.

domain: finance


Mining Sites | Industrial Buildings | Villages
The wind does not always blow and the sun does not always shine and this causes fluctuations in the grid. When the output of renewables is lower than the grid demand we burn fossil fuels in peaker plants to supplement the power supply. But, when the output of renewables is higher than the grid demand, we waste the energy which in Ontario, Canada, alone equated to over $1 billion lost by hydro/utility companies. With sufficient battery storage systems, you can store the excess energy and release it when the grid needs supplementation; effectively replacing peaker plants.


text:
This article was originally published by Airdrie Today on April 22, 2021. You can read the full article below or read the original article here.
In early April, Airdrie senior Joyce Grewcutt suffered a choking incident that led to her daughter calling her an ambulance.
For nearly 20 minutes, Grewcutt laid on the floor of her home, trying to regain her breathing, while her daughter simultaneously tended to her and called ambulance dispatch.
“We phoned 911 and the fire engine came first,” Grewcutt said. “But when I’m laying on the floor, the firemen can’t very well move me into the fire truck to take me anywhere. The ambulance isn’t coming [quickly] because it’s coming from Langdon or Calgary on 17 Avenue. That’s a long time to wait and there’s some confusion. It’s just not right.”
While Grewcutt was ultimately OK and did not require a trip to the hospital, she said the incident reminded her of an issue that has plagued Airdrie for years – the city’s dearth of health-care services.
“When you need help, they’re not there and it’s pretty terrifying,” she said, adding she’s lived in Airdrie for more than 40 years and the issue of health-care access has persisted throughout that time.
Grewcutt’s lengthy wait for an ambulance is not particularly uncommon in Airdrie. According to the Health Sciences Association of Alberta (HSAA), ambulance coverage in Calgary’s outlying communities is often under-serviced.
“Unfortunately, what I’m seeing is the current government is creating a crisis within our health-care system by not properly funding our resources,” said Mike Parker, the president of the HSAA – the union that represents Alberta’s paramedics and other health-care professionals. “When you choose to not match resources to call volume – which they’ve been doing for quite some time – this is what it looks like.”
Parker, who is also an advanced care paramedic, posted a public letter on the HSAA’s website April 8 that highlighted a similar crisis in Cochrane. According to his letter, Cochrane did not have local Emergency Medical Services (EMS) availability on at least 10 occasions between mid-January and mid-February.
The union president stated the ability of smaller communities to offer rapid ambulance service is often jeopardized when their local crews are called into incidents in Calgary, where a lack of sufficient resources and higher call volumes lead to a resource drain from rural areas.
He said ambulances are not only called into Calgary from places like Cochrane and Airdrie, but often farther-away communities such as High River and Kananaskis.
“This problem exists because the current government is choosing not to properly staff and have enough crews available to ensure the urgent needs of Calgarians and the residents of surrounding communities are met,” he wrote.
“When Calgary gets busy, crews from surrounding areas…are dispatched to the city.”
When that happens, ambulances from even smaller towns have to respond to incidents in Calgary’s bedroom communities. According to statistics supplied by HSAA, an ambulance from Strathmore responded to an incident in Airdrie on April 9 and took 22 minutes to arrive at the scene. That same night, an ambulance coming from Didsbury responded to an incident in Airdrie from 67 kilometres away, while another ambulance from Linden – approximately 70 kilometres away – also responded to an incident in Airdrie.
Airdrie’s occasional lack of ambulance availability is even more alarming, according to Parker, when considering the city still does not have its own hospital.
“When you don’t have access to those 24-7 emergency care facilities, EMS should be available to you in your community,” he said. “As populations grow and call volume increases, we’ve been under a hell of a lot of stress from things like the COVID environment. Our health system is in absolute disarray trying to match this.”
While he couldn’t say exactly how much more funding is needed to mitigate the situation, Parker said the solution to the problem is for the Province to increase funding for EMS in order to match resources to the growing call volume in large urban centres like Calgary.
“It’s black and white – stop drawing down by ensuring resources are sustained in metro cities,” he said. “That way, the responding units assigned to Airdrie are available for the citizens. We should all have equal access and opportunity for an ambulance to show up at our door in a respectable time, and an hour [wait] isn’t that.
“If you’re going to actually build an emergency service that protects the people of this province, then build one that functions,” he added. “Stop relying on a truck from Cochrane to be Airdrie’s first response unit.”
In a statement to Great West Media, Airdrie-Cochrane MLA Peter Guthrie acknowledged EMS coverage has been an issue in the riding for many years, but that steps are being taken by the Province to reduce wait times.
He said in his opinion, one of the major issues surrounding EMS is the timing of handing off a patient to the hospital for care.
“There isn’t a really good way for EMS to do that hand-over, so what ends up happening is you have ambulances and the staff waiting with that patient and they can wait with that patient up to eight hours before they can triage that patient to the hospital,” Guthrie said.
“We want to be able to make sure that the patient is able to get the attention that they need sooner. That’s the ultimate goal. Improvements are being made – I believe the services are there.”
—With files from Chelsea Kemp/Cochrane Eagle


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Bhubaneswar: As many as 354 new COVID19 cases were detected under Bhubaneswar Municipal Corporation (BMC) limits in last 24 hours, BMC said Sunday. Out of which 246 are local contact cases and 108 persons are in quarantine.
Update on newly detected #COVID19 cases in the last
24 hrs under the BMC area on 4th Oct 2020(till 9am).
Further contact tracing is still continuing.
Primary contacts & nearby houses are being quarantined, sanitized & will be under active surveillance. pic.twitter.com/7sN3TKWjcj
— BMC (@bmcbbsr) October 4, 2020


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Kids are hungry for heroes…and not the sandwich! They are passionate about super-heroes: Ninja Turtles, Superman, The Incredible Hulk,
Inspired by The Laughing Angels and packed with powerful information, You’re A Miracle…Pass It On! will enlighten, inspire and motivate kids by inviting them to learn:
Biographies and photos of people who have made (and are making) a difference in the world are combined to highlight such diverse and inspirational people as Jane Addams, Amber Coffman, Gandhi, Patch Adams, Martin Luther King, Anne Frank, Craig Kielburger, Mary Cassatt, Chief Seattle, Nkosi Johnson and many, many more! Links to organizations creating positive changes (many founded by kids) are also featured.
Today’s kids are the creators of tomorrow’s world. You’re A Miracle…Pass It On! will challenge kids to step up to the plate and make a difference in the world by facilitating awareness, presenting positive values, offering motivation and inspiration and calling kids to action. When you know something exists and how it works you can make it real in your life and pass it on!
Available at: Amazon.com, Ingram Books, Baker & Taylor, Trafford Publishing (www.trafford.com) Now also available on Amazon Kindle.
Monica Dougherty is an artist, author and art therapist, passionate about the arts and media. She has worked in television (NBC-TV in NYC) in a variety of production facilities. She also worked for a time at the Goodman Theater in Chicago. Along the way have been countless jobs as a temp (way too many to mention) and a few longer-term bread and butter jobs to pay the bills as a freelance artist and graphic designer.

domain: artistic


Gameiro at the double as PSG sink Sochaux
Gameiro, who had only played 71 minutes in the league this season, scored a first-half double as PSG confirmed they had reached cruising speed after a stuttering start.
"We were not that good in the second half, we started to get a little bit tired, it's normal," goalkeeper Salvatore Sirigu told Canal Plus. "The good thing today is the result. There is still room for improvement but we keep cool."
Gameiro had not scored in over six months.
"There are moments like that in a career," he told reporters. "Whether I start a game on the field or on the bench, I try to seize opportunities."
PSG, who spent more than 200 million euros on transfers since being bought by Qatar investors last year, were a class apart, dominating possession with a steely midfield marshalled by the impressive Marco Verratti.
Javier Pastore's delicately chipped pass into the path of Gameiro allowed the latter to make it 1-0 with a low shot after 11 minutes. Gameiro doubled the lead 12 minutes from the interval when he latched on to a Maxwell cross from the left.
PSG looked slightly weary after the break but Brazilian centre-backs Thiago Silva and Alex made sure Sirigu would be unchallenged until the final whistle.
Gameiro's double made up for Zlatan Ibrahimovic's failure to score for the first time in seven PSG appearances.
PSG, who have scored 10 goals from their last three games, face a couple of important challenges next week as they travel to Porto in the Champions League before the "classic" with Marseille.
Morocco midfielder Younes Belhanda fired home from a Karim Ait Fana pass midway through the second half and Souleymane Camara struck late to give champions Montpellier a 2-0 win at Nancy.
Montpellier's second victory of the season put them on eight points, 10 adrift of Marseille.
Lorient stayed fourth on 13 points after Gilles Sunu's late goal salvaged a 1-1 draw at Evian Thonon Gaillard.
Toulouse were fifth, one point behind, thanks to their 2-0 win at promoted Troyes.


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December 22, 2010
Here is the first trailer to Tinsel Korey’s (aka Emily Young) new movie titled ‘Give Me One Reason.’ Click the image below to be taken to the trailer. See synopsis below:
While digging a grave for a corpse in the desert, a group of gangsters discover a Pandora’s Box buried the sand. They open the old wooden mysterious box to find it filled with diamonds. They part way and are all soon dead. And so the box changes hands, from character to character everyone discovers something different in the box, and the longer they hold onto it the closer death creeps.
Thanks to xoxoTKO for the info!