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The nucleus pulposus is an aggrecan-rich, avascular tissue that permits the intervertebral disk to resist compressive loads. In the disk, nucleus pulposus cells express hypoxia-inducible factor (HIF)-1α, a transcription factor that responds to oxygen tension and regulates glycolysis. The goal of the present study was to examine the importance of HIF-1α in rat nucleus pulposus cells and to probe the function of this transcription factor in terms of regulating aggrecan gene expression. We found that HIF-1α protein levels and mRNA stability were similar at 20 and 2% O2; there was a small, but significant increase in HIF-1α transactivation domain activity in hypoxia. With respect to HIF-1α target genes GAPDH, GLUT-1, and GLUT-3, mRNA and protein levels were independent of the oxygen tension. Other than a modest increase in 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase reporter activity, the oxemic state did not change GAPDH, GLUT-1, and GLUT-3 promoter activities. Treatment of cells with 2-deoxyglucose (2-DG), a glycolytic inhibitor, resulted in a significant suppression in ATP synthesis in normoxia, whereas treatment with mitochondrial inhibitors did not affect ATP production and cell viability. However, measurement of the rate of fatty acid oxidation indicated that these cells contained functioning mitochondria. Finally, we showed that when HIF-1α was suppressed, irrespective of the oxemic state, there was a partial loss of aggrecan expression and promoter activity. Moreover, when cells were treated with 2-DG, there was inhibition in aggrecan promoter activity. Results of this study indicate that oxygen-independent stabilization of HIF-1α in nucleus pulposus cells is a metabolic adaptation that drives glycolysis and aggrecan expression.
- hypoxia
- hypoxia-inducible factor-1α
the local oxygen tension promotes cellular differentiation and regulates tissue energy metabolism. When the oxygen tension is low, there is almost complete reliance on glycolysis to generate ATP and reducing equivalents (12, 29, 30). In specialized tissues such as the intervertebral disk, the mechanism by which cells conserve energy has received little attention. Nonetheless, it is known that in the intervertebral disk, the nucleus pulposus has no direct vascular supply (8, 24, 33), and ATP is probably generated by anaerobic glycolysis (1, 10). The dependence on glycolysis reflects the low oxygen tension in the disk, which has been reported to be low as low as 2%, and the observation that modulation in the oxygen supply can cause vertebral defects (15). Regulation of glycolysis is mediated by hypoxia-inducible factor (HIF)-1α, a transcription factor that responds to the local oxygen tension, and has been shown to be expressed by nucleus pulposus cells in vivo (20).
HIF-1α is a member of the basic helix-loop-helix (bHLH)-PER-ARNT-SIM (PAS) family of proteins. It consists of a constitutively expressed β-subunit and an α-subunit that is rapidly degraded under normoxic conditions (34). The bHLH and PAS motifs are required for dimerization, and two transactivation domains (TAD), N-TAD (aa 531-575) and C-TAD (aa 786-826) bind several coactivators; whereas the downstream basic region affords specific binding to the hypoxia-response element (HRE) in target genes (9, 27). HIF-1α promotes transcription of key glycolytic enzymes, allowing a cell to adapt to low Po2 (6, 16, 21, 26, 28), and inhibits ATP generation through oxidative phosphorylation (17). In an earlier investigation, our group (22) showed that there is evidence of normoxic stabilization of this otherwise oxygen labile protein in nucleus pulposus cells.
The goal of the present study was to examine the importance of normoxic stabilization of HIF-1α in rat nucleus pulposus cells and to probe the function of this transcription factor in terms of aggrecan expression. We focused on aggrecan, since this major extracellular matrix proteoglycan permits the intervertebral disk to withstand compressive loads. We have shown that unlike other mammalian cells, under normoxic conditions, HIF-1α maintains ATP generation through glycolysis and regulates aggrecan gene expression. This finding lends strength to the view that cells of the nucleus pulposus are functionally adapted to their avascular microenvironmental niche.
Isolation of nucleus pulposus cells.
Male Wistar rats (225–250 g) were euthanized with CO2. The spinal columns were removed under aseptic conditions, and lumbar intervertebral disks were separated. The gel-like nucleus pulposus was separated from the annulus fibrosus with the use of a dissecting microscope and was treated with 0.1% collagenase and 10 U/ml hyaluronidase for 4–6 h. The partially digested nucleus pulposus tissue was maintained as an explant in Dulbecco's modified Eagle's medium (DMEM) and 10% fetal bovine serum (FBS) supplemented with antibiotics in a humidified atmosphere containing 5% CO2 at 37°C. Nucleus pulposus cells migrated out of the explant after 1 wk. When confluent, the cells were lifted using trypsin (0.25%)-EDTA (1 mM) solution and subcultured in 10-cm dishes. For some experiments, human disk tissue (minimally degenerative and regarded as surgical waste) was obtained from patients undergoing cervical spinal surgeries by following a protocol approved by the Institutional Review Board of the Thomas Jefferson University.
Cell culture in hypoxia.
Nucleus pulposus cells were cultured in an Hypoxia Work Station (Invivo2 300; Ruskinn, Brigend, UK) with a mixture of 2% O2, 5% CO2, and 93% N2 for 24–72 h. The concentration of oxygen chosen for this study was selected on the basis of our previous in vitro studies and information on the oxemic status of the disk in vivo. To induce chemical hypoxia, cells were treated with 130 μM desferrioxamine (Dfx) under normoxic conditions for 24 h. To chemically suppress HIF-1α, cells were treated with 500 mM 17-allylamino-17-demethoxygeldanamycin (17-AAG), a known HIF-1α inhibitor. In some experiments, HeLa cells were used as control and maintained in culture as described above.
Examination of HIF-1α and GLUT-1 expression by immunofluorescence analysis.
Cells were plated in flat-bottom 96-well plates and maintained in normoxia or hypoxia for 24 h. After incubation, cells were fixed with 4% paraformaldehyde on ice for 20 min and washed with phosphate-buffered saline (PBS). Cells were permeabilized with 0.5% Triton X-100 in PBS for 15 min and incubated in blocking solution (PBS containing 10% FBS) for 45 min. Cells were then incubated with anti-HIF-1α antibody (ABR, Golden, CO) or anti-GLUT-1 antibody (Abcam, Boston, MA) in blocking solution at a dilution of 1:200 at 4°C overnight. As a negative control, cells were treated with mouse isotype IgG under similar conditions. After cells were thoroughly washed with PBS, the experimental and control cells were incubated with Alexa Fluor488-conjugated anti-mouse secondary antibody (Molecular Probes, Eugene, OR) at a dilution of 1:50 for 1 h at room temperature. After washing, some cells were also incubated with 5 μM propidium iodide for 20 min at room temperature. Cells were imaged using a laser scanning confocal microscope (Olympus Fluoview, Tokyo, Japan).
Nuclear and membrane protein extraction.
Nuclear extracts were prepared from subconfluent cells according to the method of Dignam et al. (5) using the CellLytic NuCLEAR extraction kit (Sigma Aldrich, St. Louis, MO). Following culture in hypoxia or normoxia, cells were immediately placed on ice and lysed in hypotonic buffer (10 mM HEPES, pH 7.9, 1.5 mM MgCl2, 10 mM KCl, and 0.5 mM DTT) and incubated on ice for 15 min. Igepal CA-630 was added to a final concentration of 0.6%, and the mixture was vortexed vigorously for 10 s. Nuclei were recovered by centrifugation at 3,300 g for 30 s at 4°C and extracted by gentle shaking in buffer containing 20 mM HEPES, pH 7.9, 0.42 M NaCl, 25% glycerol, 1.5 mM MgC12, 0.2 mM EDTA, and 0.5 mM DTT for 30 min at 4°C. The extract was then centrifuged for 15 min at 25,000 g, and the supernatant was snap frozen at −70°C. All buffers contained a protease inhibitor cocktail [2 mM 4-(2-aminoethyl)benzenesulfonylfluoride, 1.4 pM trans-epoxysuccinyl-l-leucylamido(4-guanidino)butane, 130 pM bestatin, 1 μM leupeptin, and 0.3 pM aprotinin] and phosphatase inhibitors (5 mM NaF and 200 μM Na3VO4). Membrane proteins were isolated using the mammalian membrane protein extraction reagent kit Mem-Per (Pierce, Rockford, IL) following the manufacturer's protocol.
Western blot analysis.
Proteins were resolved on 10% SDS-polyacrylamide gels and transferred by electroblotting to nitrocellulose membranes (Bio-Rad, Hercules, CA). The membranes were blocked with 5% nonfat dry milk in TBST (50 mM Tris, pH 7.6, 150 mM NaCl, 0.1% Tween 20) and incubated overnight at 4°C in 3% nonfat dry milk in TBST with the antibodies against HIF-1α (1:500; R&D Systems, Minneapolis, MN), GLUT-1 (1:500), GLUT-3 (1:500; Abcam), or GAPDH (1:3,000; Novus, Littleton, CO) and β-actin (1:4,000) (Sigma). Immunolabeling was detected using the ECL reagent (Amersham Biosciences). Relative expression levels were determined by quantitative densitometric analysis using Kodak 1D image analysis software (Kodak, Rochester, NY).
Real-time RT-PCR analysis.
At the end of each treatment period, total RNA was extracted from cells using Trizol (Invitrogen) according to the manufacturer's protocol. RNA was further treated with RNase-free DNase I and passed through RNeasy mini columns (Qiagen). Total RNA (100 ng) was used as template for real-time PCR analysis. Reactions were set up in microcapillary tubes using 1 μl of RNA with 9 μl of a LightCycler FastStart DNA Master SYBR green I mix (Roche Diagnostics, Indianapolis, IN) to which gene-specific forward and reverse PCR primers were added. With each set of samples, no template control was included. PCR reactions were performed in LightCycler (Roche) according to the manufacturer's instructions. All the primers used were synthesized by Integrated DNA Technologies (Coralville, IA). See Table I for sequences.
Reporter plasmids.
Luciferase reporter plasmids were provided by Dr. Sherin Devaskar, University of California, Los Angels (Glut-3 promoter) (19), Dr. Oleksandr Minchenko, Palladin Institute of Biochemistry, Ukraine [6-phosphofructo-2-kinase/fructose-2,6-biphosphatase (PFKFB) promoter and HIF-1α 3′-untranslated region (UTR)] (14), and Dr. Shan Lu, University of Cincinnati (GAPDH promoter) (13). pGlut-1 wild type (Wt) and pGlut-1 mutant (Mt) were provided by Dr. Amit Maity, University of Pennsylvania (4); pGlut-1 (Wt) contains an active HRE, whereas the 4-bp mutation in pGlut-1 (Mt) disrupts HIF-1 binding. For transactivation studies of HIF-1, the binary Gal4 reporter plasmid (TAD aa 530–826) was provided by Dr. Nianli Sang (25). Backbone plasmid pM (Clonetech) contains no TAD but expresses the Gal4dbd. pFR-Luc (Stratagene) reporter contains the yeast Gal4-binding site upstream of a minimal promoter and the firefly luciferase gene. Construction of aggrecan luciferase promoter construct is described elsewhere (32). As an internal transfection control, vector pRL-TK (Promega) containing Renilla reniformis luciferase genes was used. In a previous study, we optimized the amount of transfected plasmid, the pretransfection period after seeding, and the posttransfection period before harvesting for rat nucleus pulposus cells using pSV β-galactosidase plasmid (Promega) (22).
Transfections and dual luciferase assay.
Nucleus pulposus and HeLa cells were cultured in 24-well plates at a density of 7.5 × 104 cells/well. Next day, cells were treated with 500 ng of reporter gene plasmid and 500 ng of control plasmid pTK-RL premixed with the transfection reagent Lipofectamine 2000 (Invitrogen, Carlsbad, CA). Twenty-four hours later, cells were transferred to the hypoxia chamber or maintained in normoxia for 1 additional day. The cells were then harvested, and a Dual-Luciferase reporter assay system (Promega) was used for sequential measurement of firefly and Renilla luciferase activities with specific substrates, i.e., beetle luciferin and coelenterazine, respectively. Quantification of luciferase activities and calculation of relative ratios were carried out using a luminometer (TD-20/20; Turner Designs, Sunnyvale, CA); at least three independent transfections were performed in triplicate.
Cell viability measurement by neutral red uptake.
Nucleus pulposus cells were treated with the following inhibitors: oligomycin (0.05–0.2 μM), antimycin A (10–30 μM), and atractyloside (50–200 μM) (Sigma). One day later, viability was measured using neutral red assay. Briefly, cells cultured in 24-well plates were treated with mitochondrial inhibitors in normoxia or hypoxia for 24 h. The cells were then incubated with neutral red solution dissolved in DMEM (40 μg/ml) for 3 h at 37°C. The dye solution was aspirated, and cells were washed with HBSS. The retained dye was extracted with 400 μl of acid ethanol (50% ethanol in 1% acetic acid) per well. The absorbance was read at 540 nm using a microplate reader (Tecan SpectraFluor Plus, Durham, NC). Results are presented as percent absorbance compared with untreated control values in normoxia.
ATP measurements following treatment with mitochondrial and glycolytic inhibitors.
Nucleus pulposus cells were cultured in six-well plates (2 ×105 cells/well) and treated with antimycin A (30 μM) or atractyloside (200 μM) for 24 h under normoxic (20% O2) conditions or cultured in hypoxia (2% O2). To examine the effects of inhibition of glycolysis on ATP production, cells were treated with 10 mM 2-deoxyglucose (2-DG; Sigma) for 30 min to 6 h. ATP measurement was performed using the ATP bioluminescence assay kit CLS II (Roche) per the manufacturer's protocol. Luminescence was measured at 562 nm using a luminometer (TD-20/20; Turner Designs). ATP levels were normalized to total protein, measured using the BCA assay (Pierce).
Mitochondrial fatty acid utilization by nucleus pulposus cells.
To learn whether nucleus pulposus cells possess functioning mitochondria, rat and human cells were plated in 12-well plates (2 ×105 cells/well) in complete growth medium for 24 h. Cells were then washed with PBS and suspended in preincubation medium (Krebs-Ringer bicarbonate medium containing 0.05% fatty acid-free BSA) or inhibitor medium (preincubation medium with 250 mg of glucose) for 1 h. After preincubation, the medium was removed and the cells were incubated in either 200 μl of myristate or 200 μl of palmitate incubation medium (110 μM fatty acid, 16.7 μCi/ml [3H]fatty acid, 0.05% BSA in PBS, pH 6.9). After 3 h, the medium was transferred to columns containing ∼2.9 ml of Dowex-1 ion exchange resin (no. 1X8–200; Sigma), which had been charged with 1.0 N NaOH and washed with Milli-Q water until the pH of the elute was the same as the wash water. The Dowex column serves to bind the unmetabolized fatty acids. The 3H-labeled water, produced by β-oxidation, was eluted with 2 ml of water from the Dowex. The activity of eluate was counted using a Beckmann LS 6500 liquid scintillation counter. Data were normalized to total protein measured using the Lowry method.
Measurement of aggrecan promoter activity following silencing of HIF-1α by Small Interfering RNA.
To learn whether HIF-1α activity regulates aggrecan expression, we silenced HIF-1α expression using small interfering (si)RNA technology. Briefly, nucleus pulposus cells were plated in 24-well plates and transfected with HIF-1α SMART Pool (Dharmacon) or control siRNA duplexes (Ambion) at a final concentration of 20 nM in DMEM using Lipofectamine 2000 (Invitrogen). Cells also received aggrecan or enolase-1 promoter constructs and pRL-TK plasmid at the time of transfection. Six hours after transfection, the medium was replaced with complete growth medium, and the cells were allowed to recover for 18 h. Cells were then cultured under normoxic or hypoxic conditions for an additional 24 h, and luciferase activity was assayed.
Statistical analysis.
Data are means (SD). To test for significance, data were analyzed using Student's t-test; the P values obtained are indicated in the text and legends. Each experiment was performed in triplicate.
HIF-1α protein expression and mRNA stability in nucleus pulposus cells is independent of the oxygen tension.
Figure 1A shows the relative expression levels of HIF-1α protein in rat nucleus pulposus cells under different culture conditions. At 2% O2 (hypoxia), the protein expression was similar to that measured at 20% O2 (normoxia). A similar level of expression was noted when the nucleus pulposus cells were treated with Dfx, a chemical agent used to mimic effects of hypoxia. Moreover, there was little change in HIF-1 immunofluorescence when maintained in hypoxia (Fig. 1B). To further examine the dependence of HIF-1α on oxygen tension, we analyzed mRNA expression using real-time RT-PCR. Figure 1C shows mRNA levels after exposure of nucleus pulposus cells to 2% O2 from 4 to 24 h. Compared with normoxia, no differences were apparent in HIF-1α transcript level in hypoxic nucleus pulposus cells for up to 24 h. We then analyzed 3′-UTR activity, as a measure of HIF-1α mRNA stability, in nucleus pulposus cells. We found that HIF-1α 3′-UTR activity was similar in both normoxic and hypoxic nucleus pulposus cells.
Nucleus pulposus cells show a modest increase in HIF-1α TAD activity in hypoxia.
We further assessed the activity of HIF-1α in nucleus pulposus cells by studying the activation of transactivation domain (TAD). Figure 2A shows that under hypoxic conditions, there was a modest but significant increase (40%, or 0.4-fold) in HIF-1 TAD activity in nucleus pulposus cells. HeLa cells, a model cell line widely used to evaluate the transcriptional responses of HIF-1α, was used as a control. In contrast to nucleus pulposus cells, HeLa cells displayed a significantly higher (3.5-fold) activation of HIF-1 TAD activity in hypoxia (Fig. 2B). These results indicate that hypoxia causes activation of TAD function in both nucleus pulposus and HeLa cells.
Oxemic regulation of HIF-1α target genes by nucleus pulposus cells.
Real-time RT-PCR analysis showed that expression of GLUT-1, GLUT-3, and GAPDH were unaffected by the oxemic status of the nucleus pulposus cells (Fig. 3, A–C). In terms of copy number relative to β-actin, expression of all these genes was similar under normoxic and hypoxic conditions. We also evaluated GAPDH protein levels in the nucleus pulposus cells. Figure 2D indicates that culture time in hypoxia did not change the level of expression of this glycolytic protein. Moreover, in hypoxia, Western blot followed by densitometric analysis showed that expression levels of GLUT-1 and GLUT-3 in the membrane, as well as total protein fraction, was independent of time or the oxygen tension (Fig. 3, D–F). Similarly, immunofluorescence analysis of normoxic and hypoxic nucleus pulposus cells indicated that there was a comparable level of staining of GLUT-1 (Fig. 3G).
To further evaluate the transcriptional regulation of HIF-1α target genes, we examined the promoter activities of GLUT-1 and GLUT-3 under normoxic and hypoxic conditions. For these studies, we used luciferase reporter constructs containing wild-type or mutant HRE (21). HeLa cells were used as a control. Figure 4, A and B, shows that promoter activities of GLUT-1 and GLUT-3 in nucleus pulposus cells were similar in both hypoxia and normoxia. In contrast, in hypoxia, HeLa cells evidenced a high induction of GLUT-1 (125-fold) and GLUT-3 (15-fold) promoter activities (Fig. 4, C and D). Furthermore, in nucleus pulposus cells, basal normoxic promoter activity of GLUT-1 was two- to threefold higher than background activity measured with GLUT-1 control plasmid containing mutant HRE. The normoxic basal promoter activity of HeLa cells was minimal and similar to the background activity measured with the HRE mutant GLUT-1 plasmid. Moreover, when transfected with a mutated control GLUT-1 plasmid, there was little difference between normoxic and hypoxic luciferase activities of both nucleus pulposus (Fig. 4A) and HeLa cells (Fig. 4C).
In addition, we analyzed promoter activities of two key glycolytic enzymes, GAPDH and PFKFB (Fig. 5, A–D). In accordance with GLUT-1 and GLUT-3 promoter activity, GAPDH promoter activity showed no hypoxic induction (Fig. 5A); in contrast, HeLa cells displayed a 50- to 60-fold induction of GAPDH promoter activity in hypoxia (Fig. 5C). However, unlike GLUT and GAPDH, there was a threefold induction of PFKFB promoter activity in hypoxia in nucleus pulposus cells (Fig. 5B). Hypoxic induction of PFKFB promoter activity in HeLa cells was close to 120-fold (Fig. 5D). In addition, real-time PCR analysis indicated that there was a significant increase in PFKFB mRNA expression in nucleus pulposus cells cultured in hypoxia (Fig. 5E).
Nucleus pulposus cells generate energy through glycolysis.
To investigate the mode of energy generation in nucleus pulposus cells, we measured ATP production. Under basal normoxic conditions, ATP levels in nucleus pulposus cells were between 20 and 25 nM/mg protein (Fig. 6A). When treated with 2-DG, an inhibitor of glycolysis, basal ATP levels are suppressed by almost 80% within 30 min of treatment and remained very low for 6 h (Fig. 6A). In contrast, when cells were treated with the mitochondrial function inhibitors antimycin A or atractyloside, there was no effect on basal ATP levels after 30 min or 6 h of treatment (Fig. 6B). We also evaluated cell viability after treatment with mitochondrial inhibitors in normoxia or hypoxia using the neutral red uptake assay. Figure 6C indicates that cell viability in both normoxia and hypoxia (data not shown) was unaffected.
Nucleus pulposus cells possess functioning mitochondria.
We determined that nucleus pulposus cells contained functioning mitochondria by measuring the oxidation of free fatty acids by the β-oxidation pathway. Mitochondria of both human and rat nucleus pulposus cells oxidized [3H]palmitate (C16:O) and myristate (C14:O) into CO2 and water (Fig. 7, A–C). For both human and rat nucleus pulposus cells, rates of oxidation of palmitate were comparable (Fig. 7A). Myristate was oxidized at a significantly faster rate by the rat cells compared with human nucleus pulposus cells. When excess glucose was included in the incubation medium along with 3H-labeled palmitate (Fig. 7B) and myristate (Fig. 7C), there was a significant inhibition (90–95%) of lipid oxidation and a concomitant decrease in [3H]H2O generation.
HIF-1α expression and glycolysis is required for maintaining aggrecan promoter activity in nucleus pulposus cells.
The functional significance of constitutive HIF-1α expression and glycolytic metabolism in nucleus pulposus cells was studied by measuring aggrecan expression and gene promoter activity. We suppressed HIF-1α expression and function or treated cells with 2-DG. Figure 8A shows that in normoxia, when cells were cotransfected with HIF-1α SiRNA, aggrecan promoter activity was inhibited by almost 60%. Similarly, when HIF-1α was silenced under hypoxic conditions, there was a comparable level of aggrecan promoter suppression. We confirmed that HIF-1α was silenced by determining PFKFB promoter activity in normoxic and hypoxic culture conditions. Figure 8B indicates that the silenced cells exhibited a significant decrease in the activity of the PFKFB gene promoter, a known HIF-1 target gene. Similarly, treatment with 17-AAG, a known HIF-1α inhibitor, resulted in a significant drop in aggrecan mRNA expression (Fig. 8C). In the presence of the glycolysis inhibitor 2-DG, there was ∼30% suppression of aggrecan promoter activity under normoxic conditions (Fig. 8D).
Previously, we showed that the HIF-1α isoform is expressed in the nucleus pulposus in situ, as well as in cells cultured in 20% O2 (20, 22). Since a major role of HIF-1α is to maintain cell function at a low Po2, we initially explored the mechanism by which the oxemic state regulated the expression of this critical transcription factor. Predictably, Western blot and immunofluorescence analysis showed that at 2% O2, HIF-1α was robustly expressed by nucleus pulposus cells. Surprisingly, when cells were maintained at 21% O2, there was little change in HIF-1α expression. Likewise, chemical hypoxia did not significantly increase HIF-1α expression, and when HIF-1α mRNA stability was evaluated, quantitative PCR failed to indicate differences in steady-state transcript levels in normoxia and hypoxia. Thus, from all perspectives, HIF-1α expression in nucleus pulposus cells was found to be independent of the oxygen tension. This unique finding points to substantive underlying differences between the metabolism of disk cells and cells from most other tissues. Importantly, since HIF-1α serves a critical role in regulating a host of other (nonmetabolic) target genes, it is likely that this transcription factor influences a number of other survival activities (2). In the current study, we have shown that HIF-1α is required for the synthesis of the water-binding proteoglycan molecules of the extracellular matrix.
One obvious question raised by the observation that HIF-1α was expressed in 20% O2 concerned the activity of target genes: did HIF-1α transactivate genes of the glycolytic pathway? Thus, although the transcription factor was robustly expressed by the nucleus pulposus, it was not known whether expression enhanced target gene activity. Focusing on those genes that regulate the energetic state of nucleus pulposus cells, we assessed expression of the glucose transporters GLUT-1 and GLUT-3. In the same study, we also evaluated another HIF-1 target, GAPDH, the enzyme that catalyzes the conversion of glyceraldehyde 3-phosphate into 1,3-bisphosphoglycerate (13). At 2 and 20% O2, GLUT-1, GLUT-3, and GAPDH mRNA expression levels were comparable and remained constant over time. Furthermore, at the two disparate oxygen tensions, both Western blot and immunofluorescence analysis indicated comparable GLUT-1, GLUT-3, and GAPDH protein levels. In addition, GLUT-1 and GLUT-3 promoter activities were not responsive to changes in oxygen tension, and there were no oxygen-dependent changes in GAPDH promoter activity. It is relevant to note that although these genes were not responsive to the oxemic state of the culture, we did observe a threefold induction in PFKFB promoter activity; the magnitude of the change was similar to that observed when enolase-1 promoter activity was measured in hypoxic nucleus pulposus cells (22). For these two genes, hypoxic induction of glycolytic promoter activity was probably due to a modest increase in HIF-1α transactivation activity. For comparison, our studies with HeLa cells, which are clearly HIF responsive, indicate that these modest changes in transactivation activity would not substantially increase the rate of glycolysis by nucleus pulposus cells. In the HIF-1α-responsive HeLa cell, the change from normoxia to hypoxia caused a 60- to 120-fold increase in promoter activities and a 3.5-fold elevation in TAD activity. All of these values are substantially different from those exhibited by nucleus pulposus cells. On the basis of this observation, it is concluded that the oxemic stability of the nucleus pulposus cells is optimal for survival in an environment where there are frequent shifts in vascular supply and oxygen delivery; in the intervertebral disk, these shifts may reflect minute-to-minute variations in applied biomechanical forces to the spinal units.
Since resistance to compressive forces on the spine are mediated by water molecules bound to charged sites on proteoglycans, we examined the relationship between HIF-1α expression and aggrecan promoter activity. Previous studies by Pfander et al. (18) showed that under hypoxic conditions, aggrecan mRNA and sulfation levels were significantly reduced in chondrocytes lacking HIF-1α. In a more recent study, Robins et al. (23) reported that hypoxia via a HIF-1α-dependent mechanism increased the commitment of mesenchymal cells to the chondrogenic lineage by activating Sox-9; one important aspect of commitment is the promotion of aggrecan synthesis. Studies reported presently build on these earlier observations. We have shown that irrespective of the oxemic state of the cells, when HIF-1α expression was blocked by siRNA or by pharmacological agents, aggrecan promoter activity was markedly inhibited. A similar level of inhibition was observed when glycolysis was blocked. On the basis of these observations, we suggest that HIF-1α promotes aggrecan expression directly, by elevating gene expression and protein synthesis, and, possibly, indirectly, through lineage specification as well as promotion of sulfation reactions. From a functional viewpoint, since aggrecan synthesis is dependent on HIF-1α expression, rather than oxemic status, long-term shifts in the oxemic state of the disk would be expected to exert a minimal effects on the generation of extracellular matrix components.
One of the logical consequences of stabilization of HIF-1α and the robust expression of glucose transporters and glycolytic enzymes is upregulation of anaerobic glycolysis. We found that in normoxia, basal concentrations of ATP in nucleus pulposus cells were between 20 and 25 nM/mg protein. These values are comparable to levels reported for articular chondrocytes (18), another cell type that uses glycolysis to generate energy (29, 30). In the presence of 2-DG, a potent inhibitor of glycolysis, ATP generation was suppressed almost 80%. The sensitivity of the cells to this inhibitor emphasized the reliance on glycolysis for energy generation. Together, these data confirm earlier findings that the cells of the intervertebral disk generate much, if not all, of their energy through anaerobic glycolysis (1, 10).
Although ATP generation through glycolysis is clearly the major pathway for energy generation, the possibility exists that some ATP may be generated through mitochondrial oxidative phosphorylation. We noted that when cells were treated with inhibitors of mitochondrial function, no change in ATP production, or viability, was observed. Indeed, this observation raised the question, are there functioning mitochondria in cells of the nucleus pulposus? Although, this topic has received minimal attention, Gan et al. (7) reported earlier that although nucleus pulposus cells contained mitochondria with normal architecture, the total number of organelles per cell was very low. To further address this issue, we examined fatty acid oxidation in the absence of a three-carbon source. Since cells from both human and rat tissues oxidized palmitate (23–27 pmol·min−1·mg protein−1) and myristate (25–43 pmol·min−1·mg protein−1), it was clear that the nucleus pulposus had the capacity to generate ATP through oxidative metabolism. Moreover, we showed that in concert with other tissues, if nucleus pulposus cells were provided with excess glucose, fatty acid metabolism was suppressed. These results lends strong support to the notion that although glucose and anaerobic glycolysis represent the major fuel and pathway for energy generation, respectively, the nucleus pulposus retains the capacity to metabolize fatty acids through mitochondrial oxidative metabolism.
Finally, there is now good evidence to indicate that HIF-1α plays a major role in directing the interplay between glycolysis and mitochondrial oxidative phosphorylation (11, 17). These workers have shown that HIF-1α inhibits mitochondrial function by trans-activating the gene encoding pyruvate dehydrogenase kinase 1. Since this protein suppresses pyruvate dehydrogenase, pyruvate cannot be converted into acetyl-CoA, and as a result, the TCA cycle is blocked. On the basis of these observations, it is concluded that although mitochondrial function is retained by cells of the intervertebral disk, it is reasonable to assume that normoxic expression of HIF-1α by nucleus pulposus cells serves to suppress oxidative phosphorylation, lower mitochondrial ATP generation, and promote glycolytic ATP and aggrecan synthesis. These two latter activities are of considerable functional importance and point to the role of HIF-1α in promoting the synthesis of key water-binding molecules of the extracellular matrix, complexes that are required to maintain the biomechanical properties of the intervertebral disk.
This work was supported by National Institute of Arthritis and Musculoskeletal and Skin Diseases Grant AR050087.
We thank Drs. Sherin Devaskar, Oleksandr Minchenko, Shan Lu, Amit Maity, and Nianli Sang for providing necessary plasmids.
The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
- Copyright © 2007 the American Physiological Society


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|August 19, 2012, 08:22 PM
Senior Member
Join Date: January 3, 2010
Location: NW Ohio
Posts: 455
Issues withTaurus PT145. Need input please.
Took my Taurus PT145 to the range today and while shooting the magazine frequently disengaged and slid partially out of the grip. Thought maybe I was hitting the mag release so made a point to watch that. Any idea what could cause this?
Also shot horrible. Shot several low and left and some didn't hit the paper at 15 feet! I bought it new a few months ago so I have shot it before and, although not as accurate as my upper end pistols, it didn't shoot bad. Wasn't me either today as I shot great with my Sig SP2022 and a buddy shot the PT145 with the same results.
Please no Taurus jokes and/or bashing. I have already read them all and am just trying to get some honest feedback on what the issues could be.
|August 19, 2012, 08:29 PM
Senior Member
Join Date: June 6, 1999
Location: El Paso, TX
Posts: 1,021
If you look up the magazine well you should see a spring that holds the magazine catch in. Taking that out and inspecting both it and the spring for damage would be a good place to start. Shouldn't be too much trouble with a 90 degree dental pick.
New gun, same ol' shot.
|August 19, 2012, 08:40 PM
Join Date: April 13, 2000
Location: Northern Virginia
Posts: 39,098
Unless you have specific advice on the particular issue that the OP is having with his gun, do not post in this thread.
"The gift which I am sending you is called a dog, and is in fact the most precious and valuable possession of mankind" -Theodorus Gaza
Baby Jesus cries when the fat redneck doesn't have military-grade firepower.
|August 20, 2012, 07:13 AM
Senior Member
Join Date: August 18, 2009
Posts: 826
Magazine drop-out is a fairly common problem with the PT-1xx Taurus pistols. IF not caused by inadvertent contact with the mag release button (and THAT is the MOST common cause), it is usually one (or more) of three possibilities: either the mag catch is cruded up and not engaging with the slot in the mags completely......or, the mag(s) are poorly made (the slot for the mag catch is off location just a bit and the catch doesn't engage completely.....or, the mag catch is defective. Defective mag catches are not uncommon on these (the part is plastic, I believe).
The best course of action would be to address these three possibilities in order.
So, first clean the mag catch area thoroughly. Carb cleaner, sprayed through the little straw that comes with the can, will tend to flush out any crud. You can reach inside the mag well with a toothbrush (or similar brush) and scrub a bit as well, after spraying, to dislodge any gunk.
Next, check your mags for fit. Snap in a mag, then wiggle it somewhat violently. If it pops loose (without pressing the release), then either the mag is bad, or the catch. Check to be sure that the mag goes all of the way up into the well....and that you get a solid "click" when the mag is seated all of the way. If the mag(s) are not solidly clicking in, then it COULD be that the catch slot in them is off position a bit, or burrs could be causing the catch not to fully engage. If you detect any burrs on the edges of the slot in the mag, these can be removed with a small file. If the slots are off just a bit, that can be addressed with a file as well, though I wouldn't try it unless you understand what you are doing. Also, check to see that the mag base plates are not interfering with the mag going all the way in - if that is the problem, then a bit of sandpaper will fix that, easily.
Lastly, the mag catch could be defective or broken. Shine a light down into the mag well from the top. The mag catch (the important bit) will be on the right side (opposite the release button) and should look complete (have a straight edge) and not worn. You can test your mags again, by inserting a mag with the follower and spring removed but base plate ON, then checking with a light from the top of the mag well (through the top of the mag body)....to see if the mag catch seems to engage fully. The mag catch can be removed from the gun for inspection, as well. I've never done it, so I can't advise you on how, though.
The last step, if you don't feel competent to address these possibilities, or if you can't find the problem and correct it - is to send the gun back to Taurus for repair.
|August 20, 2012, 08:34 AM
Senior Member
Join Date: October 9, 2007
Posts: 1,160
This is the gun that ended any relationship I will ever have with that company again. I had similar issues that I brought up here. I learner from the experience and now buy quality handguns.
|August 20, 2012, 12:28 PM
Senior Member
Join Date: November 19, 2005
Location: southwestern va
Posts: 765
i have a 3rd gen pt145 and its always run like a champ. I have no input on the mag release problem, but as for shooting low my gun does that as well (ive seen this problem in other posts about this gun as well). Its an easy fix, either bring up the front post higher in the sight picture, or gently sand down the front post a bit (or have a smith change it out/alter it for you).
"i got the most powerful gun in the world........an .88 magnum. It shoots thru schools......"
|August 20, 2012, 02:40 PM
Senior Member
Join Date: February 22, 2009
Location: Virginia
Posts: 1,364
My gen 3 is pretty darned accurate and I've never had the mag problem you describe.
Mine has other mag problems....good luck with the cure for your problems.
There's only one...
|August 20, 2012, 07:27 PM
Senior Member
Join Date: January 3, 2010
Location: NW Ohio
Posts: 455
Got to thinking about it and decided I'd send back to Taurus to repair. Only has about 300 rounds through it and thought why should I have to mess with it. Went back today. Hopefully it will come back repaired in a reasonable length of time.
|September 6, 2012, 04:23 PM
Senior Member
Join Date: January 3, 2010
Location: NW Ohio
Posts: 455
An update:
I got on Taurus' website, submitted a work order, and shipped the pistol back on 8/20/12. Received it back today (9/6/12), which is a two week and 3 day turnaround. Report included wih the pistol said the "mag release worn" and" barrel has irregular grooves". Both were replaced at no charge. Pretty impressed with customer service. Will shoot this weekend to see if it is indeed repaired.
|September 9, 2012, 10:09 AM
Senior Member
Join Date: May 15, 2012
Location: Texas
Posts: 1,503
Good luck. Some Tauri work maybe yours will.
|September 9, 2012, 06:28 PM
Senior Member
Join Date: January 3, 2010
Location: NW Ohio
Posts: 455
Well...took the PT145 to the range today to test. Shot a lot more accurate than prior to sending it back for repair but the mag popped out and a round stovepiped about every other shot from round 4 through round 54. Going back to Taurus again.
|September 10, 2012, 09:00 PM
Senior Member
Join Date: February 22, 2009
Location: Virginia
Posts: 1,364
My mags never fall out...they just jam.
There's only one...
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subset: commoncrawl


text:
Best buffets
Posted March 31, 2014
Raleigh, N.C. — Looking for variety when you dine? Well, checking out a buffet might be your best bet. With a chance to sample various food and even cuisines, buffets are sometimes worth the trip.
Here are our picks for the best buffets in the Triangle:
Milton's Pizza (Six Forks Road and Wakefield in Raleigh) - Their lunch buffet, available every weekday from 11 a.m. to 2 p.m., is one of my favorites! I love that it isn't just pizza. You can make your own pasta meal with various sauces and noodles available. If you aren't feeling that, you can try some grilled fish, chicken wings, corn fritters, vegetables and even ribs. Add that to their already impressive salad bar and that dessert bar (it has its own table filled with cookies, baklava, cake, cupcakes and brownies!) and you will leave there so full!
McCall's Bar-B-Q and Seafood (Goldsboro and Clayton) - We love their barbecue, so naturally we enjoy their buffet! Dessert and salad bar is included in this daily lunch buffet served from 11 a.m .to 2 p.m. everyday.
Sweet Tomatoes (Raleigh) and (Cary) - I know we don't normally put chains on these lists, but this is my go-to buffet when my in-laws visit. Their salad bar is huge and they change up their seasonal salad offerings often, so you always get something new. They also have an extensive list of soups (try their chili!) and pasta. They even offer baked potatoes. Definitely try the chocolate muffins and save room for some soft serve ice cream!
Hibachi Grill and Supreme Buffet (Raleigh) - The sheer amount of selections at this place can be overwhelming. Just take your time and hit one section at a time. From traditional buffet staples (General Tso's chicken and egg rolls) to freshly made sushi (you can watch them make it), this restaurant was a surprising delight for me and my family.
Biryani House (Cary) - Fresh flavors and just the right amount of spiciness makes this lunch buffet a must-try if you are a fan of Indian food. With only Indian cuisine offered, it gives you a chance to try out different dishes you might have not ordered. I tend to find what I like and stick with it, so this place really encourages me to sample other items. I also like that they label the food with names and descriptions, so you know what it is before you pick it up. Make sure to try the Gulab Jamun for dessert! It is a spongy milky ball soaked in a slightly sweet syrup. Perfect to help you calm down the heat!
Do you like to eat a buffets? Which ones do you recommend?


text:
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First Sighting of Jellyfish Eyes' Monsters
Two of the monster designs for Part 1 of artist Takashi Murakami's proposed live-action fantasy trilogy, Mememe no Kurage (Jellyfish Eyes) [if you need to catch up, see The Art of Monsters: Takashi Murakami and Jellyfish Eyes], have surfaced — … Continue reading
Posted in Animation, Daikaiju, Fantasy, Film, Giant Monsters, Japanese, Update Tagged daikaiju, early artwork, Jellyfish Eyes, Takashi Murakami 3 Comments


text:
I'm going to be volunteering at a polling place and need to be there before it opens. I'd like to bring something for the other poll workers. I realize that doughnuts are traditional, but I'm very reluctant to stop off to pick up fresh doughnuts because I don't want to be late.
Is there anything you can suggest that I can purchase the day before that will still be fresh the next am? Thanks for any suggestions!


text:
88 Minutes (2007) Movie Images Gallery
A thriller about a college professor who, while moonlighting as a forensic psychiatrist for the FBI, receives a death threat telling him that he has only 88 minutes to live. In narrowing down possible suspects, he frantically seeks to communicate with a problem student, an ex-girlfriend, and a serial killer on death row.
Starring: Al Pacino, Alicia Witt, Leelee Sobieski, Amy Brenneman, William Forsythe, Deborah Kara Unger, Benjamin McKenzie, Neal McDonough, Leah Cairns
Directed by: Jon Avnet
Release Date: April 18, 2008


text:
|YDS: 5.13b French: 8a Ewbanks: 29 UIAA: IX+ ZA: 30 British: E7 6c [details]
|Eric Zschiesche
|Page Views:
|Submitted By:
|Jay Knower on Jan 1, 2005
|Comments on Rubberman
|Add Comment
|Show which comments —
By Steve Sangdahl
From: eldo sprngs,co
Mar 29, 2005
|jay,is there actually any gear on this thing or should i say gear that would hold a fall?
By rich bechler
Apr 1, 2005
|hard route.gear would be very dicey.I did the 3rd ascent after eric and Dave .I have real good Betta which I will trade for narcotics.
By Jay Knower
From: Campton, NH
Apr 1, 2005
|Rich,Ok, so you do the really hard mantle move, and you stand up on the good foot. You move right and up now--but how far right? Is the right corner on? Am I thinking of the correct hold near the corner? I believe passing a small roof may be cruxy--are there any moves harder than the mantle? Any info would be appreciated.
By Jay Knower
From: Campton, NH
May 4, 2005
To answer my own questions: there are no holds anywhere near the right corner, and the corner is definitely off. I tried the route for a couple of days, and came soooo close. I just couldn't figure out how to weight the right-hand gaston after rocking up on the good foot. (anybody know the beta?)
This route is absolutely classic, with weird, balancy moves at the start, giving way to beautiful powerful moves through a small roof and up a tight corner. Plus, unlike many hard routes at DL, there are actually holds on the thing.
By Tom Anderson-Brown
From: Madison, WI
May 11, 2005
|I'd love to watch someone climb this route. Thanks so much for the great pictures Jay. Incredible!
By Remo
From: Madison, WI
Mar 17, 2009
|Rubberman felt good today, the weather was beautiful. First move felt hard(as my tips were very sore), but better than the last time I was on it. No send yet, but overall this route is super fun, and oh yeah- hard.


text:
It's February 29th. That in itself is a bit of madness. But now we enter March...and things are getting just ridiculous.
To me March Madness has little to do with basketball...other than I generally go Mad durning the Month of March from being bombarded with basketball coverage and discussion. F basketball. I used to enjoy playing it...probably still could, despite my complete lack of skill...but watching it or even listening to commentary makes my face bleed.
No, for me, there much more important events that March signifies...all of which keep me even a little more unbalanced than usual.
-Spring is fast approaching. This means 2 things:
Fishing opportunities in general will vastly re-expand as seasons (trout) and bodies of water (ice and snow) both open; Good
Remaining winter fishing opportunities are quickly waning, which may result in this year's attempt at winter trout a complete failure; Bad
-St Patrick's Day. I'm not sure what else to say about this. Last year we had three weekends of celebrations (rather than the regular 2) due to incliment weather. I love the Irish (being less than 25%, I can't consider myself Irish); instead of canceling an even due to a blizzard, let's just hold it twice! Unfortunately, after 4 years of attending the Most Commercialized Ethnic-Holiday Celebration on the Face of the Earth, I just can't do it this year. It's a shame too, because it basically feels like an RPI multi-class reunion...not to mention everything else there is to do in NY (which is why we use this day to sit in a mall and drink our faces off?). Anyway, regardless of missin out on Manhattan this year, there will be much tomfoolery around here, of that I am sure. Guinness will flow like milk, whisky will flow like Guinness, and Irish Carbombs will abound as folk, punk and Gaelic stream from the speakers. Flogging Molly's "new" single has been on EQX nonstop it seems recently...a little pop-sounding compared to their better stuff, but it's impossible to hear Molly without getting at least a little pumped. I can't wait to forcefeed the juke with whatever bills are in my pocket over the next month.
No doubt, the month of March will be a severe test of my patience, perserverance, and certainly my liver. Really, FlyingTies' header pretty much sums it all up. I just wish all of the hard days were gone.
February 29, 2008
March Madness....
It's February 29th. That in itself is a bit of madness. But now we enter March...and things are getting just ridiculous.
Posted by FoulHooked at 2:01 PM 0 comments
February 26, 2008
Kid Tested, Mother Approved...
I've heard of "cat"-fishing before, but dropping a spey in your own kid's cereal bowl seems like taking it a little far. Hell, the barb's still there, maybe he's fishing for keeps. At least I know who not to call if I ever need a babysitter.
Still, it's not as bad as it seems. I know the little buggers aren't that discerning, but there's no way that's being mistaken for a sweetened puffed ball of grain meal. If some poor sucker does fall for it, he's probably better of being eaten anyway. God that looks painful...I didn't realize cigarette smokers were such massochists.
Moral of the story...don't feed your children Kix...they are addicting and will result in DayTripper eating your babies, if not your cats.
Posted by FoulHooked at 12:50 PM 1 comments
February 20, 2008
Plans are finally being solidified. I will be spending my weekend enjoying wintertime versions of this.
I brought my camera, but was too busy enjoying the scenery (and the driving...I'm just glad there are multiple routes to/from the upper Deerfield from SR-2...two-wheel drive wasn't making it back out the way I went in), and too dumb to stop and break it out. Aside from just the terrain, there is some very fishy water back there, and a lot of choices in close proximity to one another. I may very well spend more time fishing MA than NY this summer. I even found a couple potential options, weather (and/or hangover) permitting, to test the new waders in this weekend. If those mountain spring creeks are as warm as I hope, who knows, I might even get some adorable little creature to acknowledge me this weekend.
Posted by FoulHooked at 1:43 PM 0 comments
Market Saturation
Where are these coming from? Before I had the idea to start this up last fall, I could only find a handful of fishing-related blogs, and less than five that were both interesting an applicable to me. How is it that I now have over 20 blogs in my "Outdoors" feed in Google Reader? All but 2 of those relate to fishing (generally fly); I'd like to read more general outdoor rec publication too, there just isn't enough time.
I'm going to need to cut back on my work hours just to keep up with all this stuff.
(At some point, I'll post them all, not right now...supposedly, "it is not the proper use of company time.")
Posted by FoulHooked at 11:57 AM 0 comments
February 19, 2008
Don't Fret the Weather...
...when planning your next destination trip. Germany seems to have cracked the code, now claiming the ability to predict weather patterns six months into the future.
That's amazing seeing how we have so much trouble predicting temperature from day-to-day.
"...the conscientious browser would need to mentally append "with an accuracy of 3°±2°" to the temperature prediction and realize this results in a two degree span at best and a ten degree span at worst." (OmniNerd)
CustomWeather's (which owns weather prediction service www.MyForcast.com) own data shows the best you can hope for, with a +/-3 degree margin of error, is about 60% accuracy (between 1, 2 and 3 day forcasts). Which fits with the level of accuracy for this new, incredible technology:
"The forecast for this June in Germany: a 60 per cent likelihood it will be hotter than in an average year."
60% chance it will be hotter than average? Really? In the era of global warming? Vegas oddsmakers can do better than that. I guess I'll just keep that raincoat handy whenever I head out.
Posted by FoulHooked at 4:48 PM 0 comments
February 15, 2008
Serves You Right
I've always had a bit of a penchant (unfounded?) for hating the southwest "boom," going back to the founding of Las Vegas (no, I wasn't around to despise it then, but throughout my life, it's never impressed me).
One thing that we use to distinguish ourselves from non-humans is the ability to alter our environment (sometimes drastically) to make it more hospitable. We often seem to take pride in how effeciently we do so. In this case, we made the desert bloom throughout the American Southwest. Quite an achievement, really. LA, LV, Reno, Scottsdale, Phoenix, etc., etc., the list goes on; none of these places would be able to flourish as budding and massive metropoli over the years on their own merits. They lacked the molecule of life. No, not Scotch (they didn't have that either), water. But we fixed it.
Of course, anyone with half a brain can tell you we are not nearly as effective in land transformation as we try to convince ourselves. Watering the desert has been problematic for decades, and it looks like it may actually catch up with us in the near future (via GoBLOG). [I say "us"...but you know who I'm talking about.]
There's a reason inhospitable areas don't sustain large populations (hint: it's because they can't). Certainly, if you were to go through all the trouble of altering the hydrologic regime of an entire region, and taking on all of the implications that go with it, planning future sustainability through curbing development seems prudent. Maybe that's just me.
Sometimes regulators do need to take responsibility. It's a strange concept, I know, but it would be tragic if their constituents suffered because they lost sight of the long-term. I suppose that's the same old story. Of course, it's easy to preach from this soap box living in the (relative) northeast, where water and infrastructure are both plentiful.
Perhaps it's just another scare, but let's hope somebody important at least takes note. Sustainability isn't just about looking out for our great-great-great-grandchildren. And as much fun as it is to hear people who don't belong there fighting over a resource that shouldn't be there either, I sure hope they make it work. I can't imagine the prospect of redistributing Hollywood across the rest of the country being a good thing for anyone.
On another note, Taylor & Vadney's perpetually decreasing materials stocks contained no hair or fur whatsoever. Did manage to improvise a fluffy white fur pattern out of the deteriorating scraps I still have...that's good enough for me, and kinda what it's all about anyway. I won't embarass myself by posting any pictures, but I will reccommend trying your own wire dubbing (slash-improvised hair hackle) loop I've been reading so much about. Excellent solution for making use of falling-apart-pelts.
Posted by FoulHooked at 9:47 AM 0 comments
February 14, 2008
Fly-Fishing for Humans?
Saw this guy on Colbert last night. And I thought I was alone in my ruminations on making "bugs" a larger part of our diet.
Ecologically it makes sense. Physiologically it makes sense. It's already done around the world. I mean, come on.
Just be careful while foraging. You might end up with more than you bargained for.
Talk about a reason to go barbless...
Posted by FoulHooked at 4:13 PM 0 comments
I originally lumped this in with the previous post, as it seemed a fitting subject for how I truly got started fly fishing. But I think this subject deserves it's own post...it was kind of getting lost before.
I owe somebody a large sum of gratitude, and here seems a reasonable place to put it.
Growing up I was lucky enough to be surrounded by genuinely kind elders. One person in particular, a long-time family friend, is responsible for fostering my interest in fly fishing and tying. Although it took about a decade to take hold in a major way, this man provided me with a lot. Himself an avid outdoosman (fly-fisher and bow-hunter), he knew of my interest of fishing Lake George and the surrounding area with my father, who would often brag to him about our weekend exploits. One day he simply left me stunned, with the gift of an essentially complete, self-contained, fly-tying kit consisting of one (old and clunky, but useful) vise, all the requisite tools, fur, feathers, hair, thread, chenille, hooks...you name it, it would have taken hundreds of dollars to re-create what is in that box (which itself was a custom creation...a ~2'x2', sectioned wood panel carrying case). The materials and tools were old, warn, and a bit chewed, but the gift was priceless.
As if the kit weren't enough, he continued to enlighten me with literature (which I consistently forget to grab when I visit my parents) on all aspects of angling. Without me realizing it, he kicked off a passion in me that will (hopefully) continue with me throughout my life.
Although I have not seen him in a very long time, a chance to fish with him (health permitting) would be a true joy and honor, something I have never had the chance to do. Perhaps this season....
From the very bottom of my heart, I thank you Dick.
Posted by FoulHooked at 1:15 PM 0 comments
Heeding One's Own Advice
It's never easy.
For example; finishing projects before moving on to new ones, not putting the cart before the horse and, especially, not overextending yourself for something you dont need.
I found myself pouring over tying vises this morning (not like I have the spare time to do that as it is). The DanVise is pretty tempting, especially at under $80.
See, I consider tying more or less an essential part of the fly-fishing passtime (for me it's a hobby, not a sport, art, etc.). I tied my first flies years before I learned to cast them. Everyone in my generation (of tyers) must be familiar with the yellow wooley bugger. I'm pretty sure it's the only pattern they let you tie at the 1-hour introduction to tying course. I still have 4 or 5 left from that first foray (about 12 years ago) that haven't yet rusted out. Following that, I continued with some streamers and bucktails. If memory serves me correctly, my first (and still only) LLS came trolling one of my own paltry black-nosed-dace-like creations on Lake George. [Speaking of land-locks, Happy Valentines Day Lake Champlain Salmonids]
Well, over the years, I have fallen (waaaayyyy) out of practice. Those early compilations of fur and feather far exceed anything I can put together right now. And seeing as I have been failing misereabley at getting some streamtime, getting back to tying would be the next logical step.
Unfortunately, there are always too many unfinished projects and time commitments, and I just spent a nice hunk of change on replacing my (ungainly) waders with something a little more practical and comfortable. And the truth is, I just don't need a new vice. The two I have, rough though they are, are still in working condition. Besides, it would be nice to brush up on some skills before making another investment (who knows, maybe I'll never develop a steady hand).
So what is a boy to do? Well, I can find/make time for tying (as Joe says, it isn't that you don't have enough time, you just haven't made it a priority). But for now, my aging, dilapidated, low quality vices will have to do. I can live with that. I think, though, this calls for a trip to Taylor & Vadney to see if they have any rabbit (if anyone has suggestions on additional Albany-area fly shops, I'm all ears). If I do make it to the river any time soon, it would be nice to have some custom bunny-strips with me. I hear white is the color for winter browns, but I'll take what I can get.
Posted by FoulHooked at 11:34 AM 0 comments
February 13, 2008
Of Rats and Men
"I'm not naive enough to think other people haven't figured [their own mortality] out. But to me, it seems like most of the world is in denial about this fact.... So many more people out here are in such a hurry to make a bunch of money, have a big house, drive over rated cars, etc. They hate the rat race, but they are don't know how to carry on from the sidelines."
I can't match Daytripper's way with words or clarity of vision, but I do echo his sentiments.
I think we all struggle at some point(s) in choosing what is truly important, and what are just distracting us from life. The past 14 months have put me through the wringer on this one...sometimes, even when you have a handle on it, the inability to convince someone else of the same can be what gets to you.
XKCD's comic for today didn't make pondering life any easier either.
Good luck. Get on some water. Got any good last-minute excuses that can get me out of work and on the stream Friday?
Posted by FoulHooked at 3:01 PM 0 comments
February 8, 2008
Spey Nation's "The Experience 2008"
is receiving some local press. As it should. I need something more to waste my meager income on, so I hope to attend. As for company, Pat, I'm looking at you.
Now, if we can just schedule some Lake Ontario trolling for that week, it will likely be the event of my summer.
Posted by FoulHooked at 12:13 AM 0 comments
February 7, 2008
Trout Pout-ing
I'm in a bit of a funk myself...I don't really see the resemblance though, beyond the scaly skin of course.
Been months since I've seen a trout or so much as cast a fly, weeks since the green wave-splitter even took me for a float. May not be the worst of my problems at the moment, but some QT with my rod and some rubber pants would certainly brighten the mood a bit. So far work and weather have both conspired against me. So this weekend there are several simple solutions to these mid-winter doldrums (none exclusive of any of the others):
-Get serious about tying (HA!)
-Replace my rubber pants with something shiny and new
-Sit, alone, swearing at some Tullmore Dew, as the level in the bottle slowly gets lower...and lower
-Break out the big-shoes and get some snow under me
-Head up to the VT border to feed some nymphs and buggers to Battenkill boulders
-"Sort life out"
-Rando trip to CT or scenic Hamilton, NY
Over/Under for number of above goals achieved? 0.1, now taking bets. No matter which way it goes though, There Will Be Blood (I hope).
"I'm sorry Shaun"
(Well, it's 4:45 on Friday and it looks like the most likely weekend activity is none of the above. Let's replace -Rando trip... with -Work from home.)
Posted by FoulHooked at 11:44 PM 0 comments
February 5, 2008
Reason #12...
...why I'll never be able to live in Oklahoma.
Browse around...check out the stocking schedule. Yup, OK's got their their act together. Don't ask me why I looked it up.
Posted by FoulHooked at 3:58 PM 0 comments


text:
Mark 13: 24-37
November 27, 2011
Rev. Dr. Fritz Ritsch, Preacher
I'm sure we all remember the big billboards a few months ago announcing that The Judgment was coming on May 23, 2011, so we all better be ready. And of course, it didn't, and then the purveyor of this idea, evangelist and radio host Harold Camping, said he'd made a slight math mistake—don't we all make math mistakes?–and actually the day of the Lord was to arrive on October 21, and of course, it didn't again, but I don't think people were paying as much attention, so I suppose it could have come and nobody noticed it.
But not to worry, there's no shortage of potential apocalypses. We still have the Mayan Apocalypse to look forward to in 2012. And there are promising historical trends, such as ongoing turmoil in the Middle East, always a hopeful sign that the end is near. Some people are concerned about a meteor that's going to pass awfully near the earth in a few decades, so that's a bright spot on the apocalyptic horizon. So there's still hope.
What do we make of our fascination with the End Times? It seems like even from the beginning, we were looking for the end.
Our Gospel lesson is Jesus' response to a question that His disciples have asked him about this very topic. "Master, tell us, when will this be?" To their disappointment, Jesus tells them that even He can't answer the question when. And since that's the situation, He advises them: STAY AWAKE. "For you do not know when the master of the house is coming, in the evening, or at midnight, or at cockcrow, or at dawn, or else he may find you asleep when he comes suddenly."
And then, eerily, Jesus seems to step right out of the pages of the Bible, right out of the mists of history, and speak directly to us: "And what I say to you I say to ALL: STAY AWAKE."
He speaks to his disciples, and he speaks to us. His disciples didn't understand him. They don't realize that He's predicting that the End time will come in the next few days and they will miss it. "For you do not know when the master is coming, in the evening, or at midnight, or at cockcrow, or at dawn…" he is describing the "four watches of the night in Roman reckoning" (NIB, Abingdon Press, 2003. p. 1837). On the night that Jesus is arrested, He goes to pray in Gethsemane, and his disciples are supposed "stay awake," but they do not. He comes to them three times and wakes them, and after the third He says, 'Enough! The hour is come; the Son of Man is betrayed into the hands of sinners." The hour is a biblical catch-phrase for The Apocalypse.
It gets worse. Jesus had warned Peter that Peter would deny Him three times before the cock crowed twice; and sure enough, at cockcrow, the third watch of the night, Peter had denied Jesus three times. "And Peter broke down and wept." (Mark 14: 72).
The apocalypse was right on top of them, and they did not stay awake.
Peter and the others were looking for the apocalypse of stars falling and the moon turning to blood. They completely missed the apocalypse right before their eyes, humanity putting God on trial and condemning Him to death. The apocalypse had happened on a quiet night after a good meal and a lot of alcohol. How commonplace it had seemed! How unexpected! And so they hadn't stayed awake.
The apocalypses of our lives—the apocalypse that Jesus predicts here—they often don't seem apocalyptic. They sneak up on us. A moment when another kid at school suggests that it'd be fun to pick on the fat kid. Or an older kid tells you that you'll be cool if you smoke this. The moment your boss tells you that if you cut this corner, maybe fudge the paperwork a little, it won't do any harm, and there could be a promotion in it for you. The attractive person who seems so much more interested in you than your partner ever seems.
Or maybe it's the unexpected diagnosis at a yearly medical checkup or the pink slip at work or the principal on the phone with bad news about your child.
And suddenly the world crashes in around you.
Keep watch! Stay awake! Jesus isn't simply predicting the ultimate end-time to beat all end-times! He's predicting the inevitability of apocalypse. Our faith doesn't make us immune from the common lot of human life. All of us suffer. The question is, are we ready? Have we honed our spiritual defenses in the good times so that we can find them when we need them in the bad times?
But important as it is to note that Jesus was warning his disciples that they were about to experience a personal apocalypse, we can't forget that he's also talking about the larger kind of apocalypse. In the paragraph before the Gospel lesson today, he warns his disciples "when you see the desolating sacrilege set up where it ought not to be, then those in Judea must flee to the mountains…" (Mark 13: 14). Is Jesus speaking symbolically about something that will happen in our future? Perhaps. But he is for sure talking about something that will happen in His disciples' future. In AD 41, about 10 years after Jesus tells this to his disciples, the mad emperor Caligula ordered that his statue be set up in the Jerusalem temple—he ordered it would be set up right in the Holy of Holies, the place where only the High Priest could ever enter, and then only once a year. If that had happened, it would have caused the Middle East to explode in violence. The Roman governor at the time was terrified. He delayed putting up the statue as long as he could—and then, just in time, Caligula was assassinated.
But that event pales in comparison to another event He seems to be predicting—the terrible Roman siege and ultimate destruction of Jerusalem and the Temple in AD 70.
But terrible as that event was, it too pales in comparison to the many other apocalypses that have afflicted humankind in the centuries since: the Black Plague of the Middle Ages—the Armenian Genocide—the rise of the Third Reich and the Holocaust—the Rwandan Genocide of the 1990s. All of them were apocalyptic to those who experienced them. All of them throw down the gauntlet before us as people of faith and challenge us—can your faith stand up against this? If it's happening to you, will you be ready? If it's happening to someone else, will you stay awake—or just close your eyes?
Martin Niemoller was a German war hero, a submarine captain in WWI, who became a Protestant pastor. In the aftermath of World War I, as he saw his country falling apart, he supported the rise of Adolf Hitler with enthusiasm. But as Hitler began to dabble in affairs of the church, Niemoller became cautious, then alarmed, and finally outraged. He was arrested and imprisoned in 1939. After the war, Niemoller admitted that he had made a tragic mistake—he had allowed his prejudices to blind him to the apocalypse right before his eyes. He wrote in 1946:
First they came for the communists, and I did not speak out, because I was not a communist.
Then they came for the socialists, and I did not speak out, because I was not a socialist.
Then they came for the trade unionists, and I did not speak out, because I was not a trade unionist.
Then they came for the Jews, and I did not speak out, because I was not a Jew.
And then they came for me, and there was no one left to speak for me.
When the apocalypse came, Niemoller was not ready. He was on the wrong side.
But God was bigger than the apocalypse, and God used it to change him, to enable him to grow into a deeper, more responsible faith. Niemoller spent his remaining decades as a committed Christian pacifist. His confrontation with the apocalypse changed him profoundly.
The message we often miss in our fascination with the apocalypse is this: God is bigger than the apocalypse. That's what Jesus wanted his disciples to know: that, in the end, the God of love, grace, and justice will turn out to be in charge. The sovereign Lord of Love is larger than our apocalypses.
That message is meant to change us. Every apocalypse that we as individuals face, every apocalypse that we as a people or a world face, is meant to change us. The challenge is to put the love of God ahead of our fear of death, destruction, and change itself.
The disciples learned that the Lord of Love was larger than their apocalypse. They'd run away, betrayed him, denied him. And he'd died and they thought it was the end. But he rose from the dead, he found them, and he commissioned them to save the world.
Niemoller learned it. He'd betrayed the Gospel. He'd indirectly contributed to terrible Nazi atrocities. It was as close as we've come in modern times to the true Judgment Day.
But God gave him—and the world—another chance. The real apocalypse is when we, you and I, and someday maybe the world itself—don't get another chance.
There are some guys from a local church who've been making the news lately by preaching on the street corner near Barnes and Noble downtown. They're on a soapbox preaching that you better not head down to 8.0 or be gay because the Judgment Day is coming and partying and alternative lifestyles are the very work of the devil.
A St. Stephen parishioner had a bit of a run-in with them. He is a county prosecutor. He works the gang unit. He is six-four and he used to be a gigantic and very mean college football player. He got right in the face of the guy with the megaphone and said:
"God loves you!"
The guy was taken aback, but our parishioner wasn't through. "Listen–to–me: GOD LOVES YOU!" he said.
He did not touch them. Those guys have no idea how close they came to their own personal apocalypse.
Are those guys crazy? There are true, awful apocalypses the world over and they're worried aboutFort Worthon a Saturday night?
Sometimes the good news of God's love has to get in our faces and shake us up, because we're so easily misguided by our assumptions and prejudices, and because in our troubled world, we so often find it hard to believe that it could be true that God loves us.
The terrible reality for people who take reality seriously is that apocalypse is on us daily. If it's not on us, it's on someone else. It may be as far away as the South Sudan, where the Janjaweed rebels have engaged in horrific mistreatment of a helpless populace; or it may be as close as neighborhoods terrorized by gangs on the North Side of town. We turn a blind eye to it only at great spiritual risk, because as Niemoller learned so traumatically, when we do not stand up for others, we victimize not only them, but ourselves.
It's tempting in the face of this reality to insulate ourselves and our families in a cocoon of safety. But that's not our commission as Jesus' disciples. The world is filled with apocalyptic signs, but our calling is proclaim The Good News: Jesus the Lord is larger than the apocalypse. The God of love and mercy, of Justice, Grace and Healing, is victor even over the End Times.
It's a hard message to convey; it brings its own questions and difficulties. So Martin Niemoller got another chance.Germanygot another chance. Six million Jews didn't get another chance. What do you say about that, God?
No easy answers.
But we can thank God for another chance. And pray that God helps us get it right this time.


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Hitachi SU1510 Variable Pressure SEM
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Compact, high performance and easy to use makes the SU1510 a great general purpose VPSEM for any advanced laboratory. The SU1510 is a medium size chamber Variable Pressure SEM with the same performance and features as the S-3400N and S-3700N models. Quad Bias gun electronics greatly improves low voltage performance and increases beam current well suited for today's SDD X-ray detectors. Dual high-take-off ports accommodate two EDS detectors mounted 180 degrees apart for tice the analytical data collection plus eliminates X-ray map shadows associated with rough sample surfaces. A high speed, clean, efficient TMP reduces the pumpdown time tojust over 60 seconds and eliminates the need for water cooling.


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- About
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Psychology Professor Earns National Grants to Research Treatments for Cancer Patients and Caregivers
Release Date:
Aug 27 2013
An assistant professor of psychology in the School of Science at IUPUI will apply nearly $1 million in recently approved grants to study how a telephone-based treatment process can help ease stress and anxiety issues experienced by advanced-stage lung cancer patients and the family members who care for them.
Catherine Mosher, Ph.D., Psychology
Catherine Mosher, Ph.D., said telephone-based psychological counseling is nothing new for cancer patients; however, few studies have examined how extending these services to family caregivers impacts the overall quality of care and comfort of the patient.
"Many caregivers are highly burdened and stressed as a result of the commitment it takes to care for an advanced-stage cancer patient," said Mosher. "Through this flexible approach to mental health care, we're able to tailor the process to best treat the particular problems they are experiencing."
In recent weeks, Mosher learned she had been selected to receive nearly $1 million in grant funding related to her research, including support from the National Cancer Institute and the American Cancer Society. In addition, the IUPUI Center for Cancer Population Analytics and Patient-Centered Informatics, through the IUPUI Signature Centers Initiative, pledged $300,000 to support projects for Mosher, David Haggstrom, M.D., and other campus researchers to examine ways technology can enhance cancer patient care.
All grants were for multiple years. The American Cancer Society grant awarded to Mosher in July was presented to only three people across the country. She also has two previous grants awarded by the National Institutes of Health.
The telephone-based treatment program would involve four sessions of counseling and evaluation by a trained mental health professional with both the patient and their primary caregiver. This would occur across four to eight weeks, and the focus would include delivering psychosocial and behavioral interventions that could improve symptom management and quality of life.
"Past research has shown that up to half of caregivers for cancer patients experience a high level of stress and many report health risk behaviors such as chronic sleep problems and persistent smoking. This has a marked effect on the quality of care they are able to provide these patients," Mosher said.
Mosher and her research team will gather data during the next few years with plans to implement a larger scale, randomized project in the future.
In addition to her success earning grant funding for her primary research, IUPUI students mentored by Mosher have had a successful year under her guidance as well.
(Left to right) Rebecca Adams, Joseph Winger, Catherine Mosher, Shannon Christy, Leah Van Antwerp
Doctoral psychology student Shannon Christy was one of two students to recently receive the Scott Mesh Honorary Grant, a highly competitive national award from the American Psychological Association of Graduate Students. Graduate student Joseph Winger earned the Behavioral Cooperative Oncology Group Pre-doctoral Fellowship. Graduate student Rebecca Adams earned the National Cancer Institute R25 pre-doctoral training award, and undergraduate Leah Van Antwerp earned a scholarship from Psi Chi, the international honor society for psychology.
Media ContactsCandace Gwaltney
Associate Director of Communications


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Young County TXGenWeb
Valley View Community
|The Valley View Community was a group of farm houses located north of Graham. There was a Baptist church building in the community at one time.
Nothing remains of the community today.
submitted by: Dorman Holub
© 2009
Email additions or submissions to Dorman Holub
Young County Archives | Young County Home | TXGenWeb | USGenWeb


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Right-hander Jeff Karstens probably is done for the season due to what manager John Russell described as a "cranky" arm. Karstens (3-10, 4.88) had his shoulder heavily iced in recent days.
"The doctors don't see anything structurally wrong with it," Russell said. "It's been a long year for him, and it's getting a little cranky. We don't know for sure yet if he'll be able to make another start."
Karstens last pitched Aug. 31 against the Cubs and picked up the win by allowing two runs on four hits in six innings. But he was working on extra rest, as his Aug. 25 start was skipped because of the arm troubles.
»»» To replace Karstens tomorrow against the Braves, lefty Brian Burres (2-3, 6.31) will make his eighth start of the season. If he is recalled, right-hander Brad Lincoln, who pitches tomorrow for Triple-A Indianapolis, is a possible fill-in for Karstens the rest of the month.
»»» Righty Joe Martinez was called up from Triple-A Indy, but he does not appear to be a candidate to take Karstens' rotation spot. When I asked how Martinez will be used, Russell immediately responded, "Bullpen."
Martinez went 0-1 with a 4.91 ERA in four games with the San Francisco Giants before being traded to the Pirates on July 31. "He's done OK," Russell said. "He's had some success, but also had some struggles."
The Pirates still plan to make Martinez a starter in the long term.
»»» Congrats to High-A Bradenton, which last night clinched the South Division of the Florida State League. Bradenton will face the Charlotte Stone Crabs in the first-round of the playoffs, a best-of-three series which will begin Tuesday night at McKechnie Field.
»»» The Pirates are wearing "JLB" patches on their uniform sleeves to honor former GM Joe L. Brown, who recently died.
Lineups for this afternoon's game:
Nationals (58-78) – 1. Nyjer Morgan cf, 2. Ian Desmond ss, 3. Ryan Zimmerman 3b, 4. Adam Dunn 1b, 5. Roger Bernadina lf, 6. Michael Morse rf, 7. Adam Kennedy 2b, 8. Wilson Ramos c, 9. Jason Marquis (1-7, 8.13) rhp
Pirates (45-90) – 1. Andrew McCutchen 8, 2. Jose Tabata lf, 3. Neil Walker 2b, 4. Garrett Jones 1b, 5. Pedro Alvarez 3b, 6. Ryan Doumit rf, 7. Ronny Cedeno ss, 8. Chris Snyder c, 9. Charlie Morton (1-10, 10.03) rhp


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Created for udig, for now used to browse netcdf files along a temporal axis... and since a screencast is more than many words... enjoy (as I did :) )!
...a summary of how-to-do-(mostly-GIS)-development-things in JGrasstools-gvSIG-Geopaparazzi-uDig-Eclipse-Java as they pass me by...
That's all folks... and code!
Wednesday, August 12, 2009
Friday, August 7, 2009
Time and Z navigation internals are here
Another bit of netcdf project is done. This time it was creating the internals of navigation... more about it on the project page.
Just a snappy to make this blog not look so annoying :)
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Despite the attraction of promised lower IT costs, concerns about privacy and information security, as well as concerns about access, control, auditing and compliance, continue to exert a strong influence on companies' decisions to run applications in the cloud. Recent information security issues with Microsoft, Target, and the NSA, along with well-publicized outages at public cloud providers' sites, have caused many CFOs to start scrutinizing risks of relying on the cloud and weighing these risks against costs. The cloud is no longer just a money-saving mechanism.
As more CFOs oversee technology efforts, the security and reliability of the cloud have brought into focus key questions about how enterprise applications are delivered -- both today and in the future. With so many cloud solutions commercially available, CFOs are challenged with identifying which ones are good fits for their organizations, and knowing which criteria to use in their evaluations, once the top players have been identified.
During this Webcast, we will:
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Delicious Fruit Punch Drink Mix This fruit punch drink mix packs a punch of flavor. Manufactured under the strictest quality control standards, it is a top quality product. It is approved by the USDA, FSIS and the FDA. It is fully manufactured in the USA. This drink is the perfect fruit punch recipe for any occasion. Whether you are looking for a drink to accompany a meal or a drink after a hot day this is the right one for you. When you have children over at your house and you need to give them a drink, consider this drink.
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Like most punch drinks, this drink comes with citric acid, artificial flavor, sugar, titanium dioxide, dextrose, maltodextrin, gum Arabic and food color. It is also fortified with Vitamin C in the form or ascorbic acid.


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GlaxoSmithKline's drug Avandia should stay on the market, federal health experts voted Wednesday, but the controversial diabetes pill should be subject to new restrictions due to risky heart side effects.
A panel of Food and Drug Administration advisers voted 20-12 to keep the once blockbuster pill available for diabetics. But 10 panelists also called for limiting who can receive and prescribe the much-debated medication.
The vote marks a win for British drugmaker Glaxo, which has been battered in the press and on Capitol Hill for its the handling of the drug.
The FDA is not required to follow the advice of its panelists, though it usually does. Officials said they would review the meeting transcript and make a decision on Avandia as soon as possible.
The vote also came despite an earlier ruling by the panel that Avandia appears to increase heart attack risk compared with other diabetes treatments. The panel voted 21-4 that Avandia is more likely to cause heart attack than its closest competitor, Actos. Eight panelists said there was not enough information to make a decision.
Ultimately though, panelists said the dozens of contradictory studies of Avandia didn't show strong enough evidence to justify removing a drug used by hundreds of thousands of patients.
"I would be concerned about the precedent that would be set to have this quality of data sufficient to remove a drug," said John Teerlink of the University of San Francisco.
The agency convened the two-day panel meeting to help untangle reams of conflicting data over Avandia.
The FDA has been down this road before. Three years ago a similar FDA panel voted to keep Avandia on the market, and the FDA responded by adding bolder warning labels to the drug.
"In terms of what has changed since 2007, I think the totality of evidence is much stronger," said panelist Clifford Rosen of the Maine Medical Research Institute. "It's still not absolute but it's stronger."
Despite the vote on Avandia's heart risks, panelists didn't reach a firm conclusion on whether Avandia is more likely to cause death than older drugs. Twenty panelists said it did not appear to cause death, while 12 said they didn't know. Only one panelist concluded the drug leads to death.
The panel's vote to keep Avandia on the market is a vote of confidence in FDA leadership, who have been criticized by some members of Congress for not pulling the drug earlier.
Minutes before the final vote on Avandia, the FDA's director for new drugs emphasized the high bar needed to pull a previously approved drug from the market.
"The two that have been withdrawn for cardiovascular concerns — Vioxx and Zelnorm — showed three-, four- or five-fold increase," said John Jenkins "That's the background of where we stand in terms of increased risk of very rare events."
The FDA first approved Avandia in 1999, and it quickly became the top-selling diabetes pill in the world. U.S. sales have plummeted from $2.2 billion in 2006 to $520 million last year as safety concerns swirled around the drug.
Diabetes patients face considerably elevated heart disease risks, so the 2007 study in the New England Journal of Medicine about problems with Avandia was enough to make doctors switch patients to other therapies. The American Diabetes Association even issued a warning back then, urging patients taking Avandia to talk to their doctors.
"We stopped prescribing it over the last couple of years because of the cardiovascular risks and because there was an alternative in the same class of drugs, one that works chemically in the same way, that has not been shown to have cardiovascular risks," said Dr. Brendan O'Malley, medical director of the Diabetes and Endocrinology Institute at University Community Hospital in Tampa.
"That drug is Actos. So it's a no-brainer to say, let's go to Actos and not wait until they take another look at the safety of Avandia."
Still, he said, some patients prefer Avandia and have refused to go off it.
"We have a few patients, no more than a dozen, who have continued to take Avandia even though we tell them about the risks and that there is an alternative. But some patients will say, 'I've never felt better in my life' and they don't want to switch. It's very good at controlling blood sugars."
Like other medical experts, O'Malley's major piece of advice for patients still taking Avandia is this: "Don't stop your medication without talking with your doctor and discussing alternatives."
Times medical writer Irene Maher contributed to this report from the Associated Press.


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Kukla's Korner Hockey
The Roadie
by Paul on 03/26/07 at 01:25 PM ET
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from the Pittsburgh Penguins,
Few can say they went on tour with legendary rock bands.
Joel Kwiatkowski did, though, as he and his wife followed bands Poison and Cinderella through Michigan on their "20 Years of Rock Tour" last summer.
And, it wasn't like he was just following the bands to catch some concerts. Kwiatkowski, a guitar player, actually got on stage with the bands during their sound checks.
"I am actually really into music. Chris Moody, my old [assistant equipment manager] in Florida, his best buddy is a tour manager for a bunch of bands. So, we got out with Poison and Cinderella on a little tour last summer," he said.
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Best Kindle Fire Games 2014 – Top 10
The Kindle Fire is a computer tablet version of the original Kindle and is said to be a strong competitor of apples' iPad. It has many more capabilities than the original Kindle in which playing games is one of them. The original Kindle was mostly if not completely for reading e books. Amazon has stepped it up a bit with the Kindle Fire and with a pretty low and competitive price, it should do fairly well on the market. With the ability to play games on it, there are developers from across the world working on Kindle Fire game apps that work exactly like that of the iPad. There are a ton of great games that you can play on the Kindle Fire and below we have put together a list of the top 10 Kindle Fire games available. Check them out and to be afraid to try them and leave us some feedback on your gaming experiences!
Order & Chaos Online – This game is the only available 3D massively multiplayer online role playing game for an android device (Yes, the Kindle Fire is ran on the Android operating system so it is considered and Android Device). Order & Chaos Online features a fast open world with amazing graphics and design. Players can choose their characters from 4 different races and also control gender, appearance, skills and pretty much anything else having to do with their character. There are over 1000 skills that characters can learn and over 2000 pieces of equipment. The options within this game are truly endless and it features over 500 quests for players to complete.
Assassin's Creed – Altair's Chronicles HD – Ever played Assassin's Creed on your xbox 360 or Playstation 3? Well the Amazon Kindle now features a version of this game and let me say, it is awesome. The game is played almost exactly like any other Assassin's Creed game where players master sword skills and acrobatic skills to save the world as they know it. With 6 different weapons to choose from including the grappling hook and daggers from the original games, Assassin's Creed: Altair's Chronicles brings Android Device games to a whole new level. There are even mini games available for players to complete which will put their true skills to the test.
Modern Combat 2: Black Pegasus HD – Are you looking for an Amazon Kindle game that is just like Call of Duty or Counter Strike but better? Well look no further for Modern Combat 2 is just that. This awesome first person shooter can be played against others online or by yourself. There are 12 different maps or battlefields available to play on with 4 different multiplayer modes. These include the all so popular capture the flag mode as well as a team battle mode which is exactly like team deathmatch. There are 15 different weapons available for use that are based from real military weapons. What are you waiting for? This game is one of the best if not the best first person shooter for the Kindle Fire and has a ton to offer its users.
Asphalt 6: Adrenaline – If you are looking for an awesome street-racing game with the best graphics out there then Asphalt 6 is definitely a game you need to check out. There are over 40 different exotic cars and motorcycles for you to unlock and purchase and 12 different locations for you to race them in. The game even features a multiplayer option where up to 6 players can connect and care against one another. Asphalt 6 also features unique challenges as well as different types of races including elimination and beat em all. With 11 different leagues to enter and 55 different events to check out, Asphalt 6 has a ton to offer for being a Kindle Fire game. Check it out today and start climbing the leader boards composed of players around the world.
Dead Space – Dead Space is an action packed first person/third person shooter video game with unreal graphics and design. This is the original Dead Space game made compatible for your Kindle Fire Device. The story line is the same, where players face off against the horrifying Necromorph horde. There are a ton of unique and new weapons including the Core Extractor and the Plasma Saw which is my personal favorite. Work your way through gory space ship corridors and kill everything in site for your enemies will stop at nothing to end whatever hope you had of surviving. Dead Space has a ton of awesome features and unique game content and will continue to get better for the developers are always upgrading and fixing problems.
Plants vs Zombies – Plants vs Zombies is truly the best tower defense game out there and has hit the markets hard. Finally this game is available for the Kindle Fire and brings all the same features and unique content that it did for the iPad and iPhone. Within this game players simply try to not get eaten alive by evil zombies attacking their house. You set up different types of plants in a variety of locations around your house including the roof, the front yard and the back yard which has a pool in it. There are tons of different types of zombies including zamboni drivers, zombies riding sharks and even giant troll like zombies. Plants also have different abilities including freezing zombies or even blowing them up. If you have never played Plants vs Zombies before then I encourage you to check this game out. It will keep you busy for hours and has a ton of features and game modes to check out.
BackStab HD – BackStab is a great game that mixes elements of Assassin's Creed and Prince of Persia. The story line of this game is truly something that needs to be talked about for it is full of action packed adventure and a great combat system. BackStab features a combo based combat system in an open world with tons of challenges and treasures waiting to be discovered. There are 4 unique cities for players to check out each filled with challenging quests as well as hidden secrets. Environments that players might run into include jungles, beaches and even volcanoes. In this open world, players can enhance their characters skills and abilities while finding those who betrayed them. This is an awesome game and will definitely impress you with its eye catching graphics and unique design.
MADDEN NFL 12 – Well look no further for the action packed football game that you have been wanting for your Kindle Fire. Developers wasted no timemaking Madden NFL 12 compatible for the Kindle Fire. The rosters on this game are exactly those of the NFL teams around the league with over 2500 total players making up 32 NFL teams. The stadiums you play in will be filled with thousands of fans and the game truly immerses you into the gameplay. Make trades, keep track of all your teams stats and choose your favorite plays from the play book. Madden NFL 12 allows players to have complete control over their team letting them work their way through the NFL standings and even winning the SuperBowl championship. Do you have what it takes to be a world champion?
Text Twist 2 – Text Twist 2 is the sequel to the all so popular Text Twist and has a ton more to offer its users. This is a letter twisting game where players shuffle letters to form words. The game features 5 different game modes including Timed, Untimed, Letter Mania, Lightning, and Daily Word each of which bring something new and exciting to the table. Simply twist the letters given to you to form pre-selected words to win the game. Depending on the game mode, each round may get harder or quicker putting your skills to the ultimate test. Keep track of all your statistics trying to beat your high score and win trophies to show your friends that they will never compete with you.
Angry Birds HD – Angry Birds is now available for the Kindle Fire and features all the same game content as the iPad and iPhone versions. This game features over 288 different levels each more challenging than the next. Find the best way to pass the levels to get a 3 star rating as well. There are even hidden secrets through out the levels that bare awesome treasures. Each bird has a unique ability and should be used for different purposes. Try your skills out in Angry Birds HD and see if you have what it takes to ultimately defeat the evil pigs.