A systematic review and meta-analysis of the efficacy of lixisenatide in the treatment of patients with type 2 diabetes.

The aim of this study is to assess the efficacy and safety of lixisenatide for treating type 2 diabetes. A systematic search in electronic databases (up to October 2012) was conducted and the manufacturer was contacted regarding unpublished data. Randomized controlled trials (RCTs) were included if they provided information on at least one of the following outcomes: mortality, health-related quality of life, hypoglycaemic events, adverse events, change in HbA1c, body weight, blood pressure, gastric emptying, fasting plasma glucose or 2 h postprandial glucose (PPG). Twenty-six publications and 10 unpublished study reports, relating to 14 RCTs (6156 patients) were included. Eleven studies related to placebo comparisons; active comparators were in three studies. Compared to placebo, lixisenatide significantly reduced HbA1c (-0.52%; 95% CI: -0.64 to -0.39), bodyweight (-0.65 kg; 95% CI: -0.94 to -0.37) and 2-h PPG level (-4.58 mmol/l; 95% CI: -5.88 to -3.28). There were significantly more symptomatic hypoglycaemic events among lixisenatide compared to placebo-treated patients (log OR: 0.54; 95% CI: 0.32-0.75), but significantly fewer compared to other incretin mimetics. In comparison to exenatide and liraglutide, lixisenatide was more effective in reducing 2 h-PPG with a better adverse events profile, but it showed a lower reduction in HbA1c and body weight. Lixisenatide improves HbA1c levels and moderately reduces body weight compared to placebo and showed less frequent symptomatic hypoglycaemic and gastrointestinal events and an improvement in PPG control compared to other GLP-1 agonists. Firm conclusions regarding the performance of lixisenatide compared to other incretin mimetics, however, can not yet be drawn, due to limited data.

[Chronic pain. Overview of out-patient pain treatment in Austria]. / Chronischer Schmerz. Überblick über die ambulante Schmerzversorgung in Österreich.

BACKGROUND: Chronic pain is a widespread social problem. This paper reports on the care situation for patients with chronic pain in out-patient community settings in Austria. MATERIALS AND METHODS: The study took the form of a telephone survey together with internet research. Every second out-patient pain service (from a total of 83) was contacted and 21 out of 42 agreed to participate. RESULTS: The number of community-based physicians with a certificate in pain therapy as well as the number of out-patient pain services showed considerable regional variation. Partial or full interdisciplinary teams are a feature of approximately 50% of out-patient pain units and 76% of such services use guidelines according to their own estimation. Pain perception tends to be measured using pain rating scales rather than pain questionnaires. A wide range of treatments is offered either directly or via referral. CONCLUSIONS: Quality criteria relating to the structure of care established by the Austrian Society for Pain have only been partially implemented. Potential for improvement exists particularly with regards to the prevalence of pain-specific training, interdisciplinary teamwork and the measurement of outcomes.

Optimal pacing strategy: from theoretical modelling to reality in 1500-m speed skating.

PURPOSE: Athletes are trained to choose the pace which is perceived to be correct during a specific effort, such as the 1500-m speed skating competition. The purpose of the present study was to "override" self-paced (SP) performance by instructing athletes to execute a theoretically optimal pacing profile. METHODS: Seven national-level speed-skaters performed a SP 1500-m which was analysed by obtaining velocity (every 100 m) and body position (every 200 m) with video to calculate total mechanical power output. Together with gross efficiency and aerobic kinetics, obtained in separate trials, data were used to calculate aerobic and anaerobic power output profiles. An energy flow model was applied to SP, simulating a range of pacing strategies, and a theoretically optimal pacing profile was imposed in a second race (IM). RESULTS: Final time for IM was ∼2 s slower than SP. Total power distribution per lap differed, with a higher power over the first 300 m for IM (637.0 (49.4) vs 612.5 (50.0) W). Anaerobic parameters did not differ. The faster first lap resulted in a higher aerodynamic drag coefficient and perhaps a less effective push-off. CONCLUSION: Experienced athletes have a well-developed performance template, and changing pacing strategy towards a theoretically optimal fast start protocol had negative consequences on speed-skating technique and did not result in better performance.

Child/parent-assessed population health outcome measures: a structured review.

AIMS: To identify generic measures of health-related quality of life (HRQL) for children and adolescents developed for use within general populations. Instruments are evaluated on the basis of evidence relating to their reliability and validity. METHODS: Systematic literature searches were used to identify instruments, which were then assessed against predefined criteria. Information relating to instrument content, population, reliability and validity was extracted from published papers. RESULTS: Sixteen instruments were identified that had been evaluated among a general population of children or adolescents. Four instruments had reported data on both internal consistency and test-retest reliability. All except two instruments had undergone some degree of construct validation. CONCLUSIONS: The evidence suggests that the Child Health Questionnaire (CHQ) has been the most extensively evaluated for younger populations but is available as a parent-completed measure only. The new version of the Child Health and Illness Profile (CHIP-CE) is particularly promising and has parallel child- and parent-completed versions for young ages. The weight of evidence suggests that versions of these two instruments are suitable for older children. The Warwick Child Health and Morbidity Profile could be used where information on morbidity and health service contacts is required. Once basic psychometric criteria are fulfilled, instruments should be chosen by assessing their content and design in the light of the prospective application.

Tyrphostin AG825 triggers p38 mitogen-activated protein kinase-dependent apoptosis in androgen-independent prostate cancer cells C4 and C4-2.

Prostate cancer (PCa) progression is aided by abnormal autocrine growth factor loops. We screened for small cell-permeable inhibitors of receptor tyrosine kinases that could block their signaling and trigger cell death in PCa cell lines. We found that the human epidermal growth factor receptor (HER)-2/neu inhibitor tyrphostin AG825 is preferentially toxic to PCa cells that are phenotypically androgen independent. These effects were dose and time dependent in the human LNCaP, C4, and C4-2 cell line models of progression and correlated with the inhibition of HER-2/neu phosphoactivation and its down-regulation. In addition, we show that the inhibition of HER-2/neu signaling with AG825 triggers an imbalance between extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase activation, which leads to p38-dependent apoptosis. Inhibition of HER-1 with Compound 56 had no effect. These findings suggest that the androgen-independent C4 and C4-2 cells can be killed by selectively inhibiting their HER-2/neu signaling pathway and provide insights into the mechanism of action of AG825 in PCa cells.

Role of PI3K signaling in survival and progression of LNCaP prostate cancer cells to the androgen refractory state.

The mechanisms by which prostate cancer (PCa) cells progress to a hormone refractory state are poorly understood. The progression process under androgen ablation conditions involves the survival of at least a portion of malignant cells and their eventual proliferation in an androgen-independent manner. The goal of this study was to investigate the role of PI3K signaling in such a progression. Using an in vitro model of androgen ablation, we show that after removal of androgen support, the human PCa cell line LNCaP initially arrested in G(1) and trans-differentiated into neuroendocrine-like cells that eventually resumed androgen-independent proliferation. Both acute and chronic androgen ablation resulted in an increase in basal levels of PI3K and Akt activity, which were sustained throughout the progression process. Under these conditions, inhibition of PI3K, pharmacologically or with ectopic expression of PTEN, arrested cell proliferation and blocked progression to the androgen-independent state. In contrast, LNCaP cells in the presence of androgens were marginally sensitive to PI3K inhibition. During the chronic stage of androgen deprivation, androgen-independent proliferation correlated with diminished p27(kip1) protein levels, whereas PI3K and Akt activity remained elevated. At this stage, PI3K inhibition rapidly triggered accumulation of p27(kip1), cell cycle arrest, and cell death. PI3K modulated p27(kip1) levels at least in part by regulating its rate of degradation. Taken together, these data show that androgen ablation alone can increase PI3K-Akt activation, which supports survival after acute androgen ablation and proliferation during chronic androgen deprivation. Successful progression to the androgen-independent state in the LNCaP cell line model requires intact PI3K signaling.

PTEN induces chemosensitivity in PTEN-mutated prostate cancer cells by suppression of Bcl-2 expression.

The tumor suppressor gene PTEN (MMAC1/TEP1) is lost frequently in advanced prostate cancer (PCa). However, the function of PTEN in tumorigenesis is not understood fully. In this study, we demonstrate that expression of Bcl-2 in prostate tumors correlates with loss of the PTEN protein. This finding was verified by studies in the PCa cell lines DU145, PC-3, LNCaP, and an androgen-refractory subline of LNCaP. Transient transfection of PTEN into the PTEN-null cells resulted in decreased levels of Bcl-2 mRNA and protein. These effects appear to be mediated at the level of gene transcription, since a Bcl-2 promoter-reporter construct was down-regulated by ectopic expression of PTEN in LNCaP cells. The inhibition of Bcl-2 required the lipid-phosphatase activity of PTEN and was blocked by overexpression of a constitutively active form of Akt. Moreover, the transcription-regulatory protein cAMP-response element-binding protein (CREB) may be involved, since decreased phosphorylation of CREB at Ser(133) was detected following PTEN expression, and ectopic expression of CREB repressed completely the PTEN-induced inhibition of Bcl-2 promoter activity. Furthermore, cotransfection of Bcl-2 and PTEN expression vectors rescued PTEN-induced cell death but not G(1) cell cycle arrest. Finally, forced expression of PTEN sensitized LNCaP cells to cell death induced by staurosporine, doxorubicin, and vincristine, and this chemosensitivity was attenuated by exogenous expression of Bcl-2. Taken together, these data demonstrate that loss of PTEN leads to up-regulation of the bcl-2 gene, thus contributing to survival and chemoresistance of PCa cells. These findings suggest that the PTEN gene and its regulated pathway are potential therapeutic targets in prostate cancer.

Polycyclic aromatic hydrocarbons, aliphatic hydrocarbons, trace elements, and monooxygenase activity in birds nesting on the North Platte River, Casper, Wyoming, USA.

Tree swallow (Tachycineta bicolor) and house wren (Troglodytes aedon) eggs and chicks were collected near a refinery site on the North Platte River, Casper. Wyoming, USA and at a reference site 10 km upstream. Total polycyclic aromatic hydrocarbon (PAH) concentrations in swallow and wren chicks were higher at the refinery site than at the reference site. Polycyclic aromatic hydrocarbon concentrations in sediment and chick dietary samples were consistent with these findings. The general lack of methylated PAHs in sediment, diet, and bird carcasses suggested that the PAHs were derived from combustion and not from petroleum. The predominance of odd-numbered aliphatic hydrocarbons and the low ratios (< or =0.25) of pristane:n-C17 and phytane:n-C18 in chick and diet samples also suggested that swallow and wren chicks were not being chronically exposed to petroleum. Mean ethoxyresorufin-O-dealkylase and benzyloxyresorufin-O-dealkylase activities in tree swallow livers averaged nine times higher at the refinery site than at the reference site and were probably induced by exposure to PAHs. Trace element concentrations in eggs and livers of swallows and wrens were similar or greater at the reference site than at the refinery site. Selenium, strontium, and boron concentrations were elevated in eggs and livers of swallows and wrens at both the refinery and reference sites.

Liquid chromatographic determination of para-toluenesulfonamide in edible fillet tissues from three species of fish.

Chloramine-T (N-sodium-N-chloro-p-toluene-sulfonamide) is a candidate therapeutic drug for treating bacterial gill disease, a predominant disease of a variety of fish species. Research has been initiated to obtain the U.S. Food and Drug Administration's (FDA) approval for the use of chloramine-T on a variety of fish species. An attribute of a therapeutic aquaculture drug that must be characterized before the FDA approves its use is depletion of the drug's marker residue (the drug's parent compound or metabolite of highest concentration in an edible tissue). para-Toluenesulfonamide (p-TSA) is the primary degradation product and marker residue for chloramine-T in rainbow trout. To conduct residue depletion studies for chloramine-T in fish, a robust analytical method sensitive and specific for p-TSA residues in edible fillet tissue from a variety of fish was required. Homogenized fillet tissues from rainbow trout (Oncorhynchus mykiss), walleye (Stizostedion vitreum), and channel catfish (Ictalurus punctatus) were fortified at nominal p-TSA concentrations of 17, 67, 200, 333, and 1000 ng/g. Samples were analyzed by isocratic reversed-phase liquid chromatography (LC) with absorbance detection at 226 nm. Mean recoveries of p-TSA ranged from 77 to 93.17%; relative standard deviations ranged from 1.5 to 14%; method quantitation limits ranged from 13 to 18 ng/g; and method detection limits ranged from 3.8 to 5.2 ng/g. The LC parameters produced p-TSA peaks without coelution of endogenous compounds and excluded chromatographic interference from at least 20 chemicals and drugs of potential use in aquaculture.

Characterization of a TPA-response element in the 5'-flanking region of the androgen receptor gene.

The androgen receptor (AR) protein is an important transacting factor that is necessary for mediating gene expression of androgen-responsive genes. The expression of the AR gene is regulated by androgens and agents that utilize the calcium, protein kinase A, and protein kinase C pathways. Although the role of the calcium and protein kinase A pathways in the regulation of the AR gene has been investigated, the mechanism of regulation of AR through the protein kinase C pathway is not known. We have isolated the 5'-flanking region of the mouse AR gene and identified a consensus TPA (12-O-tetradecanoylphorbol 13-acetate)-response element (TRE). Transient transfection assays indicate that the TRE sequence is sufficient to confer TPA responsiveness to cells treated with TPA. Gel retardation assays and DNA footprint analysis demonstrated specific binding of the TRE and protection of the TRE sequence. Thus, these results describe a TRE in the 5'-flanking region of the AR gene and demonstrate that the TRE is responsive to TPA treatment.

Plasticity of vascular smooth muscle alpha-actin gene transcription. Characterization of multiple, single-, and double-strand specific DNA-binding proteins in myoblasts and fibroblasts.

Transcriptional activity of the mouse vascular smooth muscle (VSM) alpha-actin promoter was governed by both cell type and developmental stage-specific mechanisms. A purine-rich motif (PrM) located as -181 to -176 in the promoter was absolutely required for activation in mouse AKR-2B embryonic fibroblasts and partially contributed to activation in undifferentiated mouse BC3H1 myoblasts. Transcriptional enhancer factor 1 recognized the PrM and cooperated with other promoter-binding proteins to regulate serum growth factor-dependent transcription in both myoblasts and fibroblasts. Two distinct protein factors (VAC-ssBF1 and VAC-ssBF2) also were identified that bound sequence-specifically to single-stranded oligonucleotide probes that spanned both the PrM and a closely positioned negative regulatory element. VAC-ssBF1 and BF2 binding activity was detected in undifferentiated myoblasts, embryonic fibroblasts, and several smooth muscle tissues in the mouse and human. A myoblast-specific protein (VAC-RF1) also was detected that bound double-stranded probes containing a CArG-like sequence that previously was shown to impart strong, cell type specific repression. The binding activity of transcription enhancer factor 1, VAC-RF1, and VAC-ssBF1 was significantly diminished when confluent BC3H1 myoblasts differentiated into myocytes and expressed VSM alpha-actin mRNA after exposure to serum-free medium. The results indicated that cell type-specific control of the VSM alpha-actin gene promoter required the participation of multiple DNA-binding proteins, including two that were enriched in smooth muscle and had preferential affinity for single-stranded DNA.

Activation of a muscle-specific actin gene promoter in serum-stimulated fibroblasts.

Treatment of AKR-2B mouse fibroblasts with serum growth factors or inhibitors of protein synthesis, such as cycloheximide, results in a stimulation of cytoskeletal beta-actin transcription but has no effect on transcription of muscle-specific isotypes, such as the vascular smooth muscle (VSM) alpha-actin gene. Deletion mapping and site-specific mutagenesis studies demonstrated that a single "CArG" element of the general form CC(A/T)6GG was necessary and possibly sufficient to impart serum and cycloheximide-inducibility to the beta-actin promoter. Although the VSM alpha-actin promoter exhibits at least three similar sequence elements, it remained refractory to serum and cycloheximide induction. However, deletion of a 33 base pair sequence between -191 and -224 relative to the transcription start site resulted in the transcriptional activation of this muscle-specific promoter in rapidly growing or serum-stimulated fibroblasts. Although the activity of this truncated promoter was potentiated by cycloheximide in a manner indistinguishable from that of the beta-actin promoter, this was dependent on a more complex array of interacting elements. These included at least one CArG box and a putative upstream activating element closely associated with the -191 to -224 inhibitory sequences. These results demonstrate that the expression of a muscle-specific actin gene in fibroblasts is suppressed by a cis-acting negative control element and that in the absence of this element, the promoter is responsive to growth factor-induced signal transduction pathways.

A mass spectroscopic method for analysis of AHH-inducing and other polychlorinated biphenyl congeners and selected pesticides in fish.

The 209 polychlorinated biphenyl (PCB) congeners exhibit a wide range in toxicity to fish, birds, and mammals. This paper discusses the use of gas chromatography/mass spectrometry negative chemical ionization (GC/MS-NCI) to quantify congeners of highly suspected toxicity such as IUPAC #77 (3,3',4,4'-tetrachlorobiphenyl) and #126 (3,3',4,4',5-pentachlorobiphenyl). GC/MS analysis time needed to produce the necessary resolution was reduced to 1 h per sample or standard, allowing an autosampler to inject 12 samples in 24 hours, plus 12 standards/QC samples. Identification and quantification of some 60+ congeners and several selected pesticides and estimation of total PCBs are also possible within the 1 h analysis. For congeners of high chlorination (penta through octa), the method exhibited excellent sensitivity, such that we could not locate a fish which exhibited PCB levels below our calibrated quantitation range. NCI was not as sensitive for mono through tri and for some tetrachlorinated PCB congeners, an exception being PCB #77, for which sensitivity was of the same order as for the more highly chlorinated biphenyls. Long term stability was excellent. Over a 6-mo period, results of replicate analyses for PCB congeners and pesticides in a composited sample of lake trout (Salvelinus namaycush) from Lake Michigan had a relative standard deviation of 12% of the mean. Over the same time period, mean recoveries for samples spiked at concentrations similar to those in Lake Michigan lake trout were 90-102%. Response was linear over a wide range of concentrations for each of the analyzed compounds. This method is now being used for routine analysis of PCB congeners and selected pesticides in our laboratory.

Development of a tool to assess suicide risk factors in urban adolescents.

Dramatic increases in the adolescent suicide rate over the past three decades have underscored the need for risk-assessment tools. The tools that do exist are oriented to older populations and their application to adolescents is questionable. A project was initiated at the University of Utah's Health Education Department to develop a pilot instrument to examine the differences between adolescents who have attempted suicide and other teenagers. Eighty-two subjects between the ages of 14 and 19 participated in the test of this instrument. Twenty-five subjects were identified by a physician or psychologist as having failed in a sincere suicide attempt within the previous 18 months. Fifty-seven nonsuicide attempters with similar demographic profiles served as a comparison group. An 86-item questionnaire was administered to both groups. Questions were generated from a review of the literature of the past three decades for problems associated with suicide in this population. Questions were sorted into three domains (family environment, social environment, and self-perceptions), with each domain having several subdomains. Statistical analysis revealed significant differences for each of the three domains and on 55 of 86 questions. The results were used to create a streamlined instrument for assessing suicide risk that can be administered in 20 minutes.

Negative regulation of serum-responsive enhancer elements.

Transcription of the c-fos proto-oncogene and the cytoskeletal actin genes is induced within minutes of the addition of serum growth factors in a variety of cell types. Inhibitors of protein synthesis such as cycloheximide have been shown to dramatically potentiate the transcriptional response, an effect termed 'superinduction'. Although the stimulatory effect of serum has been shown to be transmitted through a cis-acting enhancer sequence termed a serum response element (SRE), the sequence element(s) responsible for mediating the effect of cycloheximide has not been identified. We now report that a synthetic copy of the c-fos SRE is sufficient to confer cycloheximide-dependent inducibility upon a heterologous promoter. This does not require the presence of serum, but several mutations in the SRE that impair serum-inducibility also impair cycloheximide-inducubility. These results imply that serum-responsive enhancer elements are negatively regulated by one or more labile proteins and that both positive and negative regulators of enhancer activity require a functional 'CArG box', a sequence domain previously implicated in muscle-specific transcription.

Evidence that the functional beta-actin gene is single copy in most mice and is associated with 5' sequences capable of conferring serum- and cycloheximide-dependent regulation.

Hybridization to synthetic oligonucleotides representing conserved regions in the promoter and first intron of several vertebrate beta-actin genes was used to discriminate between what appears to be a single functional beta-actin gene and numerous pseudogenes in the mouse genome. Sequences derived from the 5' end of this gene were shown to confer serum-inducible expression upon a heterologous reporter gene when transfected into mouse fibroblasts. Moreover, these sequences rendered reporter gene expression superinducible by a combination of serum and cycloheximide. These experiments indicate that the 5' end of the mouse beta-actin gene contains sequence elements which mediate the stimulatory effects of serum growth factors and which are responsive to both positive and negative regulators of gene expression.

Specific stimulation of actin gene transcription by epidermal growth factor and cycloheximide.

Stimulation of quiescent AKR-2B mouse embryo cells with epidermal growth factor (EGF) results in a rapid and specific induction of actin mRNA sequences. These mRNAs include those coding for both beta- and gamma-cytoskeletal, but not alpha-skeletal muscle, actin isotypes. Elongation of nascent RNA chains in isolated nuclei (run-off transcription) demonstrates that the mRNA accumulation is preceded by an increase in actin gene transcription. This increase is transient, however, and is followed by a rapid attenuation of transcriptional activity. An inhibitor of protein synthesis, cycloheximide, was also found to induce beta- and gamma-actin mRNA accumulation. Furthermore, the simultaneous addition of EGF and cycloheximide produced a synergistic effect on actin sequences in both steady-state nuclear and polysomal RNA. Run-off transcription experiments demonstrate that this synergistic effect results from an increase in the magnitude and duration of actin gene transcription. It is also specific in that alpha-tubulin gene transcription is not similarly affected. These data suggest the existence of a specific labile repressor of actin gene transcription.

Polyadenylylated RNA complementary to a mouse retrovirus-like multigene family is rapidly and specifically induced by epidermal growth factor stimulation of quiescent cells.

Complementary DNA probes prepared from total polysomal poly(A)+RNA populations were used to identify clones of mouse DNA containing sequences whose expression is specifically enhanced after epidermal growth factor (EGF) stimulation of quiescent mouse embryo cells in culture. Three such clones were isolated and used to study changes in the levels of clone-specific poly(A)+RNA in the polysomes of cells after mitogenic stimulation by EGF. RNA complementary to sequences present in these clones increased approximately equal to 10-fold as a fraction of the total poly(A)+RNA by 6 hr after stimulation. All three clones were found by hybridization criteria to contain sequences related to the class of mouse retrovirus or transposon-like elements termed VL30. These VL30-related sequences were further found to be complementary to EGF-inducible poly(A)+RNAs and enhanced expression was detectable as early as 1 hr after EGF stimulation. In contrast, nine additional clones, including an AKR-type murine leukemia provirus DNA clone, contained no detectable VL30 sequence elements and were complementary to poly(A)+RNA species whose relative concentration was essentially constant in quiescent and EGF-stimulated cells. Therefore, VL30 sequence elements appear distinct in that they encompass members whose expression is specifically regulated in response to a defined peptide growth factor.