Frentizole, a Nontoxic Immunosuppressive Drug, and Its Analogs Display Antitumor Activity via Tubulin Inhibition.

Antimitotic agents are one of the more successful types of anticancer drugs, but they suffer from toxicity and resistance. The application of approved drugs to new indications (i.e., drug repurposing) is a promising strategy for the development of new drugs. It relies on finding pattern similarities: drug effects to other drugs or conditions, similar toxicities, or structural similarity. Here, we recursively searched a database of approved drugs for structural similarity to several antimitotic agents binding to a specific site of tubulin, with the expectation of finding structures that could fit in it. These searches repeatedly retrieved frentizole, an approved nontoxic anti-inflammatory drug, thus indicating that it might behave as an antimitotic drug devoid of the undesired toxic effects. We also show that the usual repurposing approach to searching for targets of frentizole failed in most cases to find such a relationship. We synthesized frentizole and a series of analogs to assay them as antimitotic agents and found antiproliferative activity against HeLa tumor cells, inhibition of microtubule formation within cells, and arrest at the G2/M phases of the cell cycle, phenotypes that agree with binding to tubulin as the mechanism of action. The docking studies suggest binding at the colchicine site in different modes. These results support the repurposing of frentizole for cancer treatment, especially for glioblastoma.

Influence of Splint Support on the Precision of Static Totally Guided Dental Implant Surgery: A Systematic Review and Network Meta-analysis.

Purpose: To assess the accuracy of totally guided implant placement with static surgical splints in relation to the different types of supporting tissues (tooth, mucosa, or bone). Materials and Methods: This review was carried out following the PRISMA guidelines. An electronic search was done of the MEDLINE (PubMed), Embase, and Cochrane Library databases, without publication year or language restrictions. Results: The literature search yielded a total of 877 articles; 18 were included in the qualitative synthesis, and 16 of these articles were included in the quantitative analysis. The included studies presented a high risk of bias, except for one randomized clinical trial. The strength of the recommendations is therefore weak. In the angular deviation treatment, statistically significant differences were observed in the accuracy of the implants with tooth vs bone support: Bone support yielded 1.31 degrees greater deviation vs tooth support (SD = 0.43; 95% CI: 0.47, 2.15, P = .002). No significant differences were observed in the linear deviations. Conclusion: Tooth support proved to be significantly more precise than bone support splints. There were no differences referring to horizontal coronal deviation, horizontal apical deviation, or vertical deviation according to the type of splint support used.

Accuracy of a guided implant system with milled surgical templates.

PURPOSE: This in vitro study analyzed the accuracy of a computer-assisted design (CAD)/computer-assisted manufacturing (CAM) guided implant surgery system by comparing linear, angular, and coronal deviations between the planned and final implant placement. METHODS: By using a fully guided surgery workflow, 32 dental implants were placed in 16 partially edentulous models. After virtual design of the restorations, radiological and CAD files were matched and implant positions were planned by using dedicated implant planning software (Galileo Implant version 1.9.2.). Templates were designed (Cerec Omnicam) and milled (Cerec MC XL) by using chairside workflow. Galileo Implant version 1.9.2. was used to evaluate accuracy. RESULTS: Mean horizontal and angular-coronal total deviation values were 0.2 mm (SD = 0.126) and 1.1º (SD = 0.834) respectively. Multivariate analysis of variance showed significant differences in horizontal and angular-coronal total deviation in the 32 implants (P = 0.0001). Multivariate analysis with one-factor interaction showed no statistical difference in implant position or implant type (P = 0.139) between eight maxilla models and eight jaw models. CONCLUSION: Horizontal and angular-coronal deviations of implants placed with chairside digital workflow were within the recommended safety margin for fully guided surgery.

Ion mobility spectrometry and mass spectrometry coupled to gas chromatography for analysis of microbial contaminated cosmetic creams.

The most commonly used technique for monitoring microbial contamination in cosmetic products is plate counting. In this contribution, headspace - gas chromatography (HS-GC) coupled to mass spectrometry (MS) or ion mobility spectrometry (IMS) is proposed as a technique to evaluate rapidly and accurately the state of microbial colonies in cosmetic creams using the volatile organic compounds produced by microorganisms (MVOC). The work focuses on monitoring two of the microorganisms that most frequently occur in such creams, Candida albicans and Staphylococcus aureus. In addition, two different types of ingredient with antimicrobial properties (a chemical preservative and a natural preservative) were added to study the behaviour of these microorganisms under different conditions. The facial creams were elaborated and inoculated with the two above microorganisms, and then sampled weekly for 4 weeks, analysing the evolution of the MVOCs by HS-GC-MS and HS-GC-IMS. In addition, microbial contamination was determined by the classical plate counting method. The pH, colour, viscosity and water activity parameters were also measured. The use of chemometric tools is essential because of the large amount of data generated, and different models based on discriminant analysis with an orthogonal projection on latent structures (OPLS-DA) were constructed. The optimal models obtained by both analytical techniques allowed differentiation between contaminated and non-contaminated creams, with a validation success rate of 94.4%. In addition, MVOC monitoring also allowed assessment of the microbial concentration.

Combined use of thermo-ultrasound and cinnamon leaf essential oil to inactivate Saccharomyces cerevisiae in culture broth and natural orange juice.

The survival of Sacharomyces cerevisiae in Trypticase Soy Broth and natural orange juice processed by combined use of thermo-ultrasound and cinnamon leaf essential oil has been evaluated and modelled. Minimal inhibitory concentration of cinnamon leaf essential oil against S. cerevisiae was determined using absorbance measurements based on the microtiter plate assay. The resistance of S. cerevisiae cells to the combined action of thermal treatment with ultrasound was analyzed in Trypticase Soy Broth with different concentrations of cinnamon leaf essential oil at 30, 40 and 50 °C. The best conditions of inactivation in TSB to study the inactivation of S. cerevisiae in natural orange juice. Experimental data were fitted by using the "shoulder + log-linear" and "Weibull" models (GInaFiT). The combined use of thermo-ultrasound and cinnamon leaf essential oil enhanced the inactivation of S. cerevisiae in TSB and natural orange juice.

Cost-effectiveness analysis of azacitidine in the treatment of high-risk myelodysplastic syndromes in Spain.

BACKGROUND: The objective of the study was to analyse whether azacitidine is a cost-effective option for the treatment of myelodysplastic syndrome in the Spanish setting compared with conventional care regimens, including best supportive care, low dose chemotherapy and standard dose chemotherapy. METHODS: A life-time Markov model was constructed to evaluate the cost-effectiveness of azacitidine compared with conventional care regimens. The health states modelled were: myelodysplastic syndrome, acute myeloid leukemia and death. Variables measured included survival rates, progression probabilities and quality of life indicators. Resource use and cost data reflect the Spanish context. The analysis was performed from the Spanish National Health System perspective, discounting both costs (in 2012 euros) and future effects at 3%. The time horizon considered was end-of-life. Results were expressed in cost per quality-adjusted life-year gained and cost per life-year gained and compared with cost-effectiveness thresholds. RESULTS: According to the current use of each conventional care regimens options in Spain, azacitidine resulted in €34,673 per quality-adjusted life-year gained (€28,891 per life-year gained) with an increase of 1.89 in quality-adjusted life-years (2.26 in life-years). Azacitidine was superior to best supportive care and low dose chemotherapy in terms of quality-adjusted life-years gained, 1.82 and 2.03, respectively (life-years 2.16 vs. best supportive care, 2.39 vs. low dose chemotherapy). Treatment with azacitidine resulted in longer survival time and thus longer treatment time and lifetime costs. The incremental cost-effectiveness ratio was €39,610 per quality-adjusted life-year gained vs. best supportive care and €30,531 per quality-adjusted life-year gained vs. low dose chemotherapy (€33,111 per life-year gained vs. best supportive care and €25,953 per life-year gained vs. low dose chemotherapy). CONCLUSIONS: The analysis showed that the use of azacitidine in the treatment of high-risk myelodysplastic syndrome is a cost-effective option compared with conventional care regimen options used in the Spanish setting and had an incremental cost-effectiveness ratio within the range of the thresholds accepted by health authorities.

[Association of FTO gene polymorphisms and morbid obesity in the population of Extremadura (Spain)]. / Asociación de polimorfismos en el gen FTO con la obesidad mórbida en la población extremeña.

OBJECTIVE: To select individuals whose morbid obesity can be attributed mainly to their individual genetic profile. After excluding patients with potential monogenic syndromes or diseases associated with obesity, we evaluated the association of the single nucleotide polymorphisms (SNPs) rs1861868 and rs9939609 of the fat-mass and obesity-associated FTO gene with an inherited predisposition to morbid obesity. PATIENTS AND METHODS: We evaluated 270 patients with morbid obesity and onset before the age of 14 years and selected 194 due to their phenotypes and family history; 289 control individuals were included. The rs1861868 and rs9939609 variants, located in the FTO gene, were genotyped. Genotype and haplotype frequencies were compared between cases and controls. RESULTS: The A allele of rs9939609 was associated with severe obesity starting in childhood among the Spanish population. The rs1861868 G/rs9939609 A haplotype of the FTO gene was also significantly associated with severe obesity in our population, with an odds ratio of 3.03 (95% confidence interval, 1.74-5.27). CONCLUSION: Analysis of the genetic basis of obesity requires rigorous selection of cases. In this study, the association of the rs9939609 SNP with obesity widely described in distinct populations was confirmed among overweight Spanish children. Genotyping rs1861868 allowed us to identify the first risk haplotype in the FTO gene, which is located in the adjacent haplotype block containing rs9939609. In-depth study of the variability of the FTO gene is essential to define its deleterious capacity.

Cardioprotective stimuli mediate phosphoinositide 3-kinase and phosphoinositide dependent kinase 1 nuclear accumulation in cardiomyocytes.

The phosphoinositide 3-kinase (PI3K)/phosphoinositide dependent kinase 1 (PDK1) signaling pathway exerts cardioprotective effects in the myocardium through activation of key proteins including Akt. Activated Akt accumulates in nuclei of cardiomyocytes suggesting that biologically relevant targets are located in that subcellular compartment. Nuclear Akt activity could be potentiated in both intensity and duration by the presence of a nuclear-associated PI3K/PDK1 signaling cascade as has been described in other non-myocyte cell types. PI3K/PDK1 distribution was determined in vitro and in vivo by immunostaining and nuclear extraction of cultured rat neonatal cardiomyocytes or transgenic mouse hearts. Results show that PI3K and PDK1 are present at a basal level in cardiomyocytes nuclei and that cardioprotective stimulation with atrial natriuretic peptide (ANP) increases their nuclear localization. In comparison, overexpression of nuclear-targeted Akt does not mediate increased translocation of either PI3K or PDK1 indicating that accumulation of Akt does not drive PI3K or PDK1 into the nuclear compartment. Furthermore, PI3K and phospho-Akt(473) show parallel temporal accumulation in the nucleus following (MI) infarction challenge. These findings demonstrate the presence of a dynamically regulated nuclear-associated signaling cascade involving PI3K and PDK that presumably influences nuclear Akt activation.

Nuclear and mitochondrial signalling Akts in cardiomyocytes.

Biological actions resulting from phosphoinositide synthesis trigger multiple downstream signalling cascades by recruiting proteins with pleckstrin homology domains, including phosphoinositide-dependent kinase-1 and protein kinase B (also known as Akt). Retrospectively, more attention has been focused on the plasma membrane-associated interactions of these molecules and resulting cytoplasmic target activation. The complex biological activities exerted by Akt activation suggest, however, that more subtle and complex subcellular control mechanisms are involved. This review examines the regulation of Akt activity from the perspective of subcellular compartmentalization and focuses specifically upon the actions of Akt activation downstream from phosphoinositide synthesis that influence cell biology by altering nuclear signalling leading to Pim-1 kinase induction as well as hexokinase phosphorylation that, together with Akt, serves to preserve mitochondrial integrity.

Myocardial induction of nucleostemin in response to postnatal growth and pathological challenge.

Stem cell-specific proteins and regulatory pathways that determine self-renewal and differentiation have become of fundamental importance in understanding regenerative and reparative processes in the myocardium. One such regulatory protein, named nucleostemin, has been studied in the context of stem cells and several cancer cell lines, where expression is associated with proliferation and maintenance of a primitive cellular phenotype. We find nucleostemin is present in young myocardium and is also induced following cardiomyopathic injury. Nucleostemin expression in cardiomyocytes is induced by fibroblast growth factor-2 and accumulates in response to Pim-1 kinase activity. Cardiac stem cells also express nucleostemin that is diminished in response to commitment to a differentiated phenotype. Overexpression of nucleostemin in cultured cardiac stem cells increases proliferation while preserving telomere length, providing a mechanistic basis for potential actions of nucleostemin in promotion of cell survival and proliferation as seen in other cell types.

Activation of Notch-mediated protective signaling in the myocardium.

The Notch network regulates multiple cellular processes, including cell fate determination, development, differentiation, proliferation, apoptosis, and regeneration. These processes are regulated via Notch-mediated activity that involves hepatocyte growth factor (HGF)/c-Met receptor and phosphatidylinositol 3-kinase/Akt signaling cascades. The impact of HGF on Notch signaling was assessed following myocardial infarction as well as in cultured cardiomyocytes. Notch1 is activated in border zone cardiomyocytes coincident with nuclear c-Met following infarction. Intramyocardial injection of HGF enhances Notch1 and Akt activation in adult mouse myocardium. Corroborating evidence in cultured cardiomyocytes shows treatment with HGF or insulin increases levels of Notch effector Hes1 in immunoblots, whereas overexpression of activated Notch intracellular domain prompts a 3-fold increase in phosphorylated Akt. Infarcted hearts injected with adenoviral vector expressing Notch intracellular domain treatment exhibit improved hemodynamic function in comparison with control mice after 4 weeks, implicating Notch signaling in a cardioprotective role following cardiac injury. These results indicate Notch activation in cardiomyocytes is mediated through c-Met and Akt survival signaling pathways, and Notch1 signaling in turn enhances Akt activity. This mutually supportive crosstalk suggests a positive survival feedback mechanism between Notch and Akt signaling in adult myocardium following injury.

Evolution of the c-kit-positive cell response to pathological challenge in the myocardium.

Cumulative evidence indicates that myocardium responds to growth or injury by recruitment of stem and/or progenitor cells that participate in repair and regenerative processes. Unequivocal identification of this population has been hampered by lack of reagents or markers specific to the recruited population, leading to controversies regarding the nature of these cells. Use of a transgenic mouse expressing green fluorescent protein driven by the c-kit promoter allows for unambiguous identification of this cell population. Green fluorescent protein (GFP) driven by the c-kit promoter labels a fraction of the c-kit+ cells recognized by antibody labeling for c-kit protein. Expression of GFP by the c-kit promoter and accumulation of GFP-positive cells in the myocardium is relatively high at birth compared with adult and declines between postnatal weeks 1 and 2, which tracks in parallel with expression of c-kit protein and c-kit-positive cells. Acute cardiomyopathic injury by infarction prompts increased expression of both GFP protein and GFP-labeled cells in the region of infarction relative to remote myocardium. Similar increases were observed for c-kit protein and cells with a slightly earlier onset and decline relative to the GFP signal. Cells coexpressing GFP, c-kit, and cardiogenic markers were apparent at 1-2 weeks postinfarction. Cardiac-resident c-kit+ cell cultures derived from the transgenic line express GFP that is diminished in parallel with c-kit by induction of differentiation. The use of genetically engineered mice validates and extends the concept of c-kit+ cells participating in the response to myocardial injury.

Akt promotes increased cardiomyocyte cycling and expansion of the cardiac progenitor cell population.

Activation of Akt is associated with enhanced cell cycling and cellular proliferation in nonmyocytes, but this effect of nuclear Akt accumulation has not been explored in the context of the myocardium. Cardiac-specific expression of nuclear-targeted Akt (Akt/nuc) in transgenics prolongs postnatal cell cycling as evidenced by increased numbers of Ki67+ cardiomyocytes at 2 to 3 weeks after birth. Similarly, nuclear-targeting of Akt promotes expansion of the presumptive cardiac progenitor cell population as assessed by immunolabeling for c-kit in combination with myocyte-specific markers Nkx 2.5 or MEF 2C. Increases in pro-proliferative cytokines, including tumor-necrosis superfamily 8, interleukin-17e, and hepatocyte growth factor, were found in nuclear-targeted Akt myocardial samples. Concurrent signaling mediated by paracrine factors downstream of Akt/nuc expression may be responsible for phenotypic effects of nuclear-targeted Akt in the myocardium, including enhanced cell proliferation and expansion of the stem cell population.

Activation of the unfolded protein response in infarcted mouse heart and hypoxic cultured cardiac myocytes.

Endoplasmic reticulum (ER) stresses that reduce ER protein folding activate the unfolded protein response (UPR). One effector of the UPR is the transcription factor X-box binding protein-1 (XBP1), which is expressed on ER stress-mediated splicing of the XBP1 mRNA. XBP1 induces certain ER-targeted proteins, eg, glucose-regulated protein 78 (GRP78), that help resolve the ER stress and foster cell survival. In this study, we determined whether hypoxia can activate the UPR in the cardiac context. Neonatal rat ventricular myocyte cultures subjected to hypoxia (16 hours) exhibited increased XBP1 mRNA splicing, XBP1 protein expression, GRP78 promoter activation, and GRP78 protein levels; however, the levels of these UPR markers declined during reoxygenation, suggesting that the UPR is activated during hypoxia but not during reoxygenation. When cells were infected with a recombinant adenovirus (AdV) encoding dominant-negative XBP1 (AdV-XBP1dn), UPR markers were reduced; however, hypoxia/reoxygenation-induced apoptosis increased. Confocal immunocytofluorescence demonstrated that hypoxia induced GRP78 in neonatal rat and isolated adult mouse ventricular myocytes. Moreover, mouse hearts subjected to in vivo myocardial infarction exhibited increased GRP78 expression in cardiac myocytes near the infarct, but not in healthy cells distal to the infarct. These results indicate that hypoxia activates the UPR in cardiac myocytes and that XBP1-inducible proteins may contribute to protecting the myocardium during hypoxic stress.

Evolución de la atención en una sala de cuidados intensivos neonatales / Evolution of care at a neonatal intensive care unit

Se estudiaron los resultados de la atención en una sala de Cuidados Intensivos Neonatales de un Hospital Pediátrico de Alta Complejidad (Hospital de Niños ¨ Superiora Sor María Ludovica ¨, La Plata) durante un periodo de 10 años, según patología asistida, Región Sanitaria de referencia, tiempo de internación y mortalidad. Para el análisis de mortalidad neonatal (MNN) se distribuyo la población en cinco categorías diagnosticas (1-Patología Quirúrgica. 2-Cardiopatías Congénitas. 3-Patología Respiratoria, asociada o no a prematurez. 4-Patología Infecciosa. 5-Patología Clínica) y por peso de ingreso ≥ 2.501 g y ≤ 2.500 g, fraccionando este ultimo grupo en intervalos de peso de 500 g. La MNN global se redujo en un 52,9 % comparando la mortalidad de 2004 con la registrada en 1995. La MNN disminuyo significativamente en cada categoría diagnostica y para cada intervalo de peso. En relación a la referencia, la Región Sanitaria XI determino el 50 % de los ingresos, siguiéndole en frecuencia la Región Sanitaria V y VI, aunque ambas con tendencia decreciente (32 % en 1995, 20 % en 2004), fundamentalmente de recién nacidos con patología no quirúrgica (68,5 % en 1995, 16,2 % en 2004). Esto condiciono un cambio en las categorías diagnosticas, no así en la complejidad de la asistencia, como lo evidencia el tiempo de internación y el número de recién nacidos que requirieron asistencia respiratoria mecánica, cifras que se mantuvieron en valores constantes.

Sarcoplasmic reticulum adenosine triphosphatase overexpression in the L-type Ca2+ channel mouse results in cardiomyopathy and Ca2+ -induced arrhythmogenesis.

BACKGROUND: Overexpression of the L-type voltage-dependent calcium channel alpha(1C)-subunit (L-VDCC OE) in transgenic mice results in adaptive hypertrophy followed by a maladaptive phase associated with a decrease in sarcoplasmic reticulum adenosine triphosphatase (SERCA)2a expression at 8 to 10 months of age. Overexpressing SERCA to manipulate calcium (Ca(2+)) cycling and prevent pathologic phenotypes in some models of heart failure has been proven to be a promising genetic strategy. OBJECTIVE: In this study we investigated whether genetic manipulation that increases Ca(2+) uptake into the sarcoplasmic reticulum by overexpressing SERCA1a (skeletal muscle specific) into the L-VDCC OE background could restore or further deteriorate Ca(2+) cycling, contractile dysfunction, and electrical remodeling in the heart failure phenotype. RESULTS: We found that the survival rate of L-VDCC OE/SERCA1a OE double transgenic mice decreased by 50%. L-VDCC OE/SERCA1a OE mice displayed an accelerated phenotype of severe dilation of both ventricles associated with deteriorated left ventricular function. Voltage clamp experiments revealed enhanced increased inward Ca(2+) current density and decreased the transient outward potassium current. Action potential duration in double transgenic ventricular myocytes was prolonged, and isoproterenol induced early after depolarization. These mice demonstrated a high incidence of spontaneous left ventricular arrhythmia. Expression of the proarrhythmic signaling protein Ca(2+)/calmodulin-dependent kinase II (CaMKII) was increased while connexin43 expression was decreased, defining an important putative mechanism in the electrophysiologic disturbances and mortality. CONCLUSIONS: Despite previous reports of improved cardiac function in heart failure models after SERCA intervention, our results advocate the need to elucidate the involvement of augmented Ca(2+) cycling in arrhythmogenesis.

Single-channel gating and regulation of human L-type calcium channels in cardiomyocytes of transgenic mice.

Overexpression of human cardiac L-type Ca(2+) channel pores (hCa(v)1.2) in mice causes heart failure. Earlier studies showed Ca(v)1.2-mRNA increase by 2.8-fold, but whole-cell current density enhancement by

Ca(2+) signaling in cardiac myocytes overexpressing the alpha(1) subunit of L-type Ca(2+) channel.

Voltage-gated L-type Ca(2+) channels (LCCs) provide Ca(2+) ingress into cardiac myocytes and play a key role in intracellular Ca(2+) homeostasis and excitation-contraction coupling. We investigated the effects of a constitutive increase of LCC density on Ca(2+) signaling in ventricular myocytes from 4-month-old transgenic (Tg) mice overexpressing the alpha(1) subunit of LCC in the heart. At this age, cells were somewhat hypertrophic as reflected by a 20% increase in cell capacitance relative to those from nontransgenic (Ntg) littermates. Whole cell I(Ca) density in Tg myocytes was elevated by 48% at 0 mV compared with the Ntg group. Single-channel analysis detected an increase in LCC density with similar conductance and gating properties. Although the overexpressed LCCs triggered an augmented SR Ca(2+) release, the "gain" function of EC coupling was uncompromised, and SR Ca(2+) content, diastolic cytosolic Ca(2+), and unitary properties of Ca(2+) sparks were unchanged. Importantly, the enhanced I(Ca) entry and SR Ca(2+) release were associated with an upregulation of the Na(+)-Ca(2+) exchange activity (indexed by the half decay time of caffeine-elicited Ca(2+) transient) by 27% and SR Ca(2+) recycling by approximately 35%. Western analysis detected a 53% increase in the Na(+)-Ca(2+) exchanger expression but no change in the abundance of ryanodine receptor (RyR), SERCA2, and phospholamban. Analysis of I(Ca) kinetics suggested that SR Ca(2+) release-dependent inactivation of LCCs remains intact in Tg cells. Thus, in spite of the modest cardiac hypertrophy, the overexpressed LCCs form functional coupling with RyRs, preserving both orthograde and retrograde Ca(2+) signaling between LCCs and RyRs. These results also suggest that a modest but sustained increase in Ca(2+) influx triggers a coordinated remodeling of Ca(2+) handling to maintain Ca(2+) homeostasis.