Internal translation of the connexin 43 transcript

Connexin 43 (Cx43), the most widely expressed gap junction protein, is associated with a number of physiological and pathological conditions. Many functions of Cx43 have been shown to be independent of gap junction formation and only require the expression of Cx43 C-terminal fragments. Recent evidence demonstrated that naturally occurring C-terminal isoforms can be generated via internal translation. Here, we confirm that C-terminal domains of Cx43, particularly the major 20-kDa isoform, can be independently generated and regulated by internal translation of the same single GJA1 gene transcript that encodes full-length Cx43. Through direct RNA transfection experiments, we provide evidence that internal translation is not due to a bona fide cap-independent IRES-mediated mechanism, as upstream ribosomal scanning or translation is required. In addition to the mTOR pathway, we show for the first time, using both inhibitors and cells from knockout mice, that the Mnk1/2 pathway regulates the translation of the main 20-kDa isoform. Internal translation of the Cx43 transcript occurs but is not cap-independent and requires translation upstream of the internal start codon. In addition to the PI3K/AKT/mTOR pathway, the major 20-kDa isoform is regulated by the Mnk1/2 pathway. Our results have major implications for past and future studies escribing gap junction-independent functions of Cx43 in cancer and other pathological conditions. This study provides further clues to the signalling pathways that regulate internal mRNA translation, an emerging mechanism that allows for increased protein diversity and functional complexity from a single mRNA transcript

Consumo de jamón curado e incidencia de eventos cardiovasculares, hipertensión arterial o ganancia de peso

Fundamento y objetivo: El jamón curado es uno de los alimentos característicos de la dieta mediterránea española. Sin embargo, no existe ningún estudio epidemiológico prospectivo que haya valorado sus efectos sobre la salud humana. Nuestro objetivo fue evaluar la asociación entre el consumo de jamón curado y la incidencia de hipertensión arterial, enfermedad cardiovascular y/o ganancia de peso. Sujetos y método: En una cohorte epidemiológica prospectiva y dinámica de 13.293 graduados universitarios (Proyecto SUN) se analizó la incidencia de enfermedad cardiovascular, hipertensión arterial y la ganancia de peso a lo largo de un seguimiento máximo de 6 años. Se ajustaron modelos de regresión de Cox para estimar hazard ratios (riesgos relativos [RR]) tras ajustar por posibles factores de confusión. Resultados: El consumo de jamón en los niveles más altos (> 4 raciones/semana) no presentaba asociación con la incidencia de eventos cardiovasculares (RR=1,02; [IC 95%: 0,44-2,39]), tras ajustar por edad, sexo, ingesta energética total y patrón dietético, en comparación con los consumos inferiores a una vez por semana. Al repetir esta comparación para la hipertensión, y tras ajustar por edad, sexo, ingesta energética total e índice de masa corporal, se encontró un RR = 0.74 (IC 95%: 0.55-1.01). En la comparación del cambio de peso medio anual entre estas mismas categorías extremas de consumo (=4 raciones) se encontró una diferencia no significativa de 0,033 kg (IC 95%: -0,041 a +0,107) en la ganancia media de peso al año tras ajustar por sexo, edad, tabaco, actividad física e índice de masa corporal inicial. Conclusiones: Los resultados de esta cohorte no proporcionan evidencia de que el consumo de jamón curado se asocie a mayor riesgo cardiovascular, de hipertensión arterial o de ganancia de peso.Background and objective: Cured ham is a characteristic food in Spanish Mediterranean diet. However, no prospective epidemiologic study assessing its effects on human health is available. Our aim was to assess the association between the consumption of cured ham and cardiovascular disease, hypertension or weight gain. Subjects and Method: In a prospective and dynamic epidemiologic cohort composed exclusively of university graduates (the SUN Project, n=13,293), we analyzed the incidence of cardiovascular disease, hypertension or average yearly weight gain after a maximum follow-up of 6 years. Cox (proportional hazards) regression models were fitted to estimate hazard ratios (relative risks [RR]) after adjusting for potential confounding. Results: No association was found between higher levels of consumption of cured ham (> 4 servings/week) and the incidence of cardiovascular disease (RR=1.02; [95%CI]: 0.44-2.39), in analyses adjusted for age, sex, total energy intake and dietary pattern, compared to the consumption of less than one serving a week. When we repeated this comparison for the incidence of hypertension, and adjusting for age, sex, total energy intake and body mass index, the RR was 0.74 (95% CI: 0.55-1.01). In the comparison of average yearly weight gain between these extreme categories of cured ham consumption (=4 servings/week) a non-significant difference of 0.033 kg (95% CI: -0.041 to 0.107) was found after adjusting for sex, age, smoking, physical activity, and baseline body mass index. Conclusions: The results of this cohort study do not support any association between the consumption of cured ham and a higher risk of cardiovascular disease, hypertension or weight gain

Heteromerization between α2A adrenoceptors and different polymorphic variants of the dopamine D4 receptor determines pharmacological and functional differences. Implications for impulsive-control disorders.

Polymorphic alleles of the human dopamine D4 receptor gene (DRD4) have been consistently associated with individual differences in personality traits and neuropsychiatric disorders, particularly between the gene encoding dopamine D4.7 receptor variant and attention deficit hyperactivity disorder (ADHD). The α2A adrenoceptor gene has also been associated with ADHD. In fact, drugs targeting the α2A adrenoceptor (α2AR), such as guanfacine, are commonly used in ADHD treatment. In view of the involvement of dopamine D4 receptor (D4R) and α2AR in ADHD and impulsivity, their concurrent localization in cortical pyramidal neurons and the demonstrated ability of D4R to form functional heteromers with other G protein-coupled receptors, in this study we evaluate whether the α2AR forms functional heteromers with D4R and weather these heteromers show different properties depending on the D4R variant involved. Using cortical brain slices from hD4.7R knock-in and wild-type mice, here, we demonstrate that α2AR and D4R heteromerize and constitute a significant functional population of cortical α2AR and D4R. Moreover, in cortical slices from wild-type mice and in cells transfected with α2AR and D4.4R, we detect a negative crosstalk within the heteromer. This negative crosstalk is lost in cortex from hD4.7R knock-in mice and in cells expressing the D4.7R polymorphic variant. We also show a lack of efficacy of D4R ligands to promote G protein activation and signaling only within the α2AR-D4.7R heteromer. Taken together, our results suggest that α2AR-D4R heteromers play a pivotal role in catecholaminergic signaling in the brain cortex and are likely targets for ADHD pharmacotherapy

Repeated administration of N-ethyl-pentedrone induces increased aggression and impairs social exploration after withdrawal in mice

N-ethyl-pentedrone (NEPD, 2-(ethylamino)-1-phenyl-1-pentanone) is one of the latest synthetic cathinone derivatives that emerged into the illicit drug market. This drug has psychostimulant properties and has been related with several intoxications and even fatalities. However, information about the consequences of its acute and repeated consumption is lacking. Thus, the aim of our study was to investigate the behavioral effects after both acute and repeated NEPD exposure as well as the neurochemical changes. Male OF1 mice were treated with an acute dose (1, 3 or 10 mg/kg, i.p.) or received repeated injections of these doses (twice/day, 5 days) of NEPD. Shortly after drug-exposure or during drug-withdrawal, anxiety-like behavior, aggressiveness, social interaction, depressive-like symptoms, body weight and temperature were assessed. Also, monoamine synthesis enzymes, levels of neurotransmitters and their precursors and main metabolites, as well as ΔFosB, were determined in striatum and prefrontal cortex from post-mortem tissue. Acute administration of NEPD induced anxiolytic effects and reduced social exploration whereas during withdrawal after repeated administration the anxiolytic effect had vanished, and the reduced social exploration was still present and accompanied with increased aggressive behavior. Moreover, NEPD (10 mg/kg) induced slight hyperthermia and reduced weight gain during the repeated administration, whereas increased locomotor activity and lack of depressive symptoms were found during withdrawal. This was accompanied by increased plasma corticosterone and decrease in striatal dopamine. Finally, the long-lasting and robust increase in ΔFosB levels found in striatum after NEPD chronic exposure suggests a high risk of dependence. The increased aggressivity and locomotor activity, together with this potential of inducing dependence justify a warning about the risks of consumption of NEPD if translated to humans. Keywords: Addiction; Aggressive behavior; Monoamine levels; N-ethyl-pentedrone; Synthetic cathinones

The commissioning of a hybrid multi-material 3D printer

Additive Manufacturing (AM) has rapidly become an important technology in both research and industry. This development has allowed the evolution of 3D printers which are able to print complex geometries at low costs and faster than traditional methods. Despite this, most of these printers are either only for using one material or one technology. This limits a lot its use in different sectors such as aeronautics, automotive or health, because multi-material prototypes are needed. For example, surgeons need surgical planning prototypes for preoperative planning. These 3D printed prototypes have mainly been manufactured using just one technology. As a result, the prototypes have some main limitations: (1) do not actually mimic the anatomical structures of the human body, (2) high costs specially for Material Jetting and Powder Bed Fusion AM technologies. Therefore, the aim of present manuscript is the design, development, and commissioning of a hybrid multi-material 3D printer.Peer ReviewedObjectius de Desenvolupament Sostenible::9 - Indústria, Innovació i InfraestructuraObjectius de Desenvolupament Sostenible::3 - Salut i BenestarPostprint (published version

Cannabinoid receptor CB2 drives HER2 pro-oncogenic signaling in breast cancer

Pharmacological activation of cannabinoid receptors elicits antitumoral responses in different models of cancer. However, the biological role of these receptors in tumor physio-pathology is still unknown. We analyzed CB2 cannabinoid receptor protein expression in two series of 166 and 483 breast tumor samples operated in the University Hospitals of Kiel, Tübingen and Freiburg between 1997 and 2010. CB2 mRNA expression was also analyzed in previously published DNA microarray datasets. The role of CB2 in oncogenesis was studied by generating a mouse line that expresses the HER2 rat ortholog (neu) and lacks CB2, and by a variety of biochemical and cell biology approaches in human breast cancer cells in culture and in vivo, upon modulation of CB2 expression by si/shRNAs and overexpression plasmids. CB2-HER2 molecular interaction was studied by co-localization, coimmunoprecipitation and proximity ligation assays. We show an association between elevated CB2 expression in HER2+ breast tumors and poor patient prognosis. We also demonstrate that genetic inactivation of CB2 impairs tumor generation and progression in MMTV-neu mice. Moreover, we show that HER2 upregulates CB2 expression by activating the transcription factor ELK1 via the ERK cascade, and that an increased CB2 expression activates the HER2 prooncogenic signaling machinery at the level of the tyrosine kinase c-SRC. Finally, HER2 and CB2 form heteromers in cancer cells. Our findings reveal an unprecedented role of CB2 as a pivotal regulator of HER2 pro-oncogenic signaling in breast cancer, and suggest that CB2 may be a biomarker with prognostic value in these tumors

Functional Selectivity of Allosteric Interactions within GPCR oligomers: the Dopamine D1-D3 Receptor Heterotetramer

The dopamine D1 receptor-D3 receptor (D1R-D3R) heteromer is being considered as a potential therapeutic target for neuropsychiatric disorders. Previous studies suggested that this heteromer could be involved in the ability of D3R agonists to potentiate locomotor activation induced by D1R agonists. It has also been postulated that its overexpression plays a role in L-dopa-induced dyskinesia and in drug addiction. However, little is known about its biochemical properties. By combining bioluminescence resonance energy transfer, bimolecular complementation techniques and cell signaling experiments in transfected cells, evidence was obtained for a tetrameric stoichiometry of the D1R-D3R heteromer, constituted by two interacting D1R and D3R homodimers coupled to Gs and GI proteins, respectively. Co-activation of both receptors led to the canonical negative interaction at the level of adenylyl cyclase signaling, to a strong recruitment of β-arrestin-1 and to a positive crosstalk of D1R and D3R agonists at the level of mitogen-activated protein kinase (MAPK) signaling. Furthermore, D1R or D3R antagonists counteracted β-arrestin-1 recruitment and MAPK activation induced by D3R and D1R agonists, respectively (cross-antagonism). Positive crosstalk and cross-antagonism at the MAPK level were counteracted by specific synthetic peptides with amino acid sequences corresponding to D1R transmembrane (TM) domains TM5 and TM6, which also selectively modified the quaternary structure of the D1R-D3R heteromer, as demonstrated by complementation of hemiproteins of YFP fused to D1R and D3R. These results demonstrate functional selectivity of allosteric modulations within the D1R-D3R heteromer, which can be involved with the reported behavioral synergism of D1R and D3R agonists

Leisure-time physical activity, sedentary behavior, and risk of breast cancer: Results from the SUN (‘Seguimiento Universidad De Navarra’) project

Evidence is still limited on the influence of sedentary lifestyles on breast cancer (BC) risk. Also, prospective information on the combined effects of both sedentariness and leisure-time physical activity (LTPA) is scarce. We aimed to assess the association of higher sedentary behavior and LTPA (separately and in combination) with the risk of BC in a middle-aged cohort of university graduates. The SUN Project is a follow-up study initiated in 1999 with recruitment permanently open. Baseline assessments included a validated questionnaire on LTPA and sedentary habits. Subsequently, participants completed biennial follow-up questionnaires. Multivariable adjusted Cox models were used to estimate the hazard ratios (HR) for incident BC according to LTPA, TV-watching, the joint classification of both, and a combined 8-item multidimensional active lifestyle score. We included 10,812 women, with 11.8 years of median follow-up of. Among 115,802 women-years of follow-up, we confirmed 101 incident cases of BC. Women in the highest category of LTPA (>16.5 MET-h/week) showed a significantly lower risk of BC (HR = 0.55; 95% CI: 0.34–0.90) compared to women in the lowest category (≤6 MET/h-week). Women watching >2 h/d of TV sh owed a higher risk (HR = 1.67; 95% CI:1.03–2.72) than those who watched TV 2 h/d may substantially increase BC risk, independently of each other

Metabolomics reveals impaired maturation of HDL particles in adolescents with hyperinsulinaemic androgen excess.

Hyperinsulinaemic androgen excess (HIAE) in prepubertal and pubertal girls usually precedes a broader pathological phenotype in adulthood that is associated with anovulatory infertility, metabolic syndrome and type 2 diabetes. The metabolic derangements that determine these long-term health risks remain to be clarified. Here we use NMR and MS-based metabolomics to show that serum levels of methionine sulfoxide in HIAE girls are an indicator of the degree of oxidation of methionine-148 residue in apolipoprotein-A1. Oxidation of apo-A1 in methionine-148, in turn, leads to an impaired maturation of high-density lipoproteins (HDL) that is reflected in a decline of large HDL particles. Notably, such metabolic alterations occur in the absence of impaired glucose tolerance, hyperglycemia and hypertriglyceridemia, and were partially restored after 18 months of treatment with a low-dose combination of pioglitazone, metformin and flutamide