Phenolic diterpenes from Rosemary supercritical extract inhibit non-small cell lung cancer lipid metabolism and synergise with therapeutic drugs in the clinic.

Lung cancer is one of the most deadly and common cancers in the world. The molecular features of patient's tumours dictate the different therapeutic decisions, which combines targeted therapy, chemotherapy, and immunotherapy. Altered cellular metabolism is one of the hallmarks of cancer. Tumour cells reprogram their metabolism to adapt to their novel requirements of growth, proliferation, and survival. Together with the Warburg effect, the role of lipid metabolism alterations in cancer development and prognosis has been highlighted. Several lipid related genes have been shown to promote transformation and progression of cancer cells and have been proposed as biomarkers for prognosis. Nevertheless, the exact mechanisms of the regulation of lipid metabolism and the biological consequences in non-small cell lung cancer (NSCLC) have not been elucidated yet. There is an urgent necessity to develop multidisciplinary and complementary strategies to improve NSCLC patients´ well-being and treatment response. Nutrients can directly affect fundamental cellular processes and some diet-derived ingredients, bioactive natural compounds and natural extracts have been shown to inhibit the tumour growth in preclinical and clinical trials. Previously, we described a supercritical extract of rosemary (SFRE) (12 - 16% composition of phenolic diterpenes carnosic acid and carnosol) as a potential antitumoral agent in colon and breast cancer due to its effects on the inhibition of lipid metabolism and DNA synthesis, and in the reduction of resistance to 5-FluoroUracil (5-FU). Herein, we demonstrate SFRE inhibits NSCLC cell bioenergetics identifying several lipid metabolism implicated targets. Moreover, SFRE synergises with standard therapeutic drugs used in the clinic, such as cisplatin, pemetrexed and pembrolizumab to inhibit of cell viability of NSCLC cells. Importantly, the clinical relevance of SFRE as a complement in the treatment of NSCLC patients is suggested based on the results of a pilot clinical trial where SFRE formulated with bioactive lipids (PCT/ES2017/070263) diminishes metabolic and inflammatory targets in peripheral-blood mononuclear cells (PBMC), such as MAPK (p=0.04), NLRP3 (p=0.044), and SREBF1 (p=0.047), which may augment the immune antitumour function. Based on these results, SFRE merits further investigation as a co-adjuvant in the treatment of NSCLC. Clinical trial registration: ClinicalTrials.gov Identifier NCT05080920.

Influence of Beetroot Juice Ingestion on Neuromuscular Performance on Semi-Professional Female Rugby Players: A Randomized, Double-Blind, Placebo-Controlled Study.

PURPOSE: Beetroot juice (BRJ) is considered an ergogenic aid with good to strong evidence for improving human performance in sport modalities with similar demands to rugby. However, most of the studies were realized in male athletes with limited evidence in female athletes. Thus, the aim of this study was to explore the acute ingestion of BRJ in female rugby players. METHODS: Fourteen semi-professional female rugby players (25.0 ± 3.7 years) belonging to a team from the First Spanish Female Rugby Division participated in this study. Participants were randomly divided into two groups that realized a neuromuscular battery after BRJ (140mL, 12.8 mmol NO3-) or placebo (PLAC, 140 mL, 0.08 mmol NO3-) ingestion on two different days separated by one week between protocols. The neuromuscular test battery consisted of a countermovement jump (CMJ), isometric handgrip strength (i.e., dominant), 10-m and 30-m sprint, agility t-test and Bronco test. Afterwards, participants reported a rate of perception scale (6-20 points) and side effects questionnaire associated with BRJ or PLAC ingestion. RESULTS: Statistically significant improvements were observed in CMJ (7.7%; p = 0.029; ES = 0.62), while no differences were reported in dominant isometric handgrip strength (-1.7%; p = 0.274; ES = -0.20); 10-m and 30-m sprint (0.5-0.8%; p = 0.441-0.588; ES = 0.03-0.18); modified agility t-test (-0.6%; p = 0.503; ES = -0.12) and Bronco test (1.94%; p = 0.459; ES = 0.16). CONCLUSIONS: BRJ ingestion could improve neuromuscular performance in the CMJ test, while no differences in sprint (10-m and 30-m sprint test), agility, isometric handgrip strength and endurance performance (i.e., Bronco test) were reported.

Acute Effects of Caffeine Intake on Psychological Responses and High-Intensity Exercise Performance.

OBJECTIVE: The aim of this study was to investigate the effects of caffeine supplementation on: (i) psychological responses of subjective vitality and mood; (ii) performance through a Wingate test; and (iii) rate of perceived exertion (RPE) reported after a Wingate test. METHODS: Fifteen male participants (22.60 ± 2.16 years) ingested 6 mg·kg-1 of caffeine or placebo (sucrose) supplementation in two experimental sessions. After 60 min from supplement intake, participants fulfilled two questionnaires, which measured subjective vitality and mood state, respectively. Subsequently, participants' performance was assessed through a Wingate test, which was followed by measurements of RPE at general, muscular, or cardiovascular level. RESULTS: Caffeine supplementation increased some components of mood, as assessed by profile of mood states (POMS) (tension and vigor dimensions) and subjective vitality profiles, which were followed by a greater maximum power, average power, and lower time needed to reach maximum power during the Wingate test. Moreover, lower RPE, both at muscular and general levels were reported by participants after the Wingate test. CONCLUSIONS: These results suggest that caffeine supplementation exerts positive effects both in psychological and physical domains in trained subjects.

A Survey on Dietary Supplement Consumption in Amateur and Professional Rugby Players.

PURPOSE: the aim of the present study was to analyse the pattern of dietary supplements (DS) consumption on federated rugby players, including the analysis of differences based on the sex and competitive level (professional vs. amateurs). MATERIAL AND METHODS: 144 rugby players (83 male and 61 female), of whom 69 were professionals and 75 amateurs, were recruited for the study. All the participants filled out a specific questionnaire about DS consumption including questions related to the consumption of DS and their effects on sport performance and health status. RESULTS: 65.3% of participants declared consuming at least one DS, with a higher prevalence in males than females (77.1% vs. 49.2%) and in professionals thanin amateur players (79.7% vs. 52.0%). The main reason for consumption was to enhance sport performance (62.3%) with differences only based on sex (74.3% males vs. 43.2% females). The most common purchase sites were the Internet (45.6%) and specialised stores (39.8%). As to the moment of ingestion, professionals did this most frequently during competition and training (56.4% vs. 28.2%), whereas amateur players did so only during competition (20.5% vs. 3.6%). Moreover, professional player intake most frequently in post-exercise (65.5% vs. 35.9%), whereas amateur during pre-exercise (30.8% vs. 5.5%). The DS most consumed included whey protein (44%), caffeine (42%), sports drinks (38%), energy bars (34%) and creatine monohydrate (31%), with a higher prevalence in male and professional players of whey protein and creatine monohydrate. CONCLUSIONS: The main reason for DS consumption is for enhancing sports performance). Professional players more frequently purchase them on the Internet and consume DS during training and competition period and in the post-exercise, whereas amateur players consume during competition and pre-exercise. Related to the main form of DS consumption, it is observed that a moderate consumption of DS could be considered ergogenic, such as whey protein, sport bar and creatine, while an absence of other DS could be considered ergogenic.

Effects of Dietary Nitrate Supplementation on Weightlifting Exercise Performance in Healthy Adults: A Systematic Review.

Dietary nitrate (NO3-) supplementation has been evidenced to induce an ergogenic effect in endurance and sprint-type exercise, which may be underpinned by enhanced muscle contractility and perfusion, particularly in type II muscle fibers. However, limited data are available to evaluate the ergogenic potential of NO3- supplementation during other exercise modalities that mandate type II fiber recruitment, such as weightlifting exercise (i.e., resistance exercise). In this systematic review, we examine the existing evidence basis for NO3- supplementation to improve muscular power, velocity of contraction, and muscular endurance during weightlifting exercise in healthy adults. We also discuss the potential mechanistic bases for any positive effects of NO3- supplementation on resistance exercise performance. Dialnet, Directory of Open Access Journals, Medline, Pubmed, Scielo, Scopus and SPORT Discus databases were searched for articles using the keywords: nitrate or beetroot and supplement or nut*r or diet and strength or "resistance exercise" or "resistance training" or "muscular power". Four articles fulfilling the inclusion criteria were identified. Two of the four studies indicated that NO3- supplementation could increase aspects of upper body weightlifting exercise (i.e., bench press) performance (increases in mean power/velocity of contraction/number of repetitions to failure), whereas another study observed an increase in the number of repetitions to failure during lower limb weightlifting exercise (i.e., back squat). Although these preliminary observations are encouraging, further research is required for the ergogenic potential of NO3- supplementation on weightlifting exercise performance to be determined.

Stimulation of Stem Cell Niches and Tissue Regeneration in Mouse Skin by Switchable Protoporphyrin IX-Dependent Photogeneration of Reactive Oxygen Species In Situ.

Here, we describe a protocol to induce switchable in vivo photogeneration of endogenous reactive oxygen species (ROS) in mouse skin. This transient production of ROS in situ efficiently activates cell proliferation in stem cell niches and stimulates tissue regeneration as strongly manifested through the acceleration of burn healing and hair follicle growth processes. The protocol is based on a regulatable photodynamic treatment that treats the tissue with precursors of the endogenous photosensitizer protoporphyrin IX and further irradiates the tissue with red light under tightly controlled physicochemical parameters. Overall, this protocol constitutes an interesting experimental tool to analyze ROS biology.

A novel zinc finger protein-based amperometric biosensor for miRNA determination.

This paper reports a simple electrochemical strategy for the determination of microRNAs (miRNAs) using a commercial His-Tag-Zinc finger protein (His-Tag-ZFP) that binds preferably (but non-sequence specifically) RNA hybrids over ssRNAs, ssDNAs, and dsDNAs. The strategy involves the use of magnetic beads (His-Tag-Isolation-MBs) as solid support to capture the conjugate formed in homogenous solution between His-Tag-ZFP and the dsRNA homohybrid formed between the target miRNA (miR-21 selected as a model) and a biotinylated synthetic complementary RNA detector probe (b-RNA-Dp) further conjugated with a streptavidin-horseradish peroxidase (Strep-HRP) conjugate. The electrochemical detection is carried out by amperometry at disposable screen-printed carbon electrodes (SPCEs) (- 0.20 V vs Ag pseudo-reference electrode) upon magnetic capture of the resultant magnetic bioconjugates and H2O2 addition in the presence of hydroquinone (HQ). The as-prepared biosensor exhibits a dynamic concentration range from 3.0 to 100 nM and a detection limit (LOD) of 0.91 nM for miR-21 in just ~ 2 h. An acceptable discrimination was achieved between the target miRNA and other non-target nucleic acids (ssDNA, dsDNA, ssRNA, DNA-RNA, miR-122, miR-205, and single central- or terminal-base mismatched sequences). The biosensor was applied to the analysis of miR-21 from total RNA (RNAt) extracted from epithelial non-tumorigenic and adenocarcinoma breast cells without target amplification, pre-concentration, or reverse transcription steps. The versatility of the methodology due to the ZFP's non-sequence-specific binding behavior makes it easily extendable to determine any target RNA only by modifying the biotinylated detector probe.

Publisher Correction: Intrinsic activation of cell growth and differentiation in ex vivo cultured human hair follicles by a transient endogenous production of ROS.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Tolerability and Safety of a Nutritional Supplement with Potential as Adjuvant in Colorectal Cancer Therapy: A Randomized Trial in Healthy Volunteers.

Bioactive supplements display relevant therapeutic properties when properly applied according to validated molecular effects. Our previous research efforts established the basis to develop a dietary supplement based on a Rosmarinus officinalis supercritical extract. This was enriched in phenolic diterpenes (RE) with proven properties against signaling pathways involved in colon tumorigenesis, and shark liver oil rich in alkylglycerols (AKG) as a bioactive lipid vehicle to improve RE bioavailability and synergize with the potential therapeutic action of the extract. Herein, we have investigated the tolerability and safety of the supplement and the biological and molecular effects from an immuno-nutritional perspective. Sixty healthy volunteers participated in a six week, double-blind, randomized parallel pilot study with two study arms: RE-AKG capsules (CR) and control capsules (CC). Mean age (±SD) of volunteers was 28.32 (±11.39) and 27.5 (±9.04) for the control and the study groups, respectively. Safety of the CR product consumption was confirmed by analyzing liver profile, vital constants, and oxidation markers (LDLox in blood and isoprostanes and thromboxanes in urine). The following were monitored: (1) the phenotyping of plasmatic leukocytes and the ex vivo response of lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear cells (PBMCs); (2) expression of genes associated with immune-modulation, inflammation, oxidative stress, lipid metabolism, and tumorigenesis; and (3) the correlation of selected genetic variants (SNPs) with the differential responses among individuals. The lack of adverse effects on liver profile and oxidation markers, together with adequate tolerability and safe immunological adaptations, provide high-quality information for the potential use of CR as co-adjuvant of therapeutic strategies against colorectal cancer.

Intrinsic activation of cell growth and differentiation in ex vivo cultured human hair follicles by a transient endogenous production of ROS.

The emerging variety of signalling roles for ROS in eukaryotic cells and tissues is currently a matter of intense research. Here we make use of ex vivo cultured single human hair follicles as an experimental model to demonstrate that a transient production of non-lethal endogenous ROS levels in these mini-organs promotes efficiently the entry into the growth phase (anagen). The stimulatory process implicates the specific activation of the hair follicle stem cell niche, encompassing the induction of stem cell differentiation markers (Ck15), overall cell proliferation and sustained growth of the tissue associated with expression of gen targets (Ccnd1) concomitant with the inhibition of Wnt signaling antagonists and repressors (Dkk1, Gsk3ß) of Wnt signaling. As a whole, this observation indicates that, once activated, ROS signalling is an intrinsic mechanism regulating the hair follicle stem cell niche independently of any external signal.

On the autonomous motion of active drops or bubbles.

Thermo-capillary stresses on the surface of a drop can be the result of a non-isothermal surface chemical conversion of a reactant dissolved in the host fluid. The strength of heat production (with e.g. absorption) on the surface is ruled by the diffusion of the reactant and depends on the state of motion of the drop. Such thermo-capillary stresses can provoke the motion of the drop or its motionless state in the presence of an external body force. If in the balance of forces, including indeed viscous drag, the net resultant force vanishes there is the possibility of autonomous motion with constant velocity of the drop. Focusing on drops with radii in the millimeter range provided here is a quantitative study of the possibility of such autonomous motion when the drop, considered as active unit, is seat of endo- or exo-thermic reactive processes that dominate its motion. The framework is restricted to Stokes flows in the hydrodynamics, negligible heat Peclet number while the solute Peclet number is considered very high. A boundary layer approximation is used in the description of reactant diffusion. Those processes eventually end up in the action being expressed by surface tension gradients and the Marangoni effect. Explicit expressions of the force acting on the drop and the velocity fields inside and outside the drop are provided. Some significant particular cases are discussed to illustrate the usefulness of the theory.

Single-Step Incubation Determination of miRNAs in Cancer Cells Using an Amperometric Biosensor Based on Competitive Hybridization onto Magnetic Beads.

This work reports an amperometric biosensor for the determination of miRNA-21, a relevant oncogene. The methodology involves a competitive DNA-target miRNA hybridization assay performed on the surface of magnetic microbeads (MBs) and amperometric transduction at screen-printed carbon electrodes (SPCEs). The target miRNA competes with a synthetic fluorescein isothiocyanate (FITC)-modified miRNA with an identical sequence for hybridization with a biotinylated and complementary DNA probe (b-Cp) immobilized on the surface of streptavidin-modified MBs (b-Cp-MBs). Upon labeling, the FITC-modified miRNA attached to the MBs with horseradish peroxidase (HRP)-conjugated anti-FITC Fab fragments and magnetic capturing of the MBs onto the working electrode surface of SPCEs. The cathodic current measured at -0.20 V (versus the Ag pseudo-reference electrode) was demonstrated to be inversely proportional to the concentration of the target miRNA. This convenient biosensing method provided a linear range between 0.7 and 10.0 nM and a limit of detection (LOD) of 0.2 nM (5 fmol in 25 µL of sample) for the synthetic target miRNA without any amplification step. An acceptable selectivity towards single-base mismatched oligonucleotides, a high storage stability of the b-Cp-MBs, and usefulness for the accurate determination of miRNA-21 in raw total RNA (RNAt) extracted from breast cancer cells (MCF-7) were demonstrated.

[Prevalence of birth defects in Risaralda, 2010-2013].

INTRODUCTION: The data regarding birth defects at local levels in developing countries like Colombia are scarce. OBJECTIVE: To describe the profile of congenital abnormalities in the province of Risaralda, Colombia. MATERIALS AND METHODS: We included the information on infants with structural and functional abnormalities at birth between June, 2010, and December, 2013, from records of the Instituto Nacional de Salud, and compared it with those of children born in the same period in a local clinic participating in the Collaborative Study of Congenital Malformations. We analyzed the data using Stata 10®. RESULTS: We found a prevalence of nine defects per 1,000 newborns from the total live births in Risaralda. The local clinic registered in the Collaborative Study of Congenital Malformations registered a prevalence of 34 defects per 1,000 births. Most frequent defects were heart defects, followed by cleft lip and palate, abdominal wall defects, skeletal dysplasia, hydrocephalus, polydactyly and Down syndrome. CONCLUSIONS: Having a baseline on the prevalence of congenital defects in Risaralda is very useful in the design of prevention policies oriented to decrease congenital defects incidence and severity. Inclusion of maternity hospitals in the Collaborative Study of Congenital Malformations strengthens national recording and reporting of birth defects.

Prevalencia de defectos congénitos en Risaralda, 2010-2013 / Prevalence of birth defects in Risaralda, 2010-2013

Resumen Introducción. Los datos sobre defectos congénitos en el ámbito regional de los países en desarrollo como Colombia son escasos. Objetivo. Describir la prevalencia de anomalías congénitas en el departamento de Risaralda, Colombia. Materiales y métodos. Se incluyeron los neonatos con defectos estructurales y funcionales entre junio de 2010 y diciembre de 2013, cuyos casos fueron notificados al Instituto Nacional de Salud por ser de interés en salud pública. Se compararon con los nacidos en el mismo periodo en una clínica de la región inscrita en el Estudio Colaborativo de Malformaciones Congénitas. Los datos se analizaron con el programa Stata 10(r). Resultados. La prevalencia entre los nacidos vivos en el periodo de estudio fue de nueve casos por cada 1.000 recién nacidos en el departamento. En la clínica inscrita en el Estudio Colaborativo de Malformaciones Congénitas, se encontró una prevalencia de 34 casos por cada 1.000 nacimientos; el primer lugar lo ocuparon las cardiopatías, seguidas por el labio y paladar hendido, los defectos de la pared abdominal (no especificados), la displasia esquelética, la hidrocefalia, la polidactilia y el síndrome de Down. Conclusión. El establecimiento de una línea de base sobre la prevalencia de los defectos congénitos en Risaralda es de gran utilidad para la adopción de políticas preventivas que lleven a la disminución de la incidencia y de la gravedad de las discapacidades; la inclusión de los hospitales materno-infantiles en la red del Estudio Colaborativo de Malformaciones Congénitas mejora el registro nacional y la notificación de los defectos congénitos.

Abstract Introduction: The data regarding birth defects at local levels in developing countries like Colombia are scarce. Objective: To describe the profile of congenital abnormalities in the province of Risaralda, Colombia. Materials and methods: We included the information on infants with structural and functional abnormalities at birth between June, 2010, and December, 2013, from records of the Instituto Nacional de Salud, and compared it with those of children born in the same period in a local clinic participating in the Collaborative Study of Congenital Malformations. We analyzed the data using Stata 10(r). Results: We found a prevalence of nine defects per 1,000 newborns from the total live births in Risaralda. The local clinic registered in the Collaborative Study of Congenital Malformations registered a prevalence of 34 defects per 1,000 births. Most frequent defects were heart defects, followed by cleft lip and palate, abdominal wall defects, skeletal dysplasia, hydrocephalus, polydactyly and Down syndrome. Conclusions: Having a baseline on the prevalence of congenital defects in Risaralda is very useful in the design of prevention policies oriented to decrease congenital defects incidence and severity. Inclusion of maternity hospitals in the Collaborative Study of Congenital Malformations strengthens national recording and reporting of birth defects.

Altered expression of P2Y2 and P2X7 purinergic receptors in the isolated rat heart mediates ischemia-reperfusion injury.

The aim of this study is to analyze the expression of purinergic receptors in the heart after ischemia-reperfusion, and their possible role in ischemia-reperfusion injury. Rat hearts were perfused according to the Langendorff technique and subjected to 30 min ischemia followed by 15 min reperfusion. Ischemia-reperfusion reduced the gene expression and protein content of purinergic receptors of the P2Y2 subtype, and increased the gene expression and protein content of the P2X7 subtype. Treatment with the agonist of the P2Y2 subtype 2-thio-UTP and with the antagonist of the P2X7 subtype Brilliant Blue improved myocardial function parameters, reduced cell death and increased the myocardial expression of antiapoptotic markers after ischemia-reperfusion. These results suggest that the myocardial expression of the protective P2Y2 subtype of purinergic receptors is reduced, whereas that of the harmful subtype P2X7 subtype is increased during coronary ischemia-reperfusion. This may contribute to myocardial injury in this condition.

Nitric oxide-mediated relaxation to lactate of coronary circulation in the isolated perfused rat heart.

The objective of this study was to analyze the effects of lactate on coronary circulation. Rat hearts were perfused in a Langendorff preparation, and the coronary response to lactate (3-30 mM) was recorded after precontracting coronary vasculature with 11-dideoxy-1a,9a-epoxymethanoprostaglandin F2α (U46619), in the presence or the absence of the inhibitor of nitric oxide synthesis, N-omega-nitro-l-arginine methyl ester (l-NAME, 10 M), the blocker of Ca-dependent potassium channels, tetraethylammonium (TEA, 10 M), or the blocker of adenosine triphosphate-sensitive potassium channels, glybenclamide (10 M). The effects of lactate were also studied in isolated segments of rat coronary arteries that were precontracted with U46619, with or without endothelium. In perfused hearts, lactate induced concentration-dependent coronary vasodilatation and a reduction in myocardial contractility (left ventricular developed pressure and dP/dt) without altering the heart rate. Coronary vasodilatation in response to lactate was reduced by l-NAME but unaffected by TEA or glybenclamide. The effects of lactate on myocardial contractility were unchanged by l-NAME, TEA, or glybenclamide. In isolated coronary artery segments, lactate also produced relaxation, an effect attenuated by removing the endothelium. Together these findings suggest that lactate exerts coronary vasodilatory effects through the release of endothelial nitric oxide, independently of potassium channels. These findings may be relevant for the regulation of coronary circulation when lactate levels are elevated.