Electrophilic Hydrosilylation of Electron-Rich Alkenes Derived from Enamines.

The low-electron count, air-stable, platinum complexes [Pt(ItBu')(ItBu)][BArF] (C1) (ItBu=1,3-di-tert-butylimidazol-2-ylidene), [Pt(SiPh)3(ItBuiPr)2][BArF] (C2) (ItBuiPr = 1-tert-butyl-3-iso-propylimidazol-2-ylidene),  [Pt(SiPh)3(ItBuMe)2][BArF] (C3), [Pt(GePh3)(ItBuiPr)2][BArF] (C4), [Pt(GePh)3(ItBuMe)2][BArF] (C5) and [Pt(GeEt)3(ItBuMe)2][BArF] (C6)  (ItBuMe = 1-tert-butyl-3-methylimidazol-2-ylidene) are efficient catalysts (particularly the germyl derivatives) in both the silylative dehydrocoupling and hydrosilylation of electron rich alkenes derived from enamines.  The steric hindrance exerted by the NHC ligand plays an important role in the selectivity of the reaction. Thus, bulky ligands are selective towards the silylative dehydrocoupling process whereas less sterically hindered promote the selective hydrosilylation reaction. The latter is, in addition, regioselective towards the ß-carbon atom of both internal and terminal enamines, leading to ß-aminosilanes. Moreover, the syn stereochemistry of the amino and silyl groups implies an anti Si‒H bond addition across the double bond. All these facts point to a mechanistic picture that, according to experimental and computational studies, involves a non-classical hydrosilylation process through an outer-sphere mechanism in which a formal nucleophilic addition of the enamine to the silicon atom of a platinum σ-SiH complex is the key step. This is in sharp contrast with the classical Chalk-Harrod mechanism prevalent in platinum chemistry.

The mineralocorticoid receptor forms higher order oligomers upon DNA binding.

The prevailing model of steroid hormone nuclear receptor function assumes ligand-induced homodimer formation followed by binding to DNA hormone response elements (HREs). This model has been challenged by evidence showing that the glucocorticoid receptor (GR) forms tetramers upon ligand and DNA binding, which then drive receptor-mediated gene transactivation and transrepression. GR and the closely-related mineralocorticoid receptors (MR) interact to transduce corticosteroid hormone signaling, but whether they share the same quaternary arrangement is unknown. Here, we used a fluorescence imaging technique, Number & Brightness, to study oligomerization in a cell system allowing real-time analysis of receptor-DNA interactions. Agonist-bound MR forms tetramers in the nucleoplasm and higher order oligomers upon binding to HREs. Antagonists form intermediate-size quaternary arrangements, suggesting that large oligomers are essential for function. Divergence between MR and GR quaternary structure is driven by different functionality of known and new multimerization interfaces, which does not preclude formation of heteromers. Thus, influencing oligomerization may be important to selectively modulate corticosteroid signaling.

Characterization of Limnospira platensis PCC 9108 R-M and CRISPR-Cas systems.

The filamentous cyanobacterium Limnospira platensis, formerly known as Arthrospira platensis or spirulina, is one of the most commercially important species of microalgae. Due to its high nutritional value, pharmacological and industrial applications it is extensively cultivated on a large commercial scale. Despite its widespread use, its precise manipulation is still under development due to the lack of effective genetic protocols. Genetic transformation of Limnospira has been attempted but the methods reported have not been generally reproducible in other laboratories. Knowledge of the transformation defense mechanisms is essential for understanding its physiology and for broadening their applications. With the aim to understand more about the genetic defenses of L. platensis, in this work we have identified the restriction-modification and CRISPR-Cas systems and we have cloned and characterized thirteen methylases. In parallel, we have also characterized the methylome and orphan methyltransferases using genome-wide analysis of DNA methylation patterns and RNA-seq. The identification and characterization of these enzymes will be a valuable resource to know how this strain avoids being genetically manipulated and for further genomics studies.

Mutation in Chek2 triggers von Hippel-Lindau hemangioblastoma growth.

PURPOSE: Von Hippel-Lindau (VHL) is a rare inherited disease mainly characterized by the growth of tumours, predominantly hemangioblastomas (Hbs) in the CNS and retina, and renal carcinomas. The natural history of VHL disease is variable, differing in the age of onset and its penetrance, even among relatives. Unfortunately, sometimes VHL shows more severe than average: the onset starts in adolescence, and surgeries are required almost every year. In these cases, the factor that triggers the appearance and growth of Hbs usually remains unknown, although additional mutations are suspected. METHODS: We present the case of a VHL patient whose first surgery was at 13 years of age. Then, along his next 8 years, he has undergone 5 surgeries for resection of 10 CNS Hbs. To clarify this severe VHL condition, DNA from a CNS Hb and white blood cells (WBC) was sequenced using next-generation sequencing technology. RESULTS: Massive DNA sequencing of the WBC (germ line) revealed a pathogenic mutation in CHEK2 and the complete loss of a VHL allele (both tumour suppressors). Moreover, in the tumour sample, several mutations, in BRAF1 and PTPN11 were found. Familiar segregation studies showed that CHEK2 mutation was in the maternal lineage, while VHL was inherited by paternal lineage. CONCLUSIONS: Finally, clinical history correlated to the different genotypes in the family, concluding that the severity of these VHL manifestations are due to both, VHL-and-CHEK2 mutations. This case report aims to notice the importance of deeper genetic analyses, in inherited rare diseases, to uncover non-expected mutations.

Intestinal Obstruction Secondary to Gallstone Ileus in a Patient with Cholangitis: Case Report

Gallstone ileus manifests as intestinal obstruction. It occurs due to the passage of a stone and its subsequent lodging in the lumen of the digestive tract. The diagnosis is confirmed by imaging; the gold standard is abdominal tomography. Management is based on the extraction of the intraluminal calculus in one or more surgical times, depending on the patient's condition. We present the case of a patient with multiple comorbidities who showed a picture of cholangitis complicated by gallstone ileus and successfully treated with enterolithotomy. Surgical management is controversial since the optimal approach for these patients has not been established.

El íleo biliar se manifiesta como una obstrucción intestinal, se presenta por el paso de un lito y su posterior alojamiento en el lumen del tubo digestivo. El diagnóstico se confirma mediante imagenología, el patrón de oro es la tomografía abdominal. El manejo se fundamenta en la extracción del cálculo intraluminal en uno o más tiempos quirúrgicos, según el estado del paciente. Se presenta el caso de un paciente con múltiples comorbilidades, que debuta con un cuadro de colangitis complicada por íleo biliar, tratado exitosamente con enterolitotomía. El manejo quirúrgico es controversial, ya que no se ha establecido el abordaje óptimo para estos pacientes.

ADAM10 Gene Variants in AD Patients and Their Relationship to CSF Protein Levels.

ADAM10 is the main α-secretase acting in the non-amyloidogenic processing of APP. We hypothesized that certain rare ADAM10 variants could increase the risk for AD by conferring the age-related downregulation of α-secretase. The ADAM10 gene was sequenced in 103 AD cases (82% familial) and 96 cognitively preserved nonagenarians. We examined rare variants (MAF < 0.01) and determined their potential association in the AD group with lower CSF protein levels, as analyzed by means of ELISA, and Western blot (species of 50 kDa, 55 kDa, and 80 kDa). Rare variants were found in 15.5% of AD cases (23% early-onset, 8% late-onset) and in 12.5% of nonagenarians, and some were group-specific. All were intronic variants except Q170H, found in three AD cases and one nonagenarian. The 3'UTR rs74016945 (MAF = 0.01) was found in 6% of the nonagenarians (OR 0.146, p = 0.057). Altogether, ADAM10 total levels or specific species were not significantly different when comparing AD with controls or carriers of rare variants versus non-carriers (except a Q170H carrier exhibiting low levels of all species), and did not differ according to the age at onset or APOE genotype. We conclude that ADAM10 exonic variants are uncommon in AD cases, and the presence of rare intronic variants (more frequent in early-onset cases) is not associated with decreased protein levels in CSF.

TRIM25 mutation (p.C168*), coding for an E3 ubiquitin ligase, is a cause of early-onset autosomal dominant dementia with amyloid load and parkinsonism.

INTRODUCTION: Patients with familial early-onset dementia (EOD) pose a unique opportunity for gene identification studies. METHODS: We present the phenotype and whole-exome sequencing (WES) study of an autosomal dominant EOD family. Candidate genes were examined in a set of dementia cases and controls (n = 3712). Western blotting was conducted of the wild-type and mutant protein of the final candidate. RESULTS: Age at disease onset was 60 years (range 56 to 63). The phenotype comprised mixed amnestic and behavioral features, and parkinsonism. Cerebrospinal fluid and plasma biomarkers, and a positron emission tomography amyloid study suggested Alzheimer's disease. WES and the segregation pattern pointed to a nonsense mutation in the TRIM25 gene (p.C168*), coding for an E3 ubiquitin ligase, which was absent in the cohorts studied. Protein studies supported a loss-of-function mechanism. DISCUSSION: This study supports a new physiopathological mechanism for brain amyloidosis. Furthermore, it extends the role of E3 ubiquitin ligases dysfunction in the development of neurodegenerative diseases. HIGHLIGHTS: A TRIM25 nonsense mutation (p.C168*) is associated with autosomal dominant early-onset dementia and parkinsonism with biomarkers suggestive of Alzheimer's disease. TRIM25 protein studies support that the mutation exerts its effect through loss of function. TRIM25, an E3 ubiquitin ligase, is known for its role in the innate immune response but this is the first report of association with neurodegeneration. The role of TRIM25 dysfunction in development of amyloidosis and neurodegeneration merits a new line of research.

Manejo de discontinuidad pélvica debido a fractura de acetábulo mediante cirugía de revisión de artroplastia de cadera y reemplazo de componente acetabular por anillo de Burch-Schneider: reporte de un caso / Treatment for pelvic discontinuity due to acetabular fracture by means of hip arthroplasty revision surgery and acetabular component replacement using a Burch-Schneider ring: a case report

Introducción. La discontinuidad pélvica es una complicación de la artroplastia total de cadera que consiste en la separación de la hemipelvis superior e inferior a través del acetábulo.Presentación del caso. Hombre de 74 años con antecedente de artroplastia total de cadera izquierda que requirió 4 cirugías de revisión de artroplastia, quien consultó al servicio de urgencias de un hospital de segundo nivel de atención de Sogamoso (Colombia) por dolor y limitación de la movilidad de la cadera izquierda durante 7 días luego de haber sufrido trauma por caída de su propia altura en la que el miembro inferior izquierdo recibió la fuerza del impacto. En el examen físico, se evidenció limitación de los arcos de movilidad de la cadera izquierda, por lo que se realizó una radiografía de cadera, en la que se observó fractura del acetábulo, y una tomografía computarizada de cadera, en la que se evidenciaron signos de aflojamiento del componente acetabular y fractura del acetábulo (tipo IIIB según clasificación de Paprosky). Teniendo en cuenta lo anterior, se diagnosticó discontinuidad pélvica y se realizó cirugía de revisión de artroplastia y reemplazo del componente acetabular por un anillo de Burch-Schneider. El paciente tuvo una adecuada evolución posoperatoria con seguimiento a 3 meses. Conclusión. Si bien hay múltiples opciones para el manejo de la discontinuidad pélvica, no hay consenso sobre cuál es la mejor; sin embargo, el uso de componentes acetabulares de refuerzo como el anillo de Burch-Schneider fue una opción con buenos resultados posoperatorios en este caso

Introduction: Pelvic discontinuity is a complication of total hip arthroplasty, consisting of a structural bone defect of the acetabulum with separation of the upper and lower hemipelvis. Case presentation: A 74-year-old male patient with a history of total left hip arthroplasty and 4 arthroplasty revision surgeries visited the emergency department of a secondary care hospital due to pain and limited mobility of the left hip over a period of 7 days after having suffered a fall from his own height in which the left lower limb received the force of the impact. Physical examination showed limitation of the range of motion of the left hip, so a hip X-ray was performed, showing a fracture of the acetabulum. A computed tomography of the hip revealed signs of loosening of the acetabular component and fracture of the acetabulum (type IIIB according to the Paprosky classification). In view of the above, pelvic discontinuity was diagnosed and revision arthroplasty and replacement of the acetabular component with a Burch-Schneider ring was performed. The patient had an adequate postoperative progression and follow-up at 3 months.Conclusion: Although there are multiple options for the treatment of pelvic discontinuity, currently, there is no conclusive data regarding the best therapeutic option. However, the use of reinforcement acetabular components, such as the Burch-Schneider ring, was an option with good postoperative outcomes in this case

The Bias of Physicians and Lack of Education in Patients of Color With Melanoma as Causes of Increased Mortality: A Scoping Review.

Minorities, particularly non-White minorities, often encounter implicit biases from healthcare professionals that may impact their standard of care and quality of life. The study of dermatology has long been based on Whites, unintentionally affecting the treatment of non-White patients. Melanoma, although mostly curable, can become fatal in those presenting with advanced stages at diagnosis. Despite being rare in racial minorities, melanoma is associated with a worse prognosis among them compared to White populations. In light of this, the objective of this study was to determine the role of education in preventing biases and improving the diagnosis and treatment of melanoma in minority groups to improve patient outcomes. This study was designed as a scoping review to gather evidence on the impact of implicit bias and lack of education on the treatment of melanoma in people of color. Following Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines, we searched for peer-reviewed studies involving melanoma, education, and treatment bias in people of color on the databases PubMed, Medline EBSCO, CINAHL, and Cochrane. The data were extracted pertaining to the following main aspects: (1) risk factors, (2) surveys of current knowledge, and 3) educational interventions. This scoping review identified socioeconomic factors, bias, and lack of education in minority populations as causes of increased mortality rates in melanoma. Moreover, because preventative dermatology is largely based on White skin types, incorporating darker skin tones into education will help dispel implicit bias. Additionally, there is evidence to indicate that current patient knowledge and understanding of skin cancer is inaccurate among many and can be significantly improved through educational interventions, such as brochures and videos. Further educational interventions may be beneficial to increase understanding of melanoma in populations of color to address health disparities in dermatological care.

Beetroot Juice Produces Changes in Heart Rate Variability and Reduces Internal Load during Resistance Training in Men: A Randomized Double-Blind Crossover.

Beetroot juice (BJ) has been used as a sport supplement, improving performance in resistance training (RT). However, its effect on the modulation of the autonomic nervous system has not yet been widely studied. Therefore, the objective of this randomized double-blind crossover study was to assess the effect of acute BJ supplementation compared to placebo in blood pressure (BP), heart rate (HR), heart rate variability (HRV) and internal load during RT measure as Root Mean Square of the Successive Differences between adjacent RR intervals Slope (RMSSD and RMSSD-Slope, respectively). Eleven men performed an incremental RT test (three sets at 60%, 70% and 80% of their repetition maximum) composed by back squat and bench press with. HR, HRV and RMSSD-Slope were measured during and post exercise. As the main results, RMSSD during exercise decrease in the BJ group compared to placebo (p = 0.023; ES = 0.999), there were no differences in RMSSD post-exercise, and there were differences in RMSSD-Slope between groups in favor of the BJ group (p = 0.025; ES = 1.104) with a lower internal load. In conclusion, BJ supplementation seems to be a valuable tool for the reduction in the internal load of exercise during RT measured as RMSSD-Slope while enhancing performance.

Performance Analysis and Architecture of a Clustering Hybrid Algorithm Called FA+GA-DBSCAN Using Artificial Datasets.

Density-Based Spatial Clustering of Applications with Noise (DBSCAN) is a widely used algorithm for exploratory clustering applications. Despite the DBSCAN algorithm being considered an unsupervised pattern recognition method, it has two parameters that must be tuned prior to the clustering process in order to reduce uncertainties, the minimum number of points in a clustering segmentation MinPts, and the radii around selected points from a specific dataset Eps. This article presents the performance of a clustering hybrid algorithm for automatically grouping datasets into a two-dimensional space using the well-known algorithm DBSCAN. Here, the function nearest neighbor and a genetic algorithm were used for the automation of parameters MinPts and Eps. Furthermore, the Factor Analysis (FA) method was defined for pre-processing through a dimensionality reduction of high-dimensional datasets with dimensions greater than two. Finally, the performance of the clustering algorithm called FA+GA-DBSCAN was evaluated using artificial datasets. In addition, the precision and Entropy of the clustering hybrid algorithm were measured, which showed there was less probability of error in clustering the most condensed datasets.

Toward a Structural Health Monitoring Methodology for Concrete Structures under Dynamic Loads Using Embedded FBG Sensors and Strain Mapping Techniques.

A data-driven-based methodology for SHM in reinforced concrete structures using embedded fiber optic sensors and pattern recognition techniques is presented. A prototype of a reinforced concrete structure was built and instrumented in a novel fashion with FBGs bonded directly to the reinforcing steel bars, which, in turn, were embedded into the concrete structure. The structure was dynamically loaded using a shaker. Superficial positive damages were induced using bonded thin steel plates. Data for pristine and damaged states were acquired. Classifiers based on Mahalanobis' distance of the covariance data matrix were developed for both supervised and unsupervised pattern recognition with an accuracy of up to 98%. It was demonstrated that the proposed sensing scheme in conjunction with the developed supervised and unsupervised pattern recognition techniques allows the detection of slight stiffness changes promoted by damages, even when strains are very small and the changes of these associated with the damage occurrence may seem negligible.

A Novel Splicing Mutation in the ACVRL1/ALK1 Gene as a Cause of HHT2.

Hereditary Hemorrhagic Telangiectasia (HHT) is a rare disorder of vascular development. Common manifestations include epistaxis, telangiectasias and arteriovenous malformations in multiple organs. Different deletions or nonsense mutations have been described in the ENG (HHT1) or ACVRL1/ALK1 (HHT2) genes, all affecting endothelial homeostasis. A novel mutation in ACVRL1/ALK1 has been identified in a Peruvian family with a clinical history compatible to HHT. Subsequently, 23 DNA samples from oral exchanges (buccal swaps) of the immediate family members were analyzed together with their clinical histories. A routine cDNA PCR followed by comparative DNA sequencing between the founder and another healthy family member showed the presence of the aforementioned specific mutation. The single mutation detected (c.525 + 1G > T) affects the consensus splice junction immediately after exon 4, provokes anomalous splicing and leads to the inclusion of intron IV between exons 4 and 5 in the ACVRL1/ALK1 mRNA and, therefore, to ALK1 haploinsufficiency. Complete sequencing determined that 10 of the 25 family members analyzed were affected by the same mutation. Notably, the approach described in this report could be used as a diagnostic technique, easily incorporated in clinical practice in developing countries and easily extrapolated to other patients carrying such a mutation.

Acute Effects of Beetroot Juice Supplements on Lower-Body Strength in Female Athletes: Double-Blind Crossover Randomized Trial.

BACKGROUND: Beetroot juice (BRJ) is used as an ergogenic aid, but no previous study has analyzed the effect this supplement has on the production of explosive force and muscular endurance in physically active women. HYPOTHESIS: BRJ improves explosive force and muscular endurance in the lower limbs of physically active women. STUDY DESIGN: Randomized double-blind crossover study. LEVEL OF EVIDENCE: Level 3. METHODS: Fourteen physically active women performed a countermovement jump (CMJ) test, a back squat test for assessing velocity and power at 50% and 75% of one-repetition maximum (1RM), and the number of repetitions on a muscular endurance test consisting of 3 sets at 75% of 1RM in a resistance training protocol comprising 3 exercises (back squat, leg press, and leg extension). The participants performed the test in 2 sessions, 150 minutes after ingesting 70 mL of either BRJ (400 mg of nitrate) or a placebo (PLA). RESULTS: A greater maximum height was achieved in the CMJ after consuming BRJ compared with a PLA (P = 0.04; effect size (ES) = 0.34). After a BRJ supplement at 50% 1RM, a higher mean velocity [+6.7%; P = 0.03; (ES) = 0.39 (-0.40 to 1.17)], peak velocity (+6%; P = 0.04; ES = 0.39 [-0.40 to 1.17]), mean power (+7.3%; P = 0.02; ES = 0.30 [-0.48 to 1.08]) and peak power (+6%; P = 0.04; ES = 0.20 [-0.59 to 0.98]) were attained in the back squat test. In the muscular endurance test, BRJ increased performance compared with the PLA (P < 0.00; ηp2 = 0.651). CONCLUSION: BRJ supplements exert an ergogenic effect on the ability to produce explosive force and muscular endurance in the lower limbs in physically active women. CLINICAL RELEVANCE: If physically active women took a BRJ supplement 120 minutes before resistance training their performance could be enhanced.

Relationship Between Gymnastic Rhythmic Practice and Body Composition, Physical Performance, and Trace Element Status in Young Girls.

This study evaluates the influence on body development of doing rhythmic gymnastics in girls from 10 to 17 years of age, the results of certain strength and flexibility abilities, and the trace element status (Ca, Fe, Zn, Cu, Mn, Cr, and Ni). The subjects were divided into three groups: (a) girls who practiced rhythmic gymnastics at a competition level (competition group); (b) girls who practiced this sport at a non-competitive level (training group); and (c) girls who do not practice any sport and with a low level of physical activity (control or sedentary group). Trace element status was determined in hair and urine samples. Results showed that doing rhythmic gymnastics does not alter the normal physical development of muscle mass, and even leads to a decrease in body fat content. Furthermore, better scores in the strength and flexibility test were obtained by the participants of this sports discipline. Statistically significant differences in urine Fe, Cu, and Mn values (p < 0.05) and in hair Cr, Cu, and Mn values (p < 0.05) were found between the two rhythmic gymnastics groups and the control group, and were higher in the competition and training groups. A principal component analysis model was performed to evaluate the possibility of cluster formation among the girls. The PCA results revealed a separation between the different groups although the separation was not perfect. PLS-DA was attempted in order to verify whether it was possible to discriminate between the groups included in this study. It was clear that the competition and control ones were very well classified (around 95% of correct predictions) but 20% of the girls belonging to the training group were misclassified as belonging to the competition one.

Atorvastatin and blood flow regulate expression of distinctive sets of genes in mouse carotid artery endothelium.

Hypercholesterolemia is a well-known pro-atherogenic risk factor and statin is the most effective anti-atherogenic drug that lowers blood cholesterol levels. However, despite systemic hypercholesterolemia, atherosclerosis preferentially occurs in arterial regions exposed to disturbed blood flow (d-flow), while the stable flow (s-flow) regions are spared. Given their predominant effects on endothelial function and atherosclerosis, we tested whether (1) statin and flow regulate the same or independent sets of genes and (2) statin can rescue d-flow-regulated genes in mouse artery endothelial cells in vivo. To test the hypotheses, C57BL/6 J mice (8-week-old male, n=5 per group) were pre-treated with atorvastatin (10mg/kg/day, Orally) or vehicle for 5 days. Thereafter, partial carotid ligation (PCL) surgery to induce d-flow in the left carotid artery (LCA) was performed, and statin or vehicle treatment was continued. The contralateral right carotid artery (RCA) remained exposed to s-flow to be used as the control. Two days or 2 weeks post-PCL surgery, endothelial-enriched RNAs from the LCAs and RCAs were collected and subjected to microarray gene expression analysis. Statin treatment in the s-flow condition (RCA+statin versus RCA+vehicle) altered the expression of 667 genes at 2-day and 187 genes at 2-week timepoint, respectively (P<0.05, fold change (FC)≥±1.5). Interestingly, statin treatment in the d-flow condition (LCA+statin versus LCA+vehicle) affected a limited number of genes: 113 and 75 differentially expressed genes at 2-day and 2-week timepoint, respectively (P<0.05, FC≥±1.5). In contrast, d-flow in the vehicle groups (LCA+vehicle versus RCA+vehicle) differentially regulated 4061 genes at 2-day and 3169 genes at 2-week timepoint, respectively (P<0.05, FC≥±1.5). Moreover, statin treatment did not reduce the number of flow-sensitive genes (LCA+statin versus RCA+statin) compared to the vehicle groups: 1825 genes at 2-day and 3788 genes at 2-week, respectively, were differentially regulated (P<0.05, FC≥±1.5). These results revealed that both statin and d-flow regulate expression of hundreds or thousands of arterial endothelial genes, respectively, in vivo. Further, statin and d-flow regulate independent sets of endothelial genes. Importantly, statin treatment did not reverse d-flow-regulated genes except for a small number of genes. These results suggest that both statin and flow play important independent roles in atherosclerosis development and highlight the need to consider their therapeutic implications for both.

Detection of Genetic Rearrangements in the Regulators of Complement Activation RCA Cluster by High-Throughput Sequencing and MLPA.

The regulators of complement activation (RCA) gene cluster in 1q31-1q32 includes most of the genes encoding complement regulatory proteins. Genetic variability in the RCA gene cluster frequently involve copy number variations (CNVs), a type of chromosome structural variation causing alterations in the number of copies of specific regions of DNA. CNVs in the RCA gene cluster often relate with gene rearrangements that result in the generation of novel genes, carrying internal duplications or deletions, and hybrid genes, resulting from the fusion or exchange of genetic material between two different genes. These gene rearrangements are strongly associated with a number of rare and common diseases characterized by complement dysregulation. Identification of CNVs in the RCA gene cluster is critical in the molecular diagnostic of these diseases. It can be done by bioinformatics analysis of DNA sequence data generated by massive parallel sequencing techniques (NGS, next generation sequencing) but often requires special techniques like multiplex ligation-dependent probe amplification (MLPA). This is because the currently used massive parallel DNA sequencing approaches do not easily identify all the structural variations in the RCA gene cluster. We will describe here how to use the MLPA assays and two computational tools to analyze NGS data, NextGENe and ONCOCNV, to detect CNVs and gene rearrangements in the RCA gene cluster.

Morning versus Evening Intake of Creatine in Elite Female Handball Players.

A great deal of evidence has been gathered on the use of creatine as an ergogenic supplement. Recent studies show greater benefits when creatine ingestion is performed close in time to training, but few studies tackle the way that circadian rhythms could influence creatine consumption. The aim of this study was therefore to observe the influence circadian rhythms exert on sports performance after creatine supplementation. Our method involved randomly assigning fourteen women players of a handball team into two groups in a single-blind study: one that consumed the supplement in the morning and one that consumed it in the evening, with both groups following a specific training program. After twelve weeks, the participants exhibited a decreased fat percentage, increased body weight and body water, and improved performance, with these results being very similar in the two groups. It is therefore concluded that, although circadian rhythms may influence performance, these appear not to affect creatine supplementation, as creatine is stored intramuscularly and is available for those moments of high energy demand, regardless of the time of day.

Endothelial Reprogramming by Disturbed Flow Revealed by Single-Cell RNA and Chromatin Accessibility Study.

Disturbed flow (d-flow) induces atherosclerosis by regulating gene expression in endothelial cells (ECs). For further mechanistic understanding, we carried out a single-cell RNA sequencing (scRNA-seq) and scATAC-seq study using endothelial-enriched single cells from the left- and right carotid artery exposed to d-flow (LCA) and stable-flow (s-flow in RCA) using the mouse partial carotid ligation (PCL) model. We find eight EC clusters along with immune cells, fibroblasts, and smooth muscle cells. Analyses of marker genes, pathways, and pseudotime reveal that ECs are highly heterogeneous and plastic. D-flow induces a dramatic transition of ECs from atheroprotective phenotypes to pro-inflammatory cells, mesenchymal (EndMT) cells, hematopoietic stem cells, endothelial stem/progenitor cells, and an unexpected immune cell-like (EndICLT) phenotypes. While confirming KLF4/KLF2 as an s-flow-sensitive transcription factor binding site, we also find those sensitive to d-flow (RELA, AP1, STAT1, and TEAD1). D-flow reprograms ECs from atheroprotective to proatherogenic phenotypes, including EndMT and potentially EndICLT.