Validating methods for testing natural molecules on molecular pathways of interest in silico and in vitro

Differentially expressed genes can serve as drug targets and are used to predict drug response and disease progression. In silico drug analysis based on the expression of these genetic biomarkers allows the detection of putative therapeutic agents, which could be used to reverse a pathological gene expression signature. Indeed, a set of bioinformatics tools can increase the accuracy of drug discovery, helping in biomarker identification. Once a drug target is identified, in vitro cell line models of disease are used to evaluate and validate the therapeutic potential of putative drugs and novel natural molecules. This study describes the development of efficacious PCR primers that can be used to identify gene expression of specific genetic pathways, which can lead to the identification of natural molecules as therapeutic agents in specific molecular pathways. For this study, genes involved in health conditions and processes were considered. In particular, the expression of genes involved in obesity, xenobiotics metabolism, endocannabinoid pathway, leukotriene B4 metabolism and signaling, inflammation, endocytosis, hypoxia, lifespan, and neurotrophins were evaluated. Exploiting the expression of specific genes in different cell lines can be useful in in vitro to evaluate the therapeutic effects of small natural molecules

Polyphenols and Lactobacillus reuteri in oral health

Oral health is one of the necessary preludes to the overall quality of life. Several medical procedures and therapies are available to treat oral diseases in general and periodontal diseases in particular, yet caries, periodontitis, oral cancer, and oral infections remain a global concern. Natural molecules, with their anti-oxidant, anti-inflammatory, and anti-microbic properties, are one of the main sources of oral health and dental health care, and should be supplemented to exploit their beneficial effects. A possible way to improve the intake of these molecules is adhering to a diet that is rich in fruits, vegetables, and probiotics, which has many beneficial properties and can improve overall health and wellbeing. The Mediterranean diet, in particular, provides several beneficial natural molecules, mainly because of the precious nutrients contained in its typical ingredients, mainly plant-based (olives, wine, citrus fruits, and many more). Its beneficial effects on several diseases and in increasing the overall wellbeing of the population are currently being studied by physicians. Among its nutrients, polyphenols (including, among other molecules, lignans, tannins, and flavonoids) seem to be of outmost importance: several studies showed their anticariogenic properties, as well as their effects in decreasing the incidence of non-communicable diseases. Therefore, plant-derived molecules – such as polyphenols – and probiotics – such as Lactobacillus reuteri – have shown a significant potential in treating and curing oral diseases, either alone or in combination, owing to their antioxidant and antimicrobial properties, respectively

Dietary supplements for the management of COVID-19 symptoms

SARS-CoV-2, the etiological agent of COVID-19, caused a pandemic in 2020, which is only recently slowing down. The symptoms of COVID-19 range from cough to fever and pneumonia and may persist beyond the active state of the infection, in a condition called post-COVID syndrome. The aim of this paper is to review the relationship between COVID-19 and nutrition and to discuss to most up-to-date dietary supplements proposed for COVID-19 treatment and prevention. Nutrition and nutritional dysregulations, such as obesity and malnutrition, are prominent risk factors for severe COVID-19. These factors exert anti-inflammatory and proinflammatory effects on the immune system, thus exacerbating or reducing the immunological response against the virus. As for the nutritional habits, the Western diet induces a chronic inflammatory state, whereas the Mediterranean diet exerts anti-inflammatory effects and has been proposed for ameliorating COVID-19 evolution and symptoms. Several vaccines have been researched and commercialized for COVID-19 prevention, whereas several drugs, although clinically tested, have not shown promising effects. To compensate for the lack of treatment, several supplements have been recommended for preventing or ameliorating COVID-19 symptoms. Thus, it is critical to review the dietary supplements proposed for COVID-19 treatment. Supplements containing α-cyclodextrin and hydroxytyrosol exhibited promising effects in several clinical trials and reduced the severity of the outcomes and the duration of the infection. Moreover, a supplement containing hydroxytyrosol, acetyl L-carnitine, and vitamins B, C, and D improved the symptoms of patients with post-COVID syndrome

Red Giant stars in the Large Magellanic Cloud clusters

We present deep J,H,Ks photometry and accurate Color Magnitude Diagrams down to K ~18.5, for a sample of 13 globular clusters in the Large Magellanic Cloud. This data set combined with the previous sample of 6 clusters published by our group gives the opportunity to study the properties of giant stars in clusters with different ages (ranging from ~80 Myr up to ~3.5 Gyr). Quantitative estimates of star population ratios (by number and luminosity) in the Asymptotic Giant Branch, the Red Giant Branch and the He-clump, have been obtained and compared with theoretical models in the framework of probing the so-called phase transitions. The AGB contribution to the total luminosity starts to be significant at ~200 Myr and reaches its maximum at ~5-600 Myr, when the RGB Phase Transition is starting. At ~900 Myr the full developing of an extended and well populated RGB has been completed. Both the occurrence of the AGB and RGB Phase Transitions are sharp events, lasting a few hundreds Myr only. These empirical results agree very well with the theoretical predictions of simple stellar population models based on canonical tracks and the fuel-consumption approach.Comment: 32 pages, 11 figures, accepted to Ap

Polymorphisms, diet and nutrigenomics

Every human being possesses an exclusive nutritional blueprint inside their genes. Bioactive food components and nutrients affect the expression of such genes. Nutrigenomics is the  science that analyzes gene-nutrient interactions (nutrigenetics), which can lead to the development of personalized nutritional recommendations to maintain optimal health and prevent disease. Genomic diversity among various ethnic groups might affect nutrients bioavailability as well as their metabolism. Nutrigenomics combines different branches of science including nutrition, bioinformatics, genomics, molecular biology, molecular medicine, and epidemiology. Genes regulate intake and metabolism of different nutrients, while nutrients positively or negatively influence the expression of a number of genes; testing of specific genetic polymorphisms may therefore become a useful tool to manage weight loss and to fully understand gene-nutrient interactions. Indeed, several approaches are used to study gene-nutrient interactions: epigenetics, the study of genome modification not related to changes in nucleotide sequence; transcriptomics, the study of tissue-specific and time-specific RNA transcripts; proteomics, the study of proteins involved in biological processes; and metabolomics, the study of changes of primary and secondary metabolites in body fluids and tissues. Hence, the use of nutrigenomics to improve and optimize a healthy, balanced diet in clinical settings could be an effective approach for long-term lifestyle changes that might lead to consistent weight loss and improve quality of life

Dietary supplements for improving nitric-oxide synthesis

Nitric oxide (NO) is an essential component of the human body, involved in blood vessel dilation, stimulation of hormone release, signaling and regulation of neurotransmission. Nitric oxide is synthesized by nitric-oxide-synthase-dependent and -independent pathways. Nitric oxide supplementation improves cardiac health, enhances performance during exercise, reduces high blood pressure during pregnancy, reduces erectile dysfunction and improves healing processes and respiratory response. Nitric-oxide-associated benefits are mostly apparent in untrained or moderately trained individuals. L-arginine and L-citrulline supplementation contributes to nitric oxide levels because L-arginine is directly involved in NO synthesis, whereas L-citrulline acts as an L-arginine precursor that is further converted to NO by a reaction catalyzed by NO synthase. L-arginine supplements increase respiratory response and enhance performance during exercise, while L-citrulline with malate and other molecules increase working capacity. Various studies involving beetroot juice have reported a significant increase in plasma nitrite levels, regarded as markers of NO, after intake of beetroot juice. Although NO supplementation may have mild to moderate side-effects, using smaller or divided doses could avoid some of these side-effects. Since nitric oxide supplementation may worsen certain health conditions and may interfere with certain medicines, it should only be taken under medical supervision

Cross-evaluation of the therapeutical methods for idiopathic congenital clubfoot (talipes equinovarus): controversies regarding the tendocalcaneous tenotomy

OBJECTIVE: There has been a lot of discussion regarding the treatment of congenital clubfoot (talipes equinovarus,) and Posenti's methodology currently seems to be the most rational, offering high rates of satisfactory results when compared to Kite's approach that prevailed in orthopedics until the end of the 90s. With the recent change of concepts, this study purports to analyze the profile of orthopedists treating this infirmity in Brazil, through a questionnaire used at the 39th Brazilian Congress of Orthopaedics and Traumatology, since such data is unknown in the Brazilian literature. METHODS: An investigative questionnaire was prepared to ascertain the treatment method used, the characteristics of the population studied, the results acquired with treatment and, particularly, how they approached the Achilles tendon. RESULTS: Of the 5,329 registered orthopedists, we acquired 539 spontaneous participations. Of these, only 88 (16.30%) orthopedists perform the treatment for congenital clubfoot; 78 (88.60%) use the Ponseti method and 9 (10.20%) use Kite's; for 47.70%, conservative treatment is performed between 4 and 6 months and for 35.30%, between 1 and 3 months; 58 (66.00%) interviewees perform the Achilles tendon tenotomy in 80% to 100% of their patients and 59 (67.05%) perform it at a surgical center due to safety conditions, sterilized environment, anesthesia, ease of access, and patient monitoring; 32 (36.36%) orthopedists present 80% of good results or more, 54 (61.36%) present 50% to 80% good results and 46 (52.27%) present a 10% relapse rate. CONCLUSIONS: Although the Ponseti Method defines that the Achilles tendon tenotomy should be performed in an outpatient setting, most of the orthopedists (59 - 67.05%) perform it in the operating room.OBJETIVO: Há muita discussão em relação ao tratamento do pé torto congênito equino-cavo-varo e atualmente a sistemática proposta por Ponseti parece ser a mais racional oferecendo altas taxas de resultados satisfatórios em detrimento à metodologia de Kite que predominou no meio ortopédico até o final da década de 90. Com a recente mudança de conceitos, este trabalho tem o intuito de analisar, por meio de questionário aplicado no 39° Congresso Brasileiro de Ortopedia e Traumatologia, o perfil dos ortopedistas no Brasil frente a esta afecção, fato este desconhecido considerando a literatura nacional. MÉTODO: Um questionário para investigação foi elaborado para verificar o método de tratamento utilizado, as características da população estudada, os resultados obtidos com o tratamento e particularmente como abordam cirurgicamente o tendão calcâneo. RESULTADOS: Do total de 5.329 ortopedistas inscritos, obtivemos 539 participações espontâneas; destes, somente 88 (16,30%) indivíduos executam o tratamento para o pé torto congênito; 78 (88,60%) aplicam o método de Ponseti; e nove (10,20%) o de Kite. Para 47,70%, o tratamento conservador é realizado entre quatro e seis meses e para 35,20% entre um e três meses; 58 (66,00%) entrevistados efetuam a tenotomia do tendão calcâneo em 80% a 100% de seus pacientes e 59 (67,05%) a realizam no centro cirúrgico devido às condições de segurança, assepsia, anestesia, facilidade de acesso e monitorização do paciente; 32 (36,36%) ortopedistas apresentam 80% ou mais de bons resultados, 54 (61,36%) apresentam 50% a 80% de bons resultados e 46 (52,27%) apresentam taxa de recidiva de 10%. CONCLUSÃO: Embora o método de Ponseti defina que a tenotomia do tendão calcâneo seja realizada ambulatorialmente com anestesia local, a maioria 59 (67,05%) dos ortopedistas a realiza no centro cirúrgico.Universidade Federal de São Paulo (UNIFESP) Departamento de Ortopedia e TraumatologiaUniversidade Federal de São Paulo (UNIFESP) Departamento de Ortopedia e Traumatologia Ortopedia PediátricaUNIFESP, Depto. de Ortopedia e TraumatologiaUNIFESP, Depto. de Ortopedia e Traumatologia Ortopedia PediátricaSciEL

Main nutritional deficiencies

Nutrition is the source of energy that is required to carry out all the processes of human body. A balanced diet is a combination of both macro- and micronutrients. “Nutritional inadequacy” involves an intake of nutrients that is lower than the estimated average requirement, whereas “nutritional deficiency” consists of severely reduced levels of one or more nutrients, making the body unable to normally perform its functions and thus leading to an increased risk of several diseases like cancer, diabetes, and heart disease. Malnutrition could be caused by environmental factors, like food scarcity, as well as disease conditions, like anorexia nervosa, fasting, swallowing inability, persistent vomiting, impaired digestion, intestinal malabsorption, or other chronic diseases. Nutritional biomarkers – like serum or plasma levels of nutrients such as folate, vitamin C, B vitamins, vitamin D, selenium, copper, zinc – could be used for the evaluation of nutrient intake and dietary exposure. Macronutrients deficiencies could cause kwashiorkor, marasmus, ketosis, growth retardation, wound healing, and increased infection susceptibility, whereas micronutrient – like iron, folate, zinc, iodine, and vitamin A – deficiencies lead to intellectual impairment, poor growth, perinatal complications, degenerative diseases associated with aging and higher morbidity and mortality. Preventing macro- and micronutrient deficiency is crucial and this could be achieved through supplementation and food-based approache

Optimization of long-range PCR protocol to prepare filaggrin exon 3 libraries for PacBio long-read sequencing

BackgroundThe filaggrin (FLG) protein, encoded by the FLG gene, is an intermediate filament-associated protein that plays a crucial role in the terminal stages of human epidermal differentiation. Loss-of-function mutations in the FLG exon 3 have been associated with skin diseases. The identification of causative mutations is challenging, due to the high sequence homology within its exon 3 (12,753 bp), which includes 10 to 12 filaggrin tandem repeats. With this study we aimed to obtain the whole FLG exon 3 sequence through PacBio technology, once 13-kb amplicons have been generated.Methods and resultsFor the preparation of SMRTbell libraries to be sequenced using PacBio technology, we focused on optimizing a 2-step long-range PCR protocol to generate 13-kb amplicons covering the whole FLG exon 3 sequence. The performance of three long-range DNA polymerases was assessed in an attempt to improve the PCR conditions required for the enzymes to function properly. We focused on optimization of the input template DNA concentration and thermocycling parameters to correctly amplify the entire FLG exon 3 sequence, minimizing non-specific amplification.ConclusionsTaken together, our findings suggested that the PrimeSTAR protocol is suitable for producing the amplicons of the 13-kb FLG whole exon 3 to prepare SMRTbell libraries. We suggest that sequencing the generated amplicons may be useful for identifying LoF variants that are causative of the patients' disorders

Clinical assessment for diet prescription

Accurate nutritional assessment based on dietary intake, physical activity, genetic makeup, and metabolites is required to prevent from developing and/or to treat people suffering from malnutrition as well as other nutrition related health issues. Nutritional screening ought to be considered as an essential part of clinical assessment for every patient on admission to healthcare setups, as well as on change in clinical conditions. Therefore, a detailed nutritional assessment must be performed every time nutritional imbalances are observed or suspected. In this review we have explored different techniques used for nutritional and physical activity assessment.  Dietary Intake (DI) assessment is a multidimensional and complex process. Traditionally, dietary intake is assessed through self-report techniques, but due to limitations like biases, random errors, misestimations, and nutrient databases-linked errors, questions arise about the adequacy of self-reporting dietary intake procedures. Despite the limitations in assessing dietary intake (DI) and physical activity (PA), new methods and improved technologies such as biomarkers analysis, blood tests, genetic assessments, metabolomic analysis, DEXA (Dual-energy X-ray absorptiometry), MRI (Magnetic resonance imaging), and CT (computed tomography) scanning procedures have made much progress in the improvement of these measures. Genes also plays a crucial role in dietary intake and physical activity. Similarly, metabolites are also involved in different nutritional pathways. This is why integrating knowledge about the genetic and metabolic markers along with the latest technologies for dietary intake (DI) and physical activity (PA) assessment holds the key for accurately assessing one’s nutritional status and prevent malnutrition and its related complications