Neonatal sepsis in low-income countries: epidemiology, diagnosis and prevention.

Introduction: Sepsis accounts for up to one-third of neonatal deaths in the world each year. The World Health Organization acknowledges neonatal sepsis as a major global health concern, and that the highest burden occurs in low- and middle-income countries (LMICs). Despite major research and clinical progress in this area, we still lack accurate diagnostic tools for neonatal sepsis, complicating the management of this condition.Areas covered: The purpose here is to review the latest data on the incidence, diagnosis, prevention, and management of neonatal sepsis in LMIC. We discuss the limitations of current diagnostic tests - including their lack of availability - and how this may influence global estimates of cases. We review the benefits of antenatal, intrapartum, and post-natal preventive measures. We briefly discuss the management, highlighting the emergence of antimicrobial resistance. Finally, we expose some high priority areas.Expert opinion: Neonatal sepsis is a challenging condition requiring a multifaceted approach to address the major diagnostic issues, but also the underlying socio-economic causes that nourish epidemic cases in LMIC. Focusing on antibiotics as a main pillar of intervention is likely to engender antimicrobial resistance, eventually hindering the appreciable gains LMICs have achieved in neonatal health outcomes.

Whole blood genome-wide transcriptome profiling and metagenomics next-generation sequencing in young infants with suspected sepsis in a low-and middle-income country: A study protocol.

Conducting collaborative and comprehensive epidemiological research on neonatal sepsis in low- and middle-income countries (LMICs) is challenging due to a lack of diagnostic tests. This prospective study protocol aims to obtain epidemiological data on bacterial sepsis in newborns and young infants at Kamuzu Central Hospital in Lilongwe, Malawi. The main goal is to determine if the use of whole blood transcriptome host immune response signatures can help in the identification of infants who have sepsis of bacterial causes. The protocol includes a detailed clinical assessment with vital sign measurements, strict aseptic blood culture protocol with state-of-the-art microbial analyses and RNA-sequencing and metagenomics evaluations of host responses and pathogens, respectively. We also discuss the directions of a brief analysis plan for RNA sequencing data. This study will provide robust epidemiological data for sepsis in neonates and young infants in a setting where sepsis confers an inordinate burden of disease.

Epididymis tuberculosis: Case report and brief review of the literature.

Tuberculosis (TB) is a secular disease caused by a bacillus, highly prevalent in Brazil. The genito-urinary tract involvement is rare, with the epididymis the most affected location. Treatment usually involves the combination of 3-4 drugs for TB for 6 months and surgery can be useful in complications.

A biosensor capable of identifying low quantities of breast cancer cells by electrical impedance spectroscopy.

Breast cancer (BC) is a malignant disease with a high prevalence worldwide. The main cause of death is not the primary tumor, but instead the spread of tumor cells to distant sites. The aim of the present study was to examine a new method for the detection of cancer cells in aqueous medium using bioimpedance spectroscopy assisted with magnetic nanoparticles (MNP's) exposure to a constant magnetic field. The spectroscopic patterns were identified for three breast cancer cell lines. Each BC cell line represents a different pathologic stage: the early stage (MCF-7), invasive phase (MDA-MB-231) and metastasis (SK-BR-3). For this purpose, bioimpedance measurements were carried out at a certain frequency range with the aid of nanoprobes, consisting of magnetic nanoparticles (MNPs) coupled to a monoclonal antibody. The antibody was specific for the predominant cell surface protein for each cell line, which was identified by using RT-qPCR and flow cytometry. Accordingly, EpCAM corresponds to MCF-7, MUC-1 to MDA-MB-231, and HER-2 to SK-BR-3. Despite their low concentrations, BC cells could be detected by impedance spectroscopy. Hence, this methodology should permit the monitoring of circulating tumor cells (CTC) and therefore help to prevent recurrences and metastatic processes during BC treatment.

Functional and histologic effects after implanting pluripotent stem cells in a murine model with sphincterotomy. / Efectos funcionales e histológicos posteriores al implante de células madre pluripotentes en un modelo murino con esfinterotomía.

INTRODUCTION AND AIMS: Fecal incontinence is a disabling condition with devastating consequences for the patients. Medical and surgical options are not very satisfactory, reason by which regenerative medicine has been considered in this field. In the present research, we analyzed functional and histologic effects after implanting pluripotent stem cells (PSCs) in a murine model with sphincterotomy. MATERIALS AND METHODS: Female Wistar rats were subjected to sphincterotomy and divided into three groups. Group 1 (control group) was treated with 300µL of balanced saline solution and group 2 (late treatment) and group 3 (early treatment) received 50,000 PSCs resuspended in 300µL of balanced saline solution. All animals were evaluated through high-resolution anorectal manometry 24hours before and after sphincterotomy and every month for three months. Finally, the rats were euthanized and histopathologic sections from the anal canal were obtained. RESULTS: All groups showed a decrease in resting anal pressure and squeeze anal pressure 24hours after sphincterotomy. At the third month, higher anal pressures in the groups treated with PSCs were detected. Regarding the histologic effects, the microscopic architecture was restored and there was a significant decrease in the inflammatory response in the groups treated with PSCs. CONCLUSION: PSCs implantation improves anal tone, as well as histologic structure, presenting better regenerative results when implanted as early treatment.

Electrical bioimpedance spectroscopy as biosensor technique to identify cells lineages and cell differentiation process.

The identification and characterization of diverse cells types and cell differentiation process requires complex techniques as flow cytometry, immunocytochemistry and the exploration of molecular markers; such techniques require infrastructure and qualified personnel. The objective of this study was to analyze the use of Electrical Bioimpedance Spectroscopy (EBIS) measurements as a non-complex alternative technique to identify populations of undifferentiated mouse Pluripotent Stem Cells (mPSCs), Mouse Embryonic Fibroblasts (MEFs) and the differentiation process from preadipocytes (3T3-L1) to mature adipocytes. EBIS measurements were compared in populations of cells which were characterized previously using microscopy. The results indicate that EBIS technique has a potential sensitivity at certain frequency range to discriminate between both evaluated cell populations and some differentiation process. Additional studies with different concentrations to evaluate quantitatively the sensitivity and specificity of the proposed technique are recommended.

Componentes Principales mediante Espectroscopia FTIR como Técnica de Caracterización Innovadora durante la Diferenciación de Células Madre Pluripotentes a Células Pancreáticas / Principal Components by FTIR Spectroscopy as Innovative Characterization Technique during Differentiation of Pluripotent Stem Cells to Pancreatic Cells

Resumen: Dos de los grandes retos en la biología de las Células Madre (CM) y la Medicina Regenerativa, son el control en la diferenciación de estas células y asegurar la pureza de las células diferenciadas, por lo que es necesario contar con técnicas rápidas, eficientes y precisas para la caracterización de CM y su diferenciación a diferentes linajes celulares. El objetivo de este trabajo fue analizar Células Madre Pluripotentes (CMP) y Células Pancreáticas Diferenciadas (CPD) mediante espectroscopía Infrarroja por Transformada de Fourier (FTIR) y Análisis de Componentes Principales (ACP). Para ello se diferenciaron CMP a CPD, caracterizando el proceso de diferenciación a los días 0, 11, 17 y 21 mediante microscopía óptica y espectroscopia vibracional. Los espectros FTIR se analizaron con el método multivariado de ACP, utilizando su segunda derivada en las regiones de proteínas, carbohidratos y ribosas. Los resultados indican que el ACP permite caracterizar y discriminar CMP y CPD en sus diferentes etapas de diferenciación en las regiones espectrales analizadas. Con lo anterior concluimos que el ACP permite caracterizar química y estructuralmente CMP y diferentes etapas de su diferenciación en una forma rápida, precisa y no invasiva.

Abstract: Two of the greatest challenges in Stem Cells (SCs) biology and regenerative medicine, are differentiation control of SCs and ensuring the purity of differentiated cells. In this sense, fast, efficient and accurate techniques for SCs characterization and their differentiation into different cell lineages are needed. The aim of this study was to analyse Pluripotent Stem Cells (PSCs) and Differentiated Pancreatic Cells (DPCs) by Fourier Transform Infrared (FTIR) spectroscopy and Principal Component Analysis (PCA). For this purpose, we differentiated PSCs toward DPCs, characterizing the differentiation process at different stages (0, 11, 17 and 21 days) through light microscopy and vibrational spectroscopy. FTIR spectra were analysed with the multivariate method of PCA, using the second derivatives in the protein, carbohydrate and ribose regions. The results indicate that the PCA allows to characterize and discriminate PSCs and DPCs at different stages of differentiation in the analysed spectral regions. From these results, we concluded that the PCA allows the chemically and structural characterization of PSCs and the different stages of their differentiation in a fast, accurate and non-invasive way.

Effect of resveratrol on mitochondrial activity in differentiated mature adipocytes / Efecto del resveratrol sobre la actividad mitocondrial en adipocitos maduros diferenciados

Obesity has reached epidemic proportions, the World Health Organization (WHO) estimates that there are more than 1,000 million overweight adults world-wide. Furthermore, obesity is characterized as an overgrowth of white adipose tissue as a result of adipocyte hypertrophy and hyperplasia. Mitochondria is considered the source of energy within the adipocyte, since it contains the molecular machinery, and it is involved in a large number of metabolic pathways, besides the transformation of chemical energy into adenosine triphosphate. Mitochondria shortage and adipocyte dysfunction result in an excessive accumulation of triacylglycerol in the cytoplasm, which determines an imbalance between energy production and energy expenditure. Resveratrol (RSV) is a polyphenol found in different plants and its effects have been associated with mitochondrial biogenesis. An adipogenesis in vitro model (3T3-L1 preadipocytes) was used, and these cells were differentiated into mature adipocytes. Subsequently the effect of RSV on the adipocytes morphology, the lipid content and mitochondrial activity was evaluated using microscopic and flow cytometry techniques. The effect of RSV on differentiated mature adipocytes, was characterized by the decrease in lipid content and the consequently declination of the mitochondrial activity. 3T3-L1 preadipocytes retained the differentiation ability until passage 18. The RSV at doses of 25 and 50 µM for 48 hours in differentiated mature adipocytes promoted the decreased in lipid content probably due to an increase in mitochondrial activity in the early hours of RSV exposure, causing the consequently declination of mitochondrial activity at the end of 48 hours.

La obesidad ha tomado dimensiones epidémicas globales y la Organización Mundial de la Salud estima que hay más de 1,000 millones de adultos con sobrepeso. Así mismo, la obesidad se ha caracterizado como la expansión del tejido adiposo blanco condicionada por la hipertrofia e/o hiperplasia de los adipocitos. La mitocondria es considerada la fuente de energía dentro del adipocito, debido a que contiene la maquinaria molecular que dirige, a través de diversas vías metabólicas, la transformación de la energía química en adenosíntrifosfato. La escasez de mitocondrias así como su disfunción en el adipocito, resulta en una acumulación excesiva de triacilgliceroles en el citoplasma, lo que condiciona un desequilibrio entre producción de energía y gasto energético. El resveratrol (RSV) es un polifenol que se encuentra en diferentes grupos de plantas y sus efectos se han asociado con la inducción de genes para la biogénesis mitocondrial. Se empleó un modelo de adipogénesis (in vitro) materializado por una línea celular de preadipocitos 3T3-L1, mismos que se diferenciaron a adipocitos maduros. Posteriormente se evaluó el efecto del RSV sobre la morfología, contenido lipídico y actividad mitocondrial en los adipocitos maduros diferenciados a través de las técnicas: microscopía invertida, confocal y citometría de flujo. El efecto del RSV sobre los adipocitos maduros diferenciados, se caracterizó por la disminución del contenido lipídico y consecuentemente de la actividad mitocondrial. Los preadipocitos 3T3-L1 conservaron la capacidad de diferenciación hasta el pase 18. Por otra parte, el resveratrol a dosis de 25 y 50 µM durante 48 horas en adipocitos maduros diferenciados, promueve una disminución en el contenido lipídico probablemente debido a un aumento de la actividad mitocondrial en las primeras horas de exposición al tratamiento, provocando la disminución de la actividad mitocondrial al término de 48 horas.

Caracterización morfológica, génica y protéica en la diferenciación de células madre embrionarias de ratón a células pancreáticas tempranas / Morphological, genetic and protein characterization in the differentiation of embryonic stem cells to early pancreatic cells

Debido al auge de la medicina regenerativa, las Células Madre (SC) representan una fuente de reemplazo celular para cualquier tejido, decidiendo emprender este trabajo de investigación con el objetivo de diferenciar células madre embrionarias de ratón (mESC) a células pancreáticas tempranas, realizando su caracterización génica y morfológica. Primeramente se cultivaron y arrestaron en su ciclo celular fibroblastos embrionarios de ratón (MEF) con mitomicina, posteriormente se expandieron las mESC y se sometieron a un protocolo de diferenciación de 21 días hacía células pancreáticas tempranas, evaluándose durante la diferenciación su morfología y expresión relativa de los genes sox-17, pdx-1, ins-1 e ins-2, determinando además la producción de las proteínas insulina y glucagón mediante inmunocitoquímica y citometría de flujo. Se obtuvieron cuerpos embrionarios (EBs) a partir de mESC, con características morfológicas diferentes de acuerdo a su diferenciación, los cuales expresaron genes de la línea germinal endodérmica (sox-17 y pdx-1) a los días 0, 11 y 17 de diferenciación, gen inductor del desarrollo embrionario pancreático (pdx-1) al día 11 de diferenciación y, genes de expresión pancreática (ins-1 e ins-2) a los días 17 y 21 de diferenciación. Finalmente se detectó la producción de proteínas insulina y glucagón en los EBs al día 21 de diferenciación. Se logró diferenciar mESC. El análisis morfológico evidenció cúmulos celulares tridimensionales correspondientes a EBs. Con el análisis de los patrones de expresión génica, se distinguieron inicialmente células con características genéticas de endodermo y posteriormente a partir del día 17 células pancreáticas tempranas, las cuales al día 21 de diferenciación expresaron las proteínas insulina y glucagón...

Due to the boom in regenerative medicine, Stem Cells (SC) represent a source of cell replacement to any tissue, we decided to undertake this research with the objective of differentiating mouse embryonic stem cells (mESC) to early pancreatic cells, developing their genetic and morphological characterization. Initially Mouse embryonic fibroblasts (MEF) were grown and arrested in their cell cycle with mitomycin, subsequently mouse embryonic SC (mESC) were expanded and subjected in to a pancreatic cell differentiation protocol of 21 days. During differentiation, morphology and the relative expression of sox-17, pdx-1, Ins-1 and Ins-2 genes were assessed, also the production of insulin and glucagon proteins was determinated by fluorescence microscopy and flow cytometry. Embryoid bodies (EBs) were obtained from mESC, with different morphological characteristics according to their differentiation, which expressed endodermal germ line genes (sox-17 y pdx-1) at days 0, 11 and 17 of differentiation, an inductor gene of embryonic pancreas development (pdx-1) was detected at day 11 of differentiation. Pancreas genes (ins-1 e ins-2) were expressed at day 17 and 21 of differentiation. Finally the production of insulin and glucagon proteins was detected on the EBS at day 21 of differentiation. In conclusion, the mESC differentiation was achieved. The morphological analysis evidenced three-dimensional cell clusters corresponding to EBs. Analysis of the gene expression patterns in the differentiation process, cells initially showed genetic characteristics of endoderm and thereafter from day 17 of differentiation characteristics of early pancreatic cells which by day 21 of differentiation expressed insulin and glucagon proteins...

In vitro anti-rotavirus activity of some medicinal plants used in Brazil against diarrhea.

Acute diarrhea, especially in children, is a very common disease with worldwide distribution and with a significant public health impact. Rotaviruses have been recognized as the major agents of diarrhea in infants and young children in developed as well as developing countries. In Brazil, diarrhea is one of the principal causes of death, mainly in the infant population. To fight diarrhea, traditional Brazilian medicine uses a great variety of plants. In this work, 12 medicinal plant species were screened for simian (SA-11) and human (HCR3) rotaviruses inhibition in vitro. At non-cytotoxic concentrations, the extracts from Artocarpus integrifolia L. (Moraceae) bark (480 microg/ml) and Spondias lutea L. (Anacardiaceae) leaves (160 microg/ml) had antiviral activity against both viruses. They showed inhibition of 99.2% and 97%, respectively, for human rotavirus, and 96.4% and 96.2% for simian rotavirus. The extracts from Myristica fragrans Houtt (Myristicaceae) seeds (160 microg/ml) and Spongias lutea bark (40 microg/ml) inhibited human rotavirus (90% and 82.2% inhibition, respectively), whereas the extracts from Anacardium occidentale L. (Anacardiaceae) leaves (4 microg/ml) and Psidium guajava L. (Myrtaceae) leaves (8 microg/ml) showed activity only against simian rotavirus (82.2% and 93.8% inhibition, respectively). Our results indicate that the extracts of Artocarpus integrifolia, Myristica fragrans and Spongias lutea can be useful in the treatment of human diarrhea if the etiologic agent is a rotavirus.

A sulphated fucan from the Laminaria abyssalis inhibits the human T cell lymphotropic virus type 1-induced syncytium formation in HeLa cells.

This work evaluated the effect of a sulphated fucan extracted from the Laminaria abyssalis marine algae on the human T cell lymphotropic virus type 1 (HTLV-1)-induced syncytium formation. The experiments were carried out in HeLa cells cocultured with a HTLV-1-infected T cell line (C91/PL cells) in the presence of the sulphated polysaccharide at concentration below that corresponding to the ED50. The sulphated fucan inhibited almost 100% of the syncytium formation at concentration of 100 microg/mI and was still active (>95%) at a concentration of 25 microg/ml. It was also observed that the best inhibition occurred when the compound was added in the first 2 h of the cell-to-cell contact. This is the first report showing that a purified sulphated polysaccharide, extracted from marine algae, is able to inhibit the cell-to-cell contact essential for the spreading of the HTLV-1.

Anti-herpes simplex virus effect of a seed extract from the tropical plant Licania tomentosa (Benth.) Fritsch (Chrysobalanaceae).

Incubation of acyclovir-resistant herpes simplex virus type 1 (ACVr-HSV1), during infection of the HEp-2 cell culture, with an extract prepared from the seeds of Licania tomentosa (Benth.) Fritsch (Chrysobalanaceae) species impaired the productive replication of this virus in a concentration-dependent manner. The extract was able to inhibit extracellular virus (virucidal effect) and also interfered with a very early event of cell infection, at a non-cytotoxic concentration.

In vitro antiviral effect of flavonoid-rich extracts of Vitex polygama (Verbenaceae) against acyclovir-resistant herpes simplex virus type 1.

Extracts and fractions rich in flavonoids from fruits and leaves of Vitex polygama Cham. (Verbenaceae) were tested against acyclovir-resistant herpes simplex virus type 1 (ACV-HSV-1). Both fruit and leaf extracts exhibited a dose-dependent antiviral activity. The extract from the leaves showed intracellular antiviral activity while the extract from the fruits had virucidal effect. A fraction from the ethyl actetate extract of the leaves inhibited virus propagation by blocking HEp-2 cell receptors.

In vitro activity of extracts of Persea americana leaves on acyclovir-resistant and phosphonoacetic resistant Herpes simplex virus.

The lyophilized aqueous crude extract (LACE) from leaves of Persea americana (Lauraceae) species showed a strong inhibitory effect on acyclovir (ACG(r)4 and dlsp TK mutants) and PAA-resistant (PAA(r)5 mutant) herpes simplex virus. After exhaustive washing of LACE using methanol, the soluble fraction was chromatographed on a reverse-phase column giving 11 fractions that were revealed by thin-layer chromatography. Analysis of the antiviral effect of the fractions showed the extract contained compounds that were able to inhibit extracellular virus and the replication of resistant acyclovir HSV. The virucidal effect was concentrated from fraction 4 up to 8. Fraction 7 mainly contains the flavonoid isoquercitrin, and fraction 8 the flavonoid quercitrin. The flavonoid afzelin that is the major substance present in the fraction 9 showed virustatic effect with no virucidal effect. These results show P. americana is a potential plant extract for treatment of herpes simplex virus infections either alone or associated with acyclovir.