Biomarcadores en fluídos biológicos y su potencial uso como indicadores de nefritis lúpica en individuos con lupus eritematoso sistémico

Lupus nephritis is one of the most severe manifestations of systemic lupus erythematous. Renal involvement in patients with systemic lupus erythematous is an important cause of morbidity and mortality. The pathogenesis of lupus nephritis involve multiple factors, wich include genetic predisposition, epigenetic regulation and environmental interaction. Conventional clinical parameters such as creatinine clearance, proteinuria, urinary sediments, antibodies anti-double- strand DNA and complement proteins they are not enough sensitive or specific to detect disease activity. In the last decades, “Omics” technologies (Proteomic, genomic, transcriptomic, metobolomic) have been used in an extensive way looking for biomarkers, which allowed to discovery variants associated with systemic lupus erythematous and lupus nephritis. Such findings have expanded our knowledge about molecular basis of disease and they have been very important to identification of potential therapeutic targets to prediction of disease and early treatment. In this review, we resume some of recent studies focused in identification of biomarkers associated with lupus nephritis in diverse biological fluids.La nefritis lúpica es una de las manifestaciones más severas del lupus eritematoso sistémico. El compromiso renal en pacientes con lupus eritematoso sistémico es un causal importante de morbilidad y mortalidad. La patogénesis de la nefritis lúpica involucra múltiples factores, entre los que se incluyen predisposición genética, regulación epigenética e interacción ambiental. Los parámetros clínicos convencionales tales como eliminación de creatinina, proteinuria, sedimentos urinarios, anticuerpos anti-ADN de doble cadena y niveles del complemento no son lo suficientemente sensibles o específicos para detectar actividad de la enfermedad.En las últimas décadas, las técnicas basadas en “Ómicas” (proteómica, genómica, transcriptómica, metabolómica) han sido utilizadas de manera extensa para la búsqueda de biomarcadores, las cuales han permitido descubrir una amplia variedad de variantes que son asociadas con lupus eritematoso sistémico y nefritis lúpica.Esos descubrimientos han expandido nuestro entendimiento de las bases moleculares de la enfermedad y han sido muy importantes para la identificación de potenciales blancos terapéuticos para predicción de la enfermedad y tratamiento temprano. En esta revisión, resumimos algunos de los estudios recientes enfocados en la identificación de biomarcadores asociados a nefritis lúpica en diversos fluidos biológicos

Índices de infestación aedica e identificación de conocimientos, actitudes y práctica sobre dengue en llanterías del Departamento del Atlántico, Colombia

Objetivo Identificar los conocimientos, actitudes y prácticas sobre dengue en propietarios y trabajadores de llantería, así como los niveles de infestación del vector en llanterías del departamento del Atlántico.Métodos Se realizó un estudio tipo descriptivo. Las variables se describieron a partir de porcentajes y medidas de tendencia central y dispersión.Se calculó el índice de infestación larvario para llanterías y el índice de depósitos en cada uno de los municipios muestreados.Se visitaron e inspeccionaron 111 llanterías; el 26,1 % (29/111) de estas se encontraron positivas para formas larvarias del vector Ae. aegypti. Los municipios de Piojó, Santo Tomas, Santa Lucia, Sabanagrande y Luruaco presentaron los índices de infestación larvaria en llanterías más altos (IIL: 50-100 %).Resultados Con respecto al dengue, el 90,9 % de los entrevistados lo consideró un problema para ellos y sus familias. El 94,6 % conoce que es transmitido por mosquitos; el 91,1 % conoce a la larva del vector como “sarapico”, el 3,6 % como “gusarapo”. El 98,2 % consideró que existe una relación entre la larva y el mosquito Ae. aegypti. El 100 % reconoce las llantas como un criadero para el mosquito. El 85,7 % consideró la fiebre como el síntoma más frecuente, el 83% manifestó asistir al puesto de salud para curar la enfermedad. El 90,8 % arroja las llantas no utilizables como basura.Conclusión En la población estudiada existen buenos conocimientos acerca del dengue y su vector; sin embargo, existen problemas de actitudes y prácticas para su prevención

From Cell to Symptoms: The Role of SARS-CoV-2 Cytopathic Effects in the Pathogenesis of COVID-19 and Long COVID

Severe Acute Respiratory Syndrome CoronaVirus 2 (SARS-CoV-2) infection triggers var-ious events from the molecular to the tissue level, which in turn is given by the intrinsic character-istics of each patient. Given the molecular diversity characteristic of each cellular phenotype, the possible cytopathic, tissue, and clinical effects are difficult to predict, which determines the hetero-geneity of COVID-19 symptoms. The purpose of this article is to provide a comprehensive review of the cytopathic effects of SARS-CoV-2 on various cell types, focusing on the development of COVID-19, which in turn may lead, in some patients, to the persistence of symptoms after recovery from the disease, a condition known as long COVID. We describe the molecular mechanisms un-derlying virus–host interactions, including alterations in protein expression, intracellular signaling pathways, and immune responses. In particular, the article highlights the potential impact of these cytopathies on cellular function and clinical outcomes, such as immune dysregulation, neuropsy-chiatric disorders, and organ damage. The article concludes by discussing future directions for re-search and implications for the management and treatment of COVID-19 and long COVID

Integrated analysis of microRNA regulation and its interaction with mechanisms of epigenetic regulation in the etiology of systemic lupus erythematosus

The aim of this study was to identity in silico the relationships among microRNAs (miRNAs) and genes encoding transcription factors, ubiquitylation, DNA methylation, and histone modifications in systemic lupus erythematosus (SLE). To identify miRNA dysregulation in SLE, we used miR2Disease and PhenomiR for information about miRNAs exhibiting differential regulation in disease and other biological processes, and HMDD for information about experimentally supported human miRNA-disease association data from genetics, epigenetics, circulating miRNAs, and miRNA-target interactions. This information was incorporated into the miRNA analysis. High-throughput sequencing revealed circulating miRNAs associated with kidney damage in patients with SLE. As the main finding of our in silico analysis of miRNAs differentially expressed in SLE and their interactions with disease-susceptibility genes, post-translational modifications, and transcription factors; we highlight 226 miRNAs associated with genes and processes. Moreover, we highlight that alterations of miRNAs such as hsa-miR-30a-5p, hsa-miR-16-5p, hsa-miR-142-5p, and hsa-miR-324-3p are most commonly associated with post-translational modifications. In addition, altered miRNAs that are most frequently associated with susceptibility-related genes are hsa-miR-16-5p, hsa-miR-374a-5p, hsa-miR-34a-5p, hsa-miR-31-5p, and hsa-miR-1-3p

Integrated analysis of microRNA regulation and its interaction with mechanisms of epigenetic regulation in the etiology of systemic lupus erythematosus

The aim of this study was to identity in silico the relationships among microRNAs (miRNAs) and genes encoding transcription factors, ubiquitylation, DNA methylation, and histone modifications in systemic lupus erythematosus (SLE). To identify miRNA dysregulation in SLE, we used miR2Disease and PhenomiR for information about miRNAs exhibiting differential regulation in disease and other biological processes, and HMDD for information about experimentally supported human miRNA-disease association data from genetics, epigenetics, circulating miRNAs, and miRNA-target interactions. This information was incorporated into the miRNA analysis. High-throughput sequencing revealed circulating miRNAs associated with kidney damage in patients with SLE. As the main finding of our in silico analysis of miRNAs differentially expressed in SLE and their interactions with disease-susceptibility genes, post-translational modifications, and transcription factors; we highlight 226 miRNAs associated with genes and processes. Moreover, we highlight that alterations of miRNAs such as hsa-miR-30a-5p, hsa-miR-16-5p, hsa-miR-142-5p, and hsa-miR-324-3p are most commonly associated with post-translational modifications. In addition, altered miRNAs that are most frequently associated with susceptibility-related genes are hsa-miR-16-5p, hsa-miR-374a-5p, hsa-miR-34a-5p, hsa-miR-31-5p, and hsa-miR-1-3p

Effective gene delivery to Trypanosoma cruzi epimastigotes through nucleofection

New opportunities have raised to study the gene function approaches of Trypanosoma cruzi after its genome sequencing in 2005. Functional genomic approaches in Trypanosoma cruzi are challenging due to the reduced tools available for genetic manipulation, as well as to the reduced efficiency of the transient transfection conducted through conventional methods. The Amaxa nucleofector device was systematically tested in the present study in order to improve the electroporation conditions in the epimastigote forms of T. cruzi. The transfection efficiency was quantified using the green fluorescent protein (GFP) as reporter gene followed by cell survival assessment. The herein used nucleofection parameters have increased the survival rates (N90%) and the transfection efficiency by approximately 35%. The small amount of epimastigotes and DNA required for the nucleofection can turn the method adopted here into an attractive tool for high throughput screening (HTS) applications, and for gene editing in parasites where genetic manipulation tools remain relatively scarce

CREditing: a tool for gene tuning in Trypanosoma cruzi

The genetic manipulation of Trypanosoma cruzi continues to be a challenge, mainly due to the lack of available and efficient molecular tools. The CRE-lox recombination system is a site-specific recombinase technology, widely used method of achieving conditional targeted deletions, inversions, insertions, gene activation, translocation, and other modifications in chromosomal or episomal DNA. In the present study, the CRE-lox system was adapted to expand the current genetic toolbox for this hard-to-manipulate parasite. For this, evaluations of whether direct protein delivery of CRE recombinase through electroporation could improve CRE-mediated recombination in T. cruzi were performed. CRE recombinase was fused to the C-terminus of T. cruzi histone H2B, which carries the nuclear localization signal and is expressed in the prokaryotic system. The fusion protein was affinity purified and directly introduced into epimastigotes and tissue culture-derived trypomastigotes. This enabled the control of gene expression as demonstrated by turning on a tdTomato (tandem dimer fluorescent protein) reporter gene that had been previously transfected into parasites, achieving CRE-mediated recombination in up to 85% of parasites. This system was further tested for its ability to turn off gene expression, remove selectable markers integrated into the genome, and conditionally knock down the nitroreductase gene, which is involved in drug resistance. Additionally, CREditing also enabled the control of gene expression in tissue culture trypomastigotes, which are more difficult to transfect than epimastigotes. The considerable advances in genomic manipulation of T. cruzi shown in this study can be used by others to aid in the greater understanding of this parasite through gain- or loss-of function approaches

Comparative Analysis of <i>In-House</i> RT-qPCR Detection of SARS-CoV-2 for Resource-Constrained Settings

We developed and standardized an efficient and cost-effective in-house RT-PCR method to detect severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We evaluated sensitivity, specificity, and other statistical parameters by different RT-qPCR methods including triplex, duplex, and simplex assays adapted from the initial World Health Organization- (WHO) recommended protocol. This protocol included the identification of the E envelope gene (E gene; specific to the Sarvecovirus genus), RdRp gene of the RNA-dependent RNA polymerase (specific for SARS-CoV-2), and RNase P gene as endogenous control. The detection limit of the E and the RdRp genes were 3.8 copies and 33.8 copies per 1 µL of RNA, respectively, in both triplex and duplex reactions. The sensitivity for the RdRp gene in the triplex and duplex RT-qPCR tests were 98.3% and 83.1%, respectively. We showed a decrease in sensitivity for the RdRp gene by 60% when the E gene acquired Ct values > 31 in the diagnostic tests. This is associated with the specific detection limit of each gene and possible interferences in the protocol. Hence, developing efficient and cost-effective methodologies that can be adapted to various health emergency scenarios is important, especially in developing countries or settings where resources are limited

High-Throughput Sequencing Reveals Circulating miRNAs as Potential Biomarkers of Kidney Damage in Patients with Systemic Lupus Erythematosus.

Renal involvement is one of the most severe manifestations of systemic lupus erythematosus (SLE). Renal biopsy is the gold standard when it comes to knowing whether a patient has lupus nephritis, and the degree of renal disease present. However, the biopsy has various complications, bleeding being the most common. Therefore, the development of alternative, non-invasive diagnostic tests for kidney disease in patients with SLE is a priority. Micro RNAs (miRNAs) are differentially expressed in various tissues, and changes in their expression have been associated with several pathological processes. The aim of this study was to identify changes in the abundance of miRNAs in plasma samples from patients with lupus nephritis that could potentially allow the diagnosis of renal damage in SLE patients. This is an observational case-control cross-sectional study, in which we characterized the differential abundance profiles of miRNAs among patients with different degrees of lupus compared with SLE patients without renal involvement and healthy control individuals. We found 89 miRNAs with changes in their abundance between lupus nephritis patients and healthy controls, and 17 miRNAs that showed significant variations between SLE patients with or without renal involvement. Validation for qPCR of a group of miRNAs on additional samples from lupus patients with or without nephritis, and from healthy individuals, showed that five miRNAs presented an average detection sensitivity of 97%, a specificity of 70.3%, a positive predictive value of 82.5%, a negative predictive value of 96% and a diagnosis efficiency of 87.9%. These results strongly suggest that miR-221-5p, miR-380-3p, miR-556-5p, miR-758-3p and miR-3074-3p are potential diagnostic biomarkers of lupus nephritis in patients with SLE. The observed differential pattern of miRNA abundance may have functional implications in the pathophysiology of SLE renal damage

Surface-enhanced Raman Spectroscopy in urinalysis of hypertension patients with kidney disease

Abstract Arterial hypertension (AH) is a multifactorial and asymptomatic disease that affects vital organs such as the kidneys and heart. Considering its prevalence and the associated severe health repercussions, hypertension has become a disease of great relevance for public health across the globe. Conventionally, the classification of an individual as hypertensive or non-hypertensive is conducted through ambulatory blood pressure monitoring over a 24-h period. Although this method provides a reliable diagnosis, it has notable limitations, such as additional costs, intolerance experienced by some patients, and interferences derived from physical activities. Moreover, some patients with significant renal impairment may not present proteinuria. Accordingly, alternative methodologies are applied for the classification of individuals as hypertensive or non-hypertensive, such as the detection of metabolites in urine samples through liquid chromatography or mass spectrometry. However, the high cost of these techniques limits their applicability for clinical use. Consequently, an alternative methodology was developed for the detection of molecular patterns in urine collected from hypertension patients. This study generated a direct discrimination model for hypertensive and non-hypertensive individuals through the amplification of Raman signals in urine samples based on gold nanoparticles and supported by chemometric techniques such as partial least squares-discriminant analysis (PLS-DA). Specifically, 162 patient urine samples were used to create a PLS-DA model. These samples included 87 urine samples from patients diagnosed with hypertension and 75 samples from non-hypertensive volunteers. In the AH group, 35 patients were diagnosed with kidney damage and were further classified into a subgroup termed (RAH). The PLS-DA model with 4 latent variables (LV) was used to classify the hypertensive patients with external validation prediction (P) sensitivity of 86.4%, P specificity of 77.8%, and P accuracy of 82.5%. This study demonstrates the ability of surface-enhanced Raman spectroscopy to differentiate between hypertensive and non-hypertensive patients through urine samples, representing a significant advance in the detection and management of AH. Additionally, the same model was then used to discriminate only patients diagnosed with renal damage and controls with a P sensitivity of 100%, P specificity of 77.8%, and P accuracy of 82.5%