Unnexins: Homologs of innexin proteins in Trypanosomatidae parasites.

Large-pore channels, including those formed by connexin, pannexin, innexin proteins, are part of a broad family of plasma membrane channels found in vertebrates and invertebrates, which share topology features. Despite their relevance in parasitic diseases such as Chagas and malaria, it was unknown whether these large-pore channels are present in unicellular organisms. We identified 14 putative proteins in Trypanosomatidae parasites as presumptive homologs of innexin proteins. All proteins possess the canonical motif of the innexin family, a pentapeptide YYQWV, and 10 of them share a classical membrane topology of large-pore channels. A sequence similarity network analysis confirmed their closeness to innexin proteins. A bioinformatic model showed that a homolog of Trypanosoma cruzi (T. cruzi) could presumptively form a stable octamer channel with a highly positive electrostatic potential in the internal cavities and extracellular entrance due to the notable predominance of residues such as Arg or Lys. In vitro dye uptake assays showed that divalent cations-free solution increases YO-PRO-1 uptake and hyperosmotic stress increases DAPI uptake in epimastigotes of T. cruzi. Those effects were sensitive to probenecid. Furthermore, probenecid reduced the proliferation and transformation of T. cruzi. Moreover, probenecid or carbenoxolone increased the parasite sensitivity to antiparasitic drugs commonly used in therapy against Chagas. Our study suggests the existence of innexin homologs in unicellular organisms, which could be protein subunits of new large-pore channels in unicellular organisms.

A Dye Uptake Assay to Measure Large-Pore Channel Activity in Trypanosoma cruzi.

Large-pore channels allow the exchange of ions and molecules between the intra- and extracellular compartments. These channels are structures formed by several protein families with little or no evolutionary linkages that include connexins (Cxs), pannexins (Panxs), innexins (Inxs), CALHM1, and LRRC8 proteins. Recently, we have described the unnexins (Unxs) proteins expressed in Trypanosoma cruzi (T. cruzi) that also is like to form large-pore channels at the plasma membrane. In this chapter, we describe a dye uptake method for evaluating the unnexin-formed channel function in T. cruzi, as well as the methods for evaluating their participation in the transformation of trypomastigotes into amastigotes. These methods can facilitate understanding the role of large-pore channels in the parasite's biology.

Unraveling the Etiology of Dilated Cardiomyopathy through Differential miRNA-mRNA Interactome.

Dilated cardiomyopathy (DCM) encompasses various acquired or genetic diseases sharing a common phenotype. The understanding of pathogenetic mechanisms and the determination of the functional effects of each etiology may allow for tailoring different therapeutic strategies. MicroRNAs (miRNAs) have emerged as key regulators in cardiovascular diseases, including DCM. However, their specific roles in different DCM etiologies remain elusive. Here, we applied mRNA-seq and miRNA-seq to identify the gene and miRNA signature from myocardial biopsies from four patients with DCM caused by volume overload (VCM) and four with ischemic DCM (ICM). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were used for differentially expressed genes (DEGs). The miRNA-mRNA interactions were identified by Pearson correlation analysis and miRNA target-prediction programs. mRNA-seq and miRNA-seq were validated by qRT-PCR and miRNA-mRNA interactions were validated by luciferase assays. We found 112 mRNAs and five miRNAs dysregulated in VCM vs. ICM. DEGs were positively enriched for pathways related to the extracellular matrix (ECM), mitochondrial respiration, cardiac muscle contraction, and fatty acid metabolism in VCM vs. ICM and negatively enriched for immune-response-related pathways, JAK-STAT, and NF-kappa B signaling. We identified four pairs of negatively correlated miRNA-mRNA: miR-218-5p-DDX6, miR-218-5p-TTC39C, miR-218-5p-SEMA4A, and miR-494-3p-SGMS2. Our study revealed novel miRNA-mRNA interaction networks and signaling pathways for VCM and ICM, providing novel insights into the development of these DCM etiologies.

Utilidad de la medición del tiempo de relajación isométrica del ventrículo izquierdo con eco/Doppler durante la administración de dipiridamol o dobutamina como maniobra provocadora de isquemia miocárdica / Utility of the measurement of the isovolumetric relaxation time of the left ventricle with echo/Doppler during the administration of dipyridamole or dobutamine as triggering maneuver of myocardial ischemia

La isquemia miocárdica prolongada la relajación ventricular. El objetivo del estudio fue valorar las alteraciones en el tiempo de relajación isométrica del ventrículo izquierdo (TRIV), como parámetro de relajación ventricular global, medido con Doppler pulsado durante la administración de Dipiridamol o Dobutamina intravenosos. Estudiamos 58 pacientes con sospecha clínica o con cardiopatía isquémica demostrada, durante la administración de fármacos como maniobra provocadora de isquemia. Se dividieron en dos grupos: 22 pacientes en el grupo Dipiridamol, que se administró a dosis de 0.84 mg/kg en 10 min y 36 pacientes en el grupo de Dobutamina, administrada a dosis de 5, 10, 20, 30 y 40 mcg/kg/min en etapas de tres minutos. A todos los pacientes se les practicó coronariografía en el mismo internamiento. Las mediciones de las velocidades máximas E y A, así como el tiempio de desaceleración de la onda E y el tiempo de hemipresión del flujo mitral, no mostraron cambios significativos en ambos grupos. En los estudios positivos por criterios de alteración de la movilidad parietal, el TRIVI corregido para la frecuencia cardiaca (TRIVI/C) se incrementó hasta en 54 por ciento (p< 0.01), sobre los valores de control del mismo paciente en el grupo de Dipiridamol. En el grupo de Dobutamina, con los mismos criterios de positividad, el TRIVI/C se incrementó en 26 por ciento (p < 0.20). En la detección de obstrucción significativa proximal de la coronaria descendente anterior o de enfermedad trivascular, en el grupo de Dipiridamol, el incremento del TRIVI/C tuvo sensibilidad, el incremento del TRIVI/C tuvo sensibilidad (S), especificidad (E) y valor predictivo positivo (VPP) de 50 por ciento y 100 por ciento, respectivamente. En el grupo de Dobutamina, la S fue de 71 por ciento, la E de 60 por ciento y el VPP de 89 por ciento. Con ninguno de los fármacos se observó prolongación significativa del TRIVI/C en ausensia de alteraciones de la movilidad o sin acentuación de las alteraciones preexistentes. La medición del TRIVI/C, en estudios con maniobras farmacológicas provocadoras de isquemia, es un parámetro útil para diferenciar los resultados positivos de los negativos, agregado a los criterios de alteraciones segmentarias de la movilidad parietal