Lysosomal cholesterol accumulation sensitizes to acetaminophen hepatotoxicity by impairing mitophagy.

The role of lysosomes in acetaminophen (APAP) hepatotoxicity is poorly understood. Here, we investigated the impact of genetic and drug-induced lysosomal cholesterol (LC) accumulation in APAP hepatotoxicity. Acid sphingomyelinase (ASMase)(-/-) mice exhibit LC accumulation and higher mortality after APAP overdose compared to ASMase(+/+) littermates. ASMase(-/-) hepatocytes display lower threshold for APAP-induced cell death and defective fusion of mitochondria-containing autophagosomes with lysosomes, which decreased mitochondrial quality control. LC accumulation in ASMase(+/+) hepatocytes caused by U18666A reproduces the susceptibility of ASMase(-/-) hepatocytes to APAP and the impairment in the formation of mitochondria-containing autolysosomes. LC extraction by 25-hydroxycholesterol increased APAP-mediated mitophagy and protected ASMase(-/-) mice and hepatocytes against APAP hepatotoxicity, effects that were reversed by chloroquine to disrupt autophagy. The regulation of LC by U18666A or 25-hydroxycholesterol did not affect total cellular sphingomyelin content or its lysosomal distribution. Of relevance, amitriptyline-induced ASMase inhibition in human hepatocytes caused LC accumulation, impaired mitophagy and increased susceptibility to APAP. Similar results were observed upon glucocerebrosidase inhibition by conduritol β-epoxide, a cellular model of Gaucher disease. These findings indicate that LC accumulation determines susceptibility to APAP hepatotoxicity by modulating mitophagy, and imply that genetic or drug-mediated ASMase disruption sensitizes to APAP-induced liver injury

Repair of a Mutation Disrupting the Guinea Pig Cytomegalovirus Pentameric Complex Acquired during Fibroblast Passage Restores Pathogenesis in Immune-Suppressed Guinea Pigs and in the Context of Congenital Infection

ABSTRACT Guinea pig cytomegalovirus (GPCMV) provides a valuable model for congenital cytomegalovirus transmission. Salivary gland (SG)-passaged stocks of GPCMV are pathogenic, while tissue culture (TC) passage in fibroblasts results in attenuation. Nonpathogenic TC-derived virus N13R10 (cloned as a bacterial artificial chromosome [BAC]) has a 4-bp deletion that disrupts GP129 , which encodes a subunit of the GPCMV pentameric complex (PC) believed to govern viral entry into select cell types, and GP130 , an overlapping open reading frame (ORF) of unknown function. To determine if this deletion contributes to attenuation of N13R10, markerless gene transfer in Escherichia coli was used to construct virus r129, a variant of N13R10 in which the 4-bp deletion is repaired. Virions from r129 were found to contain GP129 as well as two other PC subunit proteins, GP131 and GP133, whereas these three PC subunits were absent from N13R10 virions. Replication of r129 in fibroblasts appeared unaltered compared to that of N13R10. However, following experimental challenge of immunocompromised guinea pigs, r129 induced significant weight loss, longer duration of viremia, and dramatically higher (up to 1.5 × 10 6 -fold) viral loads in blood and end organs compared to N13R10. In pregnant guinea pigs, challenge with doses of r129 virus of ≥5 × 10 6 PFU resulted in levels of maternal viremia, congenital transmission, pup viral loads, intrauterine growth restriction, and pup mortality comparable to that induced by pathogenic SG virus, although higher doses of r129 were required. These results suggest that the GP129-GP130 mutation is a significant contributor to attenuation of N13R10, likely by abrogating expression of a functional PC. IMPORTANCE Tissue culture adaptation of cytomegaloviruses rapidly selects for mutations, deletions, and rearrangements in the genome, particularly for viruses passaged in fibroblast cells. Some of these mutations are focused in the region of the genome encoding components of the pentameric complex (PC), in particular homologs of human cytomegalovirus (HCMV) proteins UL128, UL130, and UL131A. These mutations can attenuate the course of infection when the virus is reintroduced into animals for vaccine and pathogenesis studies. This study demonstrates that a deletion that arose during the process of tissue culture passage can be repaired, with subsequent restoration of pathogenicity, using BAC-based mutagenesis. Restoration of pathogenicity by repair of a frameshift mutation in GPCMV gene GP129 using this approach provides a valuable genetic platform for future studies using the guinea pig model of congenital CMV infection

Osteoarthritis treatment with a novel nutraceutical acetylated ligstroside aglycone, a chemically modified extra-virgin olive oil polyphenol

Recent studies have shown that dietary patterns confer protection from certain chronic diseases related to oxidative stress, the immune system and chronic low-grade inflammatory diseases. The aim of this study was to evaluate the anti-inflammatory potential and the capacity to attenuate cartilage degradation using extra-virgin olive oil–derived polyphenols for the treatment of osteoarthritis. Results show that both nutraceuticals ligstroside aglycone and acetylated ligstroside aglycone showed an anti-inflammatory profile. Acetylated ligstroside aglycone significantly reduced the expression of pro-inflammatory genes including NOS2 and MMP13 at both RNA and protein levels; decreased nitric oxide release; and, importantly, reduced proteoglycan loss in human osteoarthritis cartilage explants. Our study demonstrated that a new synthetic acetylated ligstroside aglycone derivative offers enhanced anti-inflammatory profile than the natural nutraceutical compound in osteoarthritis. These results substantiate the role of nutraceuticals in osteoarthritis with implications for therapeutic intervention and our understanding of osteoarthritis pathophysiology.España, MINECO (CTQ2016-78703-P)España, Junta de Andalucía (FQM134

Linfosarcoma bovino en Colombia: presentación de la forma adulta de la enfermedad.

En el presente estudio se describe un caso de linfosarcoma bovino, forma adulta en una vaca Holstein, y se incluyen: la historia clínica, alteraciones en hematocrito, hemoglobina, recuento leucocitario total y diferencial, fraccionamiento electroforético en las proteínas séricas y presencia de anticuerpos específicos contra el virus del linfosarcoma bovino por la prueba de inmunodifusión. Se estudió una hembra de 5 años de edad, proveniente del municipio de Sopó, Cundinamarca, Colombia, la cual permaneció en decúbito toráxico durante una semana antes de ingresar al Laboratorio de Investigaciones Médicas Veterinarias del ICA. En el laboratorio, el animal sobrevivió en decúbito toráxico por 7 días durante los cuales se tomaron muestras de sangre con y sin anticoagulantes para determinaciones de hematocrito, hemoglobina, recuento leucocitario total y diferencial, morfología de los leucocitos, concentración de proteínas séricas totales, concentración de globulinas y presencia de anticuerpos contra el linfosarcoma bovino. Se describen las lesiones neoplásicas macro y microscópicas observadas en las porciones lumbar y sacra de la médula espinal, nervio ciático izquierdo, corazón, músculos intercostales, diafragma, hígado, abomaso, riñones y en algunos ganglios linfáticos. Macroscópicamente, el útero no presentó ninguna alteración, sin embargo microscópicamente se observó infiltración y proliferación linfoide neoplásica severa en el endometrioGanado de leche-Ganadería lech

Comparación de metodologías para la estimación de la densidad de la madera y sus implicancias en la estimación de parámetros genéticos en tres especies del género Eucalyptus

Among the characteristics that define the quality of wood, density has become the more relevant trait. In relation to the different methods available to estimate the basic wood density, the quickness, reliability and precision of each one, the objectives of this work were 1) to compare wood density values estimated by the empirical, maximum moisture content and hydrostatic balance methods in Eucalyptus globulus, E. grandis and E. viminalis, and 2) to determine if the different methods provide different genetic parameters. The average wood density ranged between 0.32 to 0.70 g/cm3, providing the empirical method the highest average value for the 3 species, tending to overestimating wood density. Heritabilities for E. globulus showed differences between the three methods, being moderate for E. viminalis and E. grandis. Given the high correlations estimated between the hydrostatic balance and maximum moisture content methods (r between 0.85 and 0.99) and the complexities inherent in their methodologies, the maximum moisture content method would be the most appropriate for the evaluated species of Eucalyptus, given its simplicity with respect to the hydrostatic balance method.Entre las diversas características que definen la calidad de la madera, la densidad es el descriptor que ha cobrado mayor relevancia. En vista a la variedad de métodos disponibles para estimar la densidad básica, de la rapidez, confiabilidad y precisión de cada uno de ellos, los objetivos propuestos para este trabajo fueron: 1) comparar las determinaciones de densidad básica de la madera obtenidas por los métodos empírico, de máximo contenido de humedad y de balanza hidrostática en Eucalyptus globulus, E. grandis y E. viminalis, y 2) determinar si diferentes métodos de estimación de densidad básica brindan diferentes estimaciones de parámetros genéticos. Los resultados mostraron valores de densidad promedio entre 0,32-0,70 g/cm3, siendo el método empírico el que estimó los mayores valores promedios para las 3 especies, mostrando una tendencia a sobreestimar los valores de densidad. Las estimaciones de heredabilidades en sentido estricto para E. globulus mostraron diferencias entre los distintos métodos, siendo moderadas las diferencias para E. viminalis y E. grandis. Dadas las altas correlaciones fenotípicas estimadas entre los métodos de balanza hidrostática y de máximo contenido de humedad (r entre 0,85 y 0,99) y las complejidades inherentes a sus metodologías, el método de máximo contenido de humedad resultaría ser el más apropiado para las especies evaluadas de Eucalyptus dada su simplicidad respecto al método de balanza hidrostática