Cyto- and genotoxic effects of metallic nanoparticles in untransformed human fibroblast

AbstractMetallic nanoparticles such as silver (Ag), cerium dioxide (CeO2) and titanium dioxide (TiO2) are produced at a large scale and included in many consumer products. It is well known that most metallic NPs are toxic to humans which raise concerns about these engineered particles. Various studies have already been published on the subject, however, almost all of these studies have been conducted in cancer or transformed cell lines. In this work we performed a comparative evaluation of these metallic NPs on normal untransformed human fibroblasts (GM07492) detecting cyto- and geno-toxic responses after exposure to these NPs. Our results showed that all three metallic NPs were able to cross the plasma membrane and were mainly found in endocytic vesicles. The Ag and TiO2 NPs affected mitochondrial enzymatic activity (XTT), increased DNA fragmentation, oxidative damage (Comet assay) and induced cell death mainly by the apoptotic pathway. Ag NPs increased GADD45α transcript levels and the phosphorylation of proteins γH2AX. Transient genotoxicity was also observed from exposure to CeO2 NPs while TiO2 NPs showed no increase in DNA damage at sub-cytotoxic concentrations. In comparison, Ag NPs were found to be the most cyto-genotoxic NPs to fibroblasts. Thus, these results support the use of normal fibroblast as a more informative tool to detect the mechanisms of action induced by metallic NPs

Provincias de La Rioja y San Juan

Fil: Fauqué, L. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Limarino, C. UBA-CONICET; ArgentinaFil: Vujovich, G. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Cegarra, M. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Yamin, M. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Tedesco, A. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Escosteguy, L. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Cardó, R. Servicio Geológico Minero Argentino - San Juan; ArgentinaFil: Díaz, I. Servicio Geológico Minero Argentino - San Juan; ArgentinaFil: Franchi, M. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaFil: Etcheverría, M. Servicio Geológico Minero Argentino - Buenos Aires; ArgentinaEn este trabajo, el apellido de Marcela Yamin aparece escrito con acento. Al comprobar que el mismo está escrito sin acento en otros trabajos de la autora, se presume que esto ha sido un pequeño error de edición, ingresándolo en el repositorio de la manera que se presume correcta.La Hoja Geológica 2969-IV Villa Unión abarca parte de la región noroccidental de la Provincia de La Rioja y el sector occidental de la Hoja comprende una porción del noreste de la provincia de San Juan. Se extiende entre los paralelos 29º y 30º de latitud sur y entre los meridianos 67º 30’ y 69º de longitud oeste, cubriendo una superfi cie aproximada de 18.000 km2. La Hoja toma el nombre de la localidad más importante del oeste riojano, Villa Unión. Otra localidad importante del mismo sector es Villa Castelli, como la anterior, a orillas del río Vinchina-Bermejo, en el límite norte de la Hoja. La localidad más occidental de la Hoja es Guandacol en la depresión tectónica recorrida por el río homónimo. Finalmente al naciente de la sierra de Famatina y dentro del área de la Hoja se ubican, de norte a sur las localidades de: Las Higueritas, Chilecito, Sañogasta y Vichigasta. La Hoja Villa Unión incluye parte de tres provincias geológicas argentinas, de este a oeste: Sistema de Famatina, Sierras Pampeanas occidentales y Precordillera. En la Hoja Villa Unión afl oran basamentos metamórfi cos muy distintos como consecuencia de las colisiones y acreciones de diferentes terrenos (Famatina, Cuyania y Chilenia) ocurridas durante el Proterozoico superior y el Paleozoico inferior. Los terrenos metamórfi cos del Sistema de Famatina son de muy bajo a bajo grado, mientras que los terrenos metamórfi cos de Sierras Pampeanas Occidentales son de grado medio a alto. El basamento correspondiente a Sierras Pampeanas Occidentales, afl orante en la Hoja Villa Unión, conforma los bloques serranos de Umango, Maz, Espinal y Las Ramaditas, elevados como consecuencia de la tectónica terciaria-cuaternaria. En ellos las litologías predominantes son: gneises, esquistos, calizas, anfi bolitas y granitoides. En la sierra de Famatina el basamento esta representado por la Formación Negro Peinado, compuesta por leptometamorfi tas. Sobre ellas se apoyan en discordancia sedimentitas ordovícicas de la Formación Suri que se interdigitan con rocas volcánicas. Estas rocas están intruidas por una secuencia de cuerpos plutónicos calcoalcalinos de la Formación Ñuñorco, caracterizada por gabros, tonalitas, granodioritas y granitos, con un clímax de actividad magmática en el Ordovícico medio a superior. Ellas representan un arco magmático desarrollado en un margen continental activo. Cuyania es un terreno compuesto, con suturas internas representadas por una faja ofi olítica de edad Grenville. Los terrenos amalgamados (Precordillera y Pie de Palo) para formar Cuyania serían posibles arcos de islas intraoceánicos representados por anfi bolitas y gneises. Sobre Cuyania se desarrolló una plataforma calcárea de edad cambro-ordovícica. Las rocas más antiguas de esta secuencia afl oran al noroeste de Guandacol, estan constituidas por depósitos continentales rojos y evaporitas de edad cámbrica inferior (Formación Cerro Totora). En discordancia se apoyan los depósitos de plataforma carbonática (Formación San Juan), además de secuencias ordovícicas clásticas (Formación Guandacol, Formación Yerba Loca) y fi nalmente secuencias clásticas con niveles conglomerádicos (Grupo Trapiche) del Ordovícico superior. Los depósitos neopaleozoicos de las cuencas Río Blanco y Paganzo se apoyan en este sector mediante discordancia angular. En la Precordillera las rocas que pertenecen a la cuenca Río Blanco comenzaron a sedimentarse en el Devónico superior-Carbonífero inferior, continuando hasta el Pérmico. Las sedimentitas de la Cuenca Paganzo que afl oran en las Sierras Pampeanas Occidentales y en la sierra de Famatina se depositaron en el Carbonífero medio y superior. Como resultado del reinicio de la subducción en el Carbonífero superior, en estas cuencas también se ha reconocido actividad magmática carbonífera y permo-triásica. En la Precordillera esta actividad magmática está representada, en la cuenca del Río Blanco, por los intrusivos del cerro Veladero (Carbonífero superior) y por los de los cerros Imán y Madril (Pérmico inferior). El magmatismo permotriásico corresponde principalmente al ámbito de la Cordillera Frontal. Como resultado de las orogenias paleozoicas quedó amalgamado un núcleo cratónico central o pangea. Esta pangea fue afectada por un régimen extensional vinculado a la completa cesación de la subducción, a lo largo del margen del supercontinente. Así, extensas áreas del basamento andino y zonas adyacentes fueron sometidas a un importante régimen extensional durante el Triásico. Los sistemas de rift ubicados en el borde occidental de las Sierras Pampeanas, se desarrollaron en el labio superior de la sutura, entre los terrenos de Cuyania y Pampia. Corresponden a las cuencas de Ischigualasto, Marayes, Salinas y Beazley. Sus secuencias están muy bien representadas, dentro de la Hoja Villa Unión, en el cerro Bola y su continuación al sudeste en el cordón de la sierra Morada. La sierra de Famatina continúa hacia el sur en la sierra de Los Tarjados, donde las unidades triásicas también poseen extensos afl oramientos. Durante el Cenozoico los movimientos del Ciclo Ándico produjeron la estructuración defi nitiva de los cordones montañosos. Como resultado de la migración del frente orogénico, los depósitos sinorogénicos terciarios asociados a este levantamiento se extienden ampliamente desde la Cordillera Frontal hasta el este del Sistema de Famatina. Estas sedimentitas se interdigitan con escasas dacitas (Formación Mogotes) y brechas y coladas ácidas y mesosilícicas (Formación Cerro Morado). Los depósitos sinorogénicos del segmento central (27º-33º S), en el que se halla la Hoja, corresponden al sector norte del segmento de subducción subhorizontal, que incluye el sistema de Cordillera Principal, Cordillera Frontal, Precordillera, Sistema de Famatina y Sierras Pampeanas, originando numerosos depocentros y las secuencias cenozoicas más potentes de la Argentina. Dentro de la Hoja se halla el extremo sur de la cuenca de Vinchina actuando como depocentro desde los 10 Ma, por lo que registra tanto los depósitos sinorogénicos relacionados con la estructuración de la Cordillera Frontal, como los más modernos, del pie oriental del Famatina. Esta cuenca registra los mayores espesores para el Neógeno de la región andina (10.260 m). La secuencia está compuesta por las Formaciones Vinchina y Toro Negro en la sierra de Los Colorados (Hoja Tinogasta) y más al sur dentro de la Hoja Villa Unión, la secuencia se compone por las Formaciones: Puesto La Flecha, Vallecito, Vinchina, Zapallar y El Corral. La Formación Santa Florentina, del Pleistoceno inferior, también representa depósitos sinorogénicos adosados al pie occidental del Famatina. Esta última sierra sufre englazamientos cuaternarios, conservándose aún sus depósitos. Continúan en el Cuaternario depósitos de niveles aluviales pedemontanos e intermontanos y depósitos recientes de cauces fl uviales, eólicos y evaporíticos. Las Sierras Pampeanas tienen una estructura de bloques limitados por fallas inversas originadas principalmente a partir del Mioceno inferior, como lo demuestran los depósitos sinorogénicos. La estructura de la sierra de Famatina es similar a la anterior, bloques de basamento precámbrico-eopaleozoico, levantados y basculados por fallas lístricas inversas durante el Cenozoico superior. La Precordillera riojana, desde el punto de vista estructural, está representada por una faja plegada y corrida epidérmica. El paisaje de la región ha resultado principalmente del accionar del proceso endógeno (tectónica) y en menor medida del modelado exógeno (fl uvial, glaciario, periglaciario, eólico y remoción en masa). El papel jugado por la estructura y la evolución tectónica regional ha sido determinante en las características morfológicas del paisaje regional. Estos aspectos geomorfológicos generales están íntimamente relacionados a lineamientos estructurales, resultado de la orogenia Andina, que han dado lugar a la formación de tres cordones montañosos, separados por depresiones tectónicas, que con algunas intermitencias han conservado sus características de cuencas en continuo hundimiento desde el terciario medio. En los cordones montañosos participan diferentes litologías: basamento precámbrico, granitoides del paleozoico inferior, sedimentitas y metamorfi tas del Paleozoico inferior, sedimentitas y rocas eruptivas del Paleozoico superior y volcanitas y sedimentitas terciarias. Cada una de estas litologías ha presentado diferentes respuestas frente al accionar de los procesos denudacionales, en particular la acción fl uvial y la meteorización. Las sierras exponen las rocas más antiguas, mientras que en los valles (depresiones tectónicas) se acumulan depósitos más modernos (terciario-cuaternarios) que resultan de la erosión de las cadenas montañosas; y cuyos materiales son llevados por los ríos y removilizados por el viento dentro de las depresiones. La Hoja Villa Unión cuenta con recursos minerales metalíferos y no metalíferos importantes. Algunos de ellos no solo tienen valor económico sino además histórico, como es el caso de la mina La Mejicana

Metallated phthalocyanines and their hydrophilic derivatives for multi-targeted oncological photodynamic therapy

Background and aim: A photosensitizer (PS) delivery and comprehensive tumor targeting platform was developed that is centered on the photosensitization of key pharmacological targets in solid tumors (cancer cells, tumor vascular endothelium, and cellular and non-cellular components of the tumor microenvironment) before photodynamic therapy (PDT). Interstitially targeted liposomes (ITLs) encapsulating zinc phthalocyanine (ZnPC) and aluminum phthalocyanine (AlPC) were formulated for passive targeting of the tumor microenvironment. In previous work it was established that the PEGylated ITLs were taken up by cultured cholangiocarcinoma cells. The aim of this study was to verify previous results in cancer cells and to determine whether the ITLs can also be used to photosensitize cells in the tumor microenvironment and vasculature. Following positive results, rudimentary in vitro and in vivo experiments were performed with ZnPC-ITLs and AlPC-ITLs as well as their water-soluble tetrasulfonated derivatives (ZnPCS4 and AlPCS4) to assemble a research dossier and bring this platform closer to clinical transition. Methods: Flow cytometry and confocal microscopy were employed to determine ITL uptake and PS distribution in cholangiocarcinoma (SK-ChA-1) cells, endothelial cells (HUVECs), fibroblasts (NIH-3T3), and macrophages (RAW 264.7). Uptake of ITLs by endothelial cells was verified under flow conditions in a flow chamber. Dark toxicity and PDT efficacy were determined by cell viability assays, while the mode of cell death and cell cycle arrest were assayed by flow cytometry. In vivo systemic toxicity was assessed in zebrafish and chicken embryos, whereas skin phototoxicity was determined in BALB/c nude mice. A PDT efficacy pilot was conducted in BALB/c nude mice bearing human triple-negative breast cancer (MDA-MB-231) xenografts. Results: The key findings were that (1) photodynamically active PSs (i.e., all except ZnPCS4) were able to effectively photosensitize cancer cells and non-cancerous cells; (2) following PDT, photodynamically active PSs were highly toxic-to-potent as per anti-cancer compound classification; (3) the photodynamically active PSs did not elicit notable systemic toxicity in zebrafish and chicken embryos; (4) ITL-delivered ZnPC and ZnPCS4 were associated with skin phototoxicity, while the aluminum-containing PSs did not exert detectable skin phototoxicity; and (5) ITL-delivered ZnPC and AlPC were equally effective in their tumor-killing capacity in human tumor breast cancer xenografts and superior to other non-phthalocyanine PSs when appraised on a per mole administered dose basis. Conclusions: AlPC(S4) are the safest and most effective PSs to integrate into the comprehensive tumor targeting and PS delivery platform. Pending further in vivo validation, these third-generation PSs may be used for multi-compartmental tumor photosensitization

Metallated phthalocyanines and their hydrophilic derivatives for multi-targeted oncological photodynamic therapy

Background and aim: A photosensitizer (PS) delivery and comprehensive tumor targeting platform was developed that is centered on the photosensitization of key pharmacological targets in solid tumors (cancer cells, tumor vascular endothelium, and cellular and non-cellular components of the tumor microenvironment) before photodynamic therapy (PDT). Interstitially targeted liposomes (ITLs) encapsulating zinc phthalocyanine (ZnPC) and aluminum phthalocyanine (AlPC) were formulated for passive targeting of the tumor microenvironment. In previous work it was established that the PEGylated ITLs were taken up by cultured cholangiocarcinoma cells. The aim of this study was to verify previous results in cancer cells and to determine whether the ITLs can also be used to photosensitize cells in the tumor microenvironment and vasculature. Following positive results, rudimentary in vitro and in vivo experiments were performed with ZnPC-ITLs and AlPC-ITLs as well as their water-soluble tetrasulfonated derivatives (ZnPCS4 and AlPCS4) to assemble a research dossier and bring this platform closer to clinical transition. Methods: Flow cytometry and confocal microscopy were employed to determine ITL uptake and PS distribution in cholangiocarcinoma (SK-ChA-1) cells, endothelial cells (HUVECs), fibroblasts (NIH-3T3), and macrophages (RAW 264.7). Uptake of ITLs by endothelial cells was verified under flow conditions in a flow chamber. Dark toxicity and PDT efficacy were determined by cell viability assays, while the mode of cell death and cell cycle arrest were assayed by flow cytometry. In vivo systemic toxicity was assessed in zebrafish and chicken embryos, whereas skin phototoxicity was determined in BALB/c nude mice. A PDT efficacy pilot was conducted in BALB/c nude mice bearing human triple-negative breast cancer (MDA-MB-231) xenografts. Results: The key findings were that (1) photodynamically active PSs (i.e., all except ZnPCS4) were able to effectively photosensitize cancer cells and non-cancerous cells; (2) following PDT, photodynamically active PSs were highly toxic-to-potent as per anti-cancer compound classification; (3) the photodynamically active PSs did not elicit notable systemic toxicity in zebrafish and chicken embryos; (4) ITL-delivered ZnPC and ZnPCS4 were associated with skin phototoxicity, while the aluminum-containing PSs did not exert detectable skin phototoxicity; and (5) ITL-delivered ZnPC and AlPC were equally effective in their tumor-killing capacity in human tumor breast cancer xenografts and superior to other non-phthalocyanine PSs when appraised on a per mole administered dose basis. Conclusions: AlPC(S4) are the safest and most effective PSs to integrate into the comprehensive tumor targeting and PS delivery platform. Pending further in vivo validation, these third-generation PSs may be used for multi-compartmental tumor photosensitization

The polymorphism L412F in TLR3 inhibits autophagy and is a marker of severe COVID-19 in males

The polymorphism L412F in TLR3 has been associated with several infectious diseases. However, the mechanism underlying this association is still unexplored. Here, we show that the L412F polymorphism in TLR3 is a marker of severity in COVID-19. This association increases in the sub-cohort of males. Impaired macroautophagy/autophagy and reduced TNF/TNFα production was demonstrated in HEK293 cells transfected with TLR3L412F-encoding plasmid and stimulated with specific agonist poly(I:C). A statistically significant reduced survival at 28 days was shown in L412F COVID-19 patients treated with the autophagy-inhibitor hydroxychloroquine (p = 0.038). An increased frequency of autoimmune disorders such as co-morbidity was found in L412F COVID-19 males with specific class II HLA haplotypes prone to autoantigen presentation. Our analyses indicate that L412F polymorphism makes males at risk of severe COVID-19 and provides a rationale for reinterpreting clinical trials considering autophagy pathways. Abbreviations: AP: autophagosome; AUC: area under the curve; BafA1: bafilomycin A1; COVID-19: coronavirus disease-2019; HCQ: hydroxychloroquine; RAP: rapamycin; ROC: receiver operating characteristic; SARS-CoV-2: severe acute respiratory syndrome coronavirus 2; TLR: toll like receptor; TNF/TNF-α: tumor necrosis factor

Genetic mechanisms of critical illness in COVID-19.

Host-mediated lung inflammation is present1, and drives mortality2, in the critical illness caused by coronavirus disease 2019 (COVID-19). Host genetic variants associated with critical illness may identify mechanistic targets for therapeutic development3. Here we report the results of the GenOMICC (Genetics Of Mortality In Critical Care) genome-wide association study in 2,244 critically ill patients with COVID-19 from 208 UK intensive care units. We have identified and replicated the following new genome-wide significant associations: on chromosome 12q24.13 (rs10735079, P = 1.65 × 10-8) in a gene cluster that encodes antiviral restriction enzyme activators (OAS1, OAS2 and OAS3); on chromosome 19p13.2 (rs74956615, P = 2.3 × 10-8) near the gene that encodes tyrosine kinase 2 (TYK2); on chromosome 19p13.3 (rs2109069, P = 3.98 ×  10-12) within the gene that encodes dipeptidyl peptidase 9 (DPP9); and on chromosome 21q22.1 (rs2236757, P = 4.99 × 10-8) in the interferon receptor gene IFNAR2. We identified potential targets for repurposing of licensed medications: using Mendelian randomization, we found evidence that low expression of IFNAR2, or high expression of TYK2, are associated with life-threatening disease; and transcriptome-wide association in lung tissue revealed that high expression of the monocyte-macrophage chemotactic receptor CCR2 is associated with severe COVID-19. Our results identify robust genetic signals relating to key host antiviral defence mechanisms and mediators of inflammatory organ damage in COVID-19. Both mechanisms may be amenable to targeted treatment with existing drugs. However, large-scale randomized clinical trials will be essential before any change to clinical practice

SARS-CoV-2 susceptibility and COVID-19 disease severity are associated with genetic variants affecting gene expression in a variety of tissues

Variability in SARS-CoV-2 susceptibility and COVID-19 disease severity between individuals is partly due to genetic factors. Here, we identify 4 genomic loci with suggestive associations for SARS-CoV-2 susceptibility and 19 for COVID-19 disease severity. Four of these 23 loci likely have an ethnicity-specific component. Genome-wide association study (GWAS) signals in 11 loci colocalize with expression quantitative trait loci (eQTLs) associated with the expression of 20 genes in 62 tissues/cell types (range: 1:43 tissues/gene), including lung, brain, heart, muscle, and skin as well as the digestive system and immune system. We perform genetic fine mapping to compute 99% credible SNP sets, which identify 10 GWAS loci that have eight or fewer SNPs in the credible set, including three loci with one single likely causal SNP. Our study suggests that the diverse symptoms and disease severity of COVID-19 observed between individuals is associated with variants across the genome, affecting gene expression levels in a wide variety of tissue types

A first update on mapping the human genetic architecture of COVID-19

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