Macrophage phagocytosis of SARS-CoV-2-infected cells mediates potent plasmacytoid dendritic cell activation.

Early and strong interferon type I (IFN-I) responses are usually associated with mild COVID-19 disease, whereas persistent or unregulated proinflammatory cytokine responses are associated with severe disease outcomes. Previous work suggested that monocyte-derived macrophages (MDMs) are resistant and unresponsive to SARS-CoV-2 infection. Here, we demonstrate that upon phagocytosis of SARS-CoV-2-infected cells, MDMs are activated and secrete IL-6 and TNF. Importantly, activated MDMs in turn mediate strong activation of plasmacytoid dendritic cells (pDCs), leading to the secretion of high levels of IFN-α and TNF. Furthermore, pDC activation promoted IL-6 production by MDMs. This kind of pDC activation was dependent on direct integrin-mediated cell‒cell contacts and involved stimulation of the TLR7 and STING signaling pathways. Overall, the present study describes a novel and potent pathway of pDC activation that is linked to the macrophage-mediated clearance of infected cells. These findings suggest that a high infection rate by SARS-CoV-2 may lead to exaggerated cytokine responses, which may contribute to tissue damage and severe disease.

Validation of a quantitative polymerase chain reaction method for human Alu gene detection in microchimeric pigs used as donors for xenotransplantation.

This work was undertaken to evaluate whether a real-time quantitative polymerase chain reaction (qPCR) is as an adequate method for detection and quantification of human-specific DNA elements (Alu gene) in tissues and blood samples of pigs in which human stem cells were engrafted. Real-time qPCR quantification was performed with the use of previously described primers. The human DNA was mixed with different quantities of porcine DNA. The primer concentration and specificity, the qPCR efficiency, the quantification variations due to different porcine DNA concentrations, and the dissociation curve produced by the assay were evaluated. The qPCR proved to be specific, robust, with a reproducible and specific bimodal melting curve. High porcine DNA concentration produced subquantification, especially with low human DNA quantity. However, the assay proved to be useful for the detection of chimeric piglets produced by human cells injected in utero, because the effect caused by the porcine DNA interference was corrected in quantification of human DNA from piglets.

Síndrome «humpy-backed» en cerdos en España / «Humpy-backed» pigs’ syndrome in Spain

El síndrome «humpy-backed» fue descrito en cerdos por primera vez en el Reino Unido en 1984. El síndrome ha sido observado en algunos países pero la etiología y la patogenia son todavía desconocidas. Este caso describe la aparición de cerdos con «humpy-backed» en la lechonera de una granja de 3.800 cerdas situada en el noreste de España. El problema afectó aproximadamente al 3% de la progenie semanal de una línea genética particular de la granja compuesta por 450 cerdas. La incidencia alcanzó picos del 9-11% en algunas semanas. Los lechones aparecían deprimidos, con pelaje hirsuto y deterioro físico progresivo. En la necropsia, a pesar de la apariencia de lordosis, no se detectaron alteraciones en los huesos y articulaciones de la columna vertebral. Microscópicamente se observó periarteritis linfoplasmocítica en corazón, bazo, intestino, hígado, riñón, músculo esquelético, pulmón y meninges. También se observó miocarditis y miositis linfoplasmocítica con degeneración de fibras musculares esqueléticas. La apariencia macroscópica de lordosis se relacionó con infiltrado celular inflamatorio multiorgánico. Se asoció a un problema de tipo genético de los animales afectados, aunque podría haber una posible implicación de circovirus porcino tipo 2 (PCV2) y el virus del síndrome reproductivo y respiratorio porcino (PRRS) en la patogenia del síndrome, debido a la seropositividad frente a los mismos detectada en la granja (AU)

«Humpy-backed» pigs’ syndrome was firstly described in the United Kingdom in 1984. The syndrome has been reported in a few countries, but the ethiology and pathogenesis remains unclear. This report describes the appearance of «humpy-backed» piglets aetiology in the nursery of a 3800-sow farm in Northeast Spain. The problem affected around 3% of the weekly offspring from one particular genetic line present on the farm composed of 450 sows. The incidence reached peaks of 9-11% in some weeks. The piglets appeared depressed, with rough hair coat and progressive clinical deterioration. In the necropsy, despite the appearance of lordosis, no defect in the bones and joint of the vertebral spine were detected. Lympho-plasmacytic periarteritis was observed in heart, spleen, gut, liver, kidney, skeletal muscle, lung and meninges. Lympho-plasmacytic myocarditis and skeletal myositis with muscular fibre degeneration was also observed. The macroscopic appearance of lordosis was associated with a multiorganic inflammatory cell infiltration. There was an association with the genetic background of the affected animals although seropositivity to porcine circovirus type 2 (PCV2) and porcine reproductive and respiratory syndrome virus (PRRSV) was also found in the farm, pointing to a possible implication of viral infection in the pathogenesis of the syndrome (AU)

Generation of human-to-pig chimerism to induce tolerance through transcutaneous in utero injection of cord blood-derived mononuclear cells or human bone marrow mesenchymals cells in a preclinical program of liver xenotransplantation: preliminary results.

OBJECTIVE: Using a percutaneous ecoguided injection system to obtain chimeric piglets through a less invasive and traumatic technique than previously reported. MATERIALS AND METHODS: The two types of human cells included umbilical cord blood mononuclear elements and mesenchymal stem cells cultured from bone marrow. Four sows at gestational day 50 were anesthetized. A needle was inserted through the skin and uterine wall to reach the peritoneal cavity of the fetuses under continuous ultrasound guidance. Fourteen piglets were injected with various cell concentrations. RESULTS: All sows carried pregnancies to term yielding 69 piglets, among which 67 were alive and two mummified. Two piglets died during the first 48 hours of life. Chimerism was detected using flow cytometry and by quantitative polymerase chain reaction (q-PCR) to detect Alu gene in blood or tissues samples. The analysis detected blood chimerism in 13 piglets (21%) by flow cytometry and the presence of the human Alu gene in 33 (51%) by q-PCR. The results suggest cell trafficking between littermates after in utero injection. CONCLUSIONS: Transcutaneous echo-guided injection succeeded to produce chimeric piglets without disadvantages to the sow or the fetuses and avoiding abortions or fetal death.

Porcine endogenous retrovirus copy number in different pig breeds is not related to genetic diversity.

The risk of zoonoses is a major obstacle to xenotransplantation. Porcine endogenous retrovirus (PERV) poses a potential risk of zoonotic infection, and its control is a prerequisite for the development of clinical xenotransplantation. The copy number of PERV varies among different breeds, and it has been suggested that the PERV integrations number is increased by inbreeding. The purpose of this study was (i) to examine the copy number of PERV in different Spanish pig breeds, Spanish wild boar and commercial cross-bred pigs from five different farms and genetic background (CCP1-CCP5) and (ii) to investigate the correlation between PERV copy number and the genetic background of the pigs in order to improve the selection of pigs for xenotransplantation. PERV copy number was determined by quantitative, real-time polymerase chain reactions. Thirty-four microsatellite markers were genotyped to describe the genetic diversity within populations (observed and expected heterozygosities, Ho and He, respectively) and the inbreeding coefficient (F). Pearson's correlation coefficient was used to determine the relationship between PERV copy number and Ho, He and F. The copy number of PERV among different pig breeds was estimated to range between three (CCP1) and 43 copies (Iberian Pig). Statistical differences were found among the studied populations concerning PERV copy number. No correlation was found between the PERV copy number and the heterozygosity (calculated at an individual level or at a population level) or the inbreeding coefficient of each population. Our data suggest that pigs inbreeding does not increase PERV copy number and support the idea that careful selection of pigs for organ donation with reduced PERV copy number will minimize the risk of retrovirus transmission to the human receptor.