Proteomic Analysis Identifies Molecular Players and Biological Processes Specific to SARS-CoV-2 Exposure in Endothelial Cells

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been responsible for the severe pandemic of acute respiratory disease, coronavirus disease 2019 (COVID-19), experienced in the 21st century. The clinical manifestations range from mild symptoms to abnormal blood coagulation and severe respiratory failure. In severe cases, COVID-19 manifests as a thromboinflammatory disease. Damage to the vascular compartment caused by SARS-CoV-2 has been linked to thrombosis, triggered by an enhanced immune response. The molecular mechanisms underlying endothelial activation have not been fully elucidated. We aimed to identify the proteins correlated to the molecular response of human umbilical vein endothelial cells (HUVECs) after exposure to SARS-CoV-2, which might help to unravel the molecular mechanisms of endothelium activation in COVID-19. In this direction, we exposed HUVECs to SARS-CoV-2 and analyzed the expression of specific cellular receptors, and changes in the proteome of HUVECs at different time points. We identified that HUVECs exhibit non-productive infection without cytopathic effects, in addition to the lack of expression of specific cell receptors known to be essential for SARS-CoV-2 entry into cells. We highlighted the enrichment of the protein SUMOylation pathway and the increase in SUMO2, which was confirmed by orthogonal assays. In conclusion, proteomic analysis revealed that the exposure to SARS-CoV-2 induced oxidative stress and changes in protein abundance and pathways enrichment that resembled endothelial dysfunction.     

Discrepancies in the Regression Modelling of Recrystallization Rate as Using the Data from Physical Simulation Tests

The analysis of numerous experimental equations published in the literature reveals a wide scatter in the predictions for the static recrystallization kinetics of steels. The powers of the deformation variables, strain and strain rate, similarly as the power of the grain size vary in these equations. These differences are highlighted and the typical values are compared between torsion and compression tests. Potential errors in physical .simulation testing are discussed.     

Aerobic industrial processing of Empeltre cv. natural black olives and product characterisation

The fermentation of natural black olives is currently carried out under anaerobic conditions. This work investigates the aerobic fermentation of this product in tanks containing 16 000 kg of Empeltre olives during two seasons. The microorganisms in brines were yeasts in all cases, whereas lactic acid bacteria growth was only observed in some tanks. Consequently, the pH during fermentation was in most cases higher than recommended 4.3 units. However, the Empeltre olive brines displayed bactericidal effect against Salmonella enterica, Sthapylococcus aureus, Escherichia coli and Listeria monocytogenes, with a 5 log population drop in one or 24 h depending on the brine age so that these olive brines are a harsh environment for the growth of these pathogens. It was found HyEDA as the major phenolic compound in both brine and olive pulp at the beginning of fermentation that slowly hydrolysed into hydroxytyrosol that reached a concentration up to 1500 mg kg⁻¹.     

Stem cell factor-induced downregulation of c-kit in human lung mast cells and HMC-1 mast cells

Recent data suggest that local overexpression of the tissue-hormone c-kit ligand (stem cell factor [SCF]) is associated with accumulation of mast cells (MCs) and a decrease in expression of c-kit in the accumulated MCs [28]. In the present study, the effects of recombinant human (rh) SCF on expression of c-kit mRNA and c-kit protein in isolated human MCs and a human mast cell line, HMC-1, were analyzed. Incubation of isolated lung MC with rhSCF (100 ng/mL) for 120 minutes resulted in decreased expression of c-kit mRNA (optical density [OD], control: 100% vs. rhSCF: 37%). Almost identical results were obtained with HMC-1 cells (OD, control: 100% vs. rhSCF: 40 to 45%). As assessed by flow cytometry and monoclonal antibodies (mAbs) to c-kit, the SCF-induced decrease of c-kit mRNA in HMC-1 was associated with a substantial decrease in surface expression of c-kit (MFI, control: 100 +/- 21%, vs. MFI in cells incubated with rhSCF [100 ng/mL at 37 degrees C for 12 hours]: 8 +/- 2%, vs. MFI in cells incubated with rhSCF, 100 ng/mL, at 4 degrees C: 34 +/- 3%). The effects of rhSCF on c-kit expression in HMC-1 cells were dose- and time-dependent with maximum effects observed with 10-100 ng/mL of rhSCF after 4 to 12 hours. The SCF-dependent loss of c-kit was also accompanied by a decreased chemotactic response to rhSCF (control: 100%; rhSCF: 71 +/- 2%). This study shows that exposure of human lung MC and HMC-1 cells to recombinant SCF results in downregulation of c-kit mRNA and surface c-kit expression. These data may explain the partial loss of c-kit on MCs in areas of SCF overexpression.     

Early ontogeny of catecholaminergic cell lineage in brain and peripheral neurons monitored by tyrosine hydroxylase-lacZ transgene

As the first and rate limiting enzyme in the biosynthetic pathway for catecholamine (CA) neurotransmitters, tyrosine hydroxylase (TH) is a specific phenotypic marker for CA cells in the central and peripheral nervous systems of adult animals. During embryogenesis, TH expression appears permanently within cells destined to be CA-secreting during adult life, and transiently in several cell types that will not express TH in adulthood. In this study, we examined the early ontogeny of TH expression in transgenic mouse embryos by following the expression of a lacZ reporter, driven by the tissue-specific promoter of the rat TH gene. The lacZ reporter product, beta-galactosidase (beta-gal), visualized by X-gal staining, first became apparent in primordia of sensory ganglia serving the glossopharyngeal (IX) and vagal (X) cranial nerves at embryonic day (E)9.0. Between E9.5 and E10.5, beta-gal expression extended to the remaining cranial sensory ganglia serving the trigeminal (V) and facial (VII) nerves, dorsal root ganglia, ventrolateral neural tube and sympathetic ganglion primordia. During that same period, the first beta-gal expression in the embryonic brain also appeared within distinct regions, such as the ventral prosencephalon, the ventral and dorsolateral mesencephalon and the rostral and caudal rhombencephalon. The level of beta-gal expression in all these tissues decreased at E13.5, but a distinct adult pattern of beta-gal expression started to emerge in the substantia nigra and ventral tegmental area in the central nervous system and the adrenal medulla in the periphery. Our findings indicate that the proximal 9.0 kb of the 5' promoter region of the rat TH gene encodes sufficient information to direct development of the appropriate catecholaminergic lineage cells in the central and most peripheral nervous systems during embryogenesis.     

Ethical analysis of health care reform: implications for diverse communities

The values driving nursing's political actions and the ethics underlying proposed health system change have not yet been thoroughly examined. This article attempts to (a) clarify ethical issues surrounding the health care debate in the United States, (b) describe the conflicts surrounding proposed reform especially the effects on disadvantaged communities and (c) earnestly entreat a critical discussion of nursing's humanistic position on health care reform. The interaction between the nursing literature and the various proposals on health care reform suggest various themes that reflect ethical issues due to conflicts in values. Our analysis is based on the belief that, in addition to political action and lobbying, the nursing community must confront critical questions about the values driving its policy making.     

Hemorrhages in muscles of broiler chickens: the relationships among blood variables at various rearing temperature regimens

Hemorrhages in muscle tissue can be considered as major quality defects of broiler carcasses. They can be induced by stunning, especially electrical stunning. The underlying mechanism, however, is considered to be multifactorial. In this study, the effect of blood circulation disturbances on the severity of hemorrhages induced by electrical stunning was investigated. The disturbances were evoked in two genetically different, fast-growing broiler strains, Ross and Hybro, by rearing the broilers at low ambient temperatures. The broilers were slaughtered by two different electrical stunning methods. Broilers reared at low temperatures showed changes in blood variables and heart weight known to be associated with blood circulation disturbances. There was no effect of rearing temperature on hemorrhage severity. Ross broilers, being the most susceptible to low temperatures, had less severe hemorrhages than Hybro broilers. There was, however, a clear effect of the stunning method on hemorrhage severity. Whole body stunning caused more severe hemorrhages than head stunning in thigh and breast muscles. These results suggest that factors interfering with blood circulation have little or no effect on the occurrence of hemorrhages induced by electrical stunning.     

B cell selection and allelic exclusion of an anti-DNA Ig transgene in MRL-lpr/lpr mice

We have used an Ig transgene (VH3H9) that increases the frequency of anti-DNA autoantibodies to address whether the production of antinuclear Abs in systemic lupus erythematosus is the consequence of a breakdown of B cell tolerance. We have shown that nonautoimmune mice regulate anti-DNA B cells, and that lupus-prone MRL-lpr/lpr mice are defective in this regulation. Here we show that a subset of anti-DNA B cells, namely those that stain nuclei in a homogeneous fashion, not only fail to be deleted in MRL-lpr/lpr mice, but undergo preferential clonal expansion. In addition, we describe a surprising finding: the VH3H9 transgene is less efficient at inhibiting endogenous heavy chain gene rearrangement on the autoimmune-prone MRL-lpr/lpr genetic background than on the nonautoimmune BALB/c background.     

The influence of sterols on the sensitivity of lipid bilayers to melittin

The sensitivity of planar lipid bilayers to the permeabalizing effect of melittin was evaluated when sterols of varying structure were incorporated into the membrane. The addition of increasing amount of cholesterol (0-50 mole %) decreased the sensitivity of membranes formed from negatively charged phospholipids to melittin but did not (in amount of up to 66 mole %) change the sensitivity of membranes formed from zwitterionic lipids. 7-Dehydrocholesterol, stigmasterol and ergosterol had the same ability as that of cholesterol to decrease the membrane sensitivity to melittin, while lanosterol had no effect on the sensitivity of membranes to melittin. The results suggest that the effect of sterols is complex and cannot be explained only by a direct interaction of melittin with cholesterol, by a decrease of membrane fluidity, or by changes in distribution of surface charge.