Dynamic glucose uptake, storage, and release by human microvascular endothelial cells

Endothelia determine blood-to-tissue solute delivery, yet glucose transit is poorly understood. To illuminate mechanisms, we tracked [3H]-2-deoxyglucose (2-DG) in human adipose-tissue microvascular endothelial cells. 2-DG uptake was largely facilitated by the glucose transporters GLUT1 and GLUT3. Once in the cytosol, >80% of 2-DG became phosphorylated and ∼20% incorporated into glycogen, suggesting that transported glucose is readily accessible to cytosolic enzymes. Interestingly, a fraction of intracellular 2-DG was released over time (15–20% over 30 min) with slower kinetics than for uptake, involving GLUT3. In contrast to intracellular 2-DG, the released 2-DG was largely unphosphorylated. Glucose release involved endoplasmic reticulum–resident translocases/phosphatases and was stimulated by adrenaline, consistent with participation of glycogenolysis and glucose dephosphorylation. Surprisingly, the fluorescent glucose derivative 2-NBD-glucose (2-NBDG) entered cells largely via fluid phase endocytosis and exited by recycling. 2-NBDG uptake was insensitive to GLUT1/GLUT3 inhibition, suggesting poor influx across membranes. 2-NBDG recycling, but not 2-DG efflux, was sensitive to N-ethyl maleimide. In sum, by utilizing radioactive and fluorescent glucose derivatives, we identified two parallel routes of entry: uptake into the cytosol through dedicated glucose transporters and endocytosis. This reveals the complex glucose handling by endothelial cells that may contribute to glucose delivery to tissues.Fil: Yazdani, Samaneh. University Of Toronto. Hospital For Sick Children; CanadáFil: Bilan, Philip J.. University Of Toronto. Hospital For Sick Children; CanadáFil: Jaldín Fincati, Javier Roberto. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Salta. Instituto de Patología Experimental. Universidad Nacional de Salta. Facultad de Ciencias de la Salud. Instituto de Patología Experimental; Argentina. University Of Toronto. Hospital For Sick Children; CanadáFil: Pang, Janice. University Of Toronto. Hospital For Sick Children; CanadáFil: Ceban, Felicia. University Of Toronto. Hospital For Sick Children; CanadáFil: Saran, Ekambir. University Of Toronto. Hospital For Sick Children; CanadáFil: Brumell, John H.. University Of Toronto. Hospital For Sick Children; Canadá. University of Toronto; CanadáFil: Freeman, Spencer A.. University Of Toronto. Hospital For Sick Children; Canadá. University of Toronto; CanadáFil: Klip, Amira. University of Toronto; Canadá. University Of Toronto. Hospital For Sick Children; Canad

Comparison between first and second wave of COVID-19 outbreak in older people. The COPE multicentre European observational cohort study

Background: Effective shielding measures and virus mutations have progressively modified the disease between the waves, likewise health care systems have adapted to the outbreak. Our aim was to compare clinical outcomes for older people with COVID-19 in Wave 1 (W1) and 2 (W2). Methods: All data, including the Clinical Frailty Scale (CFS), were collected for COVID-19 consecutive patients, aged ≥65, from thirteen hospitals, in W1 (February-June 2020) and W2 (October 2020-March 2021). The primary outcome was mortality (time to mortality and 28-day mortality). Data were analysed with multilevel Cox proportional hazards, linear and logistic regression models, adjusted for wave baseline demographic and clinical characteristics. Results: Data from 611 people admitted in W2 were added to and compared with data collected during W1 (N = 1340). Patients admitted in W2 were of similar age, median [IQR], W2 = 79 [73-84]; W1 = 80 [74-86]; had a greater proportion of men (59.4% vs 53.0%); had lower 28-day mortality (29.1% vs 40.0%), compared to W1. For combined W1-W2 sample, W2 was independently associated with improved survival: time-to-mortality aHR= 0.78 (95%CI 0.65-0.93), 28-day mortality aOR = 0.80 (95%CI 0.62-1.03). W2 was associated with increased length of hospital stay aHR = 0.69 (95%CI 0.59-0.81). Patients in W2 were less frail, CFS (adjusted mean difference [aMD]=-0.50, 95%CI -0.81, -0.18), as well as presented with lower CRP (aMD=-22.52, 95%CI -32.00, -13.04). Conclusions: COVID-19 older adults in W2 were less likely to die than during W1. Patients presented to hospital during W2 were less frail and with lower disease severity and less likely to have renal decline

Prognostic value of estimated glomerular filtration rate in hospitalised older patients (over 65) with COVID-19: a multicentre, European, observational cohort study

Background: The reduced renal function has prognostic significance in COVID-19 and it has been linked to mortality in the general population. Reduced renal function is prevalent in older age and thus we set out to better understand its effect on mortality. Methods: Patient clinical and demographic data was taken from the COVID-19 in Older People (COPE) study during two periods (February–June 2020 and October 2020–March 2021, respectively). Kidney function on admission was measured using estimated glomerular filtration rate (eGFR). The primary outcomes were time to mortality and 28-day mortality. Secondary outcome was length of hospital stay. Data were analysed with multilevel Cox proportional hazards regression, and multilevel logistic regression and adjusted for individual patient clinical and demographic characteristics. Results: One thousand eight hundred two patients (55.0% male; median [IQR] 80 [73–86] years) were included in the study. 28-day mortality was 42.3% (n = 742). 48% (n = 801) had evidence of renal impairment on admission. Using a time-to-event analysis, reduced renal function was associated with increased in-hospital mortality (compared to eGFR ≥ 60 [Stage 1&2]): eGFR 45–59 [Stage 3a] aHR = 1.26 (95%CI 1.02–1.55); eGFR 30–44 [Stage 3b] aHR = 1.41 (95%CI 1.14–1.73); eGFR 1–29 [Stage 4&5] aHR = 1.42 (95%CI 1.13–1.80). In the co-primary outcome of 28-day mortality, mortality was associated with: Stage 3a adjusted odds ratio (aOR) = 1.18 (95%CI 0.88–1.58), Stage 3b aOR = 1.40 (95%CI 1.03–1.89); and Stage 4&5 aOR = 1.65 (95%CI 1.16–2.35). Conclusion: eGFR on admission is a good independent predictor of mortality in hospitalised older patients with COVID-19 population. We found evidence of a dose-response between reduced renal function and increased mortality

Muscle cells challenged with saturated fatty acids mount an autonomous inflammatory response that activates macrophages

<p>Abstract</p> <p>Obesity is associated with chronic low-grade inflammation. Within adipose tissue of mice fed a high fat diet, resident and infiltrating macrophages assume a pro-inflammatory phenotype characterized by the production of cytokines which in turn impact on the surrounding tissue. However, inflammation is not restricted to adipose tissue and high fat-feeding is responsible for a significant increase in pro-inflammatory cytokine expression in muscle. Although skeletal muscle is the major disposer of dietary glucose and a major determinant of glycemia, the origin and consequence of muscle inflammation in the development of insulin resistance are poorly understood.</p> <p>We used a cell culture approach to investigate the vectorial crosstalk between muscle cells and macrophages upon exposure to physiological, low levels of saturated and unsaturated fatty acids. Inflammatory pathway activation and cytokine expression were analyzed in L6 muscle cells expressing myc-tagged GLUT4 (L6GLUT4<it>myc</it>) exposed to 0.2 mM palmitate or palmitoleate. Conditioned media thereof, free of fatty acids, were then tested for their ability to activate RAW264.7 macrophages.</p> <p>Palmitate -but not palmitoleate- induced IL-6, TNFα and CCL2 expression in muscle cells, through activation of the NF-κB pathway. Palmitate (0.2 mM) alone did not induce insulin resistance in muscle cells, yet conditioned media from palmitate-challenged muscle cells selectively activated macrophages towards a pro-inflammatory phenotype.</p> <p>These results demonstrate that low concentrations of palmitate activate autonomous inflammation in muscle cells to release factors that turn macrophages pro-inflammatory. We hypothesize that saturated fat-induced, low-grade muscle cell inflammation may trigger resident skeletal muscle macrophage polarization, possibly contributing to insulin resistance <it>in vivo</it>.</p