Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Fibrillar tau alters cerebral endothelial cell metabolism, vascular inflammatory activation, and barrier function in vitro and in vivo.

doi: 10.1002/alz.70077

Figure Lengend Snippet: FIGURE 1 Protofibrillar tau induces loss of barrier resistance independently of cell death. A, Transmission electron microscope analysis of tau aggregates after negative staining with 1% uranyl acetate, reveals primarily protofibrillar species (scale bars 600 nm). B, One hour of treatment with AcidSensor-labeled protofibrillar tau demonstrates early internalization in hCMECs. C, Left panel TEER (at 4000 Hz) measurement of hCMECs with the ECIS Zθ system illustrates barrier formation at about 48 hours after plating. Middle panel: TEER of hCMECs monolayers treated with 5 or 25 nM fibrillar tau, measured over time for 48 hours, indicates progressive loss of barrier resistance induced by protofibrillar tau species. Right panel: TEER of hCMECs treated with fibrillization vehicle, protofibrillar or monomeric tau (25 nM) for 48 hours (right). Data are represented as change in resistance normalized to the untreated control, n = 3 experiments with two replicates per condition. D, Apoptosis, measured as DNA fragmentation, of hCMECs treated for 24 hours with 5 or 25 nM tau. E, LDH release for hCMECs treated for 24 hours with 5 or 25 nM tau. F, Caspase 3/7 fluorescence in hCMECs treated with aggregated tau for 24 hours, or H2O2 as a positive control (left), with quantification of caspase positive cells (right). **Statistical significance established by one-way ANOVA (Cell Death ELISA, LDH); Tukey post test. Two-way ANOVA (ECIS). Significant P values are reported in the graphs. ANOVA, analysis of variance; hCMEC, human cerebral microvascular endothelial cell; LDH, lactate dehydrogenase; TEER, trans-endothelial electrical resistance.

Article Snippet: Barrier resistance experiments were performed in 8-well ECIS (8WE10+, Applied Biophysics; with 40 electrodes), or in 96-well (96W20idf Applied Biophysics; with 20 electrodes in an interdigitated configuration) gold-plated arrays pre-treated with 10 mM L-Cysteine (Alfa Aesar, A10435) for 15 minutes at room temperature then pre-coated with rat tail collagen (gibco, A10483-01), diluted 1:20 in 0.15 M NaCl according to the manufacturer’s instructions. hCMECs (400,000 cells/ml) were seeded andmonitored for≈ 48 hours until the electrical resistance reached a plateau at a frequency of 4000 Hz.

Techniques: Transmission Assay, Microscopy, Negative Staining, Labeling, Control, Fluorescence, Positive Control, Enzyme-linked Immunosorbent Assay

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Fibrillar tau alters cerebral endothelial cell metabolism, vascular inflammatory activation, and barrier function in vitro and in vivo.

doi: 10.1002/alz.70077

Figure Lengend Snippet: FIGURE 4 Modulation of glycolytic metabolism reverts fibrillar tau-mediated loss of barrier resistance. A, HA decreases tau-mediated IL-4 production in hCMECs 24 hours after treatment. B, Western blot analysis of hCMECs co-treated with tau and HA reveals a decrease in VCAM-1 expression in the presence of the glycolysis inhibitor. C, TEER measurement after co-treatment with protofibrillar tau and 100 nM HA indicates that blocking glycolysis prevents tau-mediated loss of barrier resistance. D, 2-deoxyglucose partially rescues tau-mediated loss of barrier resistance after 3 hours of tau treatment. Statistical significance established by one-way ANOVA (WB), and Tukey post hoc test, two-way ANOVA (Electrical Cell Impedance Sensing-Zθ). Significant P values are reported in the graphs. ANOVA, analysis of variance; hCMEC, human cerebral microvascular endothelial cell; HA, heptelidic acid; IL, interleukin; TEER, trans-endothelial electrical resistance; VCAM-1, vascular cell adhesion molecule 1; WB, western blot

Article Snippet: Barrier resistance experiments were performed in 8-well ECIS (8WE10+, Applied Biophysics; with 40 electrodes), or in 96-well (96W20idf Applied Biophysics; with 20 electrodes in an interdigitated configuration) gold-plated arrays pre-treated with 10 mM L-Cysteine (Alfa Aesar, A10435) for 15 minutes at room temperature then pre-coated with rat tail collagen (gibco, A10483-01), diluted 1:20 in 0.15 M NaCl according to the manufacturer’s instructions. hCMECs (400,000 cells/ml) were seeded andmonitored for≈ 48 hours until the electrical resistance reached a plateau at a frequency of 4000 Hz.

Techniques: Western Blot, Expressing, Blocking Assay

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Fibrillar tau alters cerebral endothelial cell metabolism, vascular inflammatory activation, and barrier function in vitro and in vivo.

doi: 10.1002/alz.70077

Figure Lengend Snippet: FIGURE 6 Human-derived fibrillar tau induces loss of barrier resistance and pro-inflammatory EC activation. A, TEER trace (at 4000 Hz) of hCMECs treated with 25 nM aggregated 1N4R tau or 25 nM human-derived tau fibrils. B, Representative trace for glycolysis stress test ECAR for hCMECs treated with human-derived fibrillar tau for 24 hours. C, ECAR analysis in hCMECs treated with 25 nM human-derived tau fibrils reveals increased glycolytic rates. D, Western blot analysis of VCAM-1 expression for hCMECs treated with human fibrillar tau (25 nM) for 24 hours. E, Immunocytochemical assessment of protofibrillar 1N4R tau and human-derived tau fibrils on hCMEC monolayers after 24 hours of treatment. Statistical significance established by Student t test (WB, SH) and two-way analysis of variance (Electrical Cell Impedance Sensing-Zθ). Significant P values are reported in the graphs. EC, endothelial cell; ECAR, extracellular acidification rate; hCMEC, human cerebral microvascular endothelial cell; SH, Seahorse; TEER, trans-endothelial electrical resistance; VCAM-1, vascular cell adhesion molecule 1; WB, western blot.

Article Snippet: Barrier resistance experiments were performed in 8-well ECIS (8WE10+, Applied Biophysics; with 40 electrodes), or in 96-well (96W20idf Applied Biophysics; with 20 electrodes in an interdigitated configuration) gold-plated arrays pre-treated with 10 mM L-Cysteine (Alfa Aesar, A10435) for 15 minutes at room temperature then pre-coated with rat tail collagen (gibco, A10483-01), diluted 1:20 in 0.15 M NaCl according to the manufacturer’s instructions. hCMECs (400,000 cells/ml) were seeded andmonitored for≈ 48 hours until the electrical resistance reached a plateau at a frequency of 4000 Hz.

Techniques: Derivative Assay, Activation Assay, Western Blot, Expressing