Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: LPS induces GSK-3beta activation in dose- and time-dependent manners in HPMECs. Expression of P-GSK-3beta and GSK-3beta was detected after incubation with different concentrations of LPS for 1 h (A) . The expression of P-GSK-3beta was represented as a histogram according to band intensities (B) . Expression of P-GSK-3beta and GSK-3beta was examined at indicated time points after stimulation with LPS (0.1 μg/ml) in HPMECs (C) . The Western blotting results are presented as a histogram showing the band intensity values (D) . * P < 0.05 vs. LPS un-treatment group.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Activation Assay, Expressing, Incubation, Western Blot

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: Involvement of GSK-3beta in LPS-induced GEF-H1/ROCK signaling activation. HPMECs were incubated with LPS (0.1 μg/ml) at different indicated times, and the GEF-H1 and myosin-associated phosphatase type 1 (P-MYPT 1: the substrate of ROCK) were detected by Western blot assay (A) . The expression of GEF-H1 and P-MYPT 1 were represented as a histogram according to band intensities (B) . * < 0.05 vs. LPS un-treatment group. Inhibition effect of GSK-3beta activity in HPMECs was analyzed by Western blot (C,D) . * P < 0.05 vs. the negative control group, # P < 0.05 vs. the corresponding LPS treatment group. HPMECs were pretreated with SB-216763 (20 μM) for 1 h and then were exposed to LPS (0.1 μg/ml) for 1 h. The expression of GEF-H1 and P-MYPT 1 were determined by Western blot (E) . The Western blotting results are presented as a histogram showing the band intensity values (F) . * P < 0.05 vs. the negative control group, # P < 0.05 vs. the corresponding LPS treatment group.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Activation Assay, Incubation, Western Blot, Expressing, Inhibition, Activity Assay, Negative Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: GSK-3beta signaling is involved in LPS-induced HPMECs barrier disruption. The HPMECs were plated on the gold microelectrodes. When HPMECs formed monolayers and reached stable TER values, the SB-216763 (20 μM) was added. After 1 h, the medium or LPS (0.1 μg/ml) was added for another 6 h. The HPMEC monolayers permeability was determined by real-time TER measurement (A) . The results of the 3 h LPS stimulation were represented as a histogram in (B) according to the TER curves. * P < 0.05 vs. negative control. # P < 0.05 vs. corresponding LPS-stimulated group.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Disruption, Permeability, Negative Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: LPS induces degradation of beta-catenin and ZO-1 in HPMECs monolayer. LPS (0.1 μg/ml) induced down-regulation of ZO-1 expression and increase of phosphorylated degradation of beta-catenin in a time-dependent manner (A) . The Western blotting results are presented as a histogram showing the band intensity values (B) . * P < 0.05 vs. LPS un-treatment group.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Expressing, Western Blot

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: GSK-3beta/GEF-H1/ROCK signaling is required for LPS-induced degradation of beta-catenin and ZO-1. After transfection with GEF-H1 siRNA and Control siRNA for 48 h, HPMECs were treated with SB-216763 (20 μM) and/or Y-27632 (10 μM) for another 1 h prior to LPS stimulation (0.1 μg/ml) for 3 h. The expression of ZO-1 was determined by immunoblotting, and GAPDH protein was used as loading control (A) . The Western blotting results are presented as a histogram showing the band intensity values (B) . The expression of P-beta-catenin was determined by immunoblotting, and GSK-3beta and GAPDH proteins were used as control (C) . The Western blotting results are presented as a histogram showing the band intensity values (D) . * P < 0.05 vs. negative control. # P < 0.05 vs. corresponding LPS-stimulated group. NS, no significance.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Transfection, Control, Expressing, Western Blot, Negative Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: GSK-3Beta-Dependent Activation of GEF-H1/ROCK Signaling Promotes LPS-Induced Lung Vascular Endothelial Barrier Dysfunction and Acute Lung Injury

doi: 10.3389/fcimb.2017.00357

Figure Lengend Snippet: GSK-3beta/GEF-H1/ROCK pathway is involved in LPS-induced HPMECs barrier disruption by beta-catenin and ZO-1. HPMECs monolayer was pretreated with SB-216763 (20 μM) (A) , GEF-H1 siRNA (B) , or Y-27632 (10 μM) (C) , for indicated times and then was exposed to LPS (0.1 μg/ml) for 3 h before fixation and staining with anti-beta-catenin and anti-ZO-1 antibody as described in Materials and Methods. Beta-catenin (green) and ZO-1 (green) were visualized by immunofluorescence microscopy. Red arrows not only represent the expression of beta-catenin and ZO-1 in the membrane of HPMECs but also represent the cell-cell gaps formation in the ECs monolayer.

Article Snippet: Primary human pulmonary micro-vascular endothelial cells (HPMECs) were obtained ScienCell Research Laboratories and maintained in ScienCell Endothelial Cell Medium in a humidified 37°C, 5% CO 2 incubator.

Techniques: Disruption, Staining, Immunofluorescence, Microscopy, Expressing, Membrane

Journal: Acta biomaterialia

Article Title: Probing prodrug metabolism and reciprocal toxicity with an integrated and humanized multi-tissue organ-on-a-chip platform

doi: 10.1016/j.actbio.2020.02.015

Figure Lengend Snippet: A summary of each organoid or tissue construct type with the cells making them up. Also described are the various cell sources and cell types (primary, cell line, iPSC-derived, etc.), relative percentage in each model, and documented tissue specific functionalities.

Article Snippet: Organoid or tissue construct type Cell types cell source Type of cell Percentage in model Documented functionality Citation Liver Hepatocytes Bioreclamation Primary human 75% Albumin and urea secretion; Drug metabolism; Toxicity responses [ 18 , 47 ] Stellate cells Bioreclamation Primary human 10% Kupffer cells Sekisui XenoTech Primary human 10% Liver-Derived endothelial cells Lonza primary human 5% Cardiac Cardiomyocytes Ncardia iPSC 75% Spontaneous beating; Beating changes in response to drugs; Toxicity responses [ 18 , 47 ] Cardiac fibroblasts ScienCell Primary HUMAN 20% Cardiac endothelial cells ScienCell Primary human 5% Lung (3-tissue system) A549 Alveolar epithelial cells Lonza Human cell line 100% n/a – Lung (6-tissue system) Lung fibroblasts Lonza Primary human 20% n/a – Human bronchial epithelial cells Lonza Primary human 80% Endothelial HUVEC Lonza Primary human 100% n/a – Testis Spermatogonial stem cells Donor via NDRI Primary human ~80% Androgen production; Supporting germ cell differentiation; Toxicity responses; Virus replication [ 34 – 36 ] Leydig cells Donor via NDRI Primary human ~10% Sertoli cells Donor via NDRI Primary human ~10% Peritubular cells Donor via NDRI Primary human Contaminant of SSC populations Brain Brain microvascular endothelial cells Cell Systems Primary human 30% Barrier function to the middle of the spheroid [ 37 ] Brain vascular pericytes ScienCell Primary human 15% Astrocytes ScienCell Primary human 15% Microglia Tempo Bioscience IPSC 5% Oligodendrocytes Tempo Bioscience Oligodendrocyte progenitor cells 15% Neural cells Axol Biosciences Neural stem cells 20% Open in a separate window A summary of each organoid or tissue construct type with the cells making them up.

Techniques: Construct, Cell Differentiation, Virus

Journal: Acta biomaterialia

Article Title: Probing prodrug metabolism and reciprocal toxicity with an integrated and humanized multi-tissue organ-on-a-chip platform

doi: 10.1016/j.actbio.2020.02.015

Figure Lengend Snippet: Drug toxicity assessment of ifosfamide in a 6-organoid system. L/D imaging of liver, cardiac, lung, endothelial, brain, and testis organoids under Control (no drug, a–c), Condition 1 (with ifosfamide and liver organoid, d–f), and Condition 2 (with ifosfamide and without liver organoid, g and h). In Condition 1, the metabolized drug caused downstream toxicity in the brain organoid; in Condition 2, no significant toxicity occurred. Green stain: calcein AM-stained viable cells; Red stain: ethidium homodimer 1-stained dead cells. Scale bar represents 100 μM. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Organoid or tissue construct type Cell types cell source Type of cell Percentage in model Documented functionality Citation Liver Hepatocytes Bioreclamation Primary human 75% Albumin and urea secretion; Drug metabolism; Toxicity responses [ 18 , 47 ] Stellate cells Bioreclamation Primary human 10% Kupffer cells Sekisui XenoTech Primary human 10% Liver-Derived endothelial cells Lonza primary human 5% Cardiac Cardiomyocytes Ncardia iPSC 75% Spontaneous beating; Beating changes in response to drugs; Toxicity responses [ 18 , 47 ] Cardiac fibroblasts ScienCell Primary HUMAN 20% Cardiac endothelial cells ScienCell Primary human 5% Lung (3-tissue system) A549 Alveolar epithelial cells Lonza Human cell line 100% n/a – Lung (6-tissue system) Lung fibroblasts Lonza Primary human 20% n/a – Human bronchial epithelial cells Lonza Primary human 80% Endothelial HUVEC Lonza Primary human 100% n/a – Testis Spermatogonial stem cells Donor via NDRI Primary human ~80% Androgen production; Supporting germ cell differentiation; Toxicity responses; Virus replication [ 34 – 36 ] Leydig cells Donor via NDRI Primary human ~10% Sertoli cells Donor via NDRI Primary human ~10% Peritubular cells Donor via NDRI Primary human Contaminant of SSC populations Brain Brain microvascular endothelial cells Cell Systems Primary human 30% Barrier function to the middle of the spheroid [ 37 ] Brain vascular pericytes ScienCell Primary human 15% Astrocytes ScienCell Primary human 15% Microglia Tempo Bioscience IPSC 5% Oligodendrocytes Tempo Bioscience Oligodendrocyte progenitor cells 15% Neural cells Axol Biosciences Neural stem cells 20% Open in a separate window A summary of each organoid or tissue construct type with the cells making them up.

Techniques: Imaging, Staining

Journal: Acta biomaterialia

Article Title: Probing prodrug metabolism and reciprocal toxicity with an integrated and humanized multi-tissue organ-on-a-chip platform

doi: 10.1016/j.actbio.2020.02.015

Figure Lengend Snippet: Viability quantification under insult by ifosfamide in a 6-organoid system. Quantification of live cell ratios (L/L + D) for liver, cardiac, lung, endothelial, brain, and testis organoids under Control (no drug), Condition 1 (drug with liver organoid), and Condition 2 (drug without liver organoid). A significant reduction in viability is observed for the brain organoid under Condition 1 (red). Statistical significance: ** p < 0.05. (For interpretation of the references to colour in this figure legend, the reader is referred to the web version of this article.)

Article Snippet: Organoid or tissue construct type Cell types cell source Type of cell Percentage in model Documented functionality Citation Liver Hepatocytes Bioreclamation Primary human 75% Albumin and urea secretion; Drug metabolism; Toxicity responses [ 18 , 47 ] Stellate cells Bioreclamation Primary human 10% Kupffer cells Sekisui XenoTech Primary human 10% Liver-Derived endothelial cells Lonza primary human 5% Cardiac Cardiomyocytes Ncardia iPSC 75% Spontaneous beating; Beating changes in response to drugs; Toxicity responses [ 18 , 47 ] Cardiac fibroblasts ScienCell Primary HUMAN 20% Cardiac endothelial cells ScienCell Primary human 5% Lung (3-tissue system) A549 Alveolar epithelial cells Lonza Human cell line 100% n/a – Lung (6-tissue system) Lung fibroblasts Lonza Primary human 20% n/a – Human bronchial epithelial cells Lonza Primary human 80% Endothelial HUVEC Lonza Primary human 100% n/a – Testis Spermatogonial stem cells Donor via NDRI Primary human ~80% Androgen production; Supporting germ cell differentiation; Toxicity responses; Virus replication [ 34 – 36 ] Leydig cells Donor via NDRI Primary human ~10% Sertoli cells Donor via NDRI Primary human ~10% Peritubular cells Donor via NDRI Primary human Contaminant of SSC populations Brain Brain microvascular endothelial cells Cell Systems Primary human 30% Barrier function to the middle of the spheroid [ 37 ] Brain vascular pericytes ScienCell Primary human 15% Astrocytes ScienCell Primary human 15% Microglia Tempo Bioscience IPSC 5% Oligodendrocytes Tempo Bioscience Oligodendrocyte progenitor cells 15% Neural cells Axol Biosciences Neural stem cells 20% Open in a separate window A summary of each organoid or tissue construct type with the cells making them up.

Techniques:

Journal: Diabetes

Article Title: Decreased Cerebrovascular Brain-Derived Neurotrophic Factor–Mediated Neuroprotection in the Diabetic Brain

doi: 10.2337/db10-1371

Figure Lengend Snippet: BDNF expression is reduced in the diabetic brain endothelium. Representative images of cortical sections (2-mm thick) from 6-week diabetic and age-matched nondiabetic rats immunostained for BDNF. Blood vessels (arrows) are visualized by CD31 staining ( left ). Cortex microvessels from a diabetic rat show decreased BDNF immunofluorescence when compared with those of a nondiabetic rat ( right ). Magnification 40×. (A high-quality digital representation of this figure is available in the online issue.)

Article Snippet: Primary human brain microvessel endothelial cells were purchased from Cell Systems Corporation (Kirkland, WA), mostly derived from a heterogenous mix of rapidly autopsied human brains obtained within a few hours after death.

Techniques: Expressing, Staining, Immunofluorescence

Journal: Cells

Article Title: The Anti-Inflammatory Effect of the β1-Adrenergic Receptor Antagonist Metoprolol on High Glucose Treated Human Microvascular Retinal Endothelial Cells

doi: 10.3390/cells11010051

Figure Lengend Snippet: The effects of metoprolol on the pro-inflammatory cytokines and VEGF-A factors in HREC that was stimulated with HG. The cells were cultured in NG (5 mM) or in HG (25 mM) alone or supplemented with metoprolol (Met, 10 µM), epinephrine (Epi, 1 µM), or both compounds. The data were gathered after 48 h from treatment addition. Quantitative analysis of TNF-α ( A ), IL-1β ( B ), and VEGF ( C ) mRNA levels, as evaluated by qRT-PCR. The mRNA levels of each group were normalized to the housekeeping reference gene ribosomal 18S RNA. In the histograms, the values are expressed as fold change of those that were detected in HREC that was cultured in NG condition. Each value represents the mean ± SD that was obtained from three independent experiments with four parallel samples for group in each experiment. * p < 0.05 vs. NG; # p < 0.05 vs. HG. The non-parametric Mann-Whitney test was used for pairwise comparisons.

Article Snippet: Primary human retinal endothelial cells (HREC) were purchased from Innoprot (Elexalde Derio, Spain) and were cultured with endothelial cell medium ECM that was supplemented with 5% fetal bovine serum (FBS), 1% endothelial cell growth supplement (ECGS), 100 U/mL penicillin, and 100 µg/mL streptomycin that were purchased from Innoprot.

Techniques: Cell Culture, Quantitative RT-PCR, MANN-WHITNEY

Journal: Cells

Article Title: The Anti-Inflammatory Effect of the β1-Adrenergic Receptor Antagonist Metoprolol on High Glucose Treated Human Microvascular Retinal Endothelial Cells

doi: 10.3390/cells11010051

Figure Lengend Snippet: The effects of metoprolol on Nrf2 nuclear translocation in HREC that was stimulated with high glucose levels. ( A ) The representative of immunocytochemically stained images for Nrf2 in HREC that was cultured in normal glucose levels (NG, 5 mM) or in HG, (25 mM) alone or supplemented with metoprolol (Met, 10 µM) for 48 h (in red: a′–c′ respectively). Moreover, the relative Hoechst-stained nuclei (a–c, respectively) and merged channels (a″–c″, respectively) are shown. All images were acquired with the Operetta High-Content Imaging System using a 20× magnification; scale bar = 100 µm. ( B ) High-content screening analysis was used to quantify the green fluorescence (Cy3) relative to the Nrf2 staining in the cytoplasm and nuclei. ( C ) Quantification of Nrf2 positive nuclei. The data are expressed as the mean ± SD from at least six fields/well randomly selected and each reporting more than 15 cells/field. All of the data represent the mean ± SD that were obtained from at least three independent experiments with four parallel samples for group in each experiment. * p < 0.05 vs. NG; # p < 0.05 vs. HG. The non-parametric Mann-Whitney test was used for pairwise comparisons.

Article Snippet: Primary human retinal endothelial cells (HREC) were purchased from Innoprot (Elexalde Derio, Spain) and were cultured with endothelial cell medium ECM that was supplemented with 5% fetal bovine serum (FBS), 1% endothelial cell growth supplement (ECGS), 100 U/mL penicillin, and 100 µg/mL streptomycin that were purchased from Innoprot.

Techniques: Translocation Assay, Staining, Cell Culture, Imaging, High Content Screening, Fluorescence, MANN-WHITNEY