bovine pulmonary artery endothelial cells (paec)  (Vec Technologies)

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: l-NAME attenuates hyperoxia-induced disruption of endothelial monolayer barrier integrity. PAECs were treated with and without l-NAME (3 mm) and exposed to hyperoxia for 48 h. TEER was continuously monitored as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer under normoxia and hyperoxia. B, changes in TEER of endothelial monolayer under hyperoxia with and without l-NAME. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus l-NAME+normoxia; **, p < 0.05 versus control hyperoxia.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, Control

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Uric acid prevents hyperoxia-induced disruption of lung endothelial barrier in the second phase and apoptosis. PAECs were treated with and without uric acid (3 mm) and exposed to hyperoxia for 48 h, during which TEER was continuously monitored. After exposure, apoptotic cells were detected using TUNEL assay as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer. B, alterations in the numbers of TUNEL-positive cells. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus UA+normoxia; **, p < 0.05 versus hyperoxia. UA = uric acid.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, TUNEL Assay

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT attenuates hyperoxia-induced disruption of endothelial monolayer barrier integrity. PAECs were treated with peptide P326TAT (20 μm) or control peptide PlwTAT (20 μm) and exposed to hyperoxia for 48 h. TEER was continuously monitored as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer under hyperoxia with control peptide PlwTAT. B, changes in TEER of endothelial monolayer under hyperoxia with P326TAT. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus P326TAT+normoxia or PlwTAT+normoxia; **, p < 0.05 versus hyperoxia.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, Control

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT attenuates hyperoxia-induced apoptosis of lung endothelial cells. PAECs were exposed to hyperoxia or normoxia in the presence of P326TAT (20 μm) or control peptide PlwTAT (20 μm) for 48 h, and then apoptosis was evaluated using TUNEL assay. A, representative images of TUNEL staining. WO peptide, without peptide. B, bar graph depicting the changes in the numbers of TUNEL-positive cells. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus PlwTAT+hyperoxia or hyperoxia only.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Control, TUNEL Assay, Staining

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT prevents both caspase-dependent and caspase-independent apoptosis of lung endothelial cells. PAECs were treated with peptide P326TAT (20 μm) or control peptide PlwTAT (20 μm) and exposed to hyperoxia for 48 h, after which caspase-3 activity and AIF protein level in the nuclear fraction were measured as described under “Experimental Procedures.” A, changes in caspase-3 activity. B, representative image of Western blot of AIF. WO peptide, without peptide. C, bar graph depicting changes in nuclear AIF levels of PAECs. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Control, Activity Assay, Western Blot

Journal: Transactions of the American Clinical and Climatological Association

Article Title: CIGARETTE SMOKING INCREASES THE RISK OF ACUTE RESPIRATORY DISTRESS SYNDROME

doi:

Figure Lengend Snippet: (A) Cigarette smoke extract (CSE) exacerbated LPS-induced increase in endothelial monolayer permeability. Bovine pulmonary artery endothelial cells (PAEC) were treated with vehicle (5% sham PBS) or 5% CSE in the absence or presence of LPS (0.5 mg/ml) for indicated times. Endothelial monolayer permeability was assessed by measuring electrical resistance across monolayers over time by electrical cell impedance sensor (ECIS). Data are presented as means ± SE of the normalized electrical resistance relative to the time when agents were added, as indicated by an arrow; n = 6. (B) Changes in RhoA GTPase activation in CSE-treated (S) or vehicle-treated (V) PAEC. (C) Changes in Focal Adhesion Kinase (FAK) activation in CSE-treated (S) or vehicle-treated (V) PAEC. Reproduced with permission of the American Journal of Physiology (11).

Article Snippet: Bovine pulmonary artery endothelial cells (PAEC) and rat lung microvascular endothelial cells (LMVEC) were purchased from VEC Technologies Inc. (Rensselaer, NY), used between passages 3–8, and maintained in culture ( 12 ).

Techniques: Permeability, Activation Assay

Journal: Transactions of the American Clinical and Climatological Association

Article Title: CIGARETTE SMOKING INCREASES THE RISK OF ACUTE RESPIRATORY DISTRESS SYNDROME

doi:

Figure Lengend Snippet: (A) Cigarette smoke extract (CSE) exacerbated LPS-induced increase in endothelial monolayer permeability. Bovine pulmonary artery endothelial cells (PAEC) were treated with vehicle (5% sham PBS) or 5% CSE in the absence or presence of LPS (0.5 mg/ml) for indicated times. Endothelial monolayer permeability was assessed by measuring electrical resistance across monolayers over time by electrical cell impedance sensor (ECIS). Data are presented as means ± SE of the normalized electrical resistance relative to the time when agents were added, as indicated by an arrow; n = 6. (B) Changes in RhoA GTPase activation in CSE-treated (S) or vehicle-treated (V) PAEC. (C) Changes in Focal Adhesion Kinase (FAK) activation in CSE-treated (S) or vehicle-treated (V) PAEC. Reproduced with permission of the American Journal of Physiology (11).

Article Snippet: Endothelial Cells Bovine pulmonary artery endothelial cells (PAEC) and rat lung microvascular endothelial cells (LMVEC) were purchased from VEC Technologies Inc. (Rensselaer, NY), used between passages 3–8, and maintained in culture ( 12 ).

Techniques: Permeability, Activation Assay

Journal: Pulmonary Circulation

Article Title: Sustained adenosine exposure causes endothelial mitochondrial dysfunction via equilibrative nucleoside transporters

doi: 10.1177/2045894020924994

Figure Lengend Snippet: Sustained adenosine exposure increased mitochondrial ROS in lung EC. PAECs were treated with vehicle (V) or 50 µM adenosine plus 50 µM DCF (AD) for 5, 24, 30, and 48 h. Following treatment, the cells were washed and then incubated with MitoTracker Green to stain mitochondria and MitoSOX Red to label mitochondrial ROS. The cells were visualized by fluorescence microscopy. Panel a represents three independent experiments. Panel b: after treatment, the cells were washed twice and then stained with Hoechst 33342 for 20 min to quantify cell numbers. The cells were then washed twice and stained with MitoSOX Red for 20 min. The fluorescence intensity of Hoechst 33342 and MitoSOX Red were captured by a fluorescence spectrometer with Ex 360 nm/Em 460 nm and Ex 510 nm/Em 580 nm, respectively. The relative numbers of cells in each well were normalized by the fluorescence intensity of Hoechst 33342. The fluorescence intensity of MitoSOX Red in each well was then adjusted by the relative numbers of cells. The levels of mitochondrial ROS were expressed as fold changes of the fluorescence intensity of MitoSOX Red relative to that of untreated cells. The data are presented as mean ± SD of four independent experiments ( n = 4). * p < 0.05 vs. control cells.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were purchased from Vec Technologies (Rensselaer, NY) and used between passages 3 and 7 in this report.

Techniques: Incubation, Staining, Fluorescence, Microscopy, Control

Journal: Pulmonary Circulation

Article Title: Sustained adenosine exposure causes endothelial mitochondrial dysfunction via equilibrative nucleoside transporters

doi: 10.1177/2045894020924994

Figure Lengend Snippet: ENT 1/2 and MAK kinases mediated sustained adenosine-induced mitochondrial oxidative stress. PAECs were pre-treated with ENT 1/2 inhibitors, DPM (10 µM) or NBTI (10 µM), an A 2A R antagonist, DPMX (10 µM), or an A 2B R antagonist, MRS1754 (10 µM), for 1 h and then incubated with 50 µM adenosine and 50 µM DCF (AD) in the absence or presence of individual inhibitor for 30 h (Panel a). Similarly, the cells were pre-treated with a cytosolic antioxidant, NAC (12.5 mM), a mitochondrial ROS scavenger, mitoTEMPO (5 µM), a p38 inhibitor, SB203580 (SB, 10 µM), or a JNK inhibitor, SP600125 (SP, 10 µM), and then incubated with 50 µM adenosine and 50 µM DCF (AD) in the absence or presence of individual inhibitors for 30 h (Panel b). Following the treatments, the cells were washed twice and incubated with Hoechst 33342 and MitoSOX Red to quantify cell numbers and mitochondrial ROS levels. The relative numbers of cells in each well were normalized by the fluorescence intensity of Hoechst 33342. The fluorescence intensity of MitoSOX Red in each well was then adjusted by the relative numbers of cells. The levels of mitochondrial ROS were expressed as fold changes of the fluorescence intensity of MitoSOX Red relative to that of untreated cells. The data are presented as mean ± SD of three independent experiments ( n = 3). * p < 0.05. Veh: vehicle; DPM: dipyridamole; NBTI: S-(4-Nitrobenzyl)-6-thioinosine; DPMX: 1,3-dipropyl-7-methylxanthine; MRS1754: 8-(4{[(4-cyano)phenylcarbamoylmethyl]oxy}epheyl)-1,3-di-(n-propyl)xanthine; NAC: N-acetyl-cysteine.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were purchased from Vec Technologies (Rensselaer, NY) and used between passages 3 and 7 in this report.

Techniques: Incubation, Fluorescence

Journal: Pulmonary Circulation

Article Title: Sustained adenosine exposure causes endothelial mitochondrial dysfunction via equilibrative nucleoside transporters

doi: 10.1177/2045894020924994

Figure Lengend Snippet: Sustained adenosine exposure impaired mitochondrial respiration via ENT 1/2 -mediated intracellular signaling. PAECs were pre-treated with ENT 1/2 inhibitors, DPM (10 µM) or NBTI (10 µM) for 1 h and then incubated with 50 µM adenosine and 50 µM DCF (AD) in the absence or presence of individual inhibitor for 30 h (Panel a). PAECs were pre-treated with an A 2A R antagonist, DPMX (10 µM), or an A 2B R antagonist, MRS1754 (10 µM), for 1 h and then incubated with 50 µM adenosine and 50 µM DCF (AD) in the absence or presence of DPM (10 µM) with or without DPMX (10 µM) or MRS1754 (10 µM) for 30 h (Panel b). PAECs were pre-treated with AK inhibitor, ITU (10 µM) for 1 h and then incubated with 50 µM adenosine and 50 µM DCF (AD) in the absence or presence of ITU (10 µM) for 30 h (Panel c). The cells were then washed twice with basal seahorse media and subjected to assessments of OCR using XF96 seahorse analyzer. Oligomycin, FCCP, and a mixture of antimycin A and rotenone were added to the system to assess OCR response to those inhibitors for oxidative phosphorylation. The data are presented as mean ± SD of three independent experiments ( n = 3). * p < 0.05. NBTI: S-(4-Nitrobenzyl)-6-thioinosine; DPM: dipyridamole; ITU: 5′-iodotubercidin; DPMX; 1,3-dipropyl-7-methylxanthine; MRS1754: 8-(4{[(4-cyano)phenylcarbamoylmethyl]oxy}epheyl)-1,3-di-(n-propyl)xanthine.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were purchased from Vec Technologies (Rensselaer, NY) and used between passages 3 and 7 in this report.

Techniques: Incubation, Phospho-proteomics

Journal: Pulmonary Circulation

Article Title: Sustained adenosine exposure causes endothelial mitochondrial dysfunction via equilibrative nucleoside transporters

doi: 10.1177/2045894020924994

Figure Lengend Snippet: Sustained adenosine exposure increased mitochondrial fission and mitophagy. PAECs were treated with vehicle (Ctrl) or 50 µM adenosine plus 50 µM DCF (AD) for 5, 30, and 48 h. The cell lysates were harvested for western blot analysis (Panels a and c). Densitometry analysis was performed with three independent experimental repeats (Panels b and d). Vinculin was used as a protein loading control. The data are presented as mean ± SD of three independent experiments ( n = 3). * p < 0.05.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were purchased from Vec Technologies (Rensselaer, NY) and used between passages 3 and 7 in this report.

Techniques: Western Blot, Control

Journal: Pulmonary Circulation

Article Title: Sustained adenosine exposure causes endothelial mitochondrial dysfunction via equilibrative nucleoside transporters

doi: 10.1177/2045894020924994

Figure Lengend Snippet: Sustained adenosine exposure increased mitochondrial fission and promoted cell death via mitochondrial ROS. PAECs were pre-treated with 5 µM MitoTEMPO (MT) for 1 h followed by incubation with 50 µM adenosine and 50 µM DCF (AD) in the presence or absence of 5 µM MitoTEMPO for 30 h or 48 h. The cells were then stained with MitoTracker green to visualize mitochondrial morphology (Panel a), processed for cellular morphology (Panel b), or a MTS cell viability assay (Panel c). Panels a and b are representative images from three independent experiments. In panel c, the data are presented as mean ± SD of three independent experiments ( n = 3). * p < 0.05.

Article Snippet: Primary bovine pulmonary artery endothelial cells (PAECs) were purchased from Vec Technologies (Rensselaer, NY) and used between passages 3 and 7 in this report.

Techniques: Incubation, Staining, Viability Assay

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: l-NAME attenuates hyperoxia-induced disruption of endothelial monolayer barrier integrity. PAECs were treated with and without l-NAME (3 mm) and exposed to hyperoxia for 48 h. TEER was continuously monitored as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer under normoxia and hyperoxia. B, changes in TEER of endothelial monolayer under hyperoxia with and without l-NAME. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus l-NAME+normoxia; **, p < 0.05 versus control hyperoxia.

Article Snippet: Cell Culture Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, Control

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Uric acid prevents hyperoxia-induced disruption of lung endothelial barrier in the second phase and apoptosis. PAECs were treated with and without uric acid (3 mm) and exposed to hyperoxia for 48 h, during which TEER was continuously monitored. After exposure, apoptotic cells were detected using TUNEL assay as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer. B, alterations in the numbers of TUNEL-positive cells. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus UA+normoxia; **, p < 0.05 versus hyperoxia. UA = uric acid.

Article Snippet: Cell Culture Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, TUNEL Assay

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT attenuates hyperoxia-induced disruption of endothelial monolayer barrier integrity. PAECs were treated with peptide P326TAT (20 μm) or control peptide PlwTAT (20 μm) and exposed to hyperoxia for 48 h. TEER was continuously monitored as described under “Experimental Procedures.” A, changes in TEER of endothelial monolayer under hyperoxia with control peptide PlwTAT. B, changes in TEER of endothelial monolayer under hyperoxia with P326TAT. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus P326TAT+normoxia or PlwTAT+normoxia; **, p < 0.05 versus hyperoxia.

Article Snippet: Cell Culture Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Disruption, Control

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT attenuates hyperoxia-induced apoptosis of lung endothelial cells. PAECs were exposed to hyperoxia or normoxia in the presence of P326TAT (20 μm) or control peptide PlwTAT (20 μm) for 48 h, and then apoptosis was evaluated using TUNEL assay. A, representative images of TUNEL staining. WO peptide, without peptide. B, bar graph depicting the changes in the numbers of TUNEL-positive cells. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia; #, p < 0.05 versus PlwTAT+hyperoxia or hyperoxia only.

Article Snippet: Cell Culture Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Control, TUNEL Assay, Staining

Journal: The Journal of Biological Chemistry

Article Title: Novel Peptide for Attenuation of Hyperoxia-induced Disruption of Lung Endothelial Barrier and Pulmonary Edema via Modulating Peroxynitrite Formation *

doi: 10.1074/jbc.M114.585356

Figure Lengend Snippet: Peptide P326TAT prevents both caspase-dependent and caspase-independent apoptosis of lung endothelial cells. PAECs were treated with peptide P326TAT (20 μm) or control peptide PlwTAT (20 μm) and exposed to hyperoxia for 48 h, after which caspase-3 activity and AIF protein level in the nuclear fraction were measured as described under “Experimental Procedures.” A, changes in caspase-3 activity. B, representative image of Western blot of AIF. WO peptide, without peptide. C, bar graph depicting changes in nuclear AIF levels of PAECs. Results are expressed as means ± S.E.; n = 4. *, p < 0.05 versus normoxia.

Article Snippet: Cell Culture Primary bovine pulmonary artery endothelial cells (PAECs) were obtained from Cell Applications (San Diego, CA).

Techniques: Control, Activity Assay, Western Blot