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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210306	1/8	Homo sapiens	Pulmonary artery endothelial cells	dUC	Letsiou E	2015	33%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 33% (75th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin Control condition Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD31/ CD31/ p-ERK/ t-ERK/ GAPDH/ moesin non-EV: None Proteomics yes Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Pulmonary artery endothelial cells EV-harvesting Medium Serum-containing medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 50,000 g and 100,000 g Pelleting performed Yes Pelleting: speed (g) 21000 Wash: time (min) 70 Wash: speed (g) 21000 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD31/ p-ERK/ t-ERK/ GAPDH/ moesin Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD31 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 EV concentration Yes EM EM-type Transmission-EM Image type Close-up, Wide-field

	EV210306	3/8	Homo sapiens	Pulmonary artery endothelial cells	dUC	Letsiou E	2015	22%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 22% (59th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin 18% cyclic mechanical stretch Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD31/ CD31/ p-ERK/ t-ERK/ GAPDH/ moesin non-EV: None Proteomics yes Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Pulmonary artery endothelial cells EV-harvesting Medium Serum-containing medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 21000 Wash: time (min) 70 Wash: speed (g) 21000 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD31/ p-ERK/ t-ERK/ GAPDH/ moesin Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD31 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 Report type Not Reported EV concentration Yes EM EM-type Transmission-EM Image type Close-up

	EV210306	4/8	Homo sapiens	Pulmonary artery endothelial cells	dUC	Letsiou E	2015	22%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 22% (59th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin LPS Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD31/ CD31/ p-ERK/ t-ERK/ GAPDH/ moesin non-EV: None Proteomics yes Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Pulmonary artery endothelial cells EV-harvesting Medium Serum-containing medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 21000 Wash: time (min) 70 Wash: speed (g) 21000 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD31/ p-ERK/ t-ERK/ GAPDH/ moesin Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD31 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 Report type Not Reported EV concentration Yes EM EM-type Transmission-EM Image type Close-up

	EV210306	2/8	Homo sapiens	Pulmonary artery endothelial cells	dUC	Letsiou E	2015	11%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 11% (30th percentile of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Cell culture supernatant Sample origin 5% cyclic mechanical stretch Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD31 non-EV: None Proteomics yes Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Pulmonary artery endothelial cells EV-harvesting Medium Serum-containing medium Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 21000 Wash: time (min) 70 Wash: speed (g) 21000 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins CD31 Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD31 Proteomics database No Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 Report type Not Reported EV concentration Yes

	EV210306	5/8	Mus musculus	Blood plasma	dUC	Letsiou E	2015	0%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 0% (median: 22% of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Blood plasma Sample origin Control condition Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD62E/ VE-cadherin/ moesin non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Blood plasma Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 20500 Wash: time (min) 45 Wash: speed (g) 20500 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins VE-cadherin/ moesin Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD62E Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 EV concentration Yes

	EV210306	6/8	Mus musculus	Blood plasma	dUC	Letsiou E	2015	0%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 0% (median: 22% of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Blood plasma Sample origin High tidal volume mechanical ventilation Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD62E/ VE-cadherin/ moesin non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Blood plasma Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 20500 Wash: time (min) 45 Wash: speed (g) 20500 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Western Blot Antibody details provided? No Detected EV-associated proteins VE-cadherin/ moesin Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD62E Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 EV concentration Yes

	EV210306	7/8	Mus musculus	Bronchoalveolar lavage	dUC	Letsiou E	2015	0%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 0% (median: 0% of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Bronchoalveolar lavage Sample origin Control condition Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD62E non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Bronchoalveolar lavage Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 20500 Wash: time (min) 45 Wash: speed (g) 20500 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD62E Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 EV concentration Yes

	EV210306	8/8	Mus musculus	Bronchoalveolar lavage	dUC	Letsiou E	2015	0%
Study summary Full title Pathologic mechanical stress and endotoxin exposure increases lung endothelial microparticle shedding. All authors Letsiou E, Sammani S, Zhang W, Zhou T, Quijada H, Moreno-Vinasco L, Dudek SM, Garcia JG Journal Am J Respir Cell Mol Biol Abstract Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produ (show more...)Acute lung injury (ALI) results from infectious challenges and from pathologic lung distention produced by excessive tidal volume delivered during mechanical ventilation (ventilator-induced lung injury [VILI]) and is characterized by extensive alveolar and vascular dysfunction. Identification of novel ALI therapies is hampered by the lack of effective ALI/VILI biomarkers. We explored endothelial cell (EC)-derived microparticles (EMPs) (0.1-1 μm) as potentially important markers and potential mediators of lung vascular injury in preclinical models of ALI and VILI. We characterized EMPs (annexin V and CD31 immunoreactivity) produced from human lung ECs exposed to physiologic or pathologic mechanical stress (5 or 18% cyclic stretch [CS]) or to endotoxin (LPS). EC exposure to 18% CS or to LPS resulted in increased EMP shedding compared with static cells (∼ 4-fold and ∼ 2.5-fold increases, respectively). Proteomic analysis revealed unique 18% CS-derived (n = 10) and LPS-derived EMP proteins (n = 43). VILI-challenged mice (40 ml/kg, 4 h) exhibited increased plasma and bronchoalveolar lavage CD62E (E-selectin)-positive MPs compared with control mice. Finally, mice receiving intratracheal instillation of 18% CS-derived EMPs displayed significant lung inflammation and injury. These findings indicate that ALI/VILI-producing stimuli induce significant shedding of distinct EMP populations that may serve as potential ALI biomarkers and contribute to the severity of lung injury. (hide) EV-METRIC 0% (median: 0% of all experiments on the same sample type) Reported Not reported Not applicable EV-enriched proteins Protein analysis: analysis of three or more EV-enriched proteins non EV-enriched protein Protein analysis: assessment of a non-EV-enriched protein qualitative and quantitative analysis Particle analysis: implementation of both qualitative and quantitative methods. For the quantitative method, the reporting of measured EV concentration is expected. electron microscopy images Particle analysis: inclusion of a widefield and close-up electron microscopy image density gradient Separation method: density gradient, at least as validation of results attributed to EVs EV density Separation method: reporting of obtained EV density ultracentrifugation specifics Separation method: reporting of g-forces, duration and rotor type of ultracentrifugation steps antibody specifics Protein analysis: antibody clone/reference number and dilution lysate preparation Protein analysis: lysis buffer composition Study data Sample type Bronchoalveolar lavage Sample origin High tidal volume mechanical ventilation Focus vesicles microparticle Separation protocol Separation protocol Gives a short, non-chronological overview of the different steps of the separation protocol. dUC = (Differential) (ultra)centrifugation DG = density gradient UF = ultrafiltration SEC = size-exclusion chromatography IAF = immuno-affinity capture dUC Protein markers EV: CD62E non-EV: None Proteomics no Show all info Study aim Function Sample Species Mus musculus Sample Type Bronchoalveolar lavage Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Between 800 g and 10,000 g Between 10,000 g and 50,000 g Pelleting performed Yes Pelleting: speed (g) 20500 Wash: time (min) 45 Wash: speed (g) 20500 Other Name other separation method dUC Characterization: Protein analysis Protein Concentration Method Not determined Flow cytometry Type of Flow cytometry LSR Fortessa Calibration bead size 0.8 Antibody details provided? No Detected EV-associated proteins CD62E Characterization: Lipid analysis No Characterization: Particle analysis Particle analysis: flow cytometry Flow cytometer type LSR Fortessa Hardware adjustment Calibration bead size 0.8 EV concentration Yes

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EV-TRACK ID	EV210306
species	Homo sapiens	Homo sapiens	Homo sapiens	Homo sapiens	Mus musculus	Mus musculus	Mus musculus	Mus musculus
sample type	Cell culture	Cell culture	Cell culture	Cell culture	Blood plasma	Blood plasma	Bronchoalveolar lavage	Bronchoalveolar lavage
cell type	Pulmonary artery endothelial cells	Pulmonary artery endothelial cells	Pulmonary artery endothelial cells	Pulmonary artery endothelial cells	NA	NA	NA	NA
medium	Serum-containing medium	Serum-containing medium	Serum-containing medium	Serum-containing medium	NA	NA	NA	NA
condition	Control condition	18% cyclic mechanical stretch	LPS	5% cyclic mechanical stretch	Control condition	High tidal volume mechanical ventilation	Control condition	High tidal volume mechanical ventilation
separation protocol	dUC	dUC	dUC	dUC	dUC	dUC	dUC	dUC
Exp. nr.	1	3	4	2	5	6	7	8
EV-METRIC %	33	22	22	11	0	0	0	0

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