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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV220305	1/2	Homo sapiens	Wharton's Jelly mesenchymal stem cell	(d)(U)C	Ngo NH	2022	78%
Study summary Full title Transformed extracellular vesicles with high angiogenic ability as therapeutics of distal ischemic tissues. All authors Ngo NH, Chang YH, Vuong CK, Yamashita T, Obata-Yasuoka M, Hamada H, Osaka M, Hiramatsu Y, Ohneda O Journal Front Cell Dev Biol Abstract The therapeutic effects of endothelial progenitor cells (EPC) in neovascularization have been sugges (show more...)The therapeutic effects of endothelial progenitor cells (EPC) in neovascularization have been suggested/ however, to date, few studies have been conducted on the ability of EPC-derived extracellular vesicles (EV) to rescue the ischemic tissues. In order to examine the functional sources of EV for cell-free therapy of ischemic diseases, we compared the functions of EPC-EV and those of Wharton's Jelly-derived mesenchymal stem cell (WJ-EV) in the flap mouse model. Our results demonstrated that in the intravenous injection, EPC-EV, but not WJ-EV, were uptaken by the ischemic tissues. However, EPC-EV showed poor abilities to induce neovascularization and the recovery of ischemic tissues. In addition, compared to EPC-EV, WJ-EV showed a higher ability to rescue the ischemic injury when being locally injected into the mice. In order to induce the secretion of high-functional EPC-EV, EPC were internalized with hypoxic pre-treated WJ-EV, which resulted in a transformed hwEPC. In comparison to EPC, hwEPC showed induced proliferation and upregulation of angiogenic genes and miRNAs and promoted angiogenic ability. Interestingly, hwEPC produced a modified EV (hwEPC-EV) that highly expressed miRNAs related to angiogenesis, such as miR-155, miR-183, and miR-296. Moreover, hwEPC-EV significantly induced the neovascularization of the ischemic tissues which were involved in promoting the proliferation, the expression of VEGF and miR-183, and the angiogenic functions of endothelial cells. Of note, hwEPC-EV were highly uptaken by the ischemic tissues and showed a greater effect with regard to inducing recovery from ischemic injury in the intravenous administration, compared to EPC-EV. Therefore, hwEPC-EV can be considered a functional candidate for cell-free therapy to treat the distal ischemic tissues. (hide) EV-METRIC 78% (97th 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 Hypoxia Focus vesicles extracellular vesicle 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 (Differential) (ultra)centrifugation Protein markers EV: TSG101/ CD40/ integrin beta-1 non-EV: None Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells Wharton's Jelly mesenchymal stem cell EV-harvesting Medium EV-depleted medium Preparation of EDS >=18h at >= 100,000g Cell viability (%) 99.9 Cell count 1000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: rotor type Type 70 Ti Pelleting: speed (g) 140000 Wash: volume per pellet (ml) 35 Wash: time (min) 70 Wash: Rotor Type Type 70 Ti Wash: speed (g) 140000 Characterization: Protein analysis Protein Concentration Method Bradford Protein Yield (µg) per million cells Western Blot Detected EV-associated proteins TSG101/ CD40/ integrin beta-1 Not detected contaminants ApoA1 Characterization: Lipid analysis No Characterization: Particle analysis DLS Report type Size range/distribution Reported size (nm) 0-1000 EM EM-type Transmission-EM Image type Close-up, Wide-field

	EV220305	2/2	Homo sapiens	endothelial progenitor cell	(d)(U)C	Ngo NH	2022	78%
Study summary Full title Transformed extracellular vesicles with high angiogenic ability as therapeutics of distal ischemic tissues. All authors Ngo NH, Chang YH, Vuong CK, Yamashita T, Obata-Yasuoka M, Hamada H, Osaka M, Hiramatsu Y, Ohneda O Journal Front Cell Dev Biol Abstract The therapeutic effects of endothelial progenitor cells (EPC) in neovascularization have been sugges (show more...)The therapeutic effects of endothelial progenitor cells (EPC) in neovascularization have been suggested/ however, to date, few studies have been conducted on the ability of EPC-derived extracellular vesicles (EV) to rescue the ischemic tissues. In order to examine the functional sources of EV for cell-free therapy of ischemic diseases, we compared the functions of EPC-EV and those of Wharton's Jelly-derived mesenchymal stem cell (WJ-EV) in the flap mouse model. Our results demonstrated that in the intravenous injection, EPC-EV, but not WJ-EV, were uptaken by the ischemic tissues. However, EPC-EV showed poor abilities to induce neovascularization and the recovery of ischemic tissues. In addition, compared to EPC-EV, WJ-EV showed a higher ability to rescue the ischemic injury when being locally injected into the mice. In order to induce the secretion of high-functional EPC-EV, EPC were internalized with hypoxic pre-treated WJ-EV, which resulted in a transformed hwEPC. In comparison to EPC, hwEPC showed induced proliferation and upregulation of angiogenic genes and miRNAs and promoted angiogenic ability. Interestingly, hwEPC produced a modified EV (hwEPC-EV) that highly expressed miRNAs related to angiogenesis, such as miR-155, miR-183, and miR-296. Moreover, hwEPC-EV significantly induced the neovascularization of the ischemic tissues which were involved in promoting the proliferation, the expression of VEGF and miR-183, and the angiogenic functions of endothelial cells. Of note, hwEPC-EV were highly uptaken by the ischemic tissues and showed a greater effect with regard to inducing recovery from ischemic injury in the intravenous administration, compared to EPC-EV. Therefore, hwEPC-EV can be considered a functional candidate for cell-free therapy to treat the distal ischemic tissues. (hide) EV-METRIC 78% (97th 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 treated with Hypoxia WJ-MSC derived EV Focus vesicles extracellular vesicle 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 (Differential) (ultra)centrifugation Protein markers EV: TSG101/ CD40/ Integrin beta-1 non-EV: None Proteomics no Show all info Study aim Function Sample Species Homo sapiens Sample Type Cell culture supernatant EV-producing cells endothelial progenitor cell EV-harvesting Medium EV-depleted medium Preparation of EDS >=18h at >= 100,000g Cell viability (%) 99.9 Cell count 1000000 Separation Method (Differential) (ultra)centrifugation dUC: centrifugation steps Below or equal to 800 g Between 800 g and 10,000 g Between 100,000 g and 150,000 g Pelleting performed Yes Pelleting: rotor type Type 70 Ti Pelleting: speed (g) 140000 Wash: volume per pellet (ml) 35 Wash: time (min) 70 Wash: Rotor Type Type 70 Ti Wash: speed (g) 140000 Characterization: Protein analysis Protein Concentration Method Bradford Protein Yield (µg) per million cells Western Blot Detected EV-associated proteins TSG101/ CD40/ integrin beta-1 Not detected contaminants ApoA1 Characterization: RNA analysis RNA analysis Type (RT)-(q)PCR Proteinase treatment Yes Moment of Proteinase treatment After Proteinase type Proteinase K Proteinase concentration 500 RNAse treatment Yes RNAse type RNase A RNAse concentration 0.01 Characterization: Lipid analysis No Characterization: Particle analysis DLS Report type Size range/distribution Reported size (nm) 0-1000 EM EM-type Transmission-EM Image type Close-up, Wide-field

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EV-TRACK ID	EV220305
species	Homo sapiens
sample type	Cell culture
cell type	Wharton's Jelly mesenchymal stem cell	endothelial progenitor cell
condition	Hypoxia	treated with Hypoxia WJ-MSC derived EV
separation protocol	dUC	dUC
Exp. nr.	1	2
EV-METRIC %	78	78