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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV210383	1/2	Mus musculus	Serum	ExoQuick	Mkrtchian, Soiuren	2022	75%
Study summary Full title Surgical Trauma in Mice Modifies the Content of Circulating Extracellular Vesicles All authors Souren Mkrtchian, Anette Ebberyd, Rosanne E. Veerman, María Méndez-Lago, Susanne Gabrielsson, Lars I. Eriksson, and Marta Gómez-Galán Journal Front Immunol Abstract Surgical interventions rapidly trigger a cascade of molecular, cellular, and neural signaling respon (show more...)Surgical interventions rapidly trigger a cascade of molecular, cellular, and neural signaling responses that ultimately reach remote organs, including the brain. Using a mouse model of orthopedic surgery, we have previously demonstrated hippocampal metabolic, structural, and functional changes associated with cognitive impairment. However, the nature of the underlying signals responsible for such periphery-to-brain communication remains hitherto elusive. Here we present the first exploratory study that tests the hypothesis of extracellular vesicles (EVs) as potential mediators carrying information from the injured tissue to the distal organs including the brain. The primary goal was to investigate whether the cargo of circulating EVs after surgery can undergo quantitative changes that could potentially trigger phenotypic modifications in the target tissues. EVs were isolated from the serum of the mice subjected to a tibia surgery after 6, 24, and 72 h, and the proteome and miRNAome were investigated using mass spectrometry and RNA-seq approaches. We found substantial differential expression of proteins and miRNAs starting at 6 h post-surgery and peaking at 24 h. Interestingly, one of the up-regulated proteins at 24 h was α-synuclein, a pathogenic hallmark of certain neurodegenerative syndromes. Analysis of miRNA target mRNA and corresponding biological pathways indicate the potential of post-surgery EVs to modify the extracellular matrix of the recipient cells and regulate metabolic processes including fatty acid metabolism. We conclude that surgery alters the cargo of circulating EVs in the blood, and our results suggest EVs as potential systemic signal carriers mediating remote effects of surgery on the brain. (hide) EV-METRIC 75% (98th 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 Serum Sample origin Control condition 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 ExoQuick Protein markers EV: CD63/ CD81/ HSP70 non-EV: Albumin/ Argonaute?2/ Calreticulin/ GM130/ PMP70/ Prohibitin/ Tamm-Horsfall protein Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Mus musculus Sample Type Serum Separation Method Commercial kit ExoQuick Other Name other separation method ExoQuick Characterization: Protein analysis Protein Concentration Method microBCA Protein Yield (µg) 0.75 Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD63/ CD81/ HSP70 Not detected contaminants Albumin Proteomics database Yes Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Modus Reported size (nm) 100 EM EM-type Transmission-EM Image type Close-up, Wide-field

	EV210383	2/2	Mus musculus	Serum	IAF	Mkrtchian, Soiuren	2022	75%
Study summary Full title Surgical Trauma in Mice Modifies the Content of Circulating Extracellular Vesicles All authors Souren Mkrtchian, Anette Ebberyd, Rosanne E. Veerman, María Méndez-Lago, Susanne Gabrielsson, Lars I. Eriksson, and Marta Gómez-Galán Journal Front Immunol Abstract Surgical interventions rapidly trigger a cascade of molecular, cellular, and neural signaling respon (show more...)Surgical interventions rapidly trigger a cascade of molecular, cellular, and neural signaling responses that ultimately reach remote organs, including the brain. Using a mouse model of orthopedic surgery, we have previously demonstrated hippocampal metabolic, structural, and functional changes associated with cognitive impairment. However, the nature of the underlying signals responsible for such periphery-to-brain communication remains hitherto elusive. Here we present the first exploratory study that tests the hypothesis of extracellular vesicles (EVs) as potential mediators carrying information from the injured tissue to the distal organs including the brain. The primary goal was to investigate whether the cargo of circulating EVs after surgery can undergo quantitative changes that could potentially trigger phenotypic modifications in the target tissues. EVs were isolated from the serum of the mice subjected to a tibia surgery after 6, 24, and 72 h, and the proteome and miRNAome were investigated using mass spectrometry and RNA-seq approaches. We found substantial differential expression of proteins and miRNAs starting at 6 h post-surgery and peaking at 24 h. Interestingly, one of the up-regulated proteins at 24 h was α-synuclein, a pathogenic hallmark of certain neurodegenerative syndromes. Analysis of miRNA target mRNA and corresponding biological pathways indicate the potential of post-surgery EVs to modify the extracellular matrix of the recipient cells and regulate metabolic processes including fatty acid metabolism. We conclude that surgery alters the cargo of circulating EVs in the blood, and our results suggest EVs as potential systemic signal carriers mediating remote effects of surgery on the brain. (hide) EV-METRIC 75% (98th 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 Serum Sample origin Control condition 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 Immunoaffinity capture (non-commercial) Protein markers EV: CD63/ CD81/ HSP70 non-EV: Albumin/ Argonaute?2/ Calreticulin/ GM130/ PMP70/ Prohibitin/ Tamm-Horsfall protein Proteomics yes Show all info Study aim Identification of content (omics approaches) Sample Species Mus musculus Sample Type Serum Separation Method Immunoaffinity capture Selected surface protein(s) CD9/ CD63/ CD81 Characterization: Protein analysis Protein Concentration Method microBCA Protein Yield (µg) 0.75 Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Lysis buffer provided? Yes Detected EV-associated proteins CD63/ CD81/ HSP70 Detected contaminants Albumin Proteomics database Yes Characterization: RNA analysis RNA analysis Type (RT)?(q)PCR/ RNA?sequencing Database NGI Proteinase treatment No RNAse treatment No Characterization: Lipid analysis No Characterization: Particle analysis NTA Report type Modus Reported size (nm) 100 EM EM-type Transmission-EM Image type Close-up, Wide-field

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EV-TRACK ID	EV210383
species	Mus musculus
sample type	Serum
condition	Control condition
separation protocol	ExoQuick	IAF capture (non-commercial)
Exp. nr.	1	2
EV-METRIC %	75	75