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Experiment number
  • If needed, multiple experiments were identified in a single publication based on differing sample types, separation protocols and/or vesicle types of interest.
Species
  • Species of origin of the EVs.
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
Details EV-TRACK ID Experiment nr. Species Sample type separation protocol First author Year EV-METRIC
EV150091 1/1 Homo sapiens Cell culture supernatant dUC
Total Exosome Isolation
Deng L 2015 13%

Study summary

Full title
All authors
Deng L, Blanco FJ, Stevens H, Lu R, Caudrillier A, McBride M, McClure JD, Grant J, Thomas M, Frid M, Stenmark K, White K, Seto AG, Morrell NW, Bradshaw AC, MacLean MR, Baker AH
Journal
Circ Res
Abstract
RATIONALE: The pathogenesis of pulmonary arterial hypertension (PAH) remains unclear. The 4 microRNA (show more...)RATIONALE: The pathogenesis of pulmonary arterial hypertension (PAH) remains unclear. The 4 microRNAs representing the miR-143 and miR-145 stem loops are genomically clustered. OBJECTIVE: To elucidate the transcriptional regulation of the miR-143/145 cluster and the role of miR-143 in PAH. METHODS AND RESULTS: We identified the promoter region that regulates miR-143/145 microRNA expression in pulmonary artery smooth muscle cells (PASMCs). We mapped PAH-related signaling pathways, including estrogen receptor, liver X factor/retinoic X receptor, transforming growth factor-? (Smads), and hypoxia (hypoxia response element), that regulated levels of all pri-miR stem loop transcription and resulting microRNA expression. We observed that miR-143-3p is selectively upregulated compared with miR-143-5p during PASMC migration. Modulation of miR-143 in PASMCs significantly altered cell migration and apoptosis. In addition, we found high abundance of miR-143-3p in PASMC-derived exosomes. Using assays with pulmonary arterial endothelial cells, we demonstrated a paracrine promigratory and proangiogenic effect of miR-143-3p-enriched exosomes from PASMC. Quantitative polymerase chain reaction and in situ hybridization showed elevated expression of miR-143 in calf models of PAH and in samples from PAH patients. Moreover, in contrast to our previous findings that had not supported a therapeutic role in vivo, we now demonstrate a protective role of miR-143 in experimental pulmonary hypertension in vivo in miR-143-/- and anti-miR-143-3p-treated mice exposed to chronic hypoxia in both preventative and reversal settings. CONCLUSIONS: MiR-143-3p modulated both cellular and exosome-mediated responses in pulmonary vascular cells, whereas inhibition of miR-143-3p blocked experimental pulmonary hypertension. Taken together, these findings confirm an important role for the miR-143/145 cluster in PAH pathobiology. (hide)
EV-METRIC
13% (33rd 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
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
DNF
Focus vesicles
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
dUC + Total Exosome Isolation
Protein markers
EV: CD63/ GAPDH/ CD9
non-EV:
Proteomics
no
Show all info
Study aim
Function
Sample
Species
Homo sapiens
Sample Type
Cell culture supernatant
EV-harvesting Medium
serum free
Separation Method
(Differential) (ultra)centrifugation
dUC: centrifugation steps
Between 800 g and 10,000 g
Commercial kit
Total Exosome Isolation
Characterization: Protein analysis
Western Blot
Detected EV-associated proteins
CD63/ CD9/ GAPDH
ELISA
Detected EV-associated proteins
GAPDH
NA
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