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Details	EV-TRACK ID	Experiment nr.	Species	Sample type	Separation protocol	First author	Year	EV-METRIC
	EV140093	1/1	Equus caballus	Uterine luminal fluid	ExoQuick UF	Ruiz-González I	2014	25%
Study summary Full title Exosomes, endogenous retroviruses and toll-like receptors: Pregnancy recognition in ewes All authors Ruiz-González I, Xu J, Wang X, Burghardt RC, Dunlap KA, Bazer FW Journal Reproduction Abstract Conceptus-endometrial communication during the peri-implantation period of pregnancy ensures establi (show more...)Conceptus-endometrial communication during the peri-implantation period of pregnancy ensures establishment of pregnancy. We hypothesized that this dialog involves exosomes, ovine endogenous jaagsiekte retroviruses (enJSRV) and toll-like receptors (TLR) which regulate the secretion of interferon tau (IFNT), the pregnancy recognition signal in ruminants. First, exosomes isolated from uterine flushings from cyclic and pregnant ewes were analyzed for exosomal content and uterine expression of heat shock protein 70 (HSC70). Then, conceptus trophectoderm cells (oTr1) treated with different doses of exosomes were analyzed for the expression of genes involved in TLR-mediated cell signaling. The results revealed that exosomes contain mRNAs for enJSRV-ENV, HSC70, interleukins, and interferon (IFN)-regulatory factors. Exosomal content of enJSRV-ENV mRNA and protein decreased from days 10 and 12 to day 16 of gestation, and uterine expression of HSC70 increased in pregnant ewes compared with cyclic ewes. The oTr1 cells proliferated and secreted IFNT in a dose-dependent manner in response to exosomes from cyclic ewes. The expression of CD14, CD68, IRAK1, TRAF6, IRF6, and IRF7 mRNAs that are key to TLR-mediated expression of type 1 IFNs was significantly influenced by day of pregnancy. This study demonstrated that exosomes are liberated into the uterine lumen during the estrous cycle and early pregnancy; however, in pregnant ewes, exosomes stimulate trophectoderm cells to proliferate and secrete IFNT coordinately with regulation of TLR-mediated cell signaling. These results support our hypothesis that free and/or exosomal enJSRV act on the trophectoderm via TLR to induce the secretion of IFNT in a manner similar to that for innate immune responses of macrophages and plasmacytoid dendritic cells to viral pathogens. (hide) EV-METRIC 25% (62nd 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 Uterine luminal fluid Sample origin NAY Focus vesicles exosomes 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 UF Protein markers EV: HSP70 non-EV: Cell organelle protein Proteomics no Show all info Study aim Function Sample Species Equus caballus Sample Type Uterine luminal fluid Separation Method Commercial kit ExoQuick Other Name other separation method ExoQuick Characterization: Protein analysis Western Blot Antibody details provided? Yes Antibody dilution provided? Yes Detected EV-associated proteins HSP70 Detected contaminants Cell organelle protein Characterization: Particle analysis EM EM-type transmission EM

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EV-TRACK ID	EV140093
species	Equus caballus
sample type	Uterine luminal fluid
condition	NAY
separation protocol	ExoQuick UF
Exp. nr.	1
EV-METRIC %	25