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Cell-derived exosomes (EXs) can modulate target cell differentiation via microRNAs (miRs) that they carried.Previous studies have shown that miR126 is highly expressed in hematopoietic stem cells (HSCs) and plays a role in hematopoiesis via modulating the Notch pathway that participates in progenitors’ cell fate decisions.In this study we investigated whether HSC-derived EXs (HSC-EXs) could affect the differentiation of mouse embryonic stem cells (ESCs) into HSCs.We prepared HSC-EXscon,HSC-EXssc and HSC-EXsmiR126 from control HSCs and the HSCs transfected with scramble control or miR126 mimics,respectively.HSC-EXs were isolated by ultracentrifugation and analyzed using nanoparticle tracking analysis.We incubated the collected EXs with mouse ESCs over a 10-d differentiation induction period,during which HSC-EXs and a Notch pathway activator (Jagged1,100 ng/mL) were added to the cultures every 3 d.After the 10-d differentiation period,the expression levels of miR126,SSEA1,CD117,Sca1,Notch1 and Hes1 in ESCs were assessed.The generated HSCs were validated by flow cytometry using antibodies against HSC markers (CD117,CD34 and Sca1).Our results revealed that:(1) transfection with miR126 mimics significantly increased miR126 levels in HSC-EXsmiR126.(2) HSC-EX co-culture promoted mouse ESCs differentiation into HSCs with the most prominent effect found in the HSC-EXsmiR126 co-culture.(3) HSC differentiation was verified by reduced SSEA1 expression and increased CD117 and Sca1 expression.(4) All the effects caused by HSC-EXs were accompanied by significant reduction of Notch1 and Hes1 expression,thus inhibition of the Notch1/Hes1 pathway,whereas activation of Notch by Jagged1 abolished the effects of HSC-EXsmiR126.In conclusion,HSC-EXs promote hematopoietic differentiation of mouse ESCs in vitro by inhibiting the miR126/Notch1 pathway.