Supplemental Information

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Cytotoxic effects of ATRA in NB4 cells

untreated control2μmol/L ATRA1μmol/L ATRA0.5μmol/L ATRA0.25μmol/L ATRA0.125μmol/L ATRA

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Supplemental Figure S1. Cytotoxic effects of ATRA in NB4 cells with the concentration of 1μmol of ATRA chosen for the treatment assays in this paper (see text)

Supplemental Figure S2. Top 10 ATRA up-regulated and down-regulated angiogenesis-related transcrpts detected in NB4-derived EVs (see text)

NB4 ATRA cells NB4 ATRA EVs

Supplemental Figure S3. Analysis of significant changes in the expression of angiogenesis-related transcripts between control and ATRA treated NB4 cells and their EVs. A. Venn diagram. B. Tabulated data from the Venn diagram: Common – up- or down-regulated by ATRA in both NB4 cells and their EVs; Contra-regulated – transcripts exhibiting the opposite changes in ATRA treated NB4 cell and EVs; NB4 cells – transcripts selectively up- or down-regulated by ATRA in cells but not in EVs; NB4 EVs – transcripts selectively up- or down-regulated in EVs of ATRA-treated NB4 cells but not in NB4 cells themselves. .

Common up-

regulated transcripts

Common down-

regulated transcripts

Contra-regulated transcripts

NB4 cells ATRA

up-regulated transcripts

NB4 cellsATRA

down-regulated transcripts

NB4 EVsATRA

up-regulated transcripts

NB4 EVsATRA

down-regulated transcripts

CCL2 TGFBR1 FGF1 THBS1 ANGPT2 NRP2 TYMPIL1B BAI1 COL4A3 FGF2 CXCL1 PECAM1

HPSE SERPINF1 KDR ERBB2 PF4 NRP1MDK FN1 COL18A1 PDGFA TEK TIMP2

PTGS1 ANPEP TGFB2 LECT1 CTGF VEGFACXCL10 NOTCH4 IGF1 IFNG SPHK1

IL6 JAG1 CDH5 TNFPLG EFNA1 S1PR1 ITGAVIL8 F3 EGF MMP14

CXCL6 HGF TIMP3 ANGPT1EDN1 MMP9 CCL11FIGF EPHB4 NOS3ANG MMP2 ITGB3

ANGPTL4 FGFR3 ENGTIMP1 IFNA1VEGFC EFNB2

ID1 LEPFLT1

THBS2SERPINE1

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B

NB4 ATRA EVsNB4 untreated EVs

Common up-

regulated transcripts

Common down-

regulated transcripts

Contra-regulated transcripts

EVs from untreated NB4 cells

up-regulated transcripts

EVs from untreated NB4 cells

down-regulated transcripts

EVs from ATRA treated

NB4 cellsup-regulated transcripts

EVs fromATRA treated

NB4 cells down-regulated

transcripts

FIGF PECAM1 TGFB2 IGF1 PGF CCL2 FN1PLG HGF CXCL10 THBS1 LECT1 IL6 TYMP

EDN1 VEGFA LEP NRP2 MMP2CDH5 SERPINF1 IL8 CXCL1 NRP1MDK TNF PF4 NOTCH4IFNG EFNA1 TEK TGFBR1IL1B ANGPT1 CTGF TIMP2

EFNB2 EGF ANPEPCOL18A1 TIMP3 SPHK1VEGFC CCL11 ITGAVHPSE NOS3 FGFR3CXCL6 ITGB3 JAG1S1PR1 ANG BAI1ENG TIMP1 F3KDR ANGPTL4 EPHB4

PTGS1 MMP9IFNA1 MMP14

COL4A3ID1

FLT1FGF1

THBS2SERPINE1

Supplemental Figure S4. Analysis of significant changes in the expression of angiogenesis-related transcripts between EVs from control and ATRA treated NB4 cells. A. Venn diagram. B. Tabulated data from the Venn diagram: Common – up- or down-regulated in EVs from both control and ATRA treated NB4 cells; Contra-regulated – transcripts exhibiting the opposite change in EVs of control and ATRA treated NB4 cells; EVs from untreated NB4 cells – transcripts selectively up- or down-regulated in EVs of control cells; EVs from ATRA treated NB4 cells – transcripts selectively up- or down-regulated in EVs of ATRA-treated NB4 cells but not in EVs from control cells.

A

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HUVECs t

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ated NB4 EV

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HUVECs t

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ATRA EV

s0

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100ug/ml NB4 0.45um EVs

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Supplemental Figure S5. Stimulation of HUVEC survival by NB4 derived EVs was not significantly affected by ATRA treatment. This is consistent with ATRA-related replacement of VEGF with IL-8 in the pro-angiogenic factor expression profile (see text)

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Supplemental Figure S6. Impact of ATO on vesiculation and survival of NB4 cells. A NTA displayed an altered pattern of cellular vesiculation dominated by the upsurge of larger particles post ATO treatment; B EVs derived from NB4 cells treated with 1µmol/L ATO contained more TF content than those treated with 0.25µmol/L ATO; C TF content in transferred HUVECs which is correspondent with that in NB4 EVs (see text)

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NB4 EVs

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pro

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)

** **

ns

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ns

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