Nature - USA (2020-02-13)

3

nature research | reporting summary

April 2018

1:1000), pAktS473 (4060, Cell Signaling Technology, 1:1000), tAkt (4691, Cell Signaling Technology, 1:1000), p-eNOSS1177 (9571,

Cell Signaling Technology, 1:1000), pVEGFR2Y1175 (2478, Cell Signaling Technology, 1:1000), tVEGFR2 (2479, Cell Signaling

Technology, 1:1000), t-eNOS (610296, BD Biosciences, 1:1000), p-VinculinY822 (ab200825, Abcam, 1:1000), t-Vinculin (V9131,

Sigma Aldrich, 1:1000), PI3K/P85 (06195,Upstate,1:1000), integrin αvβ3 (clone LM609, Merck, 1:1000), Shc

(ab15039,Abcam,1:1000),VE-cadherin (C-19, sc-6458,Santa Cruz Biotechnology, 1:1000), Fibronectin (HFN7.1, Developmental

Society Hybridoma Bank, 1:50), MCP1 (ab7207, Abcam, 1:50), beta-catenin (610153, BD Transduction Laboratories, 1:70),

Alexafluor 488 goat anti-mouse (A11001, Invitrogen, 1:150), Alexafluor 568 goat anti-mouse (A11061, Invitrogen, 1:150),

Alexafluor 568 goat anti-rabbit (A11011, Invitrogen, 1:150), Alexafluor 790 goat anti-rabbit (A11367, Invitrogen, 1:10000),

Alexafluor 680 goat anti-rabbit (A21076, Invitrogen, 1:10000), Alexafluor 790 goat anti-mouse (A11375, Invitrogen, 1:10000),

Alexafluor 680 goat anti-mouse (A21058, Invitrogen, 1:10000), Plexin D1 (E13, sc-46245, Santa Cruz Biotechnology), Plexin D1

(PA5-47012, ThermoFisher Scientific)

Validation Antibodies were validated by the respective suppliers for the applications described in this manuscript. Plexin D1 antibody (Santa

Cruz Biotechnology) used for magnetic force application experiments was validated for this application in-house. `

Eukaryotic cell lines

Policy information about cell lines

Cell line source(s) Bovine aortic endothelial cells were purchased from VEC Technologies; mouse endothelial cells used in this study were

isolated in-house.

Authentication Authentication was performed by western blotting and immunofluorescence for endothelial markers (PECAM, eNOS), as well

as alignment in response to laminar flow.

Mycoplasma contamination Cell lines used in this study were mycoplasma free.

Commonly misidentified lines

(See ICLAC register)

None

Animals and other organisms

Policy information about studies involving animals; ARRIVE guidelines recommended for reporting animal research

Laboratory animals All mice had a C57BL6/J background. Plexin D1fl/fl mice were obtained from J Epstein, Cdh5-Cre mice were obtained from R.

Adams, ApoE-/- mice were first generated and characterized by N. Maeda

Wild animals No wild animals were used in this study.

Field-collected samples No field-collected samples were used in this study.