Anti vascular endothelial cadherin

The As₂O₃ solution was purchased from Heilongjiang Harbin Medical Pharmaceutical Co., Ltd. (Heilongjiang, China). Matrigel was purchased from BD Biosciences (Franklin Lakes, NJ, USA), the CD105-antibody was purchased from Thermo Fisher Scientific, Inc. (Waltham, MA, USA), and anti-vascular endothelial (VE)-cadherin, anti-Bcl-2-associated X protein (Bax) and anti-B-cell lymphoma 2 (Bcl-2) antibodies were purchased from Santa Cruz Biotechnology, Inc. (Dallas, TX, USA). Matrix metalloproteinase (MMP)-2 and MMP-9 antibodies were purchased from Nanjing KeyGen Biotech Co., Ltd. (Nanjing, China), and the animals were obtained from Hunan Slack Jingda Experimental Animals Co., Ltd. (Hunan, China).

HMVEC were grown to confluence on LabTek II chamber slides (Nunc, thermofischer scientific, Waltham, MA) that had been precoated with collagen and then treated for 6 hours with LPS in the presence and absence of catecholaminergic agents, and antagonists and agonists of α- and β-AR. Cells were then fixed with 4% paraformaldehyde for 15 minutes and permeabilized with 0.5% Triton X-100 in phosphate-buffered saline (PBS). After blocking with 1% bovine serum albumin in PBS, cells were incubated with either anti–zona occludens (ZO)-1-AlexaFluor594 (Invitrogen), F-actin (rhodamine phalloidin; Invitrogen), or anti–vascular endothelial (VE)-cadherin (Santa Cruz Biotechnologies) followed by AlexaFluor488-tagged secondary antibody. Nuclei were counterstained with 4′6-diamidino-2-phenylindole. Slides were visualized with a Cytation 5 Biotek (Fischer scientific, Waltham, MA) and recorded at 20X objective on Gen5 Image+ Software (Biotek,Winooski, VT).