Modulation of monocyte adherence to endothelial cells by endothelin-1 involvement of Src (p60src) and JAK1-like kinases Lindsey J. Chisholm, MB, BS,

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Modulation of monocyte adherence to endothelial cells by endothelin-1 involvement of Src (p60src) and JAK1-like kinases Lindsey J. Chisholm, MB, BS, Peter S. Dovgan, MD, Devendra K. Agrawal, PhD, Patrick E. McGregor, MD, John D. Edwards, MD Journal of Vascular Surgery Volume 23, Issue 2, Pages 288-300 (February 1996) DOI: 10.1016/S0741-5214(96)70273-2 Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 1 Effect of ET-1 (10-7 mol/L) on monocyte adherence to HUVECs incubated with ET-1 for various time periods. Percent increase above baseline monocyte adherence (in absence of ET-1) was calculated. Values are mean ± SEM, n = 5-8. *p Value <0.05. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 2 Dose response effect of ET-1 on monocyte adherence to HUVECs. Monocytes were incubated with various doses of ET-1 for 4 hours under defined experimental conditions. Values are mean ± SEM, n = 7. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 3 Effect of phosphoramidon (10-5 mol/L) on ET-1-induced monocyte adherence to HUVECs. HUVECs were preincubated with phosphoramidon for 15 minutes before incubation with ET-1 10-8 mol/L or ET-1 10-11 mol/L for 4 hours. Values are mean ± SEM, n = 4. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 4 Effect of tyrosine kinase inhibitors on ET-1-induced monocyte adherence to HUVECs. HUVECs were incubated with genistein (G) 10-4 mol/L or herbimycin A (H) 8.75 × 10-7 mol/L for 15 minutes before addition of ET-1 10-8 mol/L or ET-1 10-11 mol/L, HUVECs were then incubated for 4 hours at 37º C in 95% air/5% CO2 in humidified chamber. Values are mean ± SEM, n = 6-8. p < 0.05; **p < 0.01. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 5 Effect of ET-1 in presence and absence of endothelin-B receptor antagonist IRL 1038 (10-5 mol/L) on monocyte adherence to HUVECs. HUVECs were incubated with ETB receptor antagonist for 15 minutes before incubation with ET-1 at various doses. Values are mean ± SEM, n = 7. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 6 Time course for effect of ET-1 (10-8 mol/L) on production of tyrosine phosphorylated proteins by HUVECs as detected by immunoblotting with PY20. Lanes marked G and H are samples where HUVECs were incubated with tyrosine kinase inhibitor genistein (G) 10-4 mol/L or herbimycin A (H) 8.75 × 10-7 mol/L for 10 minutes before addition of ET-1 10-8 mol/L for 5 minutes. Data are representative of three experiments. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 7 Dose response effect of ET-1 on production of tyrosine phosphorylated proteins by HUVECs as detected by immunoblotting with PY20. Lanes marked G and H are samples of HUVECs incubated with tyrosine kinase inhibitors genistein (G) 10-4 mol/L or herbimycin A (H) 8.75 × 10-7 mol/L for 10 minutes before addition of ET-1 10-10 mol/L for 5 minutes. Data are representative of two individual experiments. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 8 Effect of ETA and ETB receptor antagonists on tyrosine phosphorylated proteins as detected by immunoblotting with PY20. HUVECs were incubated with ETA receptor (5 × 10-8 mol/L) and ETB receptor antagonist (5 × 10-7 mol/L) for 10 minutes alone or before addition of ET-1 10-8 mol/L for 5 minutes of incubation. Lane marked G shows effect of incubating HUVECs with genistein (G) 10-4 mol/L for 10 minutes before addition of ET-1 10-8 mol/L for 5 minutse. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 9 Immunoblotting of ET-1-(10-8 mol/L) stimulated HUVECs with antibody to src, JAK1, or JAK2 or appropriate control peptide (CP). This is representative of four separate experiments. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 10 Immunoprecipitation of ET-1-stimulated and control HUVECs with antibody to p60src followed by immunoblotting with PY20 or antibody to c-src. This is representative of three separate experiments. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions

Fig. 11 Immunoprecipitation of ET-1-stimulated and control HUVECs with antibody to JAK1 followed by immunoblotting with PY20 or antibody to JAK1. This is representative of three separate experiments. Journal of Vascular Surgery 1996 23, 288-300DOI: (10.1016/S0741-5214(96)70273-2) Copyright © 1996 Society for Vascular Surgery and International Society for Cardiovascular Surgery, North American Chapter Terms and Conditions