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Effect of leptin on lymphatic endothelial cell tube formation and proliferation.

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posted on 01.07.2016, 17:49 by Akinori Sato, Ryuta Kamekura, Koji Kawata, Masaya Kawada, Sumito Jitsukawa, Keiji Yamashita, Noriyuki Sato, Tetsuo Himi, Shingo Ichimiya

(A) Representative phase contrast microscopy images showing tube formation in HDLECs treated with leptin (10, 100 ng/ml) for 17 h with or without anti-leptin antibody (10 μg/ml). Scale bar: 50 μm. (B) Histograms represent total tube length in HDLECs exposed to leptin (10, 100 ng/ml) for 17 h with or without 1 or 10 μg/ml anti-leptin antibody. Total tube length and mean length in each condition were measured using Image J software. Results are expressed as means ± SEM of four independent experiments; *p < 0.05, **p < 0.01, t-test. n.s.: not significant. (C) WST-8 assay showing proliferation of HDLECs treated with leptin (10, 100 ng/ml) for 17 h with or without 1 or 10 μg/ml anti-leptin antibody. Histograms represent absorbance at 450 nm. Results are expressed as means ± SEM of four independent experiments; *p < 0.05, t-test. n.s.: not significant. (A-C) Anti-leptin antibody was added 30 min prior to leptin treatment. Ten μg/ml mouse IgG was used as an isotype control to anti-leptin antibody. (D) Immunoblot analysis showing the expression of claudin-5, occludin, VE-cadherin, and ZO-1 in HDLECs treated with leptin (10, 100 ng/ml) for 24 h. β-actin was used as a loading control. Histograms represent relative expression of claudin-5, occludin, VE-cadherin, and ZO-1 proteins in HDLECs exposed to 10 or 100 ng/ml leptin for 24 h as determined by densitometry analysis. Results are expressed as means ± SEM of four independent experiments; t-test. n.s.: not significant.

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