MAT.Angiopoietin-1 Protects The Microcirculation From L-NAME Induced Endothelial Dysfunction

Alessio Alfieri1, Nicola J Brown1, Richard Kammerer2, Zoe LS Brookes1. 1University of Sheffield, Sheffield, UK, 2University of Manchester, Manchester, UK.

Reduced activity of nitric oxide (NO) has been implicated in the pathogenesis of several cardiovascular disorders involving endothelial dysfunction (Moncada and Higgs 2006). The growth factor angiopoietin-1 (Ang-1) signals through Tie-2 receptors expressed on vascular endothelial cells, a pathway that also reduces inflammation (Brindle et al. 2006). This study investigated the effect of the potent and stable Ang-1 variant MAT.Ang-1 (Cho et al. 2004) on the microcirculation in vivo during endothelial dysfunction induced via systemic administration of the NO inhibitor L-nitro-arginine methyl ester (L-NAME).

Aluminium dorsal microcirculatory chambers were chronically implanted into the dorsal skinfold of male C3H/HeN mice (7-10 weeks, n = 18) to expose the striated muscle microcirculation for observation with intravital microscopy over 4 hours using fluorescein isothiocyanate labelled bovine serum albumin (66 kDa FITC-BSA, 200µl.100g-1 i.v.). Animals were sedated with ketamine:xylazine (10:1 mg.kg-1 ip.) and allocated into three experimental groups: (i) control, (ii) L-NAME (10 mg.kg-1 ip.) and (iii) L-NAME + MAT.Ang-1 (33 µg per animal iv. 1 h after the NO inhibitor). Western Blotting was used to determine protein expression of eNOS and VE-cadherin in abdominal muscle.

L-NAME induced a time-dependent constriction of arterioles (34.1±6.9% vs. 11.9±3.9% in control at 4 hr; p<0.01) and this effect was reduced by co-administration with MAT.Ang-1 (18.4±10.8%). eNOS protein expression was increased by L-NAME but remained unchanged with MAT.Ang-1. On the contrary, L-NAME reduced VE-cadherin expression, being restored to control levels by co-administration with MAT.Ang-1. This effect was consistent with our previous finding that MAT.Ang-1 reduced microvascular hyperpermeability induced by L-NAME (Alfieri et al. 2009).

In summary, MAT.Ang-1 inhibits several microvascular effects of L-NAME, thus it may improve vascular reactivity and reduce inflammation by inhibiting endothelial dysfunction via NO mechanisms.

Moncada, S. and Higgs, E.A. (2006). The discovery of nitric oxide and its role in vascular biology. British Journal of Pharmacology, 147 (Supplement 1), pp. 193-201.Brindle, N.P., Saharinen, P., Alitalo, K. (2006). Signalling and functions of angiopoietin-1 in vascular protection. Circulation Research, 98, pp. 1014-1023.

Cho, C.H., Kammerer, R.A., Lee, H.J., Steinmetz, M.O., Ryu, Y.S., Lee, S.H., Yasunaga, K., Kim, K.T., Kim, I., Choi, H.H., Kim, W., Kim, S.H., Park, S.K., Lee, G.M., Koh, G.Y. (2004). COMP-Ang1: a designed angiopoietin-1 variant with nonleaky angiogenic activity. Proceedings of the National Academy of Sciences, 101, pp. 5547-52.

Alfieri, A., Brown, N.J., Kammerer, R.A., Brookes, Z.L. (2009). MAT.Angiopoietin-1 induced changes in microcirculatory haemodynamics are mediated by nitric oxide. Microcirculation, 16, p. 757.