First 11 pages of thesis. - OPUS - Universität Würzburg

3.2 eNOS deletion decreases vascular production of superoxide
in apoE ko vessels
To determine vascular superoxide production the formation of
oxyethidium from dihydroethidium was measured by HPLC. The measurements
revealed higher levels of superoxide in vessel rings of apoE ko and apoE/eNOS
dko mice, compared to C57Bl6 (9.4±0.5 AU/µg protein, n=14, p=0.000001 and
6.9±0.9 AU/µg protein, n=13, p=0.04 respectively, vs. 4.4±0.32 AU/µg protein,
n=9, Figure 9a). Total superoxide formation in apoE/eNOS dko mice was
decreased compared to apoE ko (6.9±0.9 AU/µg protein vs. 9.4±0.5 AU/µg
protein, p=0.02, Figure 9a). In addition to the experiments using chronic
deletion of eNOS, acute pharmacological inhibition of NOS by L-NAME resulted
in a significant reduction of superoxide formation in apoE ko vessels (9.4±0.5
AU/µg protein, n=14 vs. 4.8±0.6 AU/µg protein, n=14, p=0.00001, Figure 9b),
suggesting that NOS is uncoupled and contributes to superoxide formation in
apoE ko mice. Moreover, L-NIO, an eNOS specific inhibitor significantly
decreased vascular superoxide production in apoE ko mice fed with the western
diet for 24 weeks, supporting partial uncoupling of eNOS (9.3±0.6 AU/µg
protein, n=22 vs. 11.6±0.7 AU/µg protein, n=22, p=0.02, Figure 9c). Superoxide
levels in these older apoE ko mice were significantly higher than in apoE ko
mice fed with the western diet for 18 weeks (11.6±0.7 AU/µg protein, n=22,
p=0.05, Figure 9c) documenting increasing superoxide production with
progressive atherosclerosis.
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