Abstract P51: Temporal Changes In Nitric Oxide Synthase Isoform Expression And Function In Spontaneously Hypertensive Stroke Prone Rats

Abstract

Introduction: Nitric oxide (NO) has been implicated in the pathogenesis of cerebral small vessel disease (cSVD). NO is produced by three NO synthase (NOS) isoforms including endothelial (eNOS), neuronal (nNOS), and inducible (iNOS). While eNOS has been implicated in cSVD pathologies that occur in Spontaneously Hypertensive Stroke Prone Rats (SHRSP), cSVD model, the mechanistic role and temporal changes of NOS isoform expression including eNOS in relation to cSVD development in SHRSP remains unknown. We aimed to characterize the temporal changes of NOS isoforms expression in SHRSP from the asymptomatic stage until the development of cSVD lesions. Methods: 20 SHRSP and 20 Wistar Kyoto (WKY) male rats were studied. Systolic blood pressure (SBP) was measured weekly using tail-cuff plethysmography. Half of the groups were euthanized at 7 weeks of age and the remaining rats were followed until 24 weeks of age. Routine histology and immunohistochemistry were performed for eNOS, nNOS, iNOS, superoxide radicals (DHE), NO (DAF) and Iba1 for microglia activation. Results: At 7 weeks, SBP was not significantly different between the groups (WKY 106.6±5.4 vs SHRSP 120.8±5.4, P=0.06). SHRSP developed hypertension between 9-11 weeks and maintained it throughout the experiment (SBP (21 weeks): WKY 134.8±5.4 vs 168.9±5.4, P<0.0001). Brain histology was largely negative for cSVD lesions at 7 weeks. At 24 weeks, histology showed areas of demyelination, microbleed formation, arteriosclerosis and hemosiderin deposits in SHRSP. At 7 weeks, compared to WKY, eNOS and nNOS expression were 50% and 30% lower in SHRSP (P=0.0038 and 0.0211, respectively). iNOS expression was 93% higher in SHRSP (P=0.0068). Superoxide levels were 133% higher in SHRSP (P<0.0001) and NO was 40% lower in SHRSP (P<0.0008). At 24 weeks, all of these differences became larger. Iba-1 expression was 25% higher at 7 weeks and became 97% higher in SHRSP at 24 weeks (P<0.0001). Conclusions: SHRSP show significant differences in NOS isoform expression with loss of NO and gain of superoxide production early in life prior to development of cSVD lesions. Superoxide overproduction and related oxidant stress may lead to NOS dysfunction and uncoupling which may trigger cSVD onset and point towards potential future therapies.