Targeting eNOS for stroke protection Li-ping Wu

Background nitric oxide synthase (NOS) L-arginine NO  Neuroprotective:increasing cerebral Blood flow during brain ischemia  Vasodilatory  Anti-inflammatory  Antithrombotic  Antiproliferative eNOS In vascular diseases

Main isomers of NOS cNOS consitutive form iNOS inducible form  Constitutively expressed  Calcium-dependent  Including:  Induced in inflammatory and other cell types by stimuli such as endotoxin and proinflammatory cytokines  Calcium-independent Neuronal (nNOS) Endothelial (eNOS)

eNOS: neuroprotective eNOS nNOS cerebral infarction regional cerebral blood flow (knocked out)

eNOS: neuroprotective L-NAME eNOS nNOS cerebral infarction regional cerebral blood flow (lacking) (inhibiting activity)

eNOS nNOS L-arginine cerebral infarction regional cerebral blood flow (enhancing NO production) eNOS: neuroprotective

Enhanacing eNOS function Increase the number of circulating Endothelial Progenitor cells (EPCs) Contribute to neovasularization after ischemia stroke

Janus effect of NO in the brain brain ischemia nNOS NO toxic to surrounding neurons iNOS secondary damage eNOS neuroprotective

? What we should do eNOS nNOS iNOS upregulate and/or activate inhibit how

How to upregulate/activate eNOS Statins Steroid hormones Physical activity Nutrients

Statins Statins: cholesterol lowering inhibitors of hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase

Statins  Indirectly: by reducing cholesterol levels  Directly: by stability of eNOS mRNA by activating the pathway involving phosphoinositide 3-kinase (PI3K) and PKB/Akt Mechanism:

Steroid hormones corticosteroids oestrogen Directly activate eNOS

corticosteroids Transcriptional effect: modulation of target genes by glucocorticoid receptor (GR) PI3K and PKB/Akt NO bioavailability Non-nuclear effect: eNOS activation corticosteroids GR Mechanism:

corticosteroids Depending on the dose and duration of corticosteroid administration In clinical practice: not good Hyperglycemia (lactate acidosis) Ten times higher dose In animal models:

oestrogen Genomic effects: an interaction between oestrogen receptor and regulatory subunit of PI3K Nontranscriptional effects: direct binding of the oestrogen receptor  isoform to PI3K eNOS activity Mechanism:

oestrogen Strongly neuroprotective effect in animal models of stroke No effect in clinical trials of primary prevention

Physical activity Regular physical activity Mechanism:  relating to alterations in blood flow  increasing the number of circulating EPCs  increasing neovascularization

Nutrients Red wine and diets rich in flavonoids and plant polyphenols Mechanism: ?

Uncoupling of eNOS eNOS enhancing ROS production L-arginine BH4 superoxide NO NADPH oxidaseVascular damage eNOS uncoupling

eNOS – protect against stroke in human? preventive approaches chronic eNOS upregulation  regular physical activity  moderate consumption of red wine  chronic statin treatment high risk patients (high dose) primary and secondary prevention

eNOS – protect against stroke in human? acute approaches eNOS activation  PI3K-Akt-mediated eNOS activation can be achieved administration of statins,corticosteroids and oestrogens  L-arginine could be directly administered intravenously to increase NO production

Concluding remarks Although several important caveats, such as eNOS uncoupling and the dual role of NO in brain ischemia have to be considered However statins, physical activity, steroid hormones and nutrients can Lead to eNOS upregulation or activation, which protects from brain ischemic stroke Thus eNOS targeting is an attractive approach to prevent and treat stroke in humans

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