Acute augmentation of cerebral blood flow by rho-kinase inhibitors in focal cerebral ischemia is dependent on endothelial nitric oxide synthase

Endothelial nitric oxide synthase (eNOS) dysfunction is related to secondary injury and lesion expansion after cerebral ischemia. To date, there are few reports about postischemic alterations in the eNOS regulatory system. The purpose of the present study was to clarify eNOS expression, Ser1177 phosphorylation, and monomer formation after cerebral ischemia. Male Wistar rats were subjected to transient focal cerebral ischemia. Endothelial nitric oxide synthase messenger RNA (mRNA) and protein expression increased ~ 8-fold in the ischemic lesion. In the middle cerebral artery core, eNOS-Ser1177 phosphorylation increased 6 hours after ischemia; however, there was an approximately 90% decrease in eNOS-Ser1177 phosphorylation observed 24 hours after ischemia that continued until at least 7 days after ischemia. Endothelial nitric oxide synthase monomer formation also increased 24 and 48 hours after ischemia ( P<0.05), and protein nitration progressed in parallel with monomerization. To assess the effect of a neuroprotective agent on eNOS dysfunction, we evaluated the effect of fasudil, a Rho-kinase inhibitor, on eNOS phosphorylation and dimerization. Postischemic treatment with fasudil suppressed lesion expansion and dephosphorylation and monomer formation of eNOS. In conclusion, functional deterioration of eNOS progressed after cerebral ischemia. Rho-kinase inhibitors can reduce ischemic lesion expansion as well as eNOS dysfunction in the ischemic brain.

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Rapid nontranscriptional activation of endothelial nitric oxide synthase mediates increased cerebral blood flow and stroke protection by corticosteroids

Journal of Clinical Investigation ◽

10.1172/jci200315481e1 ◽

2003 ◽

Vol 111 (5) ◽

pp. 759-a-759

Author(s):

F. P. Limbourg

Keyword(s):

Nitric Oxide ◽

Blood Flow ◽

Cerebral Blood Flow ◽

Nitric Oxide Synthase ◽

Endothelial Nitric Oxide Synthase ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

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Endothelial Nitric Oxide Synthase (Enos)Uncoupling-Dependent Lowering of Cerebral Blood Flow in Prefrontal Cortex of Rats with Acute Liver Failure

Journal of Clinical and Experimental Hepatology ◽

10.1016/j.jceh.2017.01.050 ◽

2017 ◽

Vol 7 ◽

pp. S38-S39

Author(s):

Krzysztof Milewski ◽

Anna Czarnecka ◽

Inez Fresko ◽

Jan Albrecht ◽

Magdalena Zielińska

Keyword(s):

Nitric Oxide ◽

Blood Flow ◽

Cerebral Blood Flow ◽

Prefrontal Cortex ◽

Nitric Oxide Synthase ◽

Liver Failure ◽

Endothelial Nitric Oxide Synthase ◽

Enos Uncoupling ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

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Rho-kinase inhibitor can enhance endothelial nitric oxide synthase phosphorylation and suppress the infarct expansion after focal cerebral ischemia

Neuroscience Research ◽

10.1016/j.neures.2010.07.1421 ◽

2010 ◽

Vol 68 ◽

pp. e320-e321

Author(s):

Yoshiki Yagita ◽

Tsutomu Sasaki ◽

Naoki Oyama ◽

Yukio Sugiyama ◽

Yasukazu Terasaki ◽

...

Keyword(s):

Nitric Oxide ◽

Cerebral Ischemia ◽

Endothelial Nitric Oxide Synthase ◽

Kinase Inhibitor ◽

Focal Cerebral Ischemia ◽

Rho Kinase ◽

Rho Kinase Inhibitor ◽

Infarct Expansion ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

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Rho GTPase/Rho Kinase Negatively Regulates Endothelial Nitric Oxide Synthase Phosphorylation through the Inhibition of Protein Kinase B/Akt in Human Endothelial Cells

Molecular and Cellular Biology ◽

10.1128/mcb.22.24.8467-8477.2002 ◽

2002 ◽

Vol 22 (24) ◽

pp. 8467-8477 ◽

Cited By ~ 298

Author(s):

Xiu-Fen Ming ◽

Hema Viswambharan ◽

Christine Barandier ◽

Jean Ruffieux ◽

Kozo Kaibuchi ◽

...

Keyword(s):

Nitric Oxide ◽

Endothelial Cells ◽

Endothelial Nitric Oxide Synthase ◽

Rho Gtpase ◽

Protein Kinase B ◽

Rho Kinase ◽

Enos Expression ◽

Enos Phosphorylation ◽

Oxide Synthase ◽

Endothelial Nitric Oxide

ABSTRACT Endothelial nitric oxide synthase (eNOS) is an important regulator of cardiovascular homeostasis by production of nitric oxide (NO) from vascular endothelial cells. It can be activated by protein kinase B (PKB)/Akt via phosphorylation at Ser-1177. We are interested in the role of Rho GTPase/Rho kinase (ROCK) pathway in regulation of eNOS expression and activation. Using adenovirus-mediated gene transfer in human umbilical vein endothelial cells (HUVECs), we show here that both active RhoA and ROCK not only downregulate eNOS gene expression as reported previously but also inhibit eNOS phosphorylation at Ser-1177 and cellular NO production with concomitant suppression of PKB activation. Moreover, coexpression of a constitutive active form of PKB restores the phosphorylation but not gene expression of eNOS in the presence of active RhoA. Furthermore, we show that thrombin inhibits eNOS phosphorylation, as well as expression via Rho/ROCK pathway. Expression of the active PKB reverses eNOS phosphorylation but has no effect on downregulation of eNOS expression induced by thrombin. Taken together, these data demonstrate that Rho/ROCK pathway negatively regulates eNOS phosphorylation through inhibition of PKB, whereas it downregulates eNOS expression independent of PKB.

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