Abstract T P225: cGMP-Dependent Protein Kinase I in Smooth Muscle Cells Protects Against Stroke Injury in Mice

Stroke ◽  
2014 ◽  
Vol 45 (suppl_1) ◽  
Author(s):  
Dmitriy Atochin ◽  
Emmanuel Buys ◽  
Helen Swanson ◽  
Ana Dordea ◽  
Dai Fukumura ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Conditional Knockout ◽  
Tamoxifen Treatment ◽  
Dependent Protein Kinase ◽  
Doppler Flowmetry ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

The signaling pathways by which NO protects against stroke injury are mediated by cyclic guanosine monophosphate (cGMP) formation. However, the mechanisms of downstream signaling from cGMP to smooth muscle cells (SMC) remain incompletely understood. The cGMP-dependent protein kinase I (cGKI) is a key mediator of cGMP signaling. The goal of this study is to test the hypothesis that cGKI in SMC protects against stroke injury. Animals: We used the novel mouse line for highly efficient tamoxifen-inducible SMC-specific gene knockout, SMA-CreERT2, which expresses the CreERT2 recombinase under the control of the smooth muscle alpha-actin promoter (cGKI KO mice). The litter mate wild-type (WT) mice (no Cre gene) receiving the same tamoxifen treatment were used as control (cGKI WT) mice. Methods and Results: Mean blood pressure measured acutely from the common carotid artery under anesthesia (30% oxygen, 70% nitrous oxide and 1.5% isoflurane) was higher in cGKI KO mice (107±8 mmHg, Mean±SD) than in cGKI WT mice (95±1 mmHg, n=4/group, P<0.05). One hour of middle cerebral artery (MCA) occlusion by filament with 23 hours of reperfusion produced increased cerebral infarct volume (TTC staining, indirect calculation) in cGKI KO mice (123±39 mm3) as compared with cGKI WT (85±17 mm3, n=5/group, p< 0.05). Neurological deficit by 5 point scale was worse in cGKI KO mice (4.0 points) as compared with cGKI WT (2.4 points). Cerebral blood flow monitored by laser Doppler flowmetry above the core of the MCA territory indicated that reperfusion was less in cGKI KO mice than in cGKI WT mice, suggesting that cGKI mediates cerebrovascular SMC relaxation and protects against ischemic stroke. Vascular relaxation of mesenteric resistance arteries to acetylcholine, and the NO donor, NONOate after preconstriction with phenylephrine was dramatically impaired in cGKI KO mice. It suggests the importance of cGKI in the vascular SMC for NO-induced relaxation. In conclusion, stroke injury was more pronounced in cGKI SMC specific inducible conditional knockout mice than in cGKI WT mice, suggesting a protective role for cGKI in SMC against cerebral reperfusion injury. These studies identify the cGKI in cerebral vasculature as a potential target for therapies aimed at reducing stroke injury.

cGMP-dependent protein kinase mediates the reduction of Ca2+ by cAMP in vascular smooth muscle cells

1990 ◽  
Vol 258 (3) ◽  
pp. C399-C407 ◽  
Author(s):  
T. M. Lincoln ◽  
T. L. Cornwell ◽  
A. E. Taylor
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dependent Protein Kinase ◽  
Aortic Smooth Muscle ◽  
Cyclic Monophosphate ◽  
Cgmp Dependent Protein Kinase ◽  
Camp Dependent Protein Kinase ◽  
Dependent Protein

The major action of forskolin, the diterpine activator of adenylate cyclase, in primary (unpassaged) rat aortic smooth muscle cells is to reduce vasopressin-stimulated Ca2+ concentrations. In repetitively passaged cells, however, forskolin by itself increased Ca2+ levels by apparently stimulating Ca2+ uptake into the cell and had much smaller effects on inhibiting vasopressin-stimulated Ca2+ elevations. Both primary and passaged smooth muscle cells contained adenosine 3',5'-cyclic monophosphate (cAMP)-dependent protein kinase. Guanosine 3',5'-cyclic monophosphate (cGMP)-dependent protein kinase was greatly reduced or absent in passaged smooth muscle cells. The introduction of purified cGMP-dependent protein kinase into the cytoplasm of passaged cells prevented forskolin from elevating intracellular Ca2+ and restored the capacity of forskolin to reduce vasopressin-stimulated Ca2+ mobilization. Similar effects were observed for isoproterenol in passaged smooth muscle cells. When introduced into cells, the active catalytic subunit of the cAMP-dependent protein kinase did not lead to reductions in Ca2+ levels. These results suggest that cAMP elevations lead to profound changes in Ca2+ metabolism through activation of both cAMP- and cGMP-dependent protein kinases. Activation of cGMP-dependent protein kinase by cAMP leads to the reduction in intracellular Ca2+, whereas activation of cAMP-dependent protein kinase may only mediate the uptake of Ca2+ from extracellular sources.

Abstract WMP83: cGMP-Dependent Protein Kinase I in Smooth Muscle Cells Protects Against Stroke in Female Mice

Stroke ◽  
2019 ◽  
Vol 50 (Suppl_1) ◽  
Author(s):  
Maria Shvedova ◽  
Jesse D Roberts ◽  
Paul L Huang ◽  
Robert Feil ◽  
Dmitriy N Atochin
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dependent Protein Kinase ◽  
Female Mice ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

scholarly journals Heparin Responses in Vascular Smooth Muscle Cells Involve cGMP-Dependent Protein Kinase (PKG)

2014 ◽  
Vol 229 (12) ◽  
pp. 2142-2152 ◽  
Author(s):  
Albert C. Gilotti ◽  
Wutigri Nimlamool ◽  
Raymond Pugh ◽  
Joshua B. Slee ◽  
Trista C. Barthol ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Protein Kinase ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dependent Protein Kinase ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

scholarly journals Implication of microRNAs in atrial natriuretic peptide and nitric oxide signaling in vascular smooth muscle cells

2011 ◽  
Vol 301 (4) ◽  
pp. C929-C937 ◽  
Author(s):  
Kumar U. Kotlo ◽  
Bahar Hesabi ◽  
Robert S. Danziger
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Protein Kinase ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dependent Protein Kinase ◽  
Cgmp Dependent Protein Kinase ◽  
Dependent Protein

MicroRNAs (miRs) are endogenous small RNA molecules that suppress gene expression by binding to complementary sequences in the 3′ untranslated regions of their target genes. miRs have been implicated in many diseases, including heart failure, ischemic heart disease, hypertension, cardiac hypertrophy, and cancers. Nitric oxide (NO) and atrial natriuretic peptide (ANP) are potent vasorelaxants whose actions are mediated through receptor guanylyl cyclases and cGMP-dependent protein kinase. The present study examines miRs in signaling by ANP and NO in vascular smooth muscle cells. miR microarray analysis was performed on human vascular smooth muscle cells (HVSMC) treated with ANP (10 nM, 4 h) and S-nitroso- N-acetylpenicillamine (SNAP) (100 μM, 4 h), a NO donor. Twenty-two shared miRs were upregulated, and 21 shared miRs were downregulated, by both ANP and SNAP ( P < 0.05). Expression levels of four miRs (miRs-21, -26b, -98, and -1826), which had the greatest change in expression, as determined by microarray analysis, were confirmed by quantitative RT-PCR. Rp-8-Br-PET-cGMPS, a cGMP-dependent protein kinase-specific inhibitor, blocked the regulation of these miRs by ANP and SNAP. 8-bromo-cGMP mimicked the effect of ANP and SNAP on their expression. miR-21 was shown to inhibit HVSMC contraction in collagen gel lattice contraction assays. We also identified by computational algorithms and confirmed by Western blot analysis new intracellular targets of miR-21, i.e., cofilin-2 and myosin phosphatase and Rho interacting protein. Transfection with pre-miR-21 contracted cells and ANP and SNAP blocked miR-21-induced HVSMC contraction. Transfection with anti-miR-21 inhibitor reduced contractility of HVSMC ( P < 0.05). The present results implicate miRs in NO and ANP signaling in general and miR-21 in particular in cGMP signaling and vascular smooth muscle cell relaxation.

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