scholarly journals Rutaecarpine Increases Nitric Oxide Synthesis via eNOS Phosphorylation by TRPV1-Dependent CaMKII and CaMKKβ/AMPK Signaling Pathway in Human Endothelial Cells

2021 ◽  
Vol 22 (17) ◽  
pp. 9407
Author(s):  
Gi Ho Lee ◽  
Chae Yeon Kim ◽  
Chuanfeng Zheng ◽  
Sun Woo Jin ◽  
Ji Yeon Kim ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Protein Kinase ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Transient Receptor Potential ◽  
Transient Receptor Potential Vanilloid ◽  
Ampk Signaling ◽  
Compound C ◽  
Enos Phosphorylation

Rutaecarpine (RUT) is a bioactive alkaloid isolated from the fruit of Evodia rutaecarpa that exerts a cellular protective effect. However, its protective effects on endothelial cells and its mechanism of action are still unclear. In this study, we demonstrated the effects of RUT on nitric oxide (NO) synthesis via endothelial nitric oxide synthase (eNOS) phosphorylation in endothelial cells and the underlying molecular mechanisms. RUT treatment promoted NO generation by increasing eNOS phosphorylation. Additionally, RUT induced an increase in intracellular Ca2+ concentration and phosphorylation of Ca2+/calmodulin-dependent protein kinase kinase β (CaMKKβ), AMP-activated protein kinase (AMPK), and Ca2+/calmodulin-dependent kinase II (CaMKII). Inhibition of transient receptor potential vanilloid type 1 (TRPV1) attenuated RUT-induced intracellular Ca2+ concentration and phosphorylation of CaMKII, CaMKKβ, AMPK, and eNOS. Treatment with KN-62 (a CaMKII inhibitor), Compound C (an AMPK inhibitor), and STO-609 (a CaMKKβ inhibitor) suppressed RUT-induced eNOS phosphorylation and NO generation. Interestingly, RUT attenuated the expression of ICAM-1 and VCAM-1 induced by TNF-α and inhibited the inflammation-related NF-κB signaling pathway. Taken together, these results suggest that RUT promotes NO synthesis and eNOS phosphorylation via the Ca2+/CaMKII and CaM/CaMKKβ/AMPK signaling pathways through TRPV1. These findings provide evidence that RUT prevents endothelial dysfunction and benefit cardiovascular health.

scholarly journals Sipjeondaebo-tang Alleviates Oxidative Stress-Mediated Liver Injury through Activation of the CaMKK2-AMPK Signaling Pathway

2018 ◽  
Vol 2018 ◽  
pp. 1-13 ◽  
Author(s):  
Sang Mi Park ◽  
Sung Woo Kim ◽  
Eun Hye Jung ◽  
Hae Li Ko ◽  
Chae Kwang Im ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Protein Kinase ◽  
Liver Injury ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Therapeutic Potential ◽  
Glutathione Depletion ◽  
Dependent Manner ◽  
Ampk Signaling ◽  
Compound C

Sipjeondaebo-tang (SDT) is used frequently as a herbal prescription to treat deficiency syndromes in traditional Korean medicine. We investigated the hepatoprotective effects of SDT against oxidative stress and attempted to clarify the underlying molecular mechanisms. SDT pretreatment reduced arachidonic acid (AA) plus iron-mediated cytotoxicity in a concentration-dependent manner and prevented changes in apoptosis-related protein expression. In addition, SDT pretreatment significantly reduced glutathione depletion, hydrogen peroxide production, and mitochondrial dysfunction via treatment with AA plus iron. SDT increased the phosphorylation of AMP-activated protein kinase (AMPK) in accordance with the phosphorylation of Ca2+/calmodulin-dependent protein kinase kinase 2 (CaMKK2). Experiments using an AMPK chemical inhibitor (Compound C) or CaMKK2 chemical inhibitor (STO-609) suggested that the CaMKK2-AMPK signaling pathway contributes to SDT-mediated protection of mitochondria and cells. Moreover, administration of SDT for 4 consecutive days to mice significantly reduced the alanine aminotransferase and aspartate aminotransferase activities induced by carbon tetrachloride, and the numbers of degenerated hepatocytes, infiltrated inflammatory cells, nitrotyrosine-positive cells, and 4-hydroxynonenal-positive cells in liver tissue. Therefore, SDT protects hepatocytes from oxidative stress via CaMKK2-dependent AMPK activation and has the therapeutic potential to prevent or treat oxidative stress-related liver injury.

TRPV1 activation mitigates hypoxic injury in mouse cardiomyocytes by inducing autophagy through the AMPK signaling pathway

2020 ◽  
Vol 318 (5) ◽  
pp. C1018-C1029
Author(s):  
Jinyu Wei ◽  
Jiezhi Lin ◽  
Junhui Zhang ◽  
Di Tang ◽  
Fei Xiang ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Transient Receptor Potential ◽  
Calcium Influx ◽  
Receptor Potential ◽  
Cell Counting ◽  
Transient Receptor Potential Vanilloid ◽  
Hypoxic Stress ◽  
Autophagy Flux ◽  
Ampk Signaling ◽  
Hypoxic Injury

Autophagy is a highly conserved self-protection mechanism that plays a crucial role in cardiovascular diseases. Cardiomyocyte hypoxic injury promotes oxidative stress and pathological alterations in the heart, although the interplay between these effects remains elusive. The transient receptor potential vanilloid 1 (TRPV1) ion channel is a nonselective cation channel that is activated in response to a variety of exogenous and endogenous physical and chemical stimuli. Here, we investigated the effects and mechanisms of action of TRPV1 on autophagy in hypoxic cardiomyocytes. In this study, primary cardiomyocytes isolated from C57 mice were subjected to hypoxic stress, and their expression of TRPV1 and adenosine 5′-monophosphate-activated protein kinase (AMPK) was regulated. The autophagy flux was assessed by Western blotting and immunofluorescence staining, and the cell viability was determined through Cell counting kit-8 assay and Lactate dehydrogenase assays. In addition, the calcium influx after the upregulation of TRPV1 expression in cardiomyocytes was examined. The results showed that the number of autophagosomes in cardiomyocytes was higher under hypoxic stress and that the blockade of autophagy flux aggravated hypoxic damage to cardiomyocytes. Moreover, the expression of TRPV1 was induced under hypoxic stress, and its upregulation by capsaicin improved the autophagy flux and protected cardiomyocytes from hypoxic damage, whereas the silencing of TRPV1 significantly attenuated autophagy. Our observations also revealed that AMPK signaling was activated and involved in TRPV1-induced autophagy in cardiomyocytes under hypoxic stress. Overall, this study demonstrates that TRPV1 activation mitigates hypoxic injury in cardiomyocytes by improving autophagy flux through the AMPK signaling pathway and highlights TRPV1 as a novel therapeutic target for the treatment of hypoxic cardiac disease.

β-Lapachone Regulates Obesity through Modulating Thermogenesis in Brown Adipose Tissue and Adipocytes: Role of AMPK Signaling Pathway

2019 ◽  
Vol 47 (04) ◽  
pp. 803-822 ◽  
Author(s):  
Hyun Jeong Kwak ◽  
Mi-Young Jeong ◽  
Jae-Young Um ◽  
Jinbong Park
Keyword(s):  
Adipose Tissue ◽  
Protein Kinase ◽  
Brown Adipose Tissue ◽  
Signaling Pathway ◽  
Uncoupling Protein ◽  
Therapeutic Implication ◽  
Mitogen Activated Protein Kinase ◽  
Ampk Signaling ◽  
Compound C ◽  
Brown Adipose

Activation of brown adipose tissue (BAT) has been proposed as a promising target against obesity due to its increased capacity for thermogenesis. In this study, we explored the effect of [Formula: see text]-Lapachone ([Formula: see text]L), a compound obtained from the bark of the lapacho tree, against obesity. In vivo administration of [Formula: see text]L into either high fat diet (HFD)-induced obese C57BL6 mice and genetically obese Lepr[Formula: see text] mice prevented body weight gain, which was associated with tissue weight loss of white adipose tissue (WAT). In addition, [Formula: see text]L elevated thermogenic proteins including uncoupling protein 1 (UCP1) and mitochondrial count in BAT and human adipose tissue-derived mesenchymal stem cells (hAMSCs). [Formula: see text]L also induced AMP-activated protein kinase (AMPK) phosphorylation, subsequent upregulation of acetyl-CoA carboxylase (ACC) and UCP1, and these effects were diminished by AMPK inhibitor compound C, suggesting that AMPK underlies the effects of [Formula: see text]L. Mitogen-activated protein kinase pathways participated in the thermogenesis of [Formula: see text]L, specifically p38, c-Jun N-terminal kinase (JNK) and extracellular signal-regulated kinase 1/2 (ERK1/2) were activated by [Formula: see text]L treatment in hAMSCs. Additionally, inhibitors of p38/JNK/ERK1/2 abrogated the activity of [Formula: see text]L. Taken together, [Formula: see text]L exerts anti-obese effects by inducing thermogenesis mediated by AMPK signaling pathway, suggesting that [Formula: see text]L may have a potential therapeutic implication of obesity. Taken together, [Formula: see text]L exerts anti-obese effects by not only inducing thermogenesis on brown adipocytes but also inducing the browning of white adipocytes. The anti-obese effect of [Formula: see text]L is mediated by AMPK signaling pathway, suggesting that [Formula: see text]L may have potential therapeutic implication of obesity.

C1q/TNF–Related Protein-3 attenuates endothelial dysfunction in diabetic retinopathy via the Nitric Oxide signaling pathway

2020 ◽  
Author(s):  
Zheyi Yan ◽  
Xiaoming Cao ◽  
Chunfang Wang ◽  
Sha Liu ◽  
Lu Gan ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Cell Death ◽  
Diabetic Retinopathy ◽  
Endothelial Dysfunction ◽  
Signaling Pathway ◽  
Molecular Mechanisms ◽  
Apoptotic Cell ◽  
Related Protein

Abstract Background Diabetic retinopathy (DR) is a severe microvasculature complication of diabetes. Restoration of dysfunctional endothelial cells represents a promising approach to treatment of DR. It has been demonstrated that a number of CTRP (C1q/tumor necrosis factor-related protein) members improves vascular endothelial function of the aortic vasculature. However, the role of CTRPs in the treatment of DR remains largely unresolved. Therefore, the aim of this study was to determine whether members of the CTRP family improve diabetes-induced endothelial dysfunction of retinal vasculature, thus exhibiting a protective effect against diabetic injury of retina. Methods The vasoactivity of currently identified murine CTRP family members was assessed in vascular rings and the underlying molecular mechanisms elucidated in human retinal microvascular endothelial cells. We then mimicked diabetic retinopathy both in vitro and in vivo, after which they were treated with CTRP3, and the vasoactivity, apoptotic cell death and vascular leakage in the retina were evaluated. Discovery-drive approaches followed by cause-effect analysis were used to uncover the molecular mechanisms of CTRP3. Results Our results demonstrate that CTRP3, CTRP5, and CTRP9 exert vasorelaxant effects on macro- and micro-vessels, with CTRP3 being the most potent in micro-vessels. The effects of CTRP3 were found to be endothelium-dependent via the AdipoR1/AMPK/eNOS/Nitric Oxide (NO) pathway. In in vitro microvascular reactivity studies, CTRP3 successfully improved high glucose/high lipid-induced impairment of endothelium-dependent vasodilatation. Blockade of either AMPK or eNOS completely abolished the previously observed effects of CTRP3. In addition, in the murine diabetic retinopathy model, CTRP3 treatment increased endothelium-dependent relaxation and NO levels in microvessels, and inhibited apoptotic cell death and vascular leakage in the retina. Finally,blockade of NO synthesis completely abolished the effects of CTRP3 that had been measured previously. Conclusion Taken together, our findings reveal that the AdipoR1/AMPK/eNOS/NO signaling pathway, through which CTRP3 reverses endothelial dysfunction of the microvasculature by normalization of impaired vasodilatation, represents a novel intervention effective against diabetic injury of retina.

scholarly journals PKCα mediates acetylcholine-induced activation of TRPV4-dependent calcium influx in endothelial cells

2011 ◽  
Vol 301 (3) ◽  
pp. H757-H765 ◽  
Author(s):  
Ravi K. Adapala ◽  
Phani K. Talasila ◽  
Ian N. Bratz ◽  
David X. Zhang ◽  
Makoto Suzuki ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Calcium Release ◽  
Transient Receptor Potential ◽  
Calcium Influx ◽  
Specific Inhibitor ◽  
Receptor Potential ◽  
Cyclic Stretch ◽  
Mesenteric Arteries ◽  
Transient Receptor Potential Vanilloid

Transient receptor potential vanilloid channel 4 (TRPV4) is a polymodally activated nonselective cationic channel implicated in the regulation of vasodilation and hypertension. We and others have recently shown that cyclic stretch and shear stress activate TRPV4-mediated calcium influx in endothelial cells (EC). In addition to the mechanical forces, acetylcholine (ACh) was shown to activate TRPV4-mediated calcium influx in endothelial cells, which is important for nitric oxide-dependent vasodilation. However, the molecular mechanism through which ACh activates TRPV4 is not known. Here, we show that ACh-induced calcium influx and endothelial nitric oxide synthase (eNOS) phosphorylation but not calcium release from intracellular stores is inhibited by a specific TRPV4 antagonist, AB-159908. Importantly, activation of store-operated calcium influx was not altered in the TRPV4 null EC, suggesting that TRPV4-dependent calcium influx is mediated through a receptor-operated pathway. Furthermore, we found that ACh treatment activated protein kinase C (PKC) α, and inhibition of PKCα activity by the specific inhibitor Go-6976, or expression of a kinase-dead mutant of PKCα but not PKCε or downregulation of PKCα expression by chronic 12- O-tetradecanoylphorbol-13-acetate treatment, completely abolished ACh-induced calcium influx. Finally, we found that ACh-induced vasodilation was inhibited by the PKCα inhibitor Go-6976 in small mesenteric arteries from wild-type mice, but not in TRPV4 null mice. Taken together, these findings demonstrate, for the first time, that a specific isoform of PKC, PKCα, mediates agonist-induced receptor-mediated TRPV4 activation in endothelial cells.

scholarly journals Activation of transient receptor potential vanilloid 4 protects articular cartilage against IL-1β-induced inflammatory responses by regulating the CaMKK/AMPK/NF-κB signaling pathway

2021 ◽  
Author(s):  
Kyosuke Hattori ◽  
Nobunori Takahashi ◽  
Kenya Terabe ◽  
Yoshifumi Ohashi ◽  
Kenji Kishimoto ◽  
...  
Keyword(s):  
Articular Cartilage ◽  
Protein Kinase ◽  
Transient Receptor Potential ◽  
Inflammatory Responses ◽  
Receptor Potential ◽  
Cartilage Tissue ◽  
Transient Receptor Potential Vanilloid ◽  
Compound C ◽  
Transient Receptor ◽  
Safranin O

Abstract Transient receptor potential vanilloid 4 (TRPV4) plays an important role in chondrocytes via Ca2+ signaling. However, its role in the progression of osteoarthritis is unclear. This study aimed to evaluate the effects of TRPV4 activation on articular cartilage and chondrocytes stimulated with interleukin (IL)-1β. Bovine and human articular chondrocytes were stimulated with various agents, including IL-1β, GSK1016790A (GSK101; a TRPV4 agonist), Compound C (an AMP-activated protein kinase (AMPK) inhibitor), and STO-609 (a calmodulin-dependent protein kinase kinase (CaMKK) inhibitor), and were processed for Western blot analysis and real-time PCR. The dimethylmethylene blue (DMMB) assay and Safranin O staining were also performed. GSK101 reversed the IL-1β-induced increase in expression of matrix metalloproteinase (MMP)-13 and decrease in expression of aggrecan. GSK101 also decreased proteoglycan release in the DMMB assay and retained Safranin O staining of articular cartilage tissue. Furthermore, GSK101 increased AMPK phosphorylation and decreased IL-1β-induced nuclear factor kappa B (NF-κB) phosphorylation. Compound C and STO-609 reversed the suppressive effects of GSK101 on NF-κB activation and MMP-13 expression. In conclusion, TRPV4 activation had chondroprotective effects on articular cartilage stimulated with IL-1β by activating CaMKK/AMPK and suppressing the NF-κB pathway. TRPV activators may offer a promising therapeutic option for preventing the progression of osteoarthritis.

scholarly journals Role of the Transient Receptor Potential Vanilloid Type 1 (TRPV1) in the Regulation of Nitric Oxide Release in Wistar Rat Aorta

2021 ◽  
Vol 2021 ◽  
pp. 1-8
Author(s):  
Elvira Varela-López ◽  
Leonardo del Valle-Mondragón ◽  
Vicente Castrejón-Téllez ◽  
Israel Pérez-Torres ◽  
Araceli Páez Arenas ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Transient Receptor Potential ◽  
Rat Aorta ◽  
Aortic Tissue ◽  
Transient Receptor Potential Vanilloid ◽  
Aortic Endothelial Cells ◽  
Aortic Endothelial

The potential transient vanilloid receptor type 1 (TRPV1) plays important functional roles in the vascular system. In the present study, we explored the role of the TRPV1 in the production of nitric oxide (NO), biopterines (BH4 and BH2), cyclic guanosine monophosphate (cGMP), malondialdehyde (MDA), phosphodiesterase-3 (PDE-3), total antioxidant capacity (TAC), and calcitonin gene-related peptide (CGRP) in the rat aorta. Wistar rats were divided into four groups: (1) control, (2) capsaicin (CS, 20 mg/kg), (3) capsazepine (CZ, 24 mg/kg), and (4) CZ + CS. Treatments were applied daily for 4 days before removing the thoracic aortas for testing of aortic tissue and endothelial cells. TRPV1 activation produced increases in BH4 14%, cGMP 25%, NO 29%, and TAC 59.2% in comparison to the controls. BH2 and MDA increased with CZ. CGRP shows a tendency to decrease with CZ. The analysis by immunocytochemistry confirmed that the TRPV1 is present in aortic endothelial cells. Aortic endothelial cells were obtained from healthy rats and cultured to directly explore the effects of CS and CZ. The activation of the TRPV1 (CS 30 μM) produced increases in BH4 17%, NO 36.6%, TAC 56.3%, and CGRP 65%, when compared to controls. BH2 decreased with CZ + CS. CS effects were diminished by CZ in cells and in the tissue. We conclude that the TRPV1 is a structure present in the membrane of aortic endothelial cells and that it participates in the production of NO. The importance of the TRPV1 should be considered in vascular reactivity studies.

scholarly journals MAGI1 Mediates eNOS Activation and NO Production in Endothelial Cells in Response to Fluid Shear Stress

Cells ◽  
2019 ◽  
Vol 8 (5) ◽  
pp. 388 ◽  
Author(s):  
Kedar Ghimire ◽  
Jelena Zaric ◽  
Begoña Alday-Parejo ◽  
Jochen Seebach ◽  
Mélanie Bousquenaud ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Protein Kinase ◽  
Fluid Shear Stress ◽  
Adaptor Protein ◽  
No Production ◽  
Fluid Shear ◽  
Enos Phosphorylation

Fluid shear stress stimulates endothelial nitric oxide synthase (eNOS) activation and nitric oxide (NO) production through multiple kinases, including protein kinase A (PKA), AMP-activated protein kinase (AMPK), AKT and Ca2+/calmodulin-dependent protein kinase II (CaMKII). Membrane-associated guanylate kinase (MAGUK) with inverted domain structure-1 (MAGI1) is an adaptor protein that stabilizes epithelial and endothelial cell-cell contacts. The aim of this study was to assess the unknown role of endothelial cell MAGI1 in response to fluid shear stress. We show constitutive expression and co-localization of MAGI1 with vascular endothelial cadherin (VE-cadherin) in endothelial cells at cellular junctions under static and laminar flow conditions. Fluid shear stress increases MAGI1 expression. MAGI1 silencing perturbed flow-dependent responses, specifically, Krüppel-like factor 4 (KLF4) expression, endothelial cell alignment, eNOS phosphorylation and NO production. MAGI1 overexpression had opposite effects and induced phosphorylation of PKA, AMPK, and CAMKII. Pharmacological inhibition of PKA and AMPK prevented MAGI1-mediated eNOS phosphorylation. Consistently, MAGI1 silencing and PKA inhibition suppressed the flow-induced NO production. Endothelial cell-specific transgenic expression of MAGI1 induced PKA and eNOS phosphorylation in vivo and increased NO production ex vivo in isolated endothelial cells. In conclusion, we have identified endothelial cell MAGI1 as a previously unrecognized mediator of fluid shear stress-induced and PKA/AMPK dependent eNOS activation and NO production.

scholarly journals Role of the AMPK/ACC Signaling Pathway in TRPP2-Mediated Head and Neck Cancer Cell Proliferation

2020 ◽  
Vol 2020 ◽  
pp. 1-9
Author(s):  
Kun Li ◽  
Lei Chen ◽  
Zhangying Lin ◽  
Junwei Zhu ◽  
Yang Fang ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Protein Kinase ◽  
Signaling Pathways ◽  
Signaling Pathway ◽  
Transient Receptor Potential ◽  
Receptor Potential ◽  
Feedback Mechanism ◽  
Initiation Factor ◽  
Expression Levels ◽  
Compound C

Transient receptor potential polycystic 2 (TRPP2) exerts vital roles in various types of cancer; however, its underlying mechanisms remain largely unknown. This study is aimed at investigating whether knockdown of TRPP2 affected the AMP-activated protein kinase (AMPK)/acetyl-CoA carboxylase (ACC) signaling pathway and the proliferation of HN-4, cell line originating from human oral and hypopharyngeal squamous cell carcinoma. In addition, the interactions among AMPK/ACC, AMPK/protein kinase RNA-like endoplasmic reticulum kinase (PERK)/eukaryotic initiation factor 2α (eIF2α) and TRPP2/PERK/eIF2α signaling pathways, and their association with cell proliferation were also explored. The results showed that the relative expression levels of phosphorylated (p)-ACC, p-PERK, and p-eIF2α in HN-4 cells were significantly increased following treatment with 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR) and significantly decreased in cells treated with compound C. Therefore, consistent with previous studies, the AMPK/ACC and AMPK/PERK/eIF2α signaling pathways were upregulated and downregulated following treatment with an AMPK agonist and inhibitor, respectively. Furthermore, TRPP2 knockdown decreased p-PERK and p-eIF2α expression levels and increased those of p-AMPK and p-ACC. Additionally, knockdown of TRPP2 increased HN-4 cell proliferation, while treatment with an AMPK inhibitor or agonist increased or inhibited TRPP2-specific siRNA-mediated cell proliferation, respectively. In conclusion, silencing of TRPP2 expression increased HN-4 cell proliferation via inhibiting the PERK/eIF2α signaling pathway, while the AMPK/ACC signaling pathway was possibly activated by a feedback mechanism to reduce enhanced cell proliferation.

scholarly journals Cicletanine stimulates eNOS phosphorylation and NO production via Akt and MAP kinase/Erk signaling in sinusoidal endothelial cells

2013 ◽  
Vol 305 (2) ◽  
pp. G163-G171 ◽  
Author(s):  
Songling Liu ◽  
Don C. Rockey
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Protein Kinase ◽  
Map Kinase ◽  
Stellate Cell ◽  
Portal Pressure ◽  
Erk Signaling ◽  
No Production ◽  
Sinusoidal Endothelial Cells ◽  
Enos Phosphorylation

The function of the endothelial isoform of nitric oxide synthase (eNOS) and production of nitric oxide (NO) is altered in a number of disease states. Pharmacological approaches to enhancing NO synthesis and thus perhaps endothelial function could have substantial benefits in patients. We analyzed the effect of cicletanine, a synthetic pyridine with potent vasodilatory characteristics, on eNOS function and NO production in normal (liver) and injured rat sinusoidal endothelial cells, and we studied the effect of cicletanine-induced NO on stellate cell contraction and portal pressure in an in vivo model of liver injury. Sinusoidal endothelial cells were isolated from normal and injured rat livers. After exposure to cicletanine, eNOS phosphorylation, NO synthesis, and the signaling pathway regulating eNOS activation were measured. Cicletanine led to an increase in eNOS (Ser1177) phosphorylation, cytochrome c reductase activity, l-arginine conversion to l-citrulline, as well as NO production. The mechanism of the effect of cicletanine appeared to be via the protein kinase B (Akt) and MAP kinase/Erk signaling pathways. Additionally, cicletanine improved NO synthesis in injured sinusoidal endothelial cells. NO production induced by cicletanine in sinusoidal endothelial cells increased protein kinase G (PKG) activity as well as relaxation of stellate cells. Finally, administration of cicletanine to mice with portal hypertension induced by bile duct ligation led to reduction of portal pressure. The data indicate that cicletanine might improve eNOS activity in injured sinusoidal endothelial cells and likely activates hepatic stellate cell NO/PKG signaling. It raises the possibility that cicletanine could improve intrahepatic vascular function in portal hypertensive patients.

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