CCN2 Mediates S1P-Induced Upregulation of COX2 Expression in Human Granulosa-Lutein Cells

CCN1 and CCN2 are members of the CCN family and play essential roles in the regulation of multiple female reproductive functions, including ovulation. Cyclooxygenase-2 (COX2) is a critical mediator of ovulation and can be induced by sphingosine-1-phosphate (S1P) through the S1P1/3-mediated Yes-associated protein (YAP) signaling. However, it is unclear whether CCN1 or CCN2 can mediate S1P-induced upregulation of COX2 expression and increase in prostaglandin E2 (PGE2) production in human granulosa-lutein (hGL) cells. In the present study, we investigated the effects of S1P on the expressions of CCN1 and CCN2 in hGL cells. Additionally, we used a dual inhibition approach (siRNA-mediated silencing and small molecular inhibitors) to investigate the molecular mechanisms of S1P effects. Our results showed that S1P treatment significantly upregulated the expression of CCN1 and CCN2 in a concentration-dependent manner in hGL cells. Additionally, inhibition or silencing of S1P1, but not S1P3, completely abolished the S1P-induced upregulation of CCN2 expression. Furthermore, we demonstrated that S1P-induced nuclear translocation of YAP and inhibition or silencing of YAP completely abolished the S1P-induced upregulation of CCN1 and CCN2 expression. Notably, silencing of CCN2, but not CCN1, completely reversed the S1P-induced upregulation of COX2 expression and the increase in PGE2 production. Thus, CCN2 mediates the S1P-induced upregulation of COX2 expression through the S1P1-mediated signaling pathway in hGL cells. Our findings expand our understanding of the molecular mechanism underlying the S1P-mediated cellular activities in the human ovary.

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S1PR3 mediates itch and pain via distinct TRP channel-dependent pathways

10.1101/235614 ◽

2017 ◽

Author(s):

Rose Z. Hill ◽

Takeshi Morita ◽

Rachel B. Brem ◽

Diana M. Bautista

Keyword(s):

Molecular Mechanisms ◽

Dependent Manner ◽

Cellular Mechanisms ◽

Concentration Dependent Manner ◽

Major Health ◽

Sphingosine 1 Phosphate ◽

Low Levels ◽

Channel Dependent ◽

Behavioral Assays ◽

Insight Into

AbstractSphingosine 1-phosphate (S1P) is a bioactive signaling lipid associated with a variety of chronic pain and itch disorders. S1P signaling has been linked to cutaneous pain, but its role in itch has not yet been studied. Here we find that S1P triggers itch and pain in mice in a concentration-dependent manner, with low levels triggering acute itch alone, and high levels triggering both pain and itch. Calcium imaging and electrophysiological experiments revealed that S1P signals via S1PR3 and TRPA1 in a subset of pruriceptors, and via S1PR3 and TRPV1 in a subset of heat nociceptors. And in behavioral assays, S1P-evoked itch was selectively lost in mice lacking TRPA1, whereas S1P-evoked acute pain and heat hypersensitivity were selectively lost in mice lacking TRPV1. We conclude that S1P acts via different cellular and molecular mechanisms to trigger itch and pain. Our discovery elucidates the diverse roles that S1P signaling plays in somatosensation and provides insight into how itch and pain are discriminated in the periphery.Significance StatementItch and pain are major health problems with few effective treatments. Here, we show that the pro-inflammatory lipid S1P and its receptor S1PR3 trigger itch and pain behaviors via distinct molecular and cellular mechanisms. Our results provide a detailed understanding of the roles that S1P and S1PR3 play in somatosensation, highlighting their potential as targets for analgesics and antipruritics, and provide new insight into the mechanistic underpinnings of itch versus pain discrimination in the periphery.

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The Anti-Atherogenic Properties of Sesamin are Mediated via Improved Macrophage Cholesterol Efflux Through PPARγ1-LXRα and MAPK Signaling

International Journal for Vitamin and Nutrition Research ◽

10.1024/0300-9831/a000195 ◽

2014 ◽

Vol 84 (1-2) ◽

pp. 79-91 ◽

Cited By ~ 7

Author(s):

Amin F. Majdalawieh ◽

Hyo-Sung Ro

Keyword(s):

Cholesterol Efflux ◽

Transcriptional Activity ◽

Molecular Mechanisms ◽

Foam Cell ◽

Mapk Signaling ◽

Luciferase Reporter ◽

Foam Cell Formation ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Macrophage Cholesterol Efflux

Background: Foam cell formation resulting from disrupted macrophage cholesterol efflux, which is triggered by PPARγ1 and LXRα, is a hallmark of atherosclerosis. Sesamin and sesame oil exert anti-atherogenic effects in vivo. However, the exact molecular mechanisms underlying such effects are not fully understood. Aim: This study examines the potential effects of sesamin (0, 25, 50, 75, 100 μM) on PPARγ1 and LXRα expression and transcriptional activity as well as macrophage cholesterol efflux. Methods: PPARγ1 and LXRα expression and transcriptional activity are assessed by luciferase reporter assays. Macrophage cholesterol efflux is evaluated by ApoAI-specific cholesterol efflux assays. Results: The 50 μM, 75 μM, and 100 μM concentrations of sesamin up-regulated the expression of PPARγ1 (p< 0.001, p < 0.001, p < 0.001, respectively) and LXRα (p = 0.002, p < 0.001, p < 0.001, respectively) in a concentration-dependent manner. Moreover, 75 μM and 100 μM concentrations of sesamin led to 5.2-fold (p < 0.001) and 6.0-fold (p<0.001) increases in PPAR transcriptional activity and 3.9-fold (p< 0.001) and 4.2-fold (p < 0.001) increases in LXR transcriptional activity, respectively, in a concentration- and time-dependent manner via MAPK signaling. Consistently, 50 μM, 75 μM, and 100 μM concentrations of sesamin improved macrophage cholesterol efflux by 2.7-fold (p < 0.001), 4.2-fold (p < 0.001), and 4.2-fold (p < 0.001), respectively, via MAPK signaling. Conclusion: Our findings shed light on the molecular mechanism(s) underlying sesamins anti-atherogenic effects, which seem to be due, at least in part, to its ability to up-regulate PPARγ1 and LXRα expression and transcriptional activity, improving macrophage cholesterol efflux. We anticipate that sesamin may be used as a therapeutic agent for treating atherosclerosis.

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Norepinephrine induces calcium spikes and proinflammatory actions in human hepatic stellate cells

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00537.2005 ◽

2006 ◽

Vol 291 (5) ◽

pp. G877-G884 ◽

Cited By ~ 38

Author(s):

Pau Sancho-Bru ◽

Ramón Bataller ◽

Jordi Colmenero ◽

Xavier Gasull ◽

Montserrat Moreno ◽

...

Keyword(s):

Cell Proliferation ◽

Portal Hypertension ◽

Liver Fibrosis ◽

Hepatic Stellate Cells ◽

Molecular Mechanisms ◽

Nuclear Translocation ◽

Stellate Cells ◽

Luciferase Reporter ◽

Dependent Manner ◽

Cell Contraction

Catecholamines participate in the pathogenesis of portal hypertension and liver fibrosis through α1-adrenoceptors. However, the underlying cellular and molecular mechanisms are largely unknown. Here, we investigated the effects of norepinephrine (NE) on human hepatic stellate cells (HSC), which exert vasoactive, inflammatory, and fibrogenic actions in the injured liver. Adrenoceptor expression was assessed in human HSC by RT-PCR and immunocytochemistry. Intracellular Ca2+ concentration ([Ca2+]i) was studied in fura-2-loaded cells. Cell contraction was studied by assessing wrinkle formation and myosin light chain II (MLC II) phosphorylation. Cell proliferation and collagen-α1(I) expression were assessed by [3H]thymidine incorporation and quantitative PCR, respectively. NF-κB activation was assessed by luciferase reporter gene and p65 nuclear translocation. Chemokine secretion was assessed by ELISA. Normal human livers expressed α1A-adrenoceptors, which were markedly upregulated in livers with advanced fibrosis. Activated human HSC expressed α1A-adrenoceptors. NE induced multiple rapid [Ca2+]i oscillations (Ca2+ spikes). Prazosin (α1-blocker) completely prevented NE-induced Ca2+ spikes, whereas propranolol (nonspecific β-blocker) partially attenuated this effect. NE caused phosphorylation of MLC II and cell contraction. In contrast, NE did not affect cell proliferation or collagen-α1(I) expression. Importantly, NE stimulated the secretion of inflammatory chemokines (RANTES and interleukin-8) in a dose-dependent manner. Prazosin blocked NE-induced chemokine secretion. NE stimulated NF-κB activation. BAY 11-7082, a specific NF-κB inhibitor, blocked NE-induced chemokine secretion. We conclude that NE stimulates NF-κB and induces cell contraction and proinflammatory effects in human HSC. Catecholamines may participate in the pathogenesis of portal hypertension and liver fibrosis by targeting HSC.

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Zanthoxylum nitidum extract attenuates BMP-2-induced inflammation and hyperpermeability

Bioscience Reports ◽

10.1042/bsr20201098 ◽

2020 ◽

Vol 40 (10) ◽

Author(s):

Tao Hu ◽

Zhiwen Luo ◽

Kai Li ◽

Shanjin Wang ◽

Desheng Wu

Keyword(s):

Side Effects ◽

Spinal Fusion ◽

Nuclear Translocation ◽

Umbilical Vein ◽

Bone Morphogenetic Protein 2 ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Fluorescent Intensity ◽

Junction Protein ◽

Zanthoxylum Nitidum

Abstract Bone morphogenetic protein-2 (BMP-2) is commonly applied in spinal surgery to augment spinal fusion. Nevertheless, its pro-inflammatory potential could induce dangerous side effects such as vascular hyper-permeability, posing the need for manners against this condition. The present study aims to investigate the protective effect of Zanthoxylum nitidum (ZN) on BMP-2-related hyperpermeability and inflammation on the human umbilical vein endothelial cells (HUVECs). The results revealed that, in a concentration-dependent manner, BMP-2 enhanced the production of pro-inflammatory cytokines, including interleukin (IL)-1α, IL-1β, and tumor necrosis factor-α, which were, however, suppressed by ZN. ZN inhibited BMP-2-induced inflammatory response by suppressing the phosphorylation of NF-κBp65 and IκB, and the abnormal nuclear translocation of p65. Moreover, the inhibited expression intercellular tight junction protein VE-cadherin and Occludin caused by BMP-2 was blocked by ZN. The hyper-permeability of HUVECs induced by BMP-2, as expressed as the higher fluorescent intensity of dextran, was also reversed by ZN. Overall, these findings demonstrated that ZN antagonized BMP-2-induced inflammation and hyperpermeability. It could be a therapeutic candidate for the treatment of BMP-2-induced side effects during spinal fusion.

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Sphingosine Kinase as a Regulator of Calcium Entry through Autocrine Sphingosine 1-Phosphate Signaling in Thyroid FRTL-5 Cells

Endocrinology ◽

10.1210/en.2009-0288 ◽

2009 ◽

Vol 150 (11) ◽

pp. 5125-5134 ◽

Cited By ~ 11

Author(s):

Dan Gratschev ◽

Christoffer Löf ◽

Jari Heikkilä ◽

Anders Björkbom ◽

Pramod Sukumaran ◽

...

Keyword(s):

Intracellular Signaling ◽

Sphingosine Kinase ◽

Calcium Entry ◽

Dominant Negative ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Entry Pathway ◽

Sphingosine 1 Phosphate ◽

S1p Receptor ◽

In Cells

Calcium entry is one of the main regulators of intracellular signaling. Here, we have described the importance of sphingosine, sphingosine kinase 1 (SK1), and sphingosine 1-phosphate (S1P) in regulating calcium entry in thyroid FRTL-5 cells. In cells incubated with the phosphatase inhibitor calyculin A, which evokes calcium entry without mobilizing sequestered intracellular calcium, sphingosine inhibited calcium entry in a concentration-dependent manner. Furthermore, inhibiting SK1 or the ATP-binding cassette ABCC1 multidrug transporter attenuated calcium entry. The addition of exogenous S1P restored calcium entry. Neither sphingosine nor inhibition of SK1 attenuated thapsigargin-evoked calcium entry. Blocking S1P receptor 2 or phospholipase C attenuated calcium entry, whereas blocking S1P receptor 3 did not. Overexpression of wild-type SK1, but not SK2, enhanced calyculin-evoked calcium entry compared with mock-transfected cells, whereas calcium entry was decreased in cells transfected with the dominant-negative G82D SK1 mutant. Exogenous S1P restored calcium entry in G82D cells. Our results suggest that the calcium entry pathway is blocked by sphingosine and that activation of SK1 and the production of S1P, through an autocrine mechanism, facilitate calcium entry through activation of S1P receptor 2. This is a novel mechanism by which the sphingosine-S1P rheostat regulates cellular calcium homeostasis.

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Methanol Extract of Mongolian Iris bungei Maxim. Stimulates 3T3-L1 Adipocyte Differentiation

Journal of Nanoscience and Nanotechnology ◽

10.1166/jnn.2021.19160 ◽

2021 ◽

Vol 21 (7) ◽

pp. 3943-3949

Author(s):

Jaegoo Yeon ◽

Sung-Suk Suh ◽

Ui-Joung Youn ◽

Badamtsetseg Bazarragchaa ◽

Ganbold Enebish ◽

...

Keyword(s):

Bacterial Infections ◽

Fatty Acid Synthase ◽

Molecular Mechanisms ◽

Adipocyte Differentiation ◽

Methanol Extract ◽

Regulatory Element ◽

Peroxisome Proliferator ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Peroxisome Proliferator Activated Receptor

Iris bungei Maxim. (IB), which is native to China and Mongolia, is used as a traditional medicine for conditions such as inflammation, cancer, and bacterial infections. However, the effects of Iris bungei Maxim. on adipocyte differentiation have not been studied. In the present study, we first demonstrated the molecular mechanisms underlying the adipogenic activity of the methanol extract of Mongolian I. bungei Maxim. (IB). IB significantly enhanced intracellular lipid accumulation and adipocyte differentiation in 3T3-L1 preadipocytes in a concentration-dependent manner. Moreover, IB markedly stimulated the expression of genes related to adipogenesis such as peroxisome proliferator-activated receptor γ, adiponectin, and aP2. In addition, we also observed that IB induces lipogenic genes such as fatty acid synthase, sterol regulatory element binding protein 1c, stearoyl-CoA desaturase, and acetyl-CoA carboxylase. Interestingly IB regulated adipocyte differentiation in both the early and middle stages. Taken together, these adipogenic and lipogenic effects of IB suggest its efficacy for the prevention and/or treatment of type 2 diabetes.

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Betulinic Acid Suppresses Ovarian Cancer Cell Proliferation through Induction of Apoptosis

Biomolecules ◽

10.3390/biom9070257 ◽

2019 ◽

Vol 9 (7) ◽

pp. 257 ◽

Cited By ~ 1

Author(s):

Dahae Lee ◽

Seoung Rak Lee ◽

Ki Sung Kang ◽

Yuri Ko ◽

Changhyun Pang ◽

...

Keyword(s):

Ovarian Cancer ◽

Cancer Cell ◽

Betulinic Acid ◽

Molecular Mechanisms ◽

Underlying Mechanism ◽

Nuclear Staining ◽

Dependent Manner ◽

Meoh Extract ◽

Concentration Dependent Manner ◽

Induction Of Apoptosis

Ovarian cancer is one of the leading causes of cancer deaths worldwide in women, and the most malignant cancer among the different gynecological cancers. In this study, we explored potentially anticancer compounds from Cornus walteri (Cornaceae), the MeOH extract of which has been reported to show considerable cytotoxicity against several cancer cell lines. Phytochemical investigations of the MeOH extract of the stem and stem bark of C. walteri by extensive application of chromatographic techniques resulted in the isolation of 14 compounds (1–14). The isolated compounds were evaluated for inhibitory effects on the viability of A2780 human ovarian carcinoma cells and the underlying molecular mechanisms were investigated. An 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was employed to assess the anticancer effects of compounds 1–14 on A2780 cells, which showed that compound 11 (betulinic acid) reduced the viability of these cells in a concentration-dependent manner and had an half maximal (50%) inhibitory concentration (IC50) of 44.47 μM at 24 h. Nuclear staining and image-based cytometric assay were carried out to detect the induction of apoptosis by betulinic acid. Betulinic acid significantly increased the condensation of nuclei and the percentage of apoptotic cells in a concentration-dependent manner in A2780 cells. Western blot analysis was performed to investigate the underlying mechanism of apoptosis. The results indicated that the expression levels of cleaved caspase-8, -3, -9, and Bax were increased in A2780 cells treated with betulinic acid, whereas those of Bcl-2 were decreased. Thus, we provide the experimental evidence that betulinic acid can induce apoptosis in A2780 cells through both mitochondria-dependent and -independent pathways and suggest the potential use of betulinic acid in the development of novel chemotherapeutics for ovarian cancer therapy.

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Vascular Protective Effects of Xanthotoxin and Its Action Mechanism in Rat Aorta and Human Vascular Endothelial Cells

Journal of Vascular Research ◽

10.1159/000509112 ◽

2020 ◽

Vol 57 (6) ◽

pp. 313-324

Author(s):

Li-Hua Cao ◽

Ho Sub Lee ◽

Zhe-Shan Quan ◽

Yun Jung Lee ◽

Yu Jin

Keyword(s):

Endothelial Cells ◽

Cell Adhesion ◽

Adhesion Molecule ◽

Molecular Mechanisms ◽

Umbilical Vein ◽

Neuroprotective Effect ◽

Intercellular Adhesion Molecule ◽

Dependent Manner ◽

Protective Effects ◽

Concentration Dependent Manner

Objective: Xanthotoxin (XAT) is a linear furanocoumarin mainly extracted from the plants Ammi majus L. XAT has been reported the apoptosis of tumor cells, anti-convulsant, neuroprotective effect, antioxidative activity, and vasorelaxant effects. This study aimed to investigate the vascular protective effects and underlying molecular mechanisms of XAT. Methods: XAT’s activity was studied in rat thoracic aortas, isolated with aortic rings, and human umbilical vein endothelial cells (HUVECs). Results: XAT induced endothelium-dependent vasodilation in a concentration-dependent manner in the isolated rat thoracic aortas. Removal of endothelium or pretreatment of aortic rings with L-NAME, 1H-[1,2,4]-oxadiazolo-[4,3-a]-quinoxalin-1-one, and wortmannin significantly inhibited XAT-induced relaxation. In addition, treatment with thapsigargin, 2-aminoethyl diphenylborinate, Gd3+, and 4-aminopyridine markedly attenuated the XAT-induced vasorelaxation. XAT increased nitric oxide production and Akt- endothelial NOS (eNOS) phosphorylation in HUVECs. Moreover, XAT attenuated the expression of TNF-α-induced cell adhesion molecules such as intercellular adhesion molecule, vascular cell adhesion molecule-1, and E-selectin. However, this effect was attenuated by the eNOS inhibitors L-NAME and asymmetric dimethylarginine. Conclusions: This study suggests that XAT induces vasorelaxation through the Akt-eNOS-cGMP pathway by activating the KV channel and inhibiting the L-type Ca2+ channel. Furthermore, XAT exerts an inhibitory effect on vascular inflammation, which is correlated with the observed vascular protective effects.

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Suppression of the TGF-β pathway by a macrolide antibiotic decreases fibrotic responses by ocular fibroblasts in vitro

Royal Society Open Science ◽

10.1098/rsos.200441 ◽

2020 ◽

Vol 7 (9) ◽

pp. 200441

Author(s):

Thomas Stahnke ◽

Beata Gajda-Deryło ◽

Anselm G. Jünemann ◽

Oliver Stachs ◽

Katharina A. Sterenczak ◽

...

Keyword(s):

Molecular Mechanisms ◽

Transforming Growth Factor ◽

Macrolide Antibiotic ◽

Drug Repositioning ◽

Dependent Manner ◽

Glaucoma Filtration Surgery ◽

Concentration Dependent Manner ◽

Device Implantation

To elucidate and to inhibit post-surgical fibrotic processes after trabeculectomy in glaucoma therapy, we measured gene expression in a fibrotic cell culture model, based on transforming growth factor TGF-β induction in primary human tenon fibroblasts (hTFs), and used Connectivity Map (CMap) data for drug repositioning. We found that specific molecular mechanisms behind fibrosis are the upregulation of actins, the downregulation of CD34, and the upregulation of inflammatory cytokines such as IL6, IL11 and BMP6 . The macrolide antibiotic Josamycin (JM) reverses these molecular mechanisms according to data from the CMap, and we thus tested JM as an inhibitor of fibrosis. JM was first tested for its toxic effects on hTFs, where it showed no influence on cell viability, but inhibited hTF proliferation in a concentration-dependent manner. We then demonstrated that JM suppresses the synthesis of extracellular matrix (ECM) components. In hTFs stimulated with TGF-β1, JM specifically inhibited α-smooth muslce actin expression, suggesting that it inhibits the transformation of fibroblasts into fibrotic myofibroblasts. In addition, a decrease of components of the ECM such as fibronectin, which is involved in in vivo scarring, was observed. We conclude that JM may be a promising candidate for the treatment of fibrosis after glaucoma filtration surgery or drainage device implantation in vivo .

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Activation of the Transcription Factor GLI1 by WNT Signaling Underlies the Role of SULFATASE 2 as a Regulator of Tissue Regeneration

Journal of Biological Chemistry ◽

10.1074/jbc.m112.443440 ◽

2013 ◽

Vol 288 (29) ◽

pp. 21389-21398 ◽

Cited By ~ 21

Author(s):

Ikuo Nakamura ◽

Maite G. Fernandez-Barrena ◽

Maria C. Ortiz-Ruiz ◽

Luciana L. Almada ◽

Chunling Hu ◽

...

Keyword(s):

Transcription Factor ◽

Wnt Signaling ◽

Cyclin D1 ◽

Signaling Pathways ◽

Tissue Regeneration ◽

Molecular Mechanisms ◽

Nuclear Translocation ◽

Hepatocyte Proliferation ◽

Dependent Manner ◽

Gli1 Expression

Tissue regeneration requires the activation of a set of specific growth signaling pathways. The identity of these cascades and their biological roles are known; however, the molecular mechanisms regulating the interplay between these pathways remain poorly understood. Here, we define a new role for SULFATASE 2 (SULF2) in regulating tissue regeneration and define the WNT-GLI1 axis as a novel downstream effector for this sulfatase in a liver model of tissue regeneration. SULF2 is a heparan sulfate 6-O-endosulfatase, which releases growth factors from extracellular storage sites turning active multiple signaling pathways. We demonstrate that SULF2-KO mice display delayed regeneration after partial hepatectomy (PH). Mechanistic analysis of the SULF2-KO phenotype showed a decrease in WNT signaling pathway activity in vivo. In isolated hepatocytes, SULF2 deficiency blocked WNT-induced β-CATENIN nuclear translocation, TCF activation, and proliferation. Furthermore, we identified the transcription factor GLI1 as a novel target of the SULF2-WNT cascade. WNT induces GLI1 expression in a SULF2- and β-CATENIN-dependent manner. GLI1-KO mice phenocopied the SULF2-KO, showing delayed regeneration and decreased hepatocyte proliferation. Moreover, we identified CYCLIN D1, a key mediator of cell growth during tissue regeneration, as a GLI1 transcriptional target. GLI1 binds to the cyclin d1 promoter and regulates its activity and expression. Finally, restoring GLI1 expression in the liver of SULF2-KO mice after PH rescues CYCLIN D1 expression and hepatocyte proliferation to wild-type levels. Thus, together these findings define a novel pathway in which SULF2 regulates tissue regeneration in part via the activation of a novel WNT-GLI1-CYCLIN D1 pathway.

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