Yahan Liu

New paradigms in inflammatory signaling in vascular endothelial cells

Inflammation is a basic cellular process in innate and adaptive immunity. Vascular endothelial cells play an important role in the initiation, amplification, and resolution of the inflammatory response. Deregulated inflammatory response is implicated in a variety of cardiovascular diseases such as atherosclerosis, obesity, diabetes, and hypertension. Recent studies have made significant progresses in the understanding of the complex molecular pathways that mediate the pro- and anti-inflammatory signaling in endothelial cells (ECs). Specifically, a number of macromolecular complexes termed as signalosomes have been identified to integrate the proinflammatory signaling from the membrane receptors to key transcription factors such as nuclear factor-κB (NF-κB). Inflammasomes are associated with the pattern-recognition receptors such as Toll-like receptors (TLRs), nucleotide-binding oligomerization-domain (NOD)-like receptors (NLRs) to mediate innate immunity responses. Emerging evidence has also revealed that noncoding microRNAs constitute a new class of intra- and intercellular signaling molecules to modulate inflammation in ECs. Thus this article will briefly summarize these new mechanisms with a special emphasis in the context of cardiovascular diseases.

PPARδ Activation Protects Endothelial Function in Diabetic Mice

Recent evidence highlights the therapeutic potential of peroxisome proliferator–activated receptor-δ (PPARδ) agonists to increase insulin sensitivity in diabetes. However, the role of PPARδ in regulating vascular function is incompletely characterized. We investigate whether PPARδ activation improves endothelial function in diabetic and obese mice. PPARδ knockout (KO) and wild-type (WT) mice fed with high-fat diet and db/db mice were used as diabetic mouse models, compared with PPARδ KO and WT mice on normal diet and db/m+ mice. Endothelium-dependent relaxation (EDR) was measured by wire myograph. Flow-mediated vasodilatation (FMD) was measured by pressure myograph. Nitric oxide (NO) production was examined in primary endothelial cells from mouse aortae. PPARδ agonist GW1516 restored EDRs in mouse aortae under high-glucose conditions or in db/db mouse aortae ex vivo. After oral treatment with GW1516, EDRs in aortae and FMDs in mesenteric resistance arteries were improved in obese mice in a PPARδ-specific manner. The effects of GW1516 on endothelial function were mediated through phosphatidylinositol 3-kinase (PI3K) and Akt with a subsequent increase of endothelial nitric oxide synthase (eNOS) activity and NO production. The current study demonstrates an endothelial-protective effect of PPARδ agonists in diabetic mice through PI3K/Akt/eNOS signaling, suggesting the therapeutic potential of PPARδ agonists for diabetic vasculopathy.

Tumor-derived microRNA-494 promotes angiogenesis in non-small cell lung cancer

Angiogenesis, a crucial step in tumor growth and metastasis, is regulated by various pro- or anti-angiogenic factors. Recently, microRNAs have been shown to modulate angiogenic processes by modulating the expression of critical angiogenic factors. However, roles of tumor-derived microRNAs in regulating tumor vascularization remain to be elucidated. In this study, we found that delivery of miR-494 into human vascular endothelial cells (ECs) enhanced the EC migration and promoted angiogenesis. The angiogenic effect of miR-494 was mediated by the targeting of PTEN and the subsequent activation of Akt/eNOS pathway. Importantly, co-culture experiments demonstrated that a lung cancer cell line, A549, secreted and delivered miR-494 into ECs via a microvesicle-mediated route. In addition, we found that the expression of miR-494 was induced in the tumor cells in response to hypoxia, likely via a HIF-1α-mediated mechanism. Furthermore, a specific miR-494 antagomiR effectively inhibited angiogenesis and attenuated the growth of tumor xenografts in nude mice. Taken together, these results demonstrated that miR-494 is a novel tumor-derived paracrine signal to promote angiogenesis and tumor growth under hypoxic condition.

Peroxisome Proliferator-Activated Receptor-γ Ameliorates Pulmonary Arterial Hypertension by Inhibiting 5-Hydroxytryptamine 2B Receptor

An elevated plasma level of 5-hydroxytryptamine (5-HT) or upregulation of 5-HT receptor signaling or both is implicated in vascular contraction and remodeling in pulmonary arterial hypertension (PAH). Recently, peroxisome proliferator-activated receptor-&ggr; (PPAR&ggr;) agonists have been shown to ameliorate PAH. However, their effects on the 5-HT-induced contraction of pulmonary arteries remain unknown. Here, we examined the role of PPAR&ggr; in inhibiting 5-HT2B receptor (5-HT2BR) to ameliorate PAH. Pulmonary arteries from PAH rats induced by monocrotaline or chronic hypoxia showed an enhanced vasoconstriction in response to BW723C86, a specific agonist for 5-HT2BR. Expression of 5-HT2BR was also increased in pulmonary arteries from the PAH rats, accompanied by vascular remodeling and right ventricular hypertrophy. Treatment with the PPAR&ggr; agonist rosiglitazone in vivo reversed the expression and the vasocontractive effect of 5-HT2BR as well as the thickening of pulmonary arteries. In pulmonary artery smooth muscle cells, 5-HT induced the gene expression of 5-HT2BR, which was inhibited by rosiglitazone, pioglitazone, or adenovirus-mediated overexpression of constitutively activated PPAR&ggr;. The pharmacological effect of PPAR&ggr; was through the suppression of the 5-HT-induced activator protein-1 activity. These results demonstrated the beneficial effect of PPAR&ggr; on 5-HT2BR-mediated vasocontraction, providing a new mechanism for the potential use of PPAR&ggr; agonists in PAH.

Rosiglitazone Attenuated Endothelin-1-Induced Vasoconstriction of Pulmonary Arteries in the Rat Model of Pulmonary Arterial Hypertension via Differential Regulation of ET-1 Receptors

Pulmonary arterial hypertension (PAH) is a fatal disease characterized by a progressive increase in pulmonary arterial pressure leading to right ventricular failure and death. Activation of the endothelin (ET)-1 system has been demonstrated in plasma and lung tissue of PAH patients as well as in animal models of PAH. Recently, peroxisome proliferator-activated receptor γ (PPARγ) agonists have been shown to ameliorate PAH. The present study aimed to investigate the mechanism for the antivasoconstrictive effects of rosiglitazone in response to ET-1 in PAH. Sprague-Dawley rats were exposed to chronic hypoxia (10% oxygen) for 3 weeks. Pulmonary arteries from PAH rats showed an enhanced vasoconstriction in response to ET-1. Treatment with PPARγ agonist rosiglitazone (20 mg/kg per day) with oral gavage for 3 days attenuated the vasocontractive effect of ET-1. The effect of rosiglitazone was lost in the presence of L-NAME, indicating a nitric oxide-dependent mechanism. Western blotting revealed that rosiglitazone increased ETBR but decreased ETAR level in pulmonary arteries from PAH rats. ETBR antagonist A192621 diminished the effect of rosiglitazone on ET-1-induced contraction. These results demonstrated that rosiglitazone attenuated ET-1-induced pulmonary vasoconstriction in PAH through differential regulation of the subtypes of ET-1 receptors and, thus, provided a new mechanism for the therapeutic use of PPARγ agonists in PAH.

Epigallocatechin-3-O-Gallate, a Green Tea Polyphenol, Induces Expression of Pim-1 Kinase Via PPARγ in Human Vascular Endothelial Cells

Pim-1 is a serine/threonine kinase and involved in cell survival and proliferation. Recently, it has been shown that pim-1 signaling pathway plays an important role in cardiovascular protection and differentiation. In this study, we sought to explore the expression of pim-1 in human vascular endothelial cells (ECs) and its regulation by epigallocatechin-3-O-gallate (EGCG), a green tea polyphenol which has anti-oxidant, anti-inflammatory and vascular protective effects. By using quantitative reverse transcriptase PCR (qRT-PCR) and Western blotting, we showed that EGCG dose-dependently increased the expression of pim-1 in cultured umbilical vein endothelial cells. Next, we showed that EGCG activated a luciferase reporter driven by peroxisome proliferators-activated receptor (PPAR)-responsive elements. The induced expression of pim-1 was inhibited in ECs pretreated with GW9662, a specific antagonist of PPARγ. In addition, pim-1 was also up-regulated in endothelial cells treated with rosiglitazone, a specific agonist for PPARγ, or those infected with the adenovirus expressing a constitutively active PPARγ. Collectively, our results provided new evidence that pim-1 can be up-regulated by EGCG via a PPARγ-mediated mechanism and may mediate its vascular protective effects.

Peroxisome Proliferator-Activated Receptor γ Regulates the Expression of Lipid Phosphate Phosphohydrolase 1 in Human Vascular Endothelial Cells

Lipid phosphate phosphohydrolase 1 (LPP1), a membrane ectophosphohydrolase regulating the availability of bioactive lipid phosphates, plays important roles in cellular signaling and physiological processes such as angiogenesis and endothelial migration. However, the regulated expression of LPP1 remains largely unknown. Here, we aimed to examine a role of peroxisome proliferator-activated receptor γ (PPARγ) in the transcriptional control of LPP1 gene expression. In human umbilical vein endothelial cells (HUVECs), quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) demonstrated that activation of PPARγ increased the mRNA level of LPP1. Chromatin immunoprecipitation assay showed that PPARγ binds to the putative PPAR-responsive elements (PPREs) within the 5′-flanking region of the human LPP1 gene. Genomic fragment containing 1.7-kilobase of the promoter region was cloned by using PCR. The luciferase reporter assays demonstrated that overexpression of PPARγ and rosiglitazone, a specific ligand for PPARγ, could significantly upregulate the reporter activity. However, site-directed mutagenesis of the PPRE motif abolished the induction. In conclusion, our results demonstrated that PPARγ transcriptionally activated the expression of LPP1 gene in ECs, suggesting a potential role of PPARγ in the metabolism of phospholipids.

Inhibition of 5‐Hydroxytryptamine Receptor 2B Reduced Vascular Restenosis and Mitigated the β‐Arrestin2–Mammalian Target of Rapamycin/p70S6K Pathway

Background As a monoamine neurotransmitter, 5‐hydroxytryptamine (5‐HT) or serotonin modulates mood, appetite, and sleep. Besides, 5‐HT also has important peripheral functions. 5‐HT receptor 2B (5‐HT2BR) plays a key role in cardiovascular diseases, such as pulmonary arterial hypertension and cardiac valve disease. Percutaneous intervention has been used to restore blood flow in occlusive vascular disease. However, restenosis remains a significant problem. Herein, we investigated the role of 5‐HT2BR in neointimal hyperplasia, a key pathological process in restenosis. Methods and Results The expression of 5‐HT2BR was upregulated in wire‐injured mouse femoral arteries. In addition, BW723C86, a selective 5‐HT2BR agonist, promoted the injury response during restenosis. 5‐HT and BW723C86 stimulated migration and proliferation of rat aortic smooth muscle cells. Conversely, LY272015, a selective antagonist, attenuated the 5‐HT–induced smooth muscle cell migration and proliferation. In vitro study showed that the promigratory effects of 5‐HT2BR were mediated through the activation of mammalian target of rapamycin (mTOR)/p70S6K signaling in a β‐arrestin2–dependent manner. Inhibition of mammalian target of rapamycin or p70S6K mitigated 5‐HT2BR–mediated smooth muscle cell migration. Mice with deficiency of 5‐HT2BR showed significantly reduced neointimal formation in wire‐injured arteries. Conclusions These results demonstrated that activation of 5‐HT2BR and β‐arrestin2–biased downstream signaling are key pathological processes in neointimal formation, and 5‐HT2BR may be a potential target for the therapeutic intervention of vascular restenosis.

Response to Overexpression of 5-Hydroxytryptamine 2B Receptor Gene in Pulmonary Hypertension: Still a Long Way to Understand its Transcriptional Regulation

Pulmonary arterial hypertension (PAH) is a progressive and fatal disease, which involves pulmonary vasoconstriction and cardiovascular remodeling. Our present study shows that treatment with rosiglitazone, the peroxisome proliferator-activated receptor γ (PPARγ) agonist is able to attenuate 5-hydroxytryptamine (5-HT)-induced vasoconstriction of pulmonary arteries from monocrotaline- or hypoxia-treated rats and decreases the expression of serotonin receptor 2B, suggesting a potential mechanism for the benefit of PPARγ ligands in PAH.1 We thank Dr Maroteaux2 for the interest of our publication. In his Letter to the Editor, he raised several critiques of our study. Although his concern with the specificity of the 5-HT2A agonist TCB-2 is justifiable, the present study indicates a significant role of 5-HT2B pathological changes in …