Browsing by Author "Seye, Cheikh I."

Now showing 1 - 5 of 5
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    Deletion of P2Y2 receptor reveals a role for lymphotoxin-α in fatty streak formation
    (Elsevier, 2016-10) Aian, Shaomin; Hoggatt, April; Jones-Hall, Yava L.; Ware, Carl F.; Herring, Paul; Seye, Cheikh I.; Department of Cellular & Integrative Physiology, IU School of Medicine
    Background Lymphotoxin alpha (LTα) is expressed in human atherosclerotic lesions and genetic variations in the LTα pathway have been linked to myocardial infarction. Activation of the P2Y2 nucleotide receptor (P2Y2R) regulates the production of LTα. in vitro. We aimed to uncover a potential pathway linking purinergic receptor to LTα-mediated inflammatory processes pivotal to the early stages of atherosclerosis in apolipoprotein E (ApoE−/−) deficient mice. Methods and results En face immunostaining revealed that P2Y2R and VCAM-1 are preferentially expressed in the atherosclerosis prone site of the mouse aortic sinus. Deletion of the P2Y2R gene suppresses VCAM-1 expression. Compared with ApoE−/− mice, ApoE−/− mice lacking the P2Y2R gene (ApoE−/−/P2Y2R−/−) did not develop fatty streak lesions when fed a standard chow diet for 15 weeks. Systemic and CD4+ T cell production of the pro-inflammatory cytokine lymphotoxin-alpha (LTα) were specifically inhibited in ApoE−/−/P2Y2R−/−mice. Anti-LTα preventive treatment was initiated in ApoE−/− mice with intraperitoneal administration of recombinant human tumor necrosis factor receptor 1 fusion protein (TNFR1-Fc) on 5 consecutive days before the disease onset. Remarkably, none of the TNFR1:Fc-treated ApoE−/− mice exhibited atherosclerotic lesions at any developmental stage. Significance ApoE−/− mice deficient in P2Y2R exhibit low endothelial cell VCAM-1 levels, decreased production of LTα and delayed onset of atherosclerosis. These data suggest that targeting this nucleotide receptor could be an effective therapeutic approach in atherosclerosis.
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    Direct Evidence for P2Y2 Receptor Involvement in Vascular Response to Injury
    (Karger Publishers, 2016) Agca, Yuksel; Qian, Shaomin; Agca, Cansu; Seye, Cheikh I.; Cellular and Integrative Physiology, School of Medicine
    Objectives Extracellular nucleotide release at the site of arterial injury mediates proliferation and migration of vascular smooth muscle cells (SMC). Our aim was to investigate the role of the P2Y2 nucleotide receptor (P2Y2R) in neointimal hyperplasia. Approach and Results Vascular injury was induced by implantation of a polyethylene cuff around the femoral artery in wild-type and P2Y2 receptor-deficient mice (P2Y2R−/−). Electron microscopy was used to analyze monocyte and lymphocyte influx to the intima 36 hours post-injury. Compared to wild-type (WT) littermates, P2Y2R−/− mice exhibited a 3-fold decreased number of mononuclear leukocytes invading the intima (p<0.05). Concomitantly, migration of smooth muscle cells was decreased by more than 60% (p<0.05) a resulting in a sharp inhibition of intimal thickening formation in P2Y2R−/− mice (n=15) 14 days after cuff placement. In vitro, loss of P2Y2 receptor significantly impaired monocyte migration in response to nucleotide agonists. Furthermore, transgenic rats over-expressing the P2Y2R developed accelerated intimal lesions resulting in more than 95% luminal stenosis (P<0.05, n=10). Conclusions Loss-and gain-of-function approaches established a direct evidence for P2Y2 receptor involvement in neointimal hyperplasia. Specific anti-P2Y2 receptor therapies may be used against restenosis and bypass graft failure.
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    Endothelial Cell-Specific Deletion of P2Y2 Receptor Promotes Plaque Stability in Atherosclerosis-Susceptible ApoE-Null Mice
    (American Heart Association, 2017-01) Xingjuan, Chen; Qian, Shaomin; Hoggatt, April; Tang, Hongying; Hacker, Timothy A.; Obukhov, Alexander G.; Herring, Paul B.; Seye, Cheikh I.; Cellular and Integrative Physiology, School of Medicine
    OBJECTIVE: Nucleotide P2Y2 receptor (P2Y2R) contributes to vascular inflammation by increasing vascular cell adhesion molecule-1 expression in endothelial cells (EC), and global P2Y2R deficiency prevents fatty streak formation in apolipoprotein E null (ApoE-/-) mice. Because P2Y2R is ubiquitously expressed in vascular cells, we investigated the contribution of endothelial P2Y2R in the pathogenesis of atherosclerosis. APPROACH AND RESULTS: EC-specific P2Y2R-deficient mice were generated by breeding VEcadherin5-Cre mice with the P2Y2R floxed mice. Endothelial P2Y2R deficiency reduced endothelial nitric oxide synthase activity and significantly altered ATP- and UTP (uridine 5'-triphosphate)-induced vasorelaxation without affecting vasodilatory responses to acetylcholine. Telemetric blood pressure and echocardiography measurements indicated that EC-specific P2Y2R-deficient mice did not develop hypertension. We investigated the role of endothelial P2Y2R in the development of atherosclerotic lesions by crossing the EC-specific P2Y2R knockout mice onto an ApoE-/- background and evaluated lesion development after feeding a standard chow diet for 25 weeks. Histopathologic examination demonstrated reduced atherosclerotic lesions in the aortic sinus and entire aorta, decreased macrophage infiltration, and increased smooth muscle cell and collagen content, leading to the formation of a subendothelial fibrous cap in EC-specific P2Y2R-deficient ApoE-/- mice. Expression and proteolytic activity of matrix metalloproteinase-2 was significantly reduced in atherosclerotic lesions from EC-specific P2Y2R-deficient ApoE-/- mice. Furthermore, EC-specific P2Y2R deficiency inhibited nitric oxide production, leading to significant increase in smooth muscle cell migration out of aortic explants. CONCLUSIONS: EC-specific P2Y2R deficiency reduces atherosclerotic burden and promotes plaque stability in ApoE-/- mice through impaired macrophage infiltration acting together with reduced matrix metalloproteinase-2 activity and increased smooth muscle cell migration.
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    The P2Y2 nucleotide receptor is an inhibitor of vascular calcification
    (Elsevier, 2017-02) Qian, Shaomin; Regan, Jenna N.; Shelton, Maxwell T.; Hoggatt, April; Mohammad, Khalid S.; Herring, Paul B.; Seye, Cheikh I.; Cellular and Integrative Physiology, School of Medicine
    BACKGROUND AND AIMS: Mutations in the 5'-nucleotidase ecto (NT5E) gene that encodes CD73, a nucleotidase that converts AMP to adenosine, are linked to arterial calcification. However, the role of purinergic receptor signaling in the pathology of intimal calcification is not well understood. In this study, we examined whether extracellular nucleotides acting via P2Y2 receptor (P2Y2R) modulate arterial intimal calcification, a condition highly correlated with cardiovascular morbidity. METHODS: Apolipoprotein E, P2Y2R double knockout mice (ApoE-/-P2Y2R-/-) were used to determine the effect of P2Y2R deficiency on vascular calcification in vivo. Vascular smooth muscle cells (VSMC) isolated from P2Y2R-/- mice grown in high phosphate medium were used to assess the role of P2Y2R in the conversion of VSMC into osteoblasts. Luciferase-reporter assays were used to assess the effect of P2Y2R on the transcriptional activity of Runx2. RESULTS: P2Y2R deficiency in ApoE-/- mice caused extensive intimal calcification despite a significant reduction in atherosclerosis and macrophage plaque content. The ectoenzyme apyrase that degrades nucleoside di- and triphosphates accelerated high phosphate-induced calcium deposition in cultured VSMC. Expression of P2Y2R inhibits calcification in vitro inhibited the osteoblastic trans-differentiation of VSMC. Mechanistically, expression of P2Y2R inhibited Runx2 transcriptional activation of an osteocalcin promoter driven luciferase reporter gene. CONCLUSIONS: This study reveals a role for vascular P2Y2R as an inhibitor of arterial intimal calcification and provides a new mechanistic insight into the regulation of the osteoblastic trans-differentiation of SMC through P2Y2R-mediated Runx2 antagonism. Given that calcification of atherosclerotic lesions is a significant clinical problem, activating P2Y2R may be an effective therapeutic approach for treatment or prevention of vascular calcification.
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    TLR3 deficiency exacerbates the loss of epithelial barrier function during genital tract Chlamydia muridarum infection
    (PLOS, 2019-01-09) Kumar, Ramesh; Gong, Haoli; Liu, Luyao; Ramos-Solis, Nicole; Seye, Cheikh I.; Derbigny, Wilbert A.; Microbiology and Immunology, School of Medicine
    PROBLEM: Chlamydia trachomatis infections are often associated with acute syndromes including cervicitis, urethritis, and endometritis, which can lead to chronic sequelae such as pelvic inflammatory disease (PID), chronic pelvic pain, ectopic pregnancy, and tubal infertility. As epithelial cells are the primary cell type productively infected during genital tract Chlamydia infections, we investigated whether Chlamydia has any impact on the integrity of the host epithelial barrier as a possible mechanism to facilitate the dissemination of infection, and examined whether TLR3 function modulates its impact. METHOD OF STUDY: We used wild-type and TLR3-deficient murine oviduct epithelial (OE) cells to ascertain whether C. muridarum infection had any effect on the epithelial barrier integrity of these cells as measured by transepithelial resistance (TER) and cell permeability assays. We next assessed whether infection impacted the transcription and protein function of the cellular tight-junction (TJ) genes for claudins1-4, ZO-1, JAM1 and occludin via quantitative real-time PCR (qPCR) and western blot. RESULTS: qPCR, immunoblotting, transwell permeability assays, and TER studies show that Chlamydia compromises cellular TJ function throughout infection in murine OE cells and that TLR3 deficiency significantly exacerbates this effect. CONCLUSION: Our data show that TLR3 plays a role in modulating epithelial barrier function during Chlamydia infection of epithelial cells lining the genital tract. These findings propose a role for TLR3 signaling in maintaining the integrity of epithelial barrier function during genital tract Chlamydia infection, a function that we hypothesize is important in helping limit the chlamydial spread and subsequent genital tract pathology.