TNF-α induces a decrease in eNOS promoter activity

Abstract

We determined whether TNF-alpha induces a decrease in activity of the promoter for the endothelial nitric oxide synthase (eNOS) gene in pulmonary microvessel endothelial monolayers (PMEM). eNOS promoter activity was assessed in PMEM transfected with plasmids coding the wild-type (F1: -1600 nt from transcription start site) and truncated (F2: -1189, F4: -779, F5: -494, F6: -166) human eNOS promoters linked to a luciferase reporter. PMEM lysates were analyzed for the luciferase/galactosidase ratio (Luc/Gal) after incubation with TNF-alpha (50 ng/ml) for 0.5 or 4 h. TNF-alpha caused a decrease in the Luc/Gal ratio in the PMEM transfected with wild-type F1 and truncated F2, F4, and F5 plasmids but not with truncated F6 plasmid. Truncated-promoter analysis indicated the response elements (-370)CACCC, (-231)GATA, and (-186)CACCC may regulate the effect of TNF-alpha on the eNOS promoter. DNA-binding activity of (32)P-labeled oligonucleotide probes that span the GATA-binding site ((-239)-[(-231)GATA]-(-219)) and the two different CACCC-binding regions ((-379)-[(-370)CACCC]-(-358) and (-196)-[(-186) CACCC]-(-176)) were assessed using EMSA. In response to TNF-alpha treatment for 4 h, nuclear protein binding to (32)P oligonucleotides was characterized as: 1) a significant increase in binding to (-370)CACCC, 2) a significant decrease in binding to (-231)GATA, and 3) no change in (-186)CACCC binding. EMSA supershift analysis indicated that the transcription factor protein GATA-4 bound to the (-231)GATA site, and Sp3 bound to the (-370)CACCC site. Our data indicate TNF causes a decrease in eNOS promoter activity that may be mediated by GATA-4 and Sp3.

Endothelial Functionality and Echocardiographic Parameters in Patients with Rhinitis and Nasal Polyps

“…The vascular endothelium is a target of TNF-α. At the cellular level, TNF-α induces the expression of genes associated with inflammation, coagulation, and the proliferation and reduces the expression of the protein e-NOS and [31]. In addition, TNF-α induces the CAM expression on the surface of vascular endothelial cells in contrast to NO, also known to be an inhibitor of CAM expression [32].…”

ADIPONEKTIN MEREGULASI TURUN EKSPRESI iNOS PADA KULTUR HUVEC’s YANG DIPAPAR LIPO POLYSACCHARIDE (LPS)

“…Penurunan availabilitas Endothelial Nitrit Oxide (eNO) pada dinding vaskuler menyebabkan disfungsi endotel, dan endotel yang mengalami disfunsi mengekspresikan fenotip proinflamatori yang ditunjukkan dengan peningkatan ekspresi molekul adhesi sel aterogenik yaitu Cell Adhesion Molecular (CAM) sebagai akibat hilangnya eNO. Upregulasi CAM pada permukaan sel endotel menginisiasi interaksi patologis leukosit-endotel, yang mangakibatkan paparan dinding vaskuler dan jaringan sekitarnya pada aksi pengrusakan oleh leukosit (6,(11)(12)(13)(14)(15)(16)(17)(18)(19).…”

Possibility of the regression of atherosclerosis through the prevention of endothelial senescence by the regulation of nitric oxide and free radical scavengers

“…As the eNOS promoter has multiple potential cisregulatory DNA sequences, including SP1, AP1 and AP2, p53 binding regions and shear stress (or estrogen) response elements, hydrogen peroxide, protein kinase C (PKC) and estrogen can increase eNOS transcriptional regulation, and tumor necrosis factor (TNF)-a can decrease it. [5][6][7] Endothelial NO synthase mRNA has a long half-life at baseline (10-35 h), and post-transcriptional regulation may be more important ( 8 Searles CD Transcriptional and posttranscriptional regulation of endothelial nitric oxide synthase expression. Am J Physiol Cell Physiol.…”