Abstract 710: A Novel Mechanism by Which CPI-17 Is Down-regulated in Cultured Vascular Smooth Muscle Cells and Tissues by TNFα and in C57BL/6 Mice by Lipopolysaccharide

2015 ◽  
Vol 35 (suppl_1) ◽  
Author(s):  
Guogang Zhao ◽  
Shu Liu ◽  
Wen Su ◽  
Ming C Gong ◽  
Zhenheng Guo
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Promoter Activity ◽  
Muscle Cells ◽  
Mobility Shift ◽  
Down Regulation ◽  
Promoter Deletion Analysis ◽  
Gc Box

Objectives: Sepsis is characterized by a severe inflammatory response to infection, and its complications, including hypotension, can be fatal. It has long believed that hypotension in sepsis occur as a result of failure of the vascular smooth muscle cells (VSMC) to constrict. However, the molecular mechanism that links inflammation and decreases in vasoconstriction is largely unknown. Approaches and Results: CPI-17, a key regulator in vasoconstriction, was markedly decreased in mesentery arteries (MA) in mice injected with lipopolysaccharide (LPS). Incubation of cultured MA or VSMC with TNFα had similar effects on CPI-17 as LPS injection in mice. To identify transcriptional factors that respond to TNFα to suppress CPI-17 expression, we cloned a 1 kb murine CPI-17 promoter (-1,015 to +2 bp). Promoter deletion analysis revealed that a CPI-17 promoter region (-115 to -50 bp) was critical for TNFα to inhibit CPI-17 promoter activity. Bioinformatics analysis revealed this region contained 3 GC boxes that are putative binding sites for transcription factors SP1 and KLF4. Mutation of the 2nd and 3rd GC boxes, but not 1st GC box, abolished TNFα-mediated inhibitory effect on CPI-17 promoter activity. Electrophoretic mobility shift assay (EMSA) and chromatin immunoprecipitation (ChIP) illustrated that SP1 and KLF4 bound to the 2nd and 3rd GC boxes, but not 1st GC box. Interestingly, TNFα had no effect on SP1 expression but suppressed SP1 binding to the CPI-17 promoter. In contrast, TNFα increased KLF4 expression and KLF4 binding to the CPI-17 promoter. Down-regulation of either SP1 or KLF4 by siRNA abolished inhibition of CPI-17 promoter activity by TNFα but the effects were opposite: SP1 siRNA decreased CPI-17 promoter activity whereas KLF4 siRNA increased CPI-17 promoter activity. Moreover, KLF4 up-regulation, but not SP1, was also found in MA in mice injected with LPS. KLF4 up-regulation by LPS preceded CPI-17 down-regulation and correlated with a decrease in vasoconstriction and hypotension. Conclusions: These studies reveal a novel mechanism by which CPI-17 is down-regulated by LPS/TNFα through SP1 and KLF4 in vascular wall and identify SP1, KLF4, and CPI-17 as new potential therapeutic targets for treatment of hypotension in patients with sepsis.

Down-regulation of Endothelin Binding Sites in Rat Vascular Smooth Muscle Cells

1990 ◽  
Vol 3 (4) ◽  
pp. 310-312 ◽  
Author(s):  
P. Roubert ◽  
V. Gillard ◽  
P. Plus ◽  
P. E. Chabrier ◽  
P. Braquet
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Binding Sites ◽  
Muscle Cells ◽  
Down Regulation

scholarly journals Binding and receptor down-regulation of a novel vasoconstrictor endothelin in cultured rat vascular smooth muscle cells

FEBS Letters ◽  
1988 ◽  
Vol 239 (1) ◽  
pp. 13-17 ◽  
Author(s):  
Yukio Hirata ◽  
Hiroki Yoshimi ◽  
Shigeko Takaichi ◽  
Masashi Yanagisawa ◽  
Tomoh Masaki
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Down Regulation

Abstract 667: Angiotensin II-induced Osteopontin Expression in Vascular Smooth Muscle Cells involves G q/11 , Ras, ERK, Src and Ets-1.

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Keiko Abe ◽  
Mari Ishida ◽  
Mariko Sawano ◽  
Hidekatsu Nakashima ◽  
Nwe Nwe Soe ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Angiotensin Ii ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Promoter Activity ◽  
Critical Role ◽  
Muscle Cells ◽  
Separate Experiment ◽  
Type I

Background : Osteopontin (OPN), an extracellular matrix component produced by vascular smooth muscle cells (VSMC) and monocytes in response to biological stressors, is a regulator of cellular proliferation and migration. Recent studies revealed that OPN also plays a critical role in the progression of atherosclerotic plaques and that angiotensin II (AngII) is a potent upregulator of OPN expression. Therefore, the goal of the present study was to characterize the signaling mechanisms whereby AngII increases OPN expression. Methods and Results : YM254890, a specific inhibitor of G q/11 , potently suppressed AngII-induced OPN expression and ERK1/2 activation. Among DN mutants of small G proteins (Ras, Rac, Rho), only DN-Ras completely suppressed AngII-induced OPN promoter activity. Dominant negative (DN)-MEK1 inhibited AngII-induced OPN promoter activity by 54%, while DN-c-Jun N-terminal kinase (JNK) or DN-p38MAP kinase had no effect. DN-Src and Csk suppressed AngII-induced OPN promoter activity by 49% and 71%, respectively. In addition, small interfering RNA against Ets-1 (a transcriptional factor downstream of ERK1/2) suppressed AngII-induced OPN expression by 54 ± 4.3%. In a separate experiment, rats were treated with the AngII type I receptor blocker, valsartan (1 mg/kg/day), or vehicle for 2 weeks after carotid artery balloon injury. The intima/media ratio and OPN expression were significantly lower in valsartan-treated rats than in vehicle-treated rats. Conclusion : These data suggest that AngII-induced OPN expression in VSMC is mediated by signaling cascades involving G q/11 , the Ras-ERK axis, and c-Src, and by the transcription factor, Ets-1. Further, OPN may play a role in AngII-induced neointimal formation. These signaling molecules may represent therapeutic targets for the prevention of pathological vascular remodeling in patients with hypertension and atherosclerosis.

scholarly journals Transcriptional Regulation of S Phase Kinase-associated Protein 2 by NR4A Orphan Nuclear Receptor NOR1 in Vascular Smooth Muscle Cells

2011 ◽  
Vol 286 (41) ◽  
pp. 35485-35493 ◽  
Author(s):  
Florence Gizard ◽  
Yue Zhao ◽  
Hannes M. Findeisen ◽  
Hua Qing ◽  
Dianne Cohn ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Muscle Cells ◽  
Nuclear Hormone Receptor ◽  
Orphan Nuclear Receptor ◽  
Mobility Shift ◽  
Vsmc Proliferation

Members of the NR4A subgroup of the nuclear hormone receptor superfamily have emerged as key transcriptional regulators of proliferation and inflammation. NOR1 constitutes a ligand-independent transcription factor of this subgroup and induces cell proliferation; however, the transcriptional mechanisms underlying this mitogenic role remain to be defined. Here, we demonstrate that the F-box protein SKP2 (S phase kinase-associated protein 2), the substrate-specific receptor of the ubiquitin ligase responsible for the degradation of p27KIP1 through the proteasome pathway, constitutes a direct transcriptional target for NOR1. Mitogen-induced Skp2 expression is silenced in vascular smooth muscle cells (VSMC) isolated from Nor1-deficient mice or transfected with Nor1 siRNA. Conversely, adenovirus-mediated overexpression of NOR1 induces Skp2 expression in VSMC and decreases protein abundance of its target p27. Transient transfection experiments establish that NOR1 transactivates the Skp2 promoter through a nerve growth factor-induced clone B response element (NBRE). Electrophoretic mobility shift and chromatin immunoprecipitation assays further revealed that NOR1 is recruited to this NBRE site in the Skp2 promoter in response to mitogenic stimulation. In vivo Skp2 expression is increased during the proliferative response underlying neointima formation, and this transcriptional induction depends on the expression of NOR1. Finally, we demonstrate that overexpression of Skp2 rescues the proliferative arrest of Nor1-deficient VSMC. Collectively, these results characterize Skp2 as a novel NOR1-regulated target gene and detail a previously unrecognized transcriptional cascade regulating mitogen-induced VSMC proliferation.

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