Janus Kinase 3 Deficiency Promotes Vascular Reendothelialization

2021 ◽  
Author(s):  
Yung-Chun Wang ◽  
Dunpeng Cai ◽  
Xiao-Bing Cui ◽  
Ya-Hui Chuang ◽  
William P. Fay ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Injury ◽  
Intimal Hyperplasia ◽  
Carotid Arteries ◽  
Muscle Cells ◽  
Janus Kinase ◽  
Balloon Injury ◽  
Janus Kinase 3

Objective: The objective of this study is to determine the role of JAK3 (Janus kinase 3) in reendothelialization after vascular injury. Methods and Results: By using mouse carotid artery wire injury and rat balloon injury model, we found that JAK3 regulates reendothelialization and endothelial cell proliferation after vascular injury. JAK3 and phospho-JAK3 levels were increased in neointimal smooth muscle cells in response to vascular injury in mice. JAK3 deficiency dramatically attenuated the injury-induced intimal hyperplasia in carotid arteries of both male and female mice. Importantly, JAK3 deficiency caused an increased rate of reendothelialization following mechanical injury. Likewise, knockdown of JAK3 in medial smooth muscle cells elicited an accelerated reendothelialization with reduced intimal hyperplasia following balloon injury in rat carotid arteries. Interestingly, knockdown of JAK3 restored the expression of smooth muscle cell contractile protein smooth muscle α-actin in injury-induced intimal smooth muscle cells while increased the proliferating endothelial cells in the intima area. Conclusions: Our results demonstrate a novel role of JAK3 in the regeneration of endothelium after vascular injury, which may provide a new strategy to enhance reendothelialization while suppressing neointimal formation for effective vascular repair from injury.

Abstract 169: Smad3 Drives Cross-Talk Between TGFß and Wnt/ß-Catenin Signaling Pathways in Smooth Muscle Cells

2014 ◽  
Vol 34 (suppl_1) ◽  
Author(s):  
Daniel M DiRenzo ◽  
Xu Dong Shi ◽  
Lian-Wang Guo ◽  
K Craig Kent
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Wnt Signaling ◽  
Vascular Injury ◽  
Cross Talk ◽  
Intimal Hyperplasia ◽  
Muscle Cells ◽  
Arterial Injury ◽  
Protein Levels ◽  
Qrt Pcr

Restenosis (neo-intimal hyperplasia) occurs in approximately 25-50% of patients undergoing arterial interventions, primarily due to the proliferation and migration of arterial smooth muscle cells (SMCs) into the peri-luminal area. Recently, Wnt/β-catenin signaling has been shown to promote SMC proliferation and enhance neo-intimal hyperplasia but its mechanism of activation is unclear. Interestingly, Wnt/β-catenin has been shown to be activated by TGFβ in mesenchymal stem cells and fibroblasts. We have shown that TGFβ and its downstream signaling protein, Smad3, are upregulated following vascular injury and that Smad3 overexpressing SMCs display enhanced proliferation, migration, and neo-intimal hyperplasia. These results led us to hypothesize that TGFβ, through Smad3, activates Wnt/β-catenin to regulate SMC behavior following arterial injury . In primary rat SMCs, TGFβ (5ng/mL) led to β-catenin activation and relocalization from the plasma membrane to the cytoplasm / nucleus within 24 hours. Furthermore, qRT-PCR results demonstrated that expression of Wnt11 (22 fold) and Wnt9a (3.9 fold) were significantly upregulated after 24 hours of TGFβ stimulation (p<0.05, n=3). In addition, 24 hours of TGFβ stimulation in SMCs overexpressing Smad3 (TGFβ/Smad3) further enhanced the gene expression of Wnt11 (>300 fold) and Wnt9a (14 fold) and also stimulated significant increases in Wnt2b (41 fold), Wnt5a (2.9 fold), and Wnt4 (3.2 fold) (p<0.05, n=3) as measured by qRT-PCR. Western blot results demonstrated that the combined TGFβ/Smad3 stimulation increased β-catenin protein levels, suggesting that TGFβ activates canonical Wnt signaling leading to stabilization of β-catenin protein. In normal rat carotid arteries, β-catenin protein was undetectable via immunohistochemistry but could be seen in SMCs of the vessel media at 3 days post-balloon angioplasty and in neo-intimal cells at 7 and 14 days. Smad3 was also expressed in neo-intimal cells at 7 and 14 days post-angioplasty suggesting that TGFβ, through Smad3, is responsible for Wnt/β-Catenin activation during vascular injury. In conclusion, this work describes a novel cross-talk in SMCs between TGFβ and Wnt signaling which may provide a viable target for future anti-restenotic treatments.

Function and role of voltage-gated sodium channel NaV1.7 expressed in aortic smooth muscle cells

2009 ◽  
Vol 296 (1) ◽  
pp. H211-H219 ◽  
Author(s):  
Kentaro Meguro ◽  
Haruko Iida ◽  
Haruhito Takano ◽  
Toshihiro Morita ◽  
Masataka Sata ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Cell Migration ◽  
Smooth Muscle Cells ◽  
Intimal Hyperplasia ◽  
Muscle Cells ◽  
Rt Pcr ◽  
Balloon Injury ◽  
Aortic Smooth Muscle Cells ◽  
Aortic Smooth Muscle ◽  
Voltage Gated

Voltage-gated Na+ channel currents ( INa) are expressed in several types of smooth muscle cells. The purpose of this study was to evaluate the expression of INa, its functional role, pathophysiology in cultured human (hASMCs) and rabbit aortic smooth muscle cells (rASMCs), and its association with vascular intimal hyperplasia. In whole cell voltage clamp, INa was observed at potential positive to −40 mV, was blocked by tetrodotoxin (TTX), and replacing extracellular Na+ with N-methyl-d-glucamine in cultured hASMCs. In contrast to native aorta, cultured hASMCs strongly expressed SCN9A encoding NaV1.7, as determined by quantitative RT-PCR. INa was abolished by the treatment with SCN9A small-interfering (si)RNA ( P < 0.01). TTX and SCN9A siRNA significantly inhibited cell migration ( P < 0.01, respectively) and horseradish peroxidase uptake ( P < 0.01, respectively). TTX also significantly reduced the secretion of matrix metalloproteinase-2 6 and 12 h after the treatment ( P < 0.01 and P < 0.05, respectively). However, neither TTX nor siRNA had any effect on cell proliferation. L-type Ca2+ channel current was recorded, and INa was not observed in freshly isolated rASMCs, whereas TTX-sensitive INa was recorded in cultured rASMCs. Quantitative RT-PCR and immunostaining for NaV1.7 revealed the prominent expression of SCN9A in cultured rASMCs and aorta 48 h after balloon injury but not in native aorta. In conclusion, these studies show that INa is expressed in cultured and diseased conditions but not in normal aorta. The NaV1.7 plays an important role in cell migration, endocytosis, and secretion. NaV1.7 is also expressed in aorta after balloon injury, suggesting a potential role for NaV1.7 in the progression of intimal hyperplasia.

scholarly journals Inflammation and vascular smooth muscle cell dedifferentiation following carotid artery ligation

2017 ◽  
Vol 49 (3) ◽  
pp. 115-126 ◽  
Author(s):  
B. Paul Herring ◽  
April M. Hoggatt ◽  
Sarah L. Griffith ◽  
Jeanette N. McClintick ◽  
Patricia J. Gallagher
Keyword(s):  
Smooth Muscle ◽  
Carotid Artery ◽  
Smooth Muscle Cells ◽  
Vascular Injury ◽  
Carotid Arteries ◽  
Noncoding Rnas ◽  
Muscle Cells ◽  
Artery Ligation ◽  
Carotid Artery Ligation ◽  
Reporter Mice

Following vascular injury medial smooth muscle cells dedifferentiate and migrate through the internal elastic lamina where they form a neointima. The goal of the current study was to identify changes in gene expression that occur before the development of neointima and are associated with the early response to injury. Vascular injury was induced in C57BL/6 mice and in Myh11-creER(T2) mTmG reporter mice by complete ligation of the left carotid artery. Reporter mice were used to visualize cellular changes in the injured vessels. Total RNA was isolated from control carotid arteries or from carotid arteries 3 days following ligation of C57BL/6 mice and analyzed by Affymetrix microarray and quantitative RT-PCR. This analysis revealed decreased expression of mRNAs encoding smooth muscle-specific contractile proteins that was accompanied by a marked increase in a host of mRNAs encoding inflammatory cytokines following injury. There was also marked decrease in molecules associated with BMP, Wnt, and Hedgehog signaling and an increase in those associated with B cell, T cell, and macrophage signaling. Expression of a number of noncoding RNAs were also altered following injury with microRNAs 143/145 being dramatically downregulated and microRNAs 1949 and 142 upregulated. Several long noncoding RNAs showed altered expression that mirrored the expression of their nearest coding genes. These data demonstrate that following carotid artery ligation an inflammatory cascade is initiated that is associated with the downregulation of coding and noncoding RNAs that are normally required to maintain smooth muscle cells in a differentiated state.

Dihydroartemisinin ameliorates balloon injury-induced neointimal formation through suppressing autophagy in vascular smooth muscle cells

2020 ◽  
Vol 0 (0) ◽  
Author(s):  
Xiaoyuan Wang ◽  
Junpeng Wu ◽  
Haiyang Zhang ◽  
Bei Sun ◽  
Renping Huang
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Carotid Arteries ◽  
Muscle Cells ◽  
Therapeutic Effects ◽  
Balloon Injury ◽  
Neointimal Formation ◽  
Phenotypic Transition

Abstract The present study was designed to investigate the therapeutic effects of injection of dihydroartemisinin (DHA) into the balloon-injured carotid arteries on balloon injury-induced neointimal formation and to explore whether autophagy is involved in the action of DHA. Percutaneous transluminal balloon angioplasty was performed in Sprague-Dawley rats to induce neointimal formation, immediately after which DHA (100 μmol/l×1 ml) and/or Rapamycin (1 mg/100 μl), were injected into the balloon-injured carotid arteries. After 14 d, the serum samples and carotid artery tissues were harvested for analysis. Rat aortic vascular smooth muscle cells (VSMCs) were pretreated with DMSO (vehicle), DHA (1, 10, and 100 μmol/l), or 3-methyladenine (3-MA; 10 mM) for 1 h and then stimulated with plateletderived growth factor-BB (PDGF-BB; 10 ng/ml) for another 24 h. Animal experiments showed that DHA attenuated the balloon injury-induced neointimal formation, inflammation and VSMC phenotypic transition by inhibiting the balloon injury-induced autophagy activation. In vitro results showed that DHA attenuated the PDGF-BB-induced VSMC phenotypic transition, proliferation, and migration by inhibiting the PDGF-BB-induced autophagy activation. Taken together, DHA ameliorates balloon injury-induced neointimal formation through suppressing autophagy. This study provides insights into the development of a drug-eluting stent using DHA.

scholarly journals Huotan Jiedu Tongluo Decoction Inhibits Balloon-Injury-Induced Carotid Artery Intimal Hyperplasia in the Rat through the PERK-eIF2α-ATF4 Pathway and Autophagy Mediation

2021 ◽  
Vol 2021 ◽  
pp. 1-18
Author(s):  
Tenghui Tian ◽  
Keying Yu ◽  
Miao Zhang ◽  
Xiao Shao ◽  
Liping Chang ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Carotid Artery ◽  
Smooth Muscle Cells ◽  
Intimal Hyperplasia ◽  
Muscle Cells ◽  
Nuclear Antigen ◽  
Signal Molecules ◽  
Pathological Feature ◽  
Balloon Injury ◽  
Related Factors

In-stent restenosis (ISR) is the main factor affecting the outcome of percutaneous coronary intervention (PCI), and its main pathological feature is neointimal hyperplasia. Huotan Jiedu Tongluo decoction (HTJDTLD) is an effective traditional Chinese medicine (TCM) prescription for the treatment of vascular stenosis diseases. However, the precise anti-ISR mechanism of HTJDTLD remains unclear. Here, we investigated whether HTJDTLD can inhibit the excessive activation of endoplasmic reticulum stress (ERS) and reduce the level of autophagy factors through regulating the PERK-eIF2α-ATF4 pathway, thereby inhibiting the proliferation of the intima of blood vessels damaged by balloon injury (BI) and preventing the occurrence of ISR. In this study, a 2F Fogarty balloon was used to establish a common carotid artery (CCA) BI model in male Sprague-Dawley rats. Then, HTJDTLD (16.33 g/kg/d) or atorvastatin (1.19 mg/kg/d) was administered by gavage. Four weeks later, hematoxylin-eosin (HE) and Masson staining of the injured CCA were performed to observe the histological changes in the CCA. Immunohistochemistry (IHC) was used to assess the proliferation and dedifferentiation of vascular smooth muscle cells (VSMCs) in the CCA. Western blotting and RT-PCR were used to measure the expression of ERS- and autophagy-related proteins and mRNAs in the CCA. The results indicated that HTJDTLD significantly alleviated BI-induced carotid artery intimal hyperplasia and fibrosis and reduced the neointimal area (NIA) and NIA/medial area (MA) ratio. In addition, HTJDTLD inhibited the proliferation and dedifferentiation of VSMCs, reduced the expression of proliferating cell nuclear antigen (PCNA), and increased the smooth-muscle-α-actin- (SMα-actin-) positive area. HTJDTLD also significantly reduced the expression of the ERS-related factors: GRP78, p-PERK/PERK, p-eIF2α/eIF2α, ATF4, and CHOP. In addition, the expression of the autophagy-related factors, Beclin1, LC3B, and ATG12, was significantly decreased. In addition, in vitro experiments showed that HTJDTLD inhibited the above-mentioned ERS signal molecules in human umbilical vein endothelial cells (HUVEC) and rat aortic smooth muscle cells (A7R5) induced by tunicamycin (TM) and played a crucial role in protecting cells from damage. HTJDTLD may be a very promising drug for the treatment of ISR.

Effect of Activated Protein Kinase C Beta Type Mediated Phosphorylation of Signal Transducer and Activator of Transcription 4 on Proliferation and Phenotypic Transformation of Vascular Smooth Muscle Cells After Vascular Injury Induced by Nano-SiO2

2020 ◽  
Vol 20 (12) ◽  
pp. 7385-7397
Author(s):  
Ping Huang ◽  
Xiaoye Zhang ◽  
Minhua Pan ◽  
Jianneng Deng ◽  
Zhihong Chen ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Signaling Pathways ◽  
Vascular Smooth Muscle Cells ◽  
Vascular Injury ◽  
Muscle Cells ◽  
Over Expression ◽  
Core Proteins

The excessive proliferation, endothelial migration, and phenotype transformation of vascular smooth muscle cells (VSMC) lead to increased extracellular matrix secretion, which induces vascular intimal hyperplasia, which is an important restenosis mechanism after vascular injury. In our study, we verified the cytotoxicity of SiO2 nanoparticles to VSMC. To explore the role of endothelial repairs and molecular mechanisms after vascular injuries, we sequenced the transcriptome of injured vessels in the carotid artery of mice. The results showed the differentially expressed genes in normal vascular tissues, and that vascular tissues were mainly enriched with NF-κB signaling pathways, chemokine signaling pathways and other biological functions, by the leukocyte activation and adhesion of the KEGG pathway in the immune response, and by DNA binding, DNA transcription regulatory region binding, and other molecular functions. Core proteins included PRKCB, STAT4, CCL5, and BCL-2. To verify the roles of these core proteins, RT-qPCR andWestern blot techniques were used to detect their transcription and translation levels, and HE staining was used to detect morphological changes in blood vessels. To further clarify the role of core proteins in VSMC, PRKCB over expression plasmids were constructed, and the RT-qPCR and Western blot techniques were again used to detect the expression of core proteins. The results showed that the levels of transcription and translation, and of PRKCB and STAT4 phosphorylation, increased significantly after vascular injury, and then noticeably decreased three days later- and that CCL5 and Bcl-2 expression trends were consistent with this. HE staining showed that when the vascular endothelium was damaged, smooth muscle cells proliferated significantly, and that the intima thickened three days after vascular injury. After over expression of PRKCB, the expression and activation of STAT4, CCL5, and Bcl-2 significantly increased, α-SMA and Vimentin were down-regulated, OPN was up-regulated, and VSMC activity was enhanced. From these results, it could be concluded that PRKCB is activated by vascular injury, and that over-activation of PRKCB promotes activation of STAT4 and the expression of CCL5 and BCL-2—which in turn leads to enhanced VSMC activity and transformation of its contraction phenotype to the secretion phenotype. We were also able to establish that the cytotoxicity of SiO2 nanoparticles to VSMC was positively correlated with dose and time.

Abstract 237: The Role of Lymphocyte Specific Protein-1 in the Phenotypic Switch of Smooth Muscle Cells After Arterial Injury

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Mirnal A Chaudhary ◽  
Go Urabe ◽  
Alex Hayden ◽  
Sarah Franco ◽  
Xudong Shi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Western Blotting ◽  
Vascular Injury ◽  
Muscle Cells ◽  
Specific Protein ◽  
Phenotypic Switch

Background: After vascular injury, vascular smooth muscle cells (SMCs) switch from a differentiated contractile state to synthetic de-differentiated phenotype which contributes to the pathophysiology of restenosis. Experimental data generated by our lab indicate that TGF-β downregulates contractile proteins and stimulates migration. To understand how TGF-β promotes SMC phenotypic switch in injured arteries, we performed an Affymetrix Array analysis and identified Lymphocyte Specific Protein-1 (LSP1) among other upregulated genes. LSP1 is known to play a role in neutrophil extravasation, however the role of LSP1 within SMCs is unknown. We hypothesize that LSP1 contributes to SMC pathophysiological behavior through changes in cell architecture and migration in-vivo and in-vitro. Methods and Results: After carotid artery angioplasty, male Sprague-Dawley rats were sacrificed at 3, 7, and 14 days after injury for immunohistochemistry. Immunofluorescence staining revealed a unique upregulation of LSP1 within the neointima, media, and adventitia at 7 and 14 days, but not at 3 days after injury. Confocal images revealed that the LSP1 positive cells minimally express α-SMA (Pierson’s Coefficient, r=.017). Additional characterization experiments using immune cell markers CD3 and CD45 show no co-localization with LSP1 positive cells. To mimic the in-vivo neointimal cells and vascular injury induced de-differentiation in-vitro , rat A10 cells were treated with solvent or PDGF-bb (10 ng/mL). Quantitative RT-PCR demonstrated an upregulation of LSP1 mRNA after 24 hrs of PDGF-BB stimulation. Using Western Blotting, we confirm an upregulation of LSP1 protein after 48 hrs of PDGF-BB stimulation. Lastly, we performed nuclear and cytoplasmic fractionation followed by Western Blotting which demonstrated that LSP1 is remained within cytoplasmic fraction of the A10 cell after treatment with PDGF-BB. Conclusion: These results demonstrate that LSP1 is increased in-vivo after balloon injury, and in-vitro after PDGF-BB stimulation. Experiments to characterize the identity of these LSP1 cells in-vivo are in process, with future in-vitro experiments to focus on the role of LSP1 phosphorylation as a part of cytoskeletal remodeling and cellular migration.

The role of WNT/b-catenin signaling in smooth muscle cells during lung development and repair

Pneumologie ◽  
2014 ◽  
Vol 68 (06) ◽  
Author(s):  
A Moiseenko ◽  
E El Agha ◽  
B MacKenzie ◽  
S De Langhe ◽  
S Bellusci
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Lung Development ◽  
Muscle Cells
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