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Qian Y, Xiong S, Li L, Sun Z, Zhang L, Yuan W, Cai H, Feng G, Wang X, Yao H, Gao Y, Guo L, Wang Z. Spatial multiomics atlas reveals smooth muscle phenotypic transformation and metabolic reprogramming in diabetic macroangiopathy. Cardiovasc Diabetol 2024; 23:358. [PMID: 39395983 PMCID: PMC11471023 DOI: 10.1186/s12933-024-02458-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/17/2024] [Accepted: 09/25/2024] [Indexed: 10/14/2024] Open
Abstract
BACKGROUND Diabetic macroangiopathy has been the main cause of death and disability in diabetic patients. The mechanisms underlying smooth muscle cell transformation and metabolic reprogramming other than abnormal glucose and lipid metabolism remain to be further explored. METHOD Single-cell transcriptome, spatial transcriptome and spatial metabolome sequencing were performed on anterior tibial artery from 11 diabetic patients with amputation. Multi-omics integration, cell communication analysis, time series analysis, network analysis, enrichment analysis, and gene expression analysis were performed to elucidate the potential molecular features. RESULT We constructed a spatial multiomics map of diabetic blood vessels based on multiomics integration, indicating single-cell and spatial landscape of transcriptome and spatial landscape of metabolome. At the same time, the characteristics of cell composition and biological function of calcified regions were obtained by integrating spatial omics and single cell omics. On this basis, our study provides favorable evidence for the cellular fate of smooth muscle cells, which can be transformed into pro-inflammatory chemotactic smooth muscle cells, macrophage-like smooth muscle cells/foam-like smooth muscle cells, and fibroblast/chondroblast smooth muscle cells in the anterior tibial artery of diabetic patients. The smooth muscle cell phenotypic transformation is driven by transcription factors net including KDM5B, DDIT3, etc. In addition, in order to focus on metabolic reprogramming apart from abnormal glucose and lipid metabolism, we constructed a metabolic network of diabetic vascular activation, and found that HNMT and CYP27A1 participate in diabetic vascular metabolic reprogramming by combining public data. CONCLUSION This study constructs the spatial gene-metabolism map of the whole anterior tibial artery for the first time and reveals the characteristics of vascular calcification, the phenotypic transformation trend of SMCs, and the transcriptional driving network of SMCs phenotypic transformation of diabetic macrovascular disease. In the perspective of combining the transcriptome and metabolome, the study demonstrates the activated metabolic pathways in diabetic blood vessels and the key genes involved in diabetic metabolic reprogramming.
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Affiliation(s)
- Yongjiang Qian
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China
| | - Shizheng Xiong
- State Key Laboratory of Organic Electronics and Information Displays and Institute of Advanced Materials (IAM), Nanjing University of Posts and Telecommunications, Nanjing, 210023, China
| | - Lihua Li
- Department of Pathology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
| | - Zhen Sun
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China
| | - Lili Zhang
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China
| | - Wei Yuan
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China
| | - Honghua Cai
- Department of Burn and Plastic Surgery, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
| | - Guoquan Feng
- Department of Radiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
| | - Xiaoguang Wang
- Department of Joint Surgery, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
| | - Haipeng Yao
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China
| | - Yun Gao
- Department of Pathology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China
| | - Li Guo
- State Key Laboratory of Organic Electronics and Information Displays and Institute of Advanced Materials (IAM), Nanjing University of Posts and Telecommunications, Nanjing, 210023, China.
| | - Zhongqun Wang
- Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, 212001, China.
- Institue of Cardiovascular Diseases, Jiangsu University, Zhenjiang, 212001, China.
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López-Acosta O, Ruiz-Ramírez A, Barrios-Maya MÁ, Alarcon-Aguilar J, Alarcon-Enos J, Céspedes Acuña CL, El-Hafidi M. Lipotoxicity, glucotoxicity and some strategies to protect vascular smooth muscle cell against proliferative phenotype in metabolic syndrome. Food Chem Toxicol 2023; 172:113546. [PMID: 36513245 DOI: 10.1016/j.fct.2022.113546] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2022] [Revised: 11/16/2022] [Accepted: 11/29/2022] [Indexed: 12/14/2022]
Abstract
Metabolic syndrome (MetS) is a risk factor for the development of cardiovascular disease (CVD) and atherosclerosis through a mechanism that involves vascular smooth muscle cell (VSMC) proliferation, lipotoxicity and glucotoxicity. Several molecules found to be increased in MetS, including free fatty acids, fatty acid binding protein 4, leptin, resistin, oxidized lipoprotein particles, and advanced glycation end products, influence VSMC proliferation. Most of these molecules act through their receptors on VSMCs by activating several signaling pathways associated with ROS generation in various cellular compartments. ROS from NADPH-oxidase and mitochondria have been found to promote VSMC proliferation and cell cycle progression. In addition, most of the natural or synthetic substances described in this review, including pharmaceuticals with hypoglycemic and hypolipidemic properties, attenuate VSMC proliferation by their simultaneous modulation of cell signaling and their scavenging property due to the presence of a phenolic ring in their structure. This review discusses recent data in the literature on the role that several MetS-related molecules and ROS play in the change from contractile to proliferative phenotype of VSMCs. Hence the importance of proposing an appropriate strategy to prevent uncontrolled VSMC proliferation using antioxidants, hypoglycemic and hypolipidemic agents.
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Affiliation(s)
- Ocarol López-Acosta
- Depto de Biomedicina Cardiovascular, Instituto Nacional de Cardiología Ignacio Chávez, Juan Badiano No 1, Colonia Sección XVI, Tlalpan, 14080, México D.F., Mexico
| | - Angélica Ruiz-Ramírez
- Depto de Biomedicina Cardiovascular, Instituto Nacional de Cardiología Ignacio Chávez, Juan Badiano No 1, Colonia Sección XVI, Tlalpan, 14080, México D.F., Mexico
| | - Miguel-Ángel Barrios-Maya
- Depto de Biomedicina Cardiovascular, Instituto Nacional de Cardiología Ignacio Chávez, Juan Badiano No 1, Colonia Sección XVI, Tlalpan, 14080, México D.F., Mexico
| | - Javier Alarcon-Aguilar
- Laboratorio de Farmacología, Depto. de Ciencias Biológicas y de la Salud, Universidad Autónoma Metropolitana Unidad Iztapalapa, Iztapalapa, Mexico
| | - Julio Alarcon-Enos
- Departamento de Ciencias Básicas, Facultad de Ciencias, Universidad del Bio Bio, Av. Andres Bello 720, Chillan, Chile
| | - Carlos L Céspedes Acuña
- Departamento de Ciencias Básicas, Facultad de Ciencias, Universidad del Bio Bio, Av. Andres Bello 720, Chillan, Chile.
| | - Mohammed El-Hafidi
- Depto de Biomedicina Cardiovascular, Instituto Nacional de Cardiología Ignacio Chávez, Juan Badiano No 1, Colonia Sección XVI, Tlalpan, 14080, México D.F., Mexico.
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Kitagawa A, Jacob C, Gupte SA. Glucose-6-phosphate dehydrogenase and MEG3 controls hypoxia-induced expression of serum response factor (SRF) and SRF-dependent genes in pulmonary smooth muscle cell. J Smooth Muscle Res 2022; 58:34-49. [PMID: 35491127 PMCID: PMC9057900 DOI: 10.1540/jsmr.58.34] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
Although hypoxia induces aberrant gene expression and dedifferentiation of smooth muscle cells (SMCs), mechanisms that alter dedifferentiation gene expression by hypoxia remain unclear. Therefore, we aimed to gain insight into the hypoxia-controlled gene expression in SMCs. We conducted studies using SMCs cultured in 3% oxygen (hypoxia) and the lungs of mice exposed to 10% oxygen (hypoxia). Our results suggest hypoxia upregulated expression of transcription factor CP2-like protein1, krüppel-like factor 4, and E2f transcription factor 1 enriched genes including basonuclin 2 (Bcn2), serum response factor (Srf), polycomb 3 (Cbx8), homeobox D9 (Hoxd9), lysine demethylase 1A (Kdm1a), etc. Additionally, we found that silencing glucose-6-phosphate dehydrogenase (G6PD) expression and inhibiting G6PD activity downregulated Srf transcript and hypomethylation of SMC genes (Myocd, Myh11, and Cnn1) and concomitantly increased their expression in the lungs of hypoxic mice. Furthermore, G6PD inhibition hypomethylated MEG3, a long non-coding RNA, gene and upregulated MEG3 expression in the lungs of hypoxic mice and in hypoxic SMCs. Silencing MEG3 expression in SMC mitigated the hypoxia-induced transcription of SRF. These findings collectively demonstrate that MEG3 and G6PD codependently regulate Srf expression in hypoxic SMCs. Moreover, G6PD inhibition upregulated SRF-MYOCD-driven gene expression, determinant of a differentiated SMC phenotype.
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Affiliation(s)
- Atsushi Kitagawa
- Department of Pharmacology, New York Medical College, BSB 546, 15 Dana Road, Valhalla, NY 10595, USA
| | - Christina Jacob
- Department of Pharmacology, New York Medical College, BSB 546, 15 Dana Road, Valhalla, NY 10595, USA
| | - Sachin A Gupte
- Department of Pharmacology, New York Medical College, BSB 546, 15 Dana Road, Valhalla, NY 10595, USA
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Kitagawa A, Jacob C, Jordan A, Waddell I, McMurtry IF, Gupte SA. Inhibition of Glucose-6-Phosphate Dehydrogenase Activity Attenuates Right Ventricle Pressure and Hypertrophy Elicited by VEGFR Inhibitor + Hypoxia. J Pharmacol Exp Ther 2021; 377:284-292. [PMID: 33758056 PMCID: PMC11047074 DOI: 10.1124/jpet.120.000166] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2020] [Accepted: 02/16/2021] [Indexed: 12/30/2022] Open
Abstract
Pulmonary hypertension (PH) is a disease of hyperplasia of pulmonary vascular cells. The pentose phosphate pathway (PPP)-a fundamental glucose metabolism pathway-is vital for cell growth. Because treatment of PH is inadequate, our goal was to determine whether inhibition of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP, prevents maladaptive gene expression that promotes smooth muscle cell (SMC) growth, reduces pulmonary artery remodeling, and normalizes hemodynamics in experimental models of PH. PH was induced in mice by exposure to 10% oxygen (Hx) or weekly injection of vascular endothelial growth factor receptor blocker [Sugen5416 (SU); 20 mg kg-1] during exposure to hypoxia (Hx + SU). A novel G6PD inhibitor (N-[(3β,5α)-17-oxoandrostan-3-yl]sulfamide; 1.5 mg kg-1) was injected daily during exposure to Hx. We measured right ventricle (RV) pressure and left ventricle pressure-volume relationships and gene expression in lungs of normoxic, Hx, and Hx + SU and G6PD inhibitor-treated mice. RV systolic and end-diastolic pressures were higher in Hx and Hx + SU than normoxic control mice. Hx and Hx + SU decreased expression of epigenetic modifiers (writers and erasers), increased hypomethylation of the DNA, and induced aberrant gene expression in lungs. G6PD inhibition decreased maladaptive expression of genes and SMC growth, reduced pulmonary vascular remodeling, and decreased right ventricle pressures compared with untreated PH groups. Pharmacologic inhibition of G6PD activity, by normalizing activity of epigenetic modifiers and DNA methylation, efficaciously reduces RV pressure overload in Hx and Hx + SU mice and preclinical models of PH and appears to be a safe pharmacotherapeutic strategy. SIGNIFICANCE STATEMENT: The results of this study demonstrated that inhibition of a metabolic enzyme efficaciously reduces pulmonary hypertension. For the first time, this study shows that a novel inhibitor of glucose-6-phosphate dehydrogenase, the rate-limiting enzyme in the fundamental pentose phosphate pathway, modulates DNA methylation and alleviates pulmonary artery remodeling and dilates pulmonary artery to reduce pulmonary hypertension.
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Affiliation(s)
- Atsushi Kitagawa
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
| | - Christina Jacob
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
| | - Allan Jordan
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
| | - Ian Waddell
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
| | - Ivan F McMurtry
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
| | - Sachin A Gupte
- Department of Pharmacology, New York Medical College, Valhalla, New York (A.K., C.J., S.A.G.); Drug Discovery Unit, Cancer Research UK Manchester Institute, University of Manchester, Macclesfield, United Kingdom (A.J., I.W.); and Departments of Pharmacology and Internal Medicine and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama (I.F.M.)
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Zhao M, Wang W, Lu Y, Wang N, Kong D, Shan L. MicroRNA‑153 attenuates hypoxia‑induced excessive proliferation and migration of pulmonary arterial smooth muscle cells by targeting ROCK1 and NFATc3. Mol Med Rep 2021; 23:194. [PMID: 33495839 PMCID: PMC7809904 DOI: 10.3892/mmr.2021.11833] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/01/2020] [Accepted: 12/01/2020] [Indexed: 12/20/2022] Open
Abstract
The aim of the present study was to explore the effect of microRNA (miR)‑153 on the proliferation and migration of pulmonary artery smooth muscle cells (PASMCs) in a hypoxic condition by targeting ρ‑associated, coiled‑coil‑containing protein kinase 1 (ROCK1) and nuclear factor of activated T cells cytoplasmic 3 (NFATc3). The right ventricular systolic pressure, right ventricular hypertrophy index, medial wall thickness and medial wall area were studied at different time‑points after rats were exposed to hypoxia. Western blot analysis was used to detect ROCK1 and NFATc3 protein levels. In addition, reverse transcription‑quantitative (RT‑q) PCR was performed to confirm the mRNA levels of miR‑153, ROCK1 and NFATc3 in human (H)PASMCs under hypoxic conditions. Transfected cells were then used to evaluate the effect of miR‑153 on cell proliferation and migration abilities. The association between miR‑153 and ROCK1 or NFATc3 was identified through double luciferase assays. Hypoxia induced pulmonary vascular remodeling and pulmonary arterial hypertension, which resulted from the abnormal proliferation of HPASMCs. ROCK1 and NFATc3 were the target genes of miR‑153 and miR‑153 mimic inhibited the protein expressions of ROCK1 and NFATc3 in HPASMCs and further inhibited cell proliferation and migration under hypoxic conditions. By contrast, the miR‑153 inhibitor promoted the proliferation and migration of HPASMCs. miR‑153 regulated the proliferation and migration of HPASMCs under hypoxia by targeting ROCK1 and NFATc3.
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Affiliation(s)
- Minjie Zhao
- Department of Respiratory Disease, The First Affiliated Hospital of Jinzhou Medical University, Jinzhou, Liaoning 121001, P.R. China
| | - Wei Wang
- Department of Respiratory Disease, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning 110000, P.R. China
| | - Ya Lu
- Department of Respiratory Disease, The First Affiliated Hospital of Jinzhou Medical University, Jinzhou, Liaoning 121001, P.R. China
| | - Nan Wang
- Department of Respiratory Disease, The First Affiliated Hospital of Jinzhou Medical University, Jinzhou, Liaoning 121001, P.R. China
| | - Delei Kong
- Department of Respiratory Disease, The First Affiliated Hospital of China Medical University, Shenyang, Liaoning 110000, P.R. China
| | - Lina Shan
- Department of Respiratory Disease, The First Affiliated Hospital of Jinzhou Medical University, Jinzhou, Liaoning 121001, P.R. China
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Hou J, Liu B, Zhu B, Wang D, Qiao Y, Luo E, Nawabi AQ, Yan G, Tang C. Role of integrin-linked kinase in the hypoxia-induced phenotypic transition of pulmonary artery smooth muscle cells: Implications for hypoxic pulmonary hypertension. Exp Cell Res 2019; 382:111476. [PMID: 31255599 DOI: 10.1016/j.yexcr.2019.06.021] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2019] [Revised: 06/13/2019] [Accepted: 06/19/2019] [Indexed: 11/21/2022]
Abstract
The phenotypic transition of pulmonary artery smooth muscle cells (PASMCs) from a contractile/differentiated to synthetic/de-differentiated phenotype is an important mechanism for the occurrence and development of hypoxic pulmonary hypertension (HPH). Integrin-linked kinase (ILK) is an early hypoxic response factor whose kinase activity is significantly affected during early hypoxia. Myocardin and ETS-like protein 1 (Elk-1) are co-activators of serum response factor (SRF) and can bind to SRF to mediate the phenotypic transition of PASMCs. However, little is known about the role of ILK on the phenotypic transition of these PASMCs. Thus, in our study, we explored the role of ILK in this process. We found that the expression of ILK and myocardin decreased gradually with the increase in hypoxia exposure time in the pulmonary arteries of rats. We observed that hypoxia exposure for 1 h caused an increase in the phosphorylation of Elk-1 but did not affect the expression of ILK, myocardin, or SRF. Exposure to hypoxic treatment for 1 h decreased ILK kinase activity and caused Elk-1 to suppress myocardin binding to SRF and the smooth muscle (SM) α-actin gene promoters. In addition, hypoxia exposure for 24 h decreased the expression of ILK, myocardin, SM α-actin, and calponin but increased the expression of osteopontin. Silencing of the myocardin gene significantly decreased the expression of SM α-actin and calponin but increased the expression of osteopontin. Silencing of the ILK gene significantly decreased the expression of myocardin, SM α-actin, and calponin but increased the expression of osteopontin. ILK overexpression reversed the effects of 24 h of hypoxia on the expression of myocardin, SM α-actin, calponin, and osteopontin and reversed the decrease in binding of myocardin to the SM α-actin promoter caused by 24 h of hypoxia exposure. Thus, our results suggest that ILK initiates the phenotypic transition of PASMCs. The underlying mechanism may involve hypoxia downregulating ILK kinase activity and protein expression, causing Elk-1 to compete with myocardin for binding to the SM α-actin promoter, which downregulates the expression of the downstream target myocardin and results in the phenotypic transition of PASMCs from a contractile to a synthetic phenotype. This may be an important mechanism in the development of HPH.
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MESH Headings
- Actins/genetics
- Actins/metabolism
- Animals
- Biomarkers/metabolism
- Calcium-Binding Proteins/metabolism
- Cell Hypoxia/genetics
- Cobalt/pharmacology
- Down-Regulation/genetics
- Hemodynamics/genetics
- Hypertension, Pulmonary/complications
- Hypertension, Pulmonary/enzymology
- Hypertension, Pulmonary/pathology
- Hypertension, Pulmonary/physiopathology
- Hypoxia/complications
- Hypoxia/enzymology
- Hypoxia/pathology
- Male
- Microfilament Proteins/metabolism
- Models, Biological
- Myocytes, Smooth Muscle/enzymology
- Myocytes, Smooth Muscle/pathology
- Nuclear Proteins/metabolism
- Osteopontin/metabolism
- Phenotype
- Phosphorylation
- Promoter Regions, Genetic/genetics
- Protein Binding
- Protein Serine-Threonine Kinases/metabolism
- Pulmonary Artery/pathology
- Pulmonary Artery/physiopathology
- RNA, Messenger/genetics
- RNA, Messenger/metabolism
- Rats, Sprague-Dawley
- Serum Response Factor/metabolism
- Trans-Activators/metabolism
- Vascular Remodeling/genetics
- ets-Domain Protein Elk-1/metabolism
- Calponins
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Affiliation(s)
- Jiantong Hou
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Bo Liu
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Boqian Zhu
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Dong Wang
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Yong Qiao
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Erfei Luo
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Abdul Qadir Nawabi
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China
| | - Gaoliang Yan
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China.
| | - Chengchun Tang
- Department of Cardiology, Zhongda Hospital of Southeast University Medical School, Nanjing, China.
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Li Q, Zhou X, Zhou X. Downregulation of miR‑98 contributes to hypoxic pulmonary hypertension by targeting ALK1. Mol Med Rep 2019; 20:2167-2176. [PMID: 31322216 PMCID: PMC6691262 DOI: 10.3892/mmr.2019.10482] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2018] [Accepted: 05/31/2019] [Indexed: 12/21/2022] Open
Abstract
Chronic hypoxia is one of the most common causes of secondary pulmonary hypertension, the mechanisms of which remain unclear. MicroRNAs (miRNAs) are small, noncoding RNAs that inhibit the translation or accelerate the degradation of mRNA. Previous studies have demonstrated that deregulated miRNA expression contributes to various cellular processes including cell apoptosis and proliferation, which are mediated by hypoxia. In the present study, the expression of miR‑98 was identified to be decreased in the lung tissue of a hypoxic pulmonary hypertension (HPH) rat model and pulmonary artery (PA) smooth muscle cells (PASMCs), which was induced by hypoxia. By transfecting miR‑98 mimics into PASMCs, the high expression of miR‑98 inhibited cell proliferation, but upregulated hypoxia‑induced PASMCs apoptosis. However, these effects of miR‑98 mimics on PASMCs were reversed by ALK1 (activin receptor‑like kinase‑1) overexpression. ALK1 was identified as a candidate target of miR‑98. In addition, overexpressing miR‑98 markedly decreased the pulmonary artery wall thickness and the right ventricular systolic pressure in rats induced by hypoxia. These results provided clear evidence that miR‑98 was a direct regulator of ALK1, and that the downregulation of miR‑98 contributed to the pathogenesis of HPH. These results provide a novel potential therapeutic strategy for the treatment of HPH.
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Affiliation(s)
- Qingling Li
- Department of Respiratory Medicine, Xuzhou Municipal Hospital Affiliated to Xuzhou Medical University, Xuzhou, Jiangsu 221002, P.R. China
| | - Xincan Zhou
- Department of Respiratory Medicine, Xuzhou Municipal Hospital Affiliated to Xuzhou Medical University, Xuzhou, Jiangsu 221002, P.R. China
| | - Xianghui Zhou
- Department of Respiratory Medicine, Xuzhou Municipal Hospital Affiliated to Xuzhou Medical University, Xuzhou, Jiangsu 221002, P.R. China
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Wang F, Xu X, Tang W, Min L, Yang J. Rab6A GTPase contributes to phenotypic modulation in pulmonary artery smooth muscle cells under hypoxia. J Cell Biochem 2019; 120:7858-7867. [PMID: 30417421 DOI: 10.1002/jcb.28060] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/13/2018] [Accepted: 10/22/2018] [Indexed: 01/24/2023]
Abstract
Previous studies have demonstrated that hypoxia can induce phenotypic modulation of pulmonary smooth muscle cells; however, the mechanisms remain unclear. The present study aimed to investigate the effect of the GTPase Rab6A-mediated phenotypic modulation and other activities of rat pulmonary artery smooth muscle cells (RPASMCs). We revealed that Rab6A was induced by hypoxia (1% O2 ) and was involved in a hypoxia-induced phenotypic switch and endoplasmic reticulum stress (ERS) in RPASMCs. After 48 hours of hypoxia, the expression of the phenotype marker protein smooth muscle actin was downregulated and vimentin (VIM) expression was upregulated. Rab6A was upregulated after 48 hours of hypoxia, and the level of glucose-regulated protein, 78 kDa (GRP78) after 12 hours of hypoxic stimulation was also increased. After transfection with a Rab6A short interfering RNA under hypoxic conditions, the expression levels of GRP78 and VIM in RPASMCs were downregulated. Overall, hypoxia-induced RPASMCs to undergo ERS followed by phenotypic transformation. Rab6A is involved in this hypoxia-induced phenotypic modulation and ERS in RPASMCs.
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Affiliation(s)
- Fang Wang
- Department of Respiratory Medicine, Northern Jiangsu People's Hospital, Clinical Medical College of Yangzhou University, Yangzhou, China
| | - Xingxiang Xu
- Department of Respiratory Medicine, Northern Jiangsu People's Hospital, Clinical Medical College of Yangzhou University, Yangzhou, China
| | - Weian Tang
- Department of Respiratory Medicine, Northern Jiangsu People's Hospital, Clinical Medical College of Yangzhou University, Yangzhou, China
| | - Lingfeng Min
- Department of Respiratory Medicine, Northern Jiangsu People's Hospital, Clinical Medical College of Yangzhou University, Yangzhou, China
| | - Junjun Yang
- Department of Respiratory Medicine, Northern Jiangsu People's Hospital, Clinical Medical College of Yangzhou University, Yangzhou, China
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Liu A, Liu Y, Li B, Yang M, Liu Y, Su J. Role of miR-223-3p in pulmonary arterial hypertension via targeting ITGB3 in the ECM pathway. Cell Prolif 2018; 52:e12550. [PMID: 30507047 PMCID: PMC6496671 DOI: 10.1111/cpr.12550] [Citation(s) in RCA: 36] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2018] [Revised: 10/08/2018] [Accepted: 10/10/2018] [Indexed: 12/17/2022] Open
Abstract
Objectives To investigate the functions of miR‐223‐3p and ITGB3 in pulmonary arterial hypertension (PAH). Materials and Methods Microarray analysis was used to detect differentially expressed genes and microRNAs. In in vitro models, the expressions of miR‐223‐3p and ITGB3 were detected by qRT‐PCR and Western blot. α‐SMA expression and cell proliferation were analysed by immunofluorescence and MTT assay, respectively. In in vivo models, PAH progressions were determined by measuring the levels of mPAP and RVSP. Lung and myocardial tissues were subjected to HE staining and Masson and Sirius red‐saturated carbazotic acid staining to investigate the pathological features. Results The microarray analysis revealed that ITGB3 was upregulated, while hsa‐miR‐223‐3p was downregulated in PAH. After the induction of hypoxia, miR‐223‐3p was downregulated and ITGB3 was upregulated in PASMCs. Hypoxia induction promoted cell proliferation and inhibited α‐SMA expression in PASMCs. Both the upregulation of miR‐223‐3p and the downregulation of ITGB3 attenuated the aberrant proliferation induced by hypoxia conditions. After approximately 4 weeks, the mPAP and RVSP levels of rats injected with MCT were decreased by the overexpression of miR‐223‐3p or the silencing of ITGB3. The staining results revealed that both miR‐223‐3p overexpression and ITGB3 knockdown alleviated the pulmonary vascular remodelling and improved the PAH pathological features of rats. Conclusions MiR‐223‐3p alleviated the progression of PAH by suppressing the expression of ITGB3, a finding which provides novel targets for clinical treatment.
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Affiliation(s)
- Aijun Liu
- Department of Pediatric Cardiac Surgery Center, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Yifan Liu
- Weifang Medicial University, Weifang, China
| | - Bin Li
- Department of Pediatric Cardiac Surgery Center, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Ming Yang
- Department of Pediatric Cardiac Surgery Center, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Yang Liu
- Department of Pediatric Cardiac Surgery Center, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
| | - Junwu Su
- Department of Pediatric Cardiac Surgery Center, Beijing Anzhen Hospital, Capital Medical University, Beijing, China
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10
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Raczkowski F, Rissiek A, Ricklefs I, Heiss K, Schumacher V, Wundenberg K, Haag F, Koch-Nolte F, Tolosa E, Mittrücker HW. CD39 is upregulated during activation of mouse and human T cells and attenuates the immune response to Listeria monocytogenes. PLoS One 2018; 13:e0197151. [PMID: 29742141 PMCID: PMC5942830 DOI: 10.1371/journal.pone.0197151] [Citation(s) in RCA: 47] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2017] [Accepted: 04/27/2018] [Indexed: 11/26/2022] Open
Abstract
The ectoenzymes CD39 and CD73 degrade extracellular ATP to adenosine. ATP is released by stressed or damaged cells and provides pro-inflammatory signals to immune cells through P2 receptors. Adenosine, on the other hand, suppresses immune cells by stimulating P1 receptors. Thus, CD39 and CD73 can shape the quality of immune responses. Here we demonstrate that upregulation of CD39 is a consistent feature of activated conventional CD4+ and CD8+ T cells. Following stimulation in vitro, CD4+ and CD8+ T cells from human blood gained surface expression of CD39 but displayed only low levels of CD73. Activated human T cells from inflamed joints largely presented with a CD39+CD73— phenotype. In line, in spleens of mice with acute Listeria monocytogenes, listeria-specific CD4+ and CD8+ T cells acquired a CD39+CD73— phenotype. To test the function of CD39 in control of bacterial infection, CD39-deficient (CD39-/-) mice were infected with L. monocytogenes. CD39-/- mice showed better initial control of L. monocytogenes, which was associated with enhanced production of inflammatory cytokines. In the late stage of infection, CD39-/- mice accumulated more listeria-specific CD8+ T cells in the spleen than wildtype animals suggesting that CD39 attenuates the CD8+ T-cell response to infection. In conclusion, our results demonstrate that CD39 is upregulated on conventional CD4+ and CD8+ T cells at sites of acute infection and inflammation, and that CD39 dampens responses to bacterial infection.
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Affiliation(s)
- Friederike Raczkowski
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Anne Rissiek
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Isabell Ricklefs
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Kirsten Heiss
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Valéa Schumacher
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Kira Wundenberg
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Friedrich Haag
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Friedrich Koch-Nolte
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Eva Tolosa
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
| | - Hans-Willi Mittrücker
- Institute for Immunology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
- * E-mail:
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11
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Zhang W, Zhu T, Wu W, Ge X, Xiong X, Zhang Z, Hu C. LOX-1 mediated phenotypic switching of pulmonary arterial smooth muscle cells contributes to hypoxic pulmonary hypertension. Eur J Pharmacol 2017; 818:84-95. [PMID: 29069578 DOI: 10.1016/j.ejphar.2017.10.037] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/28/2017] [Revised: 10/19/2017] [Accepted: 10/20/2017] [Indexed: 12/13/2022]
Abstract
In pulmonary hypertension (PH), pulmonary arterial smooth muscle cells (PASMCs) are dedifferentiated, undergoing a contractile-to-synthetic phenotypic switching. Lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) plays diverse roles in the cardiovascular system, but its contribution to PH remains to be fully defined. The present study was undertaken to explore the role of LOX-1 in PASMCs dedifferentiation in hypoxia-induced pulmonary vascular remodeling and PH. In a rat model of hypoxic PH, pulmonary vascular remodeling was accompanied by increased expression of LOX-1 in pulmonary arteries. In primary rat PASMCs, hypoxia-induced PASMCs dedifferentiation occurred concomitantly with LOX-1 upregulation. Inhibition of LOX-1 by either siRNA knockdown or neutralizing antibody significantly ameliorated PASMCs dedifferentiation. Mechanistically, LOX-1 promotes PASMCs dedifferentiation under hypoxic conditions via ERK1/2-Elk-1/MRTF-A/SRF signaling pathway. In conclusion, our data uncovers an important role of LOX-1 in the maintenance of PASMCs phenotype. Therapeutic targeting of LOX-1/ERK1/2-Elk-1/MRTF-A/SRF signaling axis would be exploited to treat hypoxic PH.
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Affiliation(s)
- Weifang Zhang
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China; Department of Pharmacy, the Second Affiliated Hospital of Nanchang University, Nanchang, Jiangxi 330006, China
| | - Tiantian Zhu
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China
| | - Weihua Wu
- School of pharmaceutical Sciences, Hunan University of Medicine, Huaihua, Hunan 418000, China
| | - Xiaoyue Ge
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China
| | - Xiaoming Xiong
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China; Hunan Provincial Key Laboratory of Cardiovascular Research, Central South University, Changsha, Hunan 410078, China
| | - Zheng Zhang
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China; Hunan Provincial Key Laboratory of Cardiovascular Research, Central South University, Changsha, Hunan 410078, China.
| | - Changping Hu
- Department of Pharmacology, Xiangya School of Pharmaceutical Sciences, Central South University, Changsha, Hunan 410078, China; Hunan Provincial Key Laboratory of Cardiovascular Research, Central South University, Changsha, Hunan 410078, China.
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12
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Chen D, Gao W, Wang S, Ni B, Gao Y. Critical effects of epigenetic regulation in pulmonary arterial hypertension. Cell Mol Life Sci 2017; 74:3789-3808. [PMID: 28573430 PMCID: PMC11107652 DOI: 10.1007/s00018-017-2551-8] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2017] [Revised: 05/14/2017] [Accepted: 05/29/2017] [Indexed: 12/11/2022]
Abstract
Pulmonary arterial hypertension (PAH) is characterized by persistent pulmonary vasoconstriction and pulmonary vascular remodeling. The pathogenic mechanisms of PAH remain to be fully clarified and measures of effective prevention are lacking. Recent studies; however, have indicated that epigenetic processes may exert pivotal influences on PAH pathogenesis. In this review, we summarize the latest research findings regarding epigenetic regulation in PAH, focusing on the roles of non-coding RNAs, histone modifications, ATP-dependent chromatin remodeling and DNA methylation, and discuss the potential of epigenetic-based therapies for PAH.
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Affiliation(s)
- Dewei Chen
- Department of Pathophysiology and High Altitude Pathology, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, People's Republic of China
- Key Laboratory of High Altitude Medicine of PLA, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, 400038, People's Republic of China
| | - Wenxiang Gao
- Department of Pathophysiology and High Altitude Pathology, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, People's Republic of China
- Key Laboratory of High Altitude Medicine of PLA, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, 400038, People's Republic of China
| | - Shouxian Wang
- Department of Pathophysiology and High Altitude Pathology, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, People's Republic of China
- Key Laboratory of High Altitude Medicine of PLA, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, 400038, People's Republic of China
| | - Bing Ni
- Department of Pathophysiology and High Altitude Pathology, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, People's Republic of China.
- Key Laboratory of High Altitude Medicine of PLA, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, 400038, People's Republic of China.
| | - Yuqi Gao
- Institute of Medicine and Hygienic Equipment for High Altitude Region, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, People's Republic of China.
- Key Laboratory of High Altitude Medicine of PLA, College of High Altitude Military Medicine, Third Military Medical University, Chongqing, 400038, People's Republic of China.
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13
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Yu H, Jia Q, Feng X, Chen H, Wang L, Ni X, Kong W. Hypoxia decrease expression of cartilage oligomeric matrix protein to promote phenotype switching of pulmonary arterial smooth muscle cells. Int J Biochem Cell Biol 2017; 91:37-44. [PMID: 28860005 DOI: 10.1016/j.biocel.2017.08.007] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/13/2017] [Revised: 07/30/2017] [Accepted: 08/10/2017] [Indexed: 02/03/2023]
Abstract
Extracellular matrix proteins play important roles in the development of pulmonary hypertension(pH). However, the role of Cartilage oligomeric matrix protein (COMP) in the development of hypoxia-induced pH is largely unknown. We tested the hypothesis that COMP deficiency induced by hypoxia leads to the phenotype switching of pulmonary arterial smooth muscle cells (PASMCs). The expression of COMP decreased in a chronic hypoxia rat pH model (P<0.05) and in PASMCs under hypoxia (3%O2) (P<0.05). The expressions of differentiated marker proteins reduced in the pulmonary arteries from 5 month old COMP-/- mice and in PASMCs under hypoxia or with the siRNA of COMP treatment under normoxia, but increased in PASMCs with adenovirus-increased COMP under hypoxia. The absorbance of cell counting kit-8 at 450nm and the expressions of proliferating cell nuclear antigen (PCNA) and osteopontin increased in PASMCs with the siRNA of COMP under normoxia (P<0.05). PCNA and osteopontin decreased in PASMCs with adenovirus-increased COMP under hypoxia (P<0.05). Additionally, the expression of bone morphogenetic protein receptor 2 (BMPR2) was reduced in COMP-/- mice (P<0.01). Both mRNA and protein levels of bone morphogenetic protein 2 (BMP2) were lower in PASMCs with the siRNA of COMP (P<0.05). The protein level of BMP2 could be reversed by adenovirus-increased COMP under hypoxia (P<0.05). These data suggest that COMP could normally have a protective role against PASMC phenotype switching and maintain BMP2/BMPR2 signaling, and these protective actions could be lost as a result of hypoxia promoting a depletion of COMP.
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Affiliation(s)
- Hang Yu
- Department of Physiology, Harbin Medical University-Daqing, Daqing, Heilongjiang Province, China.
| | - Qingbo Jia
- Department of Chest Surgery, the Fifth Affiliated Hospital of Harbin Medical University, Daqing, Heilongjiang Province, China.
| | - Xiaoqian Feng
- Department of Anatomy, Harbin Medical University-Daqing, Daqing, Heilongjiang Province, China.
| | - Hongxia Chen
- Department of Physiology and Pathophysiology, Harbin Medical University-Daqing, Daqing, Heilongjiang Province, China.
| | - Liang Wang
- Department of Anatomy, Harbin Medical University-Daqing, Daqing, Heilongjiang Province, China.
| | - Xiuqin Ni
- Department of Anatomy, Harbin Medical University-Daqing, Daqing, Heilongjiang Province, China.
| | - Wei Kong
- Department of Physiology and Pathophysiology, Basic Medical College of Peking University, Beijing, China.
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14
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Thorpe RB, Hubbell MC, Silpanisong J, Williams JM, Pearce WJ. Chronic hypoxia attenuates the vasodilator efficacy of protein kinase G in fetal and adult ovine cerebral arteries. Am J Physiol Heart Circ Physiol 2017; 313:H207-H219. [PMID: 28550175 DOI: 10.1152/ajpheart.00480.2016] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/06/2016] [Revised: 03/30/2017] [Accepted: 04/25/2017] [Indexed: 11/22/2022]
Abstract
Long-term hypoxia (LTH) attenuates nitric oxide-induced vasorelaxation in ovine middle cerebral arteries. Because cGMP-dependent protein kinase (PKG) is an important mediator of NO signaling in vascular smooth muscle, we tested the hypothesis that LTH diminishes the ability of PKG to interact with target proteins and cause vasorelaxation. Prominent among proteins that regulate vascular tone is the large-conductance Ca2+-sensitive K+ (BK) channel, which is a substrate for PKG and is responsive to phosphorylation on multiple serine/threonine residues. Given the influence of these proteins, we also examined whether LTH attenuates PKG and BK channel protein abundances and PKG activity. Middle cerebral arteries were harvested from normoxic and hypoxic (altitude of 3,820 m for 110 days) fetal and adult sheep. These arteries were denuded and equilibrated with 95% O2-5% CO2 in the presence of N-nitro-l-arginine methyl ester (l-NAME) to inhibit potential confounding influences of events upstream from PKG. Expression and activity of PKG-I were not significantly affected by chronic hypoxia in either fetal or adult arteries. Pretreatment with the BK inhibitor iberiotoxin attenuated vasorelaxation induced by 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphate in normoxic but not LTH arteries. The spatial proximities of PKG with BK channel α- and β1-proteins were examined using confocal microscopy, which revealed a strong dissociation of PKG with these proteins after LTH. These results support our hypothesis that hypoxia reduces the ability of PKG to attenuate vasoconstriction in part through suppression of the ability of PKG to associate with and thereby activate BK channels in arterial smooth muscle.NEW & NOTEWORTHY Using measurements of contractility, protein abundance, kinase activity, and confocal colocalization in fetal and adult ovine cerebral arteries, the present study demonstrates that long-term hypoxia diminishes the ability of cGMP-dependent protein kinase (PKG) to cause vasorelaxation through suppression of its colocalization and interaction with large-conductance Ca2+-sensitive K+ (BK) channel proteins in cerebrovascular smooth muscle. These experiments are among the first to demonstrate hypoxic changes in BK subunit abundances in fetal cerebral arteries and also introduce the use of advanced methods of confocal colocalization to study interaction between PKG and its targets.
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Affiliation(s)
- Richard B Thorpe
- Center for Perinatal Biology and Divisions of Physiology, Pharmacology, and Biochemistry, Loma Linda University School of Medicine, Loma Linda, California
| | - Margaret C Hubbell
- Center for Perinatal Biology and Divisions of Physiology, Pharmacology, and Biochemistry, Loma Linda University School of Medicine, Loma Linda, California
| | - Jinjutha Silpanisong
- Center for Perinatal Biology and Divisions of Physiology, Pharmacology, and Biochemistry, Loma Linda University School of Medicine, Loma Linda, California
| | - James M Williams
- Center for Perinatal Biology and Divisions of Physiology, Pharmacology, and Biochemistry, Loma Linda University School of Medicine, Loma Linda, California
| | - William J Pearce
- Center for Perinatal Biology and Divisions of Physiology, Pharmacology, and Biochemistry, Loma Linda University School of Medicine, Loma Linda, California
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15
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Chettimada S, Joshi SR, Dhagia V, Aiezza A, Lincoln TM, Gupte R, Miano JM, Gupte SA. Vascular smooth muscle cell contractile protein expression is increased through protein kinase G-dependent and -independent pathways by glucose-6-phosphate dehydrogenase inhibition and deficiency. Am J Physiol Heart Circ Physiol 2016; 311:H904-H912. [PMID: 27521420 PMCID: PMC5114469 DOI: 10.1152/ajpheart.00335.2016] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/02/2016] [Accepted: 07/06/2016] [Indexed: 11/22/2022]
Abstract
Homeostatic control of vascular smooth muscle cell (VSMC) differentiation is critical for contractile activity and regulation of blood flow. Recently, we reported that precontracted blood vessels are relaxed and the phenotype of VSMC is regulated from a synthetic to contractile state by glucose-6-phosphate dehydrogenase (G6PD) inhibition. In the current study, we investigated whether the increase in the expression of VSMC contractile proteins by inhibition and knockdown of G6PD is mediated through a protein kinase G (PKG)-dependent pathway and whether it regulates blood pressure. We found that the expression of VSMC-restricted contractile proteins, myocardin (MYOCD), and miR-1 and miR-143 are increased by G6PD inhibition or knockdown. Importantly, RNA-sequence analysis of aortic tissue from G6PD-deficient mice revealed uniform increases in VSMC-restricted genes, particularly those regulated by the MYOCD-serum response factor (SRF) switch. Conversely, expression of Krüppel-like factor 4 (KLF4) is decreased by G6PD inhibition. Interestingly, the G6PD inhibition-induced expression of miR-1 and contractile proteins was blocked by Rp-β-phenyl-1,N2-etheno-8-bromo-guanosine-3',5'-cyclic monophosphorothioate, a PKG inhibitor. On the other hand, MYOCD and miR-143 levels are increased by G6PD inhibition through a PKG-independent manner. Furthermore, blood pressure was lower in the G6PD-deficient compared with wild-type mice. Therefore, our results suggest that the expression of VSMC contractile proteins induced by G6PD inhibition occurs via PKG1α-dependent and -independent pathways.
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MESH Headings
- Animals
- Aorta/drug effects
- Aorta/metabolism
- Blotting, Western
- Cattle
- Chromatography, Liquid
- Contractile Proteins/drug effects
- Contractile Proteins/genetics
- Contractile Proteins/metabolism
- Cyclic GMP-Dependent Protein Kinase Type I/antagonists & inhibitors
- Cyclic GMP-Dependent Protein Kinase Type I/metabolism
- Cyclic GMP-Dependent Protein Kinases/antagonists & inhibitors
- Cyclic GMP-Dependent Protein Kinases/metabolism
- Gene Knockdown Techniques
- Glucosephosphate Dehydrogenase/antagonists & inhibitors
- Glucosephosphate Dehydrogenase/genetics
- Immunoprecipitation
- Kruppel-Like Factor 4
- Kruppel-Like Transcription Factors/drug effects
- Kruppel-Like Transcription Factors/genetics
- Kruppel-Like Transcription Factors/metabolism
- Mice
- MicroRNAs/drug effects
- MicroRNAs/genetics
- Muscle, Smooth, Vascular/cytology
- Muscle, Smooth, Vascular/drug effects
- Muscle, Smooth, Vascular/metabolism
- Myocytes, Smooth Muscle/drug effects
- Myocytes, Smooth Muscle/metabolism
- Nuclear Proteins/drug effects
- Nuclear Proteins/genetics
- Nuclear Proteins/metabolism
- Polymerase Chain Reaction
- Rats
- Serum Response Factor/drug effects
- Serum Response Factor/genetics
- Serum Response Factor/metabolism
- Tandem Mass Spectrometry
- Trans-Activators/drug effects
- Trans-Activators/genetics
- Trans-Activators/metabolism
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Affiliation(s)
- Sukrutha Chettimada
- Biochemistry & Molecular Biology, University of South Alabama, Mobile, Alabama; Pharmacology, New York Medical College, Valhalla, New York
| | - Sachindra Raj Joshi
- Biochemistry & Molecular Biology, University of South Alabama, Mobile, Alabama; Pharmacology, New York Medical College, Valhalla, New York
| | - Vidhi Dhagia
- Pharmacology, New York Medical College, Valhalla, New York
| | - Alessandro Aiezza
- Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, New York; and
| | | | - Rakhee Gupte
- Biochemistry & Molecular Biology, University of South Alabama, Mobile, Alabama; Pharmacology, New York Medical College, Valhalla, New York
| | - Joseph M Miano
- Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, New York; and
| | - Sachin A Gupte
- Biochemistry & Molecular Biology, University of South Alabama, Mobile, Alabama; Pharmacology, New York Medical College, Valhalla, New York
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16
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Gao Y, Chen T, Raj JU. Endothelial and Smooth Muscle Cell Interactions in the Pathobiology of Pulmonary Hypertension. Am J Respir Cell Mol Biol 2016; 54:451-60. [PMID: 26744837 DOI: 10.1165/rcmb.2015-0323tr] [Citation(s) in RCA: 95] [Impact Index Per Article: 10.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
In the pulmonary vasculature, the endothelial and smooth muscle cells are two key cell types that play a major role in the pathobiology of pulmonary vascular disease and pulmonary hypertension. The normal interactions between these two cell types are important for the homeostasis of the pulmonary circulation, and any aberrant interaction between them may lead to various disease states including pulmonary vascular remodeling and pulmonary hypertension. It is well recognized that the endothelial cell can regulate the function of the underlying smooth muscle cell by releasing various bioactive agents such as nitric oxide and endothelin-1. In addition to such paracrine regulation, other mechanisms exist by which there is cross-talk between these two cell types, including communication via the myoendothelial injunctions and information transfer via extracellular vesicles. Emerging evidence suggests that these nonparacrine mechanisms play an important role in the regulation of pulmonary vascular tone and the determination of cell phenotype and that they are critically involved in the pathobiology of pulmonary hypertension.
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Affiliation(s)
- Yuansheng Gao
- 1 Department of Physiology and Pathophysiology, Health Science Center, Peking University, Beijing, China; and
| | - Tianji Chen
- 2 Department of Pediatrics, College of Medicine, University of Illinois at Chicago, Chicago, Illinois
| | - J Usha Raj
- 2 Department of Pediatrics, College of Medicine, University of Illinois at Chicago, Chicago, Illinois
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17
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Sahoo S, Meijles DN, Al Ghouleh I, Tandon M, Cifuentes-Pagano E, Sembrat J, Rojas M, Goncharova E, Pagano PJ. MEF2C-MYOCD and Leiomodin1 Suppression by miRNA-214 Promotes Smooth Muscle Cell Phenotype Switching in Pulmonary Arterial Hypertension. PLoS One 2016; 11:e0153780. [PMID: 27144530 PMCID: PMC4856285 DOI: 10.1371/journal.pone.0153780] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2015] [Accepted: 04/04/2016] [Indexed: 12/22/2022] Open
Abstract
BACKGROUND Vascular hyperproliferative disorders are characterized by excessive smooth muscle cell (SMC) proliferation leading to vessel remodeling and occlusion. In pulmonary arterial hypertension (PAH), SMC phenotype switching from a terminally differentiated contractile to synthetic state is gaining traction as our understanding of the disease progression improves. While maintenance of SMC contractile phenotype is reportedly orchestrated by a MEF2C-myocardin (MYOCD) interplay, little is known regarding molecular control at this nexus. Moreover, the burgeoning interest in microRNAs (miRs) provides the basis for exploring their modulation of MEF2C-MYOCD signaling, and in turn, a pro-proliferative, synthetic SMC phenotype. We hypothesized that suppression of SMC contractile phenotype in pulmonary hypertension is mediated by miR-214 via repression of the MEF2C-MYOCD-leiomodin1 (LMOD1) signaling axis. METHODS AND RESULTS In SMCs isolated from a PAH patient cohort and commercially obtained hPASMCs exposed to hypoxia, miR-214 expression was monitored by qRT-PCR. miR-214 was upregulated in PAH- vs. control subject hPASMCs as well as in commercially obtained hPASMCs exposed to hypoxia. These increases in miR-214 were paralleled by MEF2C, MYOCD and SMC contractile protein downregulation. Of these, LMOD1 and MEF2C were directly targeted by the miR. Mir-214 overexpression mimicked the PAH profile, downregulating MEF2C and LMOD1. AntagomiR-214 abrogated hypoxia-induced suppression of the contractile phenotype and its attendant proliferation. Anti-miR-214 also restored PAH-PASMCs to a contractile phenotype seen during vascular homeostasis. CONCLUSIONS Our findings illustrate a key role for miR-214 in modulation of MEF2C-MYOCD-LMOD1 signaling and suggest that an antagonist of miR-214 could mitigate SMC phenotype changes and proliferation in vascular hyperproliferative disorders including PAH.
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Affiliation(s)
- Sanghamitra Sahoo
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Daniel N. Meijles
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Imad Al Ghouleh
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Manuj Tandon
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Eugenia Cifuentes-Pagano
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - John Sembrat
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Division of Pulmonary, Allergy and Critical Care Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Mauricio Rojas
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Division of Pulmonary, Allergy and Critical Care Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Elena Goncharova
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Division of Pulmonary, Allergy and Critical Care Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
| | - Patrick J. Pagano
- Department of Pharmacology and Chemical Biology University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- Vascular Medicine Institute, University of Pittsburgh, Pittsburgh, Pennsylvania, 15261, United States of America
- * E-mail:
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18
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Bai X, Dee R, Mangum KD, Mack CP, Taylor JM. RhoA signaling and blood pressure: The consequence of failing to “Tone it Down”. World J Hypertens 2016; 6:18-35. [DOI: 10.5494/wjh.v6.i1.18] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/30/2015] [Revised: 11/24/2015] [Accepted: 01/22/2016] [Indexed: 02/06/2023] Open
Abstract
Uncontrolled high blood pressure is a major risk factor for heart attack, stroke, and kidney failure and contributes to an estimated 25% of deaths worldwide. Despite numerous treatment options, estimates project that reasonable blood pressure (BP) control is achieved in only about half of hypertensive patients. Improvements in the detection and management of hypertension will undoubtedly be accomplished through a better understanding of the complex etiology of this disease and a more comprehensive inventory of the genes and genetic variants that influence BP regulation. Recent studies (primarily in pre-clinical models) indicate that the small GTPase RhoA and its downstream target, Rho kinase, play an important role in regulating BP homeostasis. Herein, we summarize the underlying mechanisms and highlight signaling pathways and regulators that impart tight spatial-temporal control of RhoA activity. We also discuss known allelic variations in the RhoA pathway and consider how these polymorphisms may affect genetic risk for hypertension and its clinical manifestations. Finally, we summarize the current (albeit limited) clinical data on the efficacy of targeting the RhoA pathway in hypertensive patients.
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19
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Rafikov R, Sun X, Rafikova O, Louise Meadows M, Desai AA, Khalpey Z, Yuan JXJ, Fineman JR, Black SM. Complex I dysfunction underlies the glycolytic switch in pulmonary hypertensive smooth muscle cells. Redox Biol 2015; 6:278-286. [PMID: 26298201 PMCID: PMC4556771 DOI: 10.1016/j.redox.2015.07.016] [Citation(s) in RCA: 78] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2015] [Revised: 07/21/2015] [Accepted: 07/28/2015] [Indexed: 01/21/2023] Open
Abstract
ATP is essential for cellular function and is usually produced through oxidative phosphorylation. However, mitochondrial dysfunction is now being recognized as an important contributing factor in the development cardiovascular diseases, such as pulmonary hypertension (PH). In PH there is a metabolic change from oxidative phosphorylation to mainly glycolysis for energy production. However, the mechanisms underlying this glycolytic switch are only poorly understood. In particular the role of the respiratory Complexes in the mitochondrial dysfunction associated with PH is unresolved and was the focus of our investigations. We report that smooth muscle cells isolated from the pulmonary vessels of rats with PH (PH-PASMC), induced by a single injection of monocrotaline, have attenuated mitochondrial function and enhanced glycolysis. Further, utilizing a novel live cell assay, we were able to demonstrate that the mitochondrial dysfunction in PH-PASMC correlates with deficiencies in the activities of Complexes I-III. Further, we observed that there was an increase in mitochondrial reactive oxygen species generation and mitochondrial membrane potential in the PASMC isolated from rats with PH. We further found that the defect in Complex I activity was due to a loss of Complex I assembly, although the assembly of Complexes II and III were both maintained. Thus, we conclude that loss of Complex I assembly may be involved in the switch of energy metabolism in smooth muscle cells to glycolysis and that maintaining Complex I activity may be a potential therapeutic target for the treatment of PH.
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Affiliation(s)
- Ruslan Rafikov
- Division of Translational and Regenerative Medicine, The University of Arizona, Tucson, AZ, USA; Department of Medicine, The University of Arizona, Tucson, AZ, USA.
| | - Xutong Sun
- Division of Translational and Regenerative Medicine, The University of Arizona, Tucson, AZ, USA; Department of Medicine, The University of Arizona, Tucson, AZ, USA
| | - Olga Rafikova
- Division of Translational and Regenerative Medicine, The University of Arizona, Tucson, AZ, USA; Department of Medicine, The University of Arizona, Tucson, AZ, USA
| | | | - Ankit A Desai
- Department of Medicine, The University of Arizona, Tucson, AZ, USA
| | - Zain Khalpey
- Department of Surgery, The University of Arizona, Tucson, AZ, USA
| | - Jason X-J Yuan
- Division of Translational and Regenerative Medicine, The University of Arizona, Tucson, AZ, USA; Department of Medicine, The University of Arizona, Tucson, AZ, USA
| | - Jeffrey R Fineman
- Department of Pediatrics and the University of California San Francisco, San Francisco, CA, USA; Cardiovascular Research Institute, University of California San Francisco, San Francisco, CA, USA
| | - Stephen M Black
- Division of Translational and Regenerative Medicine, The University of Arizona, Tucson, AZ, USA; Department of Medicine, The University of Arizona, Tucson, AZ, USA.
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Manso PH, Figueira RL, Prado CM, Gonçalves FL, Simões ALB, Ramos SG, Sbragia L. Early neonatal echocardiographic findings in an experimental rabbit model of congenital diaphragmatic hernia. ACTA ACUST UNITED AC 2015; 48:234-9. [PMID: 25651459 PMCID: PMC4381943 DOI: 10.1590/1414-431x20144184] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2014] [Accepted: 11/13/2014] [Indexed: 12/22/2022]
Abstract
This study aimed to demonstrate that congenital diaphragmatic hernia (CDH) results in
vascular abnormalities that are directly associated with the severity of pulmonary
hypoplasia and hypertension. These events increase right ventricle (RV) afterload and
may adversely affect disease management and patient survival. Our objective was to
investigate cardiac function, specifically right ventricular changes, immediately
after birth and relate them to myocardial histological findings in a CDH model.
Pregnant New Zealand rabbits underwent the surgical procedure at 25 days of gestation
(n=14). CDH was created in one fetus per horn (n=16), and the other fetuses were used
as controls (n=20). At term (30 days), fetuses were removed, immediately dried and
weighed before undergoing four-parameter echocardiography. The lungs and the heart
were removed, weighed, and histologically analyzed. CDH animals had smaller total
lung weight (P<0.005), left lung weight (P<0.005), and lung-to-body ratio
(P<0.005). Echocardiography revealed a smaller left-to-right ventricle ratio
(LV/RV, P<0.005) and larger diastolic right ventricle size (DRVS, P<0.007).
Histologic analysis revealed a larger number of myocytes undergoing mitotic division
(186 vs 132, P<0.05) in CDH hearts. Immediate RV dilation of CDH
hearts is related to myocyte mitosis increase. This information may aid the design of
future strategies to address pulmonary hypertension in CDH.
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Affiliation(s)
- P H Manso
- Departamento de Pediatria, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - R L Figueira
- Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - C M Prado
- Departamento de Patologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - F L Gonçalves
- Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - A L B Simões
- Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - S G Ramos
- Departamento de Patologia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
| | - L Sbragia
- Departamento de Cirurgia e Anatomia, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil
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Chettimada S, Gupte R, Rawat D, Gebb SA, McMurtry IF, Gupte SA. Hypoxia-induced glucose-6-phosphate dehydrogenase overexpression and -activation in pulmonary artery smooth muscle cells: implication in pulmonary hypertension. Am J Physiol Lung Cell Mol Physiol 2015; 308:L287-300. [PMID: 25480333 PMCID: PMC4338932 DOI: 10.1152/ajplung.00229.2014] [Citation(s) in RCA: 65] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2014] [Accepted: 12/01/2014] [Indexed: 11/22/2022] Open
Abstract
Severe pulmonary hypertension is a debilitating disease with an alarmingly low 5-yr life expectancy. Hypoxia, one of the causes of pulmonary hypertension, elicits constriction and remodeling of the pulmonary arteries. We now know that pulmonary arterial remodeling is a consequence of hyperplasia and hypertrophy of pulmonary artery smooth muscle (PASM), endothelial, myofibroblast, and stem cells. However, our knowledge about the mechanisms that cause these cells to proliferate and hypertrophy in response to hypoxic stimuli is still incomplete, and, hence, the treatment for severe pulmonary arterial hypertension is inadequate. Here we demonstrate that the activity and expression of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the pentose phosphate pathway, are increased in hypoxic PASM cells and in lungs of chronic hypoxic rats. G6PD overexpression and -activation is stimulated by H2O2. Increased G6PD activity contributes to PASM cell proliferation by increasing Sp1 and hypoxia-inducible factor 1α (HIF-1α), which directs the cells to synthesize less contractile (myocardin and SM22α) and more proliferative (cyclin A and phospho-histone H3) proteins. G6PD inhibition with dehydroepiandrosterone increased myocardin expression in remodeled pulmonary arteries of moderate and severe pulmonary hypertensive rats. These observations suggest that altered glucose metabolism and G6PD overactivation play a key role in switching the PASM cells from the contractile to synthetic phenotype by increasing Sp1 and HIF-1α, which suppresses myocardin, a key cofactor that maintains smooth muscle cell in contractile state, and increasing hypoxia-induced PASM cell growth, and hence contribute to pulmonary arterial remodeling and pathogenesis of pulmonary hypertension.
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Affiliation(s)
- Sukrutha Chettimada
- Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, Alabama
| | - Rakhee Gupte
- Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, Alabama
| | - Dhwajbahadur Rawat
- Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, Alabama
| | - Sarah A Gebb
- Department of Cell Biology and Neurosciences, College of Medicine, University of South Alabama, Mobile, Alabama
| | - Ivan F McMurtry
- Department of Pharmacology, College of Medicine, University of South Alabama, Mobile, Alabama; Department of Medicine, College of Medicine, University of South Alabama, Mobile, Alabama; and Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama
| | - Sachin A Gupte
- Department of Biochemistry and Molecular Biology, College of Medicine, University of South Alabama, Mobile, Alabama; Center for Lung Biology, College of Medicine, University of South Alabama, Mobile, Alabama
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Abstract
Myocardin (MYOCD) is a potent transcriptional coactivator that functions primarily in cardiac muscle and smooth muscle through direct contacts with serum response factor (SRF) over cis elements known as CArG boxes found near a number of genes encoding for contractile, ion channel, cytoskeletal, and calcium handling proteins. Since its discovery more than 10 years ago, new insights have been obtained regarding the diverse isoforms of MYOCD expressed in cells as well as the regulation of MYOCD expression and activity through transcriptional, post-transcriptional, and post-translational processes. Curiously, there are a number of functions associated with MYOCD that appear to be independent of contractile gene expression and the CArG-SRF nucleoprotein complex. Further, perturbations in MYOCD gene expression are associated with an increasing number of diseases including heart failure, cancer, acute vessel disease, and diabetes. This review summarizes the various biological and pathological processes associated with MYOCD and offers perspectives to several challenges and future directions for further study of this formidable transcriptional coactivator.
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Affiliation(s)
- Joseph M Miano
- Aab Cardiovascular Research Institute, University of Rochester School of Medicine and Dentistry, Rochester, NY, USA
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Shan F, Li J, Huang QY. HIF-1 alpha-induced up-regulation of miR-9 contributes to phenotypic modulation in pulmonary artery smooth muscle cells during hypoxia. J Cell Physiol 2014; 229:1511-20. [PMID: 24615545 DOI: 10.1002/jcp.24593] [Citation(s) in RCA: 44] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2013] [Accepted: 02/20/2014] [Indexed: 12/12/2022]
Abstract
Pulmonary artery smooth muscle cells (PASMCs) are associated with the development of hypoxic pulmonary hypertension (HPH). Recent studies have implicated a critical role for microRNAs (miRNAs) in HPH; however, their expression and regulation in hypoxia-mediated phenotypic modulation of PASMCs remains largely unclear. Here, we report that miR-9 was induced in hypoxia and involved in a hypoxia-induced phenotypic switch in rat primary PASMCs. Knockdown of miR-9 followed by hypoxia exposure attenuated PASMCs proliferation and enhanced the expression of contractile genes in vascular smooth muscle cells (VSMCs), while overexpression of miR-9 in normoxia promoted a proliferative phenotype in PASMCs. The primary transcripts of miR-9-1 and miR-9-3, but not miR-9-2, increased dramatically after hypoxia, whereas silencing of the hypoxia-associated transcription factor HIF-1α following hypoxia exposure abolished the enhancement of both primary transcripts in PASMCs. Using in silico analysis, we found three putative HIF-1α binding motifs on miR-9-1 and one motif on miR-9-3 located within the 5-kb region upstream of the transcriptional start sites. Chromatin immunoprecipitation assay revealed that hypoxia enhanced the direct interaction between HIF-1α and the regulatory elements of miR-9-1 and miR-9-3. Reporter assays showed that the regulatory regions of miR-9-1 and miR-9-3 behaved as enhancers in a HIF-1α-dependent manner during hypoxia. Taken together, our data uncover a regulatory mechanism involving HIF-1α-mediated up-regulation of miR-9, which plays a role in the hypoxia-induced phenotypic switch of PASMCs.
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Affiliation(s)
- Fabo Shan
- Department of Pathophysiology and High Altitude Physiology, College of High Altitude Military Medicine, Chongqing, China; Key Laboratory of High Altitude Medicine, Ministry of Education, Chongqing, China; Key Laboratory of High Altitude Medicine, PLA, Third Military Medical University, Chongqing, China
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24
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Xiao Y, Christou H, Liu L, Visner G, Mitsialis SA, Kourembanas S, Liu H. Endothelial indoleamine 2,3-dioxygenase protects against development of pulmonary hypertension. Am J Respir Crit Care Med 2014; 188:482-91. [PMID: 23822766 DOI: 10.1164/rccm.201304-0700oc] [Citation(s) in RCA: 45] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
Abstract
RATIONALE A proliferative and apoptosis-resistant phenotype in pulmonary arterial smooth muscle cells (PASMCs) is key to pathologic vascular remodeling in pulmonary hypertension (PH). Expression of indoleamine-2,3-dioxygenase (IDO) by vascular endothelium is a newly identified vasomotor-regulatory mechanism also involved in molecular signaling cascades governing vascular smooth muscle cell (vSMC) plasticity. OBJECTIVES To investigate the therapeutic potential of enhanced endothelial IDO in development of PH and its associated vascular remodeling. METHODS We used loss and gain of function in vivo studies to establish the role and determine the therapeutic effect of endothelial IDO in hypoxia-induced PH in mice and monocrotaline-induced PH in rats. We also studied PASMC phenotype in an IDO-high in vivo and in vitro tissue microenvironment. MEASUREMENTS AND MAIN RESULTS The endothelium was the primary site for endogenous IDO production within mouse lung, and the mice lacking this gene had exaggerated hypoxia-induced PH. Conversely, augmented pulmonary endothelial IDO expression, through a human IDO-encoding Sleeping Beauty (SB)-based nonviral gene-integrating approach, halted and attenuated the development of PH, right ventricular hypertrophy, and vascular remodeling in both preclinical models of PH. IDO derived from endothelial cells promoted apoptosis in PH-PASMCs through depolarization of mitochondrial transmembrane potential and down-regulated PH-PASMC proliferative/synthetic capacity through enhanced binding of myocardin to CArG box DNA sequences present within the promoters of vSMC differentiation-specific genes. CONCLUSIONS Enhanced endothelial IDO ameliorates PH and its associated vascular structural remodeling through paracrine phenotypic modulation of PH-PASMCs toward a proapoptotic and less proliferative/synthetic state.
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Affiliation(s)
- Yongguang Xiao
- Department of Surgery, Boston Children’s Hospital, Boston, MA, USA
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25
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Zheng XL. Myocardin and smooth muscle differentiation. Arch Biochem Biophys 2014; 543:48-56. [DOI: 10.1016/j.abb.2013.12.015] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2013] [Revised: 12/15/2013] [Accepted: 12/18/2013] [Indexed: 01/08/2023]
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Abstract
Hypoxic pulmonary hypertension of the newborn is characterized by elevated pulmonary vascular resistance and pressure due to vascular remodeling and increased vessel tension secondary to chronic hypoxia during the fetal and newborn period. In comparison to the adult, the pulmonary vasculature of the fetus and the newborn undergoes tremendous developmental changes that increase susceptibility to a hypoxic insult. Substantial evidence indicates that chronic hypoxia alters the production and responsiveness of various vasoactive agents such as endothelium-derived nitric oxide, endothelin-1, prostanoids, platelet-activating factor, and reactive oxygen species, resulting in sustained vasoconstriction and vascular remodeling. These changes occur in most cell types within the vascular wall, particularly endothelial and smooth muscle cells. At the cellular level, suppressed nitric oxide-cGMP signaling and augmented RhoA-Rho kinase signaling appear to be critical to the development of hypoxic pulmonary hypertension of the newborn.
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Affiliation(s)
- Yuansheng Gao
- Department of Physiology and Pathophysiology, Peking University, Health Science Center, Beijing, China
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27
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Protein kinase G-I deficiency induces pulmonary hypertension through Rho A/Rho kinase activation. THE AMERICAN JOURNAL OF PATHOLOGY 2012; 180:2268-75. [PMID: 22632818 DOI: 10.1016/j.ajpath.2012.02.016] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/08/2011] [Revised: 01/13/2012] [Accepted: 02/13/2012] [Indexed: 02/07/2023]
Abstract
Protein kinase G (PKG) plays an important role in the regulation of vascular smooth cell contractility and is a critical mediator of nitric oxide signaling, which regulates cardiovascular homeostasis. PKG-I-knockout (Prkg1(-/-)) mice exhibit impaired nitric oxide/cGMP-dependent vasorelaxation and systemic hypertension. However, it remains unknown whether PKG-I deficiency induces pulmonary hypertension. In this study, we characterized the hypertensive pulmonary phenotypes in Prkg1(-/-) mice and delineated the underlying molecular basis. We observed a significant increase in right ventricular systolic pressure in Prkg1(-/-) mice in the absence of systemic hypertension and left-sided heart dysfunction. In addition, we observed marked muscularization of distal pulmonary vessels in Prkg1(-/-) mice. Microangiography revealed impaired integrity of the pulmonary vasculature in Prkg1(-/-) mice. Mechanistically, PKG-I-mediated phosphorylation of Rho A Ser188 was markedly decreased, and the resultant Rho A activation was significantly increased in Prkg1(-/-) lung tissues, which resulted in Rho kinase activation. The i.t. administration of fasudil, a Rho kinase inhibitor, reversed the hypertensive pulmonary phenotype in Prkg1(-/-) mice. Taken together, these data show that PKG-I deficiency induces pulmonary hypertension through Rho A/Rho kinase activation-mediated vasoconstriction and pulmonary vascular remodeling.
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28
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Singh DK, Sarkar J, Raghavan A, Reddy SP, Raj JU. Hypoxia modulates the expression of leucine zipper-positive MYPT1 and its interaction with protein kinase G and Rho kinases in pulmonary arterial smooth muscle cells. Pulm Circ 2012; 1:487-98. [PMID: 22530104 PMCID: PMC3329079 DOI: 10.4103/2045-8932.93548] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/23/2022] Open
Abstract
We have shown previously that acute hypoxia downregulates protein kinase G (PKG) expression and activity in ovine fetal pulmonary vessels and pulmonary arterial smooth muscle cells (SMC). Here, we report that acute hypoxia also reduces the expression of leucinezipper-positive MYPT1 (LZ+MYPT1), a subunit of myosin light chain (MLC) phosphatase, in ovine fetal pulmonary arterial SMC. We found that in hypoxia, there is greater interaction between LZ+ MYPT1 and RhoA and Rho kinase 1 (ROCK1)/Rho kinase 2 (ROCK2) and decreased interaction between LZ+ MYPT1 and PKG, resulting in increased MLC20 phosphorylation, a higher pMLC20/MLC20 ratio and SMC contraction. In normoxic SMC PKG overexpression, LZ+ MYPT1 expression is upregulated while PKG knockdown had an opposite effect. LZ+ MYPT1 overexpression enhanced the interaction between PKG and LZ+ MYPT1. Overexpression of a mutant LZ- MYPT1 isoform in SMC mimicked the effects of acute hypoxia and decreased pMLC20/MLC20 ratio. Collectively, our data suggest that hypoxia downregulates LZ+ MYPT1 expression by suppressing PKG levels, reduces the interaction of LZ+ MYPT1 with PKG and promotes LZ+ MYPT1 interaction with RhoA or ROCK1/ROCK2, thereby promoting pulmonary arterial SMC contraction.
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Affiliation(s)
- Dev K Singh
- Department of Pediatrics, Division of Developmental Biology and Basic Research, University of Illinois at Chicago, Children's Hospital University of Illinois, Chicago, IL, USA, 1 & 2 Author contributed equally
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29
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Regulation of S100A4 expression via the JAK2–STAT3 pathway in rhomboid-phenotype pulmonary arterial smooth muscle cells exposure to hypoxia. Int J Biochem Cell Biol 2012; 44:1337-45. [DOI: 10.1016/j.biocel.2012.04.017] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2012] [Revised: 04/02/2012] [Accepted: 04/22/2012] [Indexed: 01/27/2023]
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30
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Gupte SA, Wolin MS. Relationships between vascular oxygen sensing mechanisms and hypertensive disease processes. Hypertension 2012; 60:269-75. [PMID: 22710643 DOI: 10.1161/hypertensionaha.112.190702] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Affiliation(s)
- Sachin A Gupte
- Department of Biochemistry and Molecular Biology, University of South Alabama, Mobile, AL, USA
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31
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Chettimada S, Rawat DK, Dey N, Kobelja R, Simms Z, Wolin MS, Lincoln TM, Gupte SA. Glc-6-PD and PKG contribute to hypoxia-induced decrease in smooth muscle cell contractile phenotype proteins in pulmonary artery. Am J Physiol Lung Cell Mol Physiol 2012; 303:L64-74. [PMID: 22582112 DOI: 10.1152/ajplung.00002.2012] [Citation(s) in RCA: 32] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Persistent hypoxic pulmonary vasoconstriction (HPV) plays a significant role in the pathogenesis of pulmonary hypertension, which is an emerging clinical problem around the world. We recently showed that hypoxia-induced activation of glucose-6-phosphate dehydrogenase (Glc-6-PD) in pulmonary artery smooth muscle links metabolic changes within smooth muscle cells to HPV and that inhibition of Glc-6PD reduces acute HPV. Here, we demonstrate that exposing pulmonary arterial rings to hypoxia (20-30 Torr) for 12 h in vitro significantly (P < 0.05) reduces (by 30-50%) SM22α and smooth muscle myosin heavy chain expression and evokes HPV. Glc-6-PD activity was also elevated in hypoxic pulmonary arteries. Inhibition of Glc-6-PD activity prevented the hypoxia-induced reduction in SM22α expression and inhibited HPV by 80-90% (P < 0.05). Furthermore, Glc-6-PD and protein kinase G (PKG) formed a complex in pulmonary artery, and Glc-6-PD inhibition increased PKG-mediated phosphorylation of VASP (p-VASP). In turn, increasing PKG activity upregulated SM22α expression and attenuated HPV evoked by Glc-6-PD inhibition. Increasing passive tension (from 0.8 to 3.0 g) in hypoxic arteries for 12 h reduced Glc-6-PD, increased p-VASP and SM22α levels, and inhibited HPV. The present findings indicate that increases in Glc-6-PD activity influence PKG activity and smooth muscle cell phenotype proteins, all of which affect pulmonary artery contractility and remodeling.
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Affiliation(s)
- Sukrutha Chettimada
- Department of Biochemistry & Molecular Biology, College of Medicine, University of South Alabama, Mobile, AL 36688, USA
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Yi B, Cui J, Ning J, Gu J, Wang G, Bai L, Qian G, Lu K. cGMP-dependent protein kinase Iα transfection inhibits hypoxia-induced migration, phenotype modulation and annexins A1 expression in human pulmonary artery smooth muscle cells. Biochem Biophys Res Commun 2012; 418:598-602. [PMID: 22293199 DOI: 10.1016/j.bbrc.2012.01.034] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2011] [Accepted: 01/08/2012] [Indexed: 01/06/2023]
Abstract
Our previous work has demonstrated that the cellular phenotype changes of human pulmonary artery smooth muscle cells (PASMCs) play an important role during pulmonary vascular remodelling. However, little is known about the role of PASMCs phenotype modulation in the course of hypoxia-induced migration and its behind molecular mechanisms. In this study, we have shown that cGMP-dependent protein kinase (PKG) Iα transfection significantly attenuated the hypoxia-induced down-regulation of the expressions of SM-α-actin, MHC and calponin. Hypoxia-induced PASMC migration was also suppressed by PKGIα overexpression. Furthermore, this overexpression attenuated ANX A1 upregulation under hypoxic conditions. All those effects were reversed by a PKG inhibitor KT5823. Our data indicate that manipulating upstream entity e.g., PKGIa, may have a potential therapeutic value to prevent hypoxia-associated pulmonary arterial remodeling for pulmonary hypertension development.
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Affiliation(s)
- Bin Yi
- Department of Anesthesia, Southwest Hospital, The Third Military Medical University, Chongqing 400038, China
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33
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Yi B, Cui J, Ning JN, Wang GS, Qian GS, Lu KZ. Over-expression of PKGIα inhibits hypoxia-induced proliferation, Akt activation, and phenotype modulation of human PASMCs: The role of phenotype modulation of PASMCs in pulmonary vascular remodeling. Gene 2012; 492:354-60. [DOI: 10.1016/j.gene.2011.11.010] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2011] [Revised: 10/12/2011] [Accepted: 11/03/2011] [Indexed: 11/16/2022]
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34
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Ramchandran R, Pilipenko E, Bach L, Raghavan A, Reddy SP, Raj JU. Hypoxic regulation of pulmonary vascular smooth muscle cyclic guanosine monophosphate-dependent kinase by the ubiquitin conjugating system. Am J Respir Cell Mol Biol 2011; 46:323-30. [PMID: 21997485 DOI: 10.1165/rcmb.2011-0165oc] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
We previously reported that hypoxia attenuates nitric oxide-cyclic guanosine monophosphate (NO-cGMP)-mediated fetal pulmonary vessel relaxation by inhibiting cGMP-dependent protein kinase 1 (PKG1) activity, but not all the mechanisms by which acute hypoxia inhibits PKG1 activity have been delineated. Here we demonstrate for the first time, to the best of our knowledge, that acute hypoxia induces an accumulation of ubiquitinated PKG1 in ovine fetal and newborn pulmonary artery smooth muscle cells. Such a modification was not evident in ovine fetal systemic (cerebral) artery smooth muscle cells. The accumulation of polyubiquitinated PKG1 observed after 4 hours of hypoxia was affected neither by the activation of PKG1 kinase activity with the cell-permeable cGMP analogue 8-bromo-cGMP, nor by its inhibition with DT-3 in fetal pulmonary artery smooth muscle cells. Ubiquitinated PKG1α was unable to bind the cGMP analogue 8-(2-aminoethyl)thioguanosine-3',5' (AET)-cGMP, a ligand for the unmodified protein. Inhibition of the proteasomal complex with MG132 led to the accumulation of polyubiquitinated PKG1 in normoxia, indicating the involvement of the ubiquitin-26S proteasomal system in degradation and clearance of this protein under normoxic conditions. The ubiquitinated PKG1 under hypoxic conditions, however, was not predominantly targeted for proteasomal degradation. Importantly, reoxygenation reversed the acute hypoxia-induced accumulation of ubiquitinated PKG1. Our results suggest that the PKG1 ubiquitination induced by acute hypoxia plays a unique role in the regulation of the pulmonary vascular smooth muscle cell vasoreactivity and relaxation mediated by the NO-cGMP-PKG1 pathway.
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Affiliation(s)
- Ramaswamy Ramchandran
- Department of Pediatrics, College of Medicine, University of Illinois at Chicago, 840 S. Wood St., M/C 856, Chicago, IL 60612, USA.
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35
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Yin H, Li Q, Qian G, Wang Y, Li Y, Wu G, Wang G. Rab1 GTPase regulates phenotypic modulation of pulmonary artery smooth muscle cells by mediating the transport of angiotensin II type 1 receptor under hypoxia. Int J Biochem Cell Biol 2011; 43:401-8. [PMID: 21095238 PMCID: PMC3072814 DOI: 10.1016/j.biocel.2010.11.010] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/18/2010] [Revised: 11/03/2010] [Accepted: 11/14/2010] [Indexed: 12/30/2022]
Abstract
Previous studies have demonstrated that Rab1 is involved in the export of angiotensin II (Ang II) type 1 receptor (AT1R) to the cell surface in endothelial cells and cardiomyocytes. The aim of this study was to evaluate whether the modification of Rab1-mediated endoplasmic reticulum (ER) to the Golgi body transport alters the cell surface expression and function of endogenous AT1R and AT1R-mediated phenotypic modulation in primary cultures of pulmonary artery smooth muscle cells (PASMCs). Lentiviral expression of wild-type Rab1 (Rab1WT) significantly increased cell surface expression of endogenous AT1R. However, Rab1 siRNA had the opposite effect, and attenuated downregulation of the expression of PASMCs phenotype markers, α smooth muscle actin (α-SMA) and vimentin (VIM) in rat pulmonary artery smooth muscle cells (RPASMCs) during hypoxia. Analysis of the subcellular localization of AT1R revealed that Rab1 regulated AT1R transport from the ER to the Golgi in PASMCs. Consistent with their effects on AT1R export, Rab1 modified the AT1R-mediated cell growth and the phosphorylation of signal transducing activator of transcription 3 (STAT3) during hypoxia. We found that hypoxia promoted Rab1 expression and strongly correlated with the repressed expression of PASMC phenotype markers in RPASMCs. These data strongly indicate that Rab1 modulates PASMCs function by manipulating AT1R traffic from the ER to the Golgi and provide the first evidence implicating ER-to-Golgi transport as a regulatory step for the control of RPASMCs growth.
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Affiliation(s)
- Hongjin Yin
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
| | - Qi Li
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
| | - Guisheng Qian
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
| | - Yaoli Wang
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
| | - Yuncheng Li
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
| | - Guangyu Wu
- Department of Pharmacology and Experimental Therapeutics, Louisiana State University Health Sciences Center, New Orleans, LA 70112, USA
| | - Guansong Wang
- Institute of Respiratory Diseases, the Second Affiliated Hospital of the Third Military Medical University, Chongqing 400037, PR China
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You JJ, Yang CM, Chen MS, Yang CH. Regulation of Cyr61/CCN1 expression by hypoxia through cooperation of c-Jun/AP-1 and HIF-1α in retinal vascular endothelial cells. Exp Eye Res 2010; 91:825-36. [PMID: 21029732 DOI: 10.1016/j.exer.2010.10.006] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/21/2010] [Revised: 09/24/2010] [Accepted: 10/18/2010] [Indexed: 12/14/2022]
Abstract
Hypoxia is the most important factor in the pathogenesis of diabetic retinopathy. Cysteine-rich 61 (Cyr61) is one of the angiogenic factors involved in the development of proliferative diabetic retinopathy (PDR). The aim of this study was to investigate the mechanism of hypoxia-induced Cyr61 expression in retinal vascular endothelial cells. The hypoxia-induced expression of mRNA and protein of Cyr61 was studied in monkey choroidal retinal vascular endothelial (RF/6A) cells. Luciferase reporter assays and electrophoretic mobility shift assays were used to identify the hypoxia responsible region and transcription factors in the Cyr61 promoter. Chromatin immunoprecipitation and immunoprecipitation were performed to study the role of hypoxia-inducible factor (HIF)-1α and c-Jun/activator protein-1 (AP-1) in Cyr61 transcriptional regulation. The results showed that hypoxia significantly induced Cyr61 mRNA and protein expression in RF/6A cells. The effect was mediated through phosphorylation of c-Jun. Luciferase assays, electrophoretic mobility shift assays, chromatin immunoprecipitation and immunoprecipitation showed that HIF-1α interacted with c-Jun/AP-1 and their binding on the AP-1 binding motif within the Cyr61 promoter induced the expression of Cyr61. In conclusion, hypoxia controlled the transcriptional regulation of the Cyr61 gene in RF/6A cells by cooperation of HIF-1α and c-Jun/AP-1. Cyr61 might play an important role in ischemic retinal diseases, such as PDR.
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Affiliation(s)
- Jian-Jang You
- Keelung General Hospital, Department of Health, The Executive Yuan, Keelung, Taiwan
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Abstract
During the development of the pulmonary vasculature in the fetus, many structural and functional changes occur to prepare the lung for the transition to air breathing. The development of the pulmonary circulation is genetically controlled by an array of mitogenic factors in a temporo-spatial order. With advancing gestation, pulmonary vessels acquire increased vasoreactivity. The fetal pulmonary vasculature is exposed to a low oxygen tension environment that promotes high intrinsic myogenic tone and high vasocontractility. At birth, a dramatic reduction in pulmonary arterial pressure and resistance occurs with an increase in oxygen tension and blood flow. The striking hemodynamic differences in the pulmonary circulation of the fetus and newborn are regulated by various factors and vasoactive agents. Among them, nitric oxide, endothelin-1, and prostaglandin I2 are mainly derived from endothelial cells and exert their effects via cGMP, cAMP, and Rho kinase signaling pathways. Alterations in these signaling pathways may lead to vascular remodeling, high vasocontractility, and persistent pulmonary hypertension of the newborn.
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Affiliation(s)
- Yuansheng Gao
- Department of Physiology and Pathophysiology, Peking University, Health Science Center, Beijing, China; and Department of Pediatrics, University of Illinois, College of Medicine at Chicago, Chicago, Illinois
| | - J. Usha Raj
- Department of Physiology and Pathophysiology, Peking University, Health Science Center, Beijing, China; and Department of Pediatrics, University of Illinois, College of Medicine at Chicago, Chicago, Illinois
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Choi C, Sellak H, Brown FM, Lincoln TM. cGMP-dependent protein kinase and the regulation of vascular smooth muscle cell gene expression: possible involvement of Elk-1 sumoylation. Am J Physiol Heart Circ Physiol 2010; 299:H1660-70. [PMID: 20802137 DOI: 10.1152/ajpheart.00677.2010] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/05/2023]
Abstract
Although the regulation of smooth muscle cell (SMC) gene expression by cGMP-dependent protein kinase (PKG) is now recognized, the mechanisms underlying these effects are not fully understood. In this study, we report that PKG-I stimulates myocardin/serum response factor (SRF)-dependent gene expression in vascular SMCs. The expression of PKG in PKG-deficient cells enhanced myocardin-induced SM22 promoter activity in a concentration-dependent fashion. However, neither SRF nor myocardin expression was affected. To investigate alternative mechanisms, we examined whether PKG affects the phosphorylation of E26-like protein-1 (Elk-1), a SRF/myocardin transcription antagonist. The activation of PKG caused an increase in a higher molecular mass form of phospho-Elk-1 that was determined to be small ubiquitin-related modifier (sumo)ylated Elk-1. PKG increased Elk-1 sumoylation twofold compared with the PKG-deficient cells, and Elk-1 sumoylation was reduced using dominant-negative sumo-conjugating enzyme, DN-Ubc9, confirming PKG-dependent sumoylation of phospho-Elk-1 in vascular SMCs. In addition, PKG stimulated Elk-1 sumoylation in COS-7 cells overexpressing Elk-1, sumo-1, and PKG-I. The increased expression of PKG in vascular SMCs inhibited Elk-1 binding to SMC-specific promoters, SM22 and smooth muscle myosin heavy chain, as measured by EMSA and chromatin immunoprecipitation assay, and PKG suppressed the Elk-1 inhibition of SM22 reporter gene expression. Taken together, these data suggest that PKG-I decreases Elk-1 activity by sumo modification of Elk-1, thereby increasing myocardin-SRF activity on SMC-specific gene expression.
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Affiliation(s)
- ChungSik Choi
- Department of Physiology, College of Medicine, University of South Alabama, Mobile, Alabama 36609, USA
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Jie W, Guo J, Shen Z, Wang X, Zheng S, Wang G, Ao Q. Contribution of myocardin in the hypoxia-induced phenotypic switching of rat pulmonary arterial smooth muscle cells. Exp Mol Pathol 2010; 89:301-6. [PMID: 20621093 DOI: 10.1016/j.yexmp.2010.06.010] [Citation(s) in RCA: 29] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/12/2010] [Revised: 06/27/2010] [Accepted: 06/29/2010] [Indexed: 12/19/2022]
Abstract
BACKGROUND Hypoxic exposure contributes to the phenotypic switching of smooth muscle cells (SMCs), while the mechanisms involved in this process is not yet fully elucidated. Myocardin as a co-actor of serum reaction factor plays a crucial role in differentiation of SMCs. This study was aimed to investigate the role of myocardin in hypoxia-induced phenotypic switching of rat pulmonary arterial SMCs (PASMCs). METHODS Primary PASMCs were cultured under normoxia and hypoxia (3%O(2), 48 h) respectively, and then the cell proliferation was assessed and the expression of SM22α, osteopontin (contractile and synthetic marker of SMCs, respectively), myocardin and platelet-derived growth factor-BB (PDGF-BB) were detected. After pGCSIL-GFP-shMYOCD lentviral vector was transduced to the PASMCs, the expression of myocardin and SM22α were examined. Moreover, myocardin expression in PASMCs treated with medium enriched with PDGF-BB and conditional medium (CM) from normoxia- and hypoxia-exposed PASMCs was assessed. RESULTS Exposing PASMCs to hypoxia led to an increased cell numbers and the up-regulation of proliferating cell nuclear antigen (PCNA), osteopontin and PDGF-BB; moreover, a significant down-regulation of SM22α and myocardin was identified. Further analysis revealed that knock-down of myocardin with pGCSIL-GFP-shMYOCD vector followed by a decreased SM22α in the PASMCs, and treatment of PASMCs with either exogenous PDGF-BB or hypoxic CM led to a marked decrease of myocardin. CONCLUSIONS Our findings suggest that the decrease of myocardin in PASMCs exposed to hypoxia is partly regulated by the increase of PDGF-BB, which contributes to the phenotypic switching of PASMCs in hypoxic condition.
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Affiliation(s)
- Wei Jie
- Institute of Pathology, Tongji Hospital, Huazhong University of Science and Technology, Wuhan 430030, PR China
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Puetz S, Lubomirov LT, Pfitzer G. Regulation of smooth muscle contraction by small GTPases. Physiology (Bethesda) 2010; 24:342-56. [PMID: 19996365 DOI: 10.1152/physiol.00023.2009] [Citation(s) in RCA: 113] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023] Open
Abstract
Next to changes in cytosolic [Ca(2+)], members of the Rho subfamily of small GTPases, in particular Rho and its effector Rho kinase, also known as ROK or ROCK, emerged as key regulators of smooth muscle function in health and disease. In this review, we will focus on the regulation of the contractile machinery by Rho/ROK signaling and its interaction with PKC and cyclic nucleotide signaling. We will briefly discuss the emerging evidence that remodeling of cortical actin is necessary for contraction.
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Affiliation(s)
- Sandra Puetz
- Institut für Vegetative Physiologie, Universitaet Koeln, Koeln, Germany,
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