Discussion on the Inhibitory Mechanism of Lanthanum Chloride on Vascular Smooth Muscle Cell Calcification Induced by High Phosphorus Based on NF-κB Signaling Pathway / 中国药房

OBJECTIVE：To discuss the inhibitory effect of lanthanum chloride on the calcification of vascular smooth muscle cells（VSMCs）induced by high phosphorus and its mechanism. METHODS ：On the basis of screening the action concentration and time of lanthanum chloride by MTT method ，human VSMCs were divided into control group （1 mmol/L phosphorus solution ）， lanthanum chloride high concentration control group （1 mmol/L phosphorus solution+ 60 μmol/L lanthanum chloride），model group （3 mmol/L phosphorus solution ），sodium chloride group （3 mmol/L phosphorus solution+ 180 μmol/L sodium chloride），nuclear factor κB（NF-κB）signaling pathway agonist+lanthanum chloride group （3 mmol/L phosphorus solution+ 1 μg/mL lipopolysaccharide+ 60 μmol/L lanthanum chloride），positive control group （3 mmol/L phosphorus solution+ 100 μmol/L sodium pyrophosphate），and lanthanum chloride low ，medium，and high concentration groups （3 mmol/L phosphorus solution+ 15，30，60 μmol/L lanthanum chloride）. Alizarin red S staining and Von Kossa staining were used to detect cell calcification in each group after treated with phosphorus solution for 6 d and relevant medicine for 2 d. Western blot assay was used to detect the protein expression of TNF-α receptor associated protein 6（TRAF6），nuclear factor κB inhibitor protein α（IκBα），NF-κB p65，bone morphogenetic protein 2 （BMP-2），smooth muscle 22 α（SM22α）and Runt related transcription factor 2（Runx2）. Real-time fluorescence quantitative polymerase chain reaction was used to detect mRNA expression of TRAF 6，IκBα，BMP-2，SM22α and Runx2. RESULTS ： Compared with control group ，no cell calcification was observed in the lanthanum chloride high concentration control group ，while obvious cell calcification and significant increase of OD value were observed in model group and sodium chloride group （P＜ 0.01）；protein and mRNA expression of TRAF 6 and BMP- 2 in cytoplasm as well as mRNA expression of Runx 2，protein expression of NF-κB p65 and Runx 2 in nucleus were significantly increased （P＜0.01）；protein and mRNA expression of IκBα and SM22α as well as protein expression of NF-κB p65 in cytoplasm were significantly decreased （P＜0.01）. Compared with model group，cell calcification was significantly improved in lanthanum chloride groups and positive control group ，while OD values were significantly reduced ；the expression levels of the above-mentioned protein and mRNA were reversed to varying degrees （P＜0.05 or P＜0.01）. Compared with lanthanum chloride high concentration group ，obvious cell calcification was observed in NF-κB signaling pathway agonist + lanthanum chloride group ，and OD value was significantly increased ；the above indexes were significantly reversed in cytoplasm and nucleus （P＜0.05 or P＜0.01）. CONCLUSIONS ：Lanthanum chloride can inhibit the calcification of VSMCs induced by high phosphorus ，and its mechanism may be related to the inhibition of NF-κB signaling pathway activation.

rBMSCs/ITGA5B1 Promotes Human Vascular Smooth Muscle Cell Differentiation via Enhancing Nitric Oxide Production

BACKGROUND AND OBJECTIVES: Previous studies have shown that integrins alpha5beta1 (ITGA5B1) gene-modified rat bone marrow mesenchymal stem cells (rBMSCs) could prevent cell anoikis and increase the nitric oxide (NO) production. Here we examined the capability of rBMSCs/ITGA5B1 on the phenotype modulation of Human Pulmonary Artery Smooth Muscle Cell (HPASMC) in vitro. METHODS AND RESULTS: The synthetic (dedifferentiated) phenotype of HPASMC was induced by monocrotaline (MCT, 1μM) for 24 h and then co-cultured with rBMSCs/ITGA5B1 in a transwell culture system. The activation of NO/cGMP (nitric oxide/Guanosine-3′, 5′-cyclic monophosphate) signaling was investigated in HPASMC. The changes of pro-inflammatory factors, oxidative stress, vasodilator, vasoconstrictor, contractile and synthetic genes, and the morphological changes of HPASMC were investigated. The results of this study showed that the NO/cGMP signal, endothelial nitric oxide synthase (eNOS) expression, the expression of the vasoprotective genes heme oxygenase-1 (HMOX1) and prostaglandin-endoperoxide synthase 2 (PTGS2) were increased, but the expression of transforming growth factor-β1 (TGF-β1), CCAAT/enhancer-binding proteins delta (Cebpd), Krüppel-like factor 4 (KLF4), and activating transcription factor 4 (ATF4) were reduced in MCT treated HPASMC co-cultured with rBMSCs/ITGA5B1. The synthetic smooth muscle cells (SMCs) phenotype markers thrombospondin-1, epiregulin and the vasoconstrictor endothelin (ET)-1, thromboxane A2 receptor (TbxA2R) were down-regulated, whereas the contractile SMCs phenotype marker transgelin expression was up-regulated by rBMSCs/ITGA5B1. Furthermore, rBMSCs/ITGA5B1 promoted the morphological restoration from synthetic (dedifferentiation) to contractile (differentiation) phenotype in MCT treated HPASMC. CONCLUSIONS: rBMSCs/ITGA5B1 could inhibit inflammation and oxidative stress related genes to promote the HPASMC cell differentiation by activation NO/cGMP signal.

Adipophilin increases lipid accumulation in human vascular smooth muscle cells / 中华内分泌代谢杂志

Objective To observe the effect of acetylated low-density lipoprotein (AcLDL) on the expression of adipophilin and the effect of adipophilin on AcLDL uptake and lipid accumulation in human vascular smooth muscle cells (HVSMCs)in order to approach the role played by adipophilin in genesis of macroangiopathy. Methodse HVSMCs were treated with various amount of AcLDL. Adipophilin expression levels were detected by Northern blot and Western blot. The effects of adipophilin on AcLDL uptake and lipid accumulation in HVSMCs were observed by the methods of siRNA, flow cytometry, enzymatic method and oil red stain. Results AcLDL dose-dependently increased adipophilin expression in HVSMCs. Silence adipophilin by siRNA decreased AcLDL uptake (decreasing by 38.7%, P<0. 05) and lipids accumulation (tfiglyceride and total cholesterol decreasing by 30.6% and 29.8% respectively, both P<0. 01) in HVSMCs, Conclusion Adipophilin is able to increase AcLDL uptake and lipid accumulation in HVSMCs, suggesting that it might play a role in enhancing atherosclerosis.

Effect of Mycophenolic Acid on Human Vascular Smooth Muscle Cell Proliferation and Its Signal Transduction

PURPOSE: Vascular smooth muscle cells (VSMCs) migration and proliferation play important roles in chronic allograft rejection. Mycophenolic acid (MPA) inhibits the proliferation of VSMCs, glomerular mesangial cells and fibroblasts as well as lymphocytes. Since reactive oxygen species (ROS) and mitogen-activated protein kinase (MAPK) play important roles in the proliferation of VSMCs, the present study examined the effects of MPA on intracellular ROS generation, activation of ERK and p38 MAPK, and the proliferation of VSMCs cultured under platelet derived growth factor (PDGF). METHODS: Human VSMCs obtained from ATCC were cultured with RPMI-1640 containing 10% fetal bovine serum. Near confluent VSMCs were incubated with serum-free media for 48 hours to arrest and synchronize the cell growth. MPA was administered 1 hour before the addition of PDGF. 5-(and-6)- chloromethyl-2',7'-dichlorodihydrofluorescein (DCF)-sensitive intracellular ROS was detected by FACS. Activations of ERK1/ERK2 and p38 MAPK were measured by Western blot analysis. Proliferation of VSMC was assessed by [(3)]. RESULTS: PDGF administered at 10 ng/ml, which induced human VSMCs proliferation, rapidly increased intracellular ROS by 1.6-fold (P<0.05), ERK1/ERK2 activation by 2.1-fold, (P<0.05) and p38 MAPK activation by 1.9-fold (P<0.05), respectively, as compared to the control. MPA 1 and 10nM effectively inhibited PDGF-induced human VSMCs proliferation. MPA also effectively inhibited PDGF-induced intracellular ROS generation as well as ERK1/ERK2 and p38 MAPK activation. CONCLUSION: The present study suggests that MPA inhibits PDGF-induced human VSMCs proliferation, possibly by inhibiting intracellular ROS generation and the phosphorylation of ERK1/ERK2 and p38 MAPK. H]-thymidine incorporation.