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Objective::To investigate the protective effect of Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma and Chuanxiong Rhizoma extracts on vascular calcification induced by high glucose in mice by observing the expression of osteopontin (OPN) and smooth muscle 22α (SM22α) as well as vascular calcium deposition in the common carotid artery and thoracic aorta of mice. Method::Totally 130 male C57BL/6 mice were randomly divided into normal control group and high glucose group. The mice in high glucose group were intraperitoneally injected with streptozotocin(STZ), and fed on a high-fat diet for 7 months. Then, the mice were randomly divided into model group, low-dose and high-dose Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma and Chuanxiong Rhizoma extracts groups (0.819, 1.638 g·kg-1), and metformin group (150 mg·kg-1). Each group was intragastrically administered once a day for 9 weeks. The changes in blood glucose were measured. Seven days before the end of the administration, a group of 4-week old male C57BL/6 mice were purchased and fed normally for one week as a youth group. At the end of the administration, the common carotid artery and thoracic aorta tissues of the mice were collected. Von Kossa staining was used to determine the degree of calcium deposition in the common carotid artery and thoracic aorta. The expression levels of OPN and SM22α protein in the common carotid artery and thoracic aorta were detected by immunohistochemistry. The expression of OPN and SM22α protein in the common carotid artery of mice was determined by Western blot. Result::As compared with the young group, the blood glucose of the normal control group was slightly increased without statistical difference, the common carotid artery and thoracic aorta were uniformly stained, and no black granular precipitate was observed. As compared with the normal control group, the blood glucose of the model group was increased (P<0.01), with a large amount of brown-black particles deposited in the intimal elastic fibers, showing obvious calcium salt deposition. As compared with the model group, blood glucose was significantly decreased in each administration group (P<0.05, P<0.01), and the degree of vascular calcium salt deposition was significantly reduced. There were no significant changes in expression levels of OPN protein and SM22α protein in the common carotid artery and thoracic aorta between the youth group and normal control group. As compared with the normal control group, the expression of intimal OPN protein in the common carotid artery and thoracic aorta of the model group was positive, SM22α protein expression was weakly positive, and the gray value of OPN protein expression in the common carotid artery was significantly increased (P<0.01), while the gray value of SM22α protein was decreased significantly (P<0.01). As compared with the model group, the expression levels of intimal OPN protein and SM22α protein in the common carotid artery and thoracic aorta of each administration group were significantly improved, and the gray value of OPN protein expression in the common carotid artery was reduced (P<0.05, P<0.01), while SM22α protein expression was significantly increased (P<0.01). Conclusion::High glucose can induce calcification of common carotid artery and thoracic aorta in mice and accelerate vascular aging. This formation process may be related to the expression of OPN and SM22α. Ginseng Radix et Rhizoma, Notoginseng Radix et Rhizoma and Chuanxiong Rhizoma extracts can reduce vascular calcification and delay vascular aging by regulating the expression of OPN and SM22α.
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AIM:To investigate the effect of 27nt-microRNA (27nt-miRNA) on the expression of smooth muscle 22α protein (SM22α) and the cell viability, migration and phenotypic changes of vascular smooth muscle cells (VSMCs).METHODS:The highly expression plasmids of 27nt-miRNA, and anti-27nt-miRNA and negative control plasmids were constructed, packaged with lentivirus and transfected into the rat primary VSMCs.Platelet-derived growth factor BB (PDGF-BB) was added to induce VSMCs phenotype conversion.The cell viability was measured by MTT assay.The migration ability was detected by scratch assay.The mRNA and protein expression of SM22αwas determined by RT-PCR, immunocytochemical staining and Western blot.RESULTS:Compared with normal group, the cell viability in PDGF-BB group was increased (P<0.05) , the migration ability was increased (P<0.05) and the expression of SM22αat mRNA and protein level was decreased (P<0.05).Compared with negative control lentiviral group, the cell viability in 27ntmiRNA over-expression group was decreased (P<0.05) , the migration ability was decreased (P<0.05) , and the mRNA and protein expression of SM22αwas increased (P<0.05).While in anti-27nt-miRNA group, the cell viability was increased (P<0.05) , the migration ability was increased (P<0.05) , and the mRNA and protein expression of SM22αwas decreased (P<0.05).CONCLUSION:27nt-miRNA significantly increases the expression of SM22α, while inhibits the viability and migration ability of VSMCs, and inhibits its phenotypic shift from contractile to synthetic.
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To investigate the effect of 27nt-miRNA on the differentiation of mesenchymal stem cells into vascular smooth muscle cells. The highly expression plasmids of 27nt-miRNA and anti-27nt-miRNA, and negative control plasmids were constructed, packaged with lentivirus and transfected into human umbilical cord mesenchymal stem cells (hUCMSCs). Collagen IV was added to induce hUCMSCs differentiation into blood vessel smooth muscle cells (VSMCs). The cell viability was measured by MTT assay. The expression of SMA, SM22α at mRNA and protein levels was determined by RT-PCR, immunocytochemical staining and Western blotting. Compared with the negative control group, the viability of the 27nt-miRNA overexpression group was decreased by 20.48% (P<0.05), and the expression of SMA mRNA and SM22α mRNA and protein was significantly increased (P<0.05); the viability of Anti-27nt-miRNA group was increased 18.07% (P<0.05), and the expression of SMA mRNA and SM22α mRNA and protein was decreased (P<0.05). In summary, 27nt-miRNA promotes mesenchymal stem cells differentiation into vascular smooth muscle cells and inhibits cells viability.
Subject(s)
Humans , Cell Differentiation , Cells, Cultured , Mesenchymal Stem Cells , MicroRNAs , Muscle, Smooth, Vascular , Myocytes, Smooth MuscleABSTRACT
Objective To evaluate smooth muscle protein of 22 kDa (SM22) in the diagnosis of acute intestinal ischemia.Methods 96 healthy adult SD rats were evenly divided into experimental group and control group,with each group subdivided into 6 subgroups,subject respectively to superior mesenteric artery ligation or sham operation.The venous blood samples were extracted from each group rats' right heart atO.5,1,2,4,8,12 h after the operation,for SM22 testing and small intestines tissues for direct immunofluorescence staining of SM22.Results The serum SM22 concentration reached a peak at 4 h (265 ± 15) mg/L,then gradually decreased (P ＜ 0.05).The I-FABP was mainly expressed in the epithelium of intestinal mucosa.During the 4 hours of intestinal ischemia,The number of SM22 positive particles did not change.After 4 hours,the number of SM22 positive granules had gradually decreased compared with the control group (all P ＜ 0.05).Conclusion SM22 mainly exists in the smooth muscle of intestinal,during the ischemic necrosis of the intestinal muscle layer SM22 leaks into blood stream,resulting in high serum levels of SM22 facilitating early diagnosis of acute intestinal ischemia.
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Objective: To examine the serum level of IL-22 in asthmatic patients and the expression of IL-22R1 in human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts, so as to explore the target cells of IL-22. Methods: The serum levels of IL-22 and IL-17 in 36 asthmatic patients and 20 normal control subjects were measured by enzyme-linked immunosorbent assay (ELISA). And lung function of the asthmatic patients was assessed by Gaeger spirometry. According to the values of FEV1/FVC and FEV1%, the patients were divided into two groups, bronchodilation test positive group (19 patients) and bronchial provocation test positive group (17 patients). The expression of IL-22R1 mRNA in human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts was examined using real-time PCR, and IL-22R1 protein expression was detected by immunofluorescence staining and Western blotting analysis. Results: There was no significant difference in serum IL-22 and IL-17 levels between the asthmatic patients and normal controls. The serum IL-22 and IL-17 levels in asthmatic patients positive for bronchodilation test was significantly higher than those positive for bronchial provocation test(P<0.05). IL-22R1 mRNA and protein were detected in all the 3 types of cells. Conclusion: IL-22 may be involved in the pathogenesis of asthma, and human airway epithelial cells, human airway smooth muscle cells, and lung fibroblasts may all be the target cells of IL-22.