Objective To explore the expression and significance of hypoxia-inducible factor 1α (HIF-1α) in endplate chondrocytes, and to study the relations between HIF-1α expression and endplate chondrocytes apoptosis. Methods Eight Sprague Dawley rats were selected to obtain the L1-5 intervertebral disc endplate; the endplate chondrocytes were isolated by enzyme digestion method, and the endplate chondrocytes at passage 3 were cultured under 20% O2 condition (group A), and under 0.5% O2 condition (group B). Cell morphology was observed by inverted phase contrast microscope and cell apoptosis was detected using flow cytometry after cultured for 24 hours; the mRNA expression of HIF-1α was detected by real-time fluorescent quantitative PCR, the protein expressions of HIF-1α, Bax, and Bcl-2 by Western blot. Gene clone technology to design and synthesize two siRNAs based on the sequence of HIF-1α mRNA. HIF-1α specific RNAi sequence compound was constructed and transfected into cells. The transfected endplate chondrocytes at passage 3 were cultured under 0.5% O2 condition in group C and group D (HIF-1α gene was silenced). After cultured for 24 hours, cells were observed via immunofluorescence staining of HIF-1α, and cell apoptosis was detected using flow cytometry. Meanwhile, the mRNA expressions of HIF-1α, collagen type II (COL II), Aggrecan, and SOX9 were detected by real-time fluorescent quantitative PCR, and the protein expressions of HIF-1α, Bax, and Bcl-2 by Western blot. Results At 24 hours after culture, small amount of vacuoles necrotic cells could be observed in group A and group B; there was no significant difference in apoptosis rate between groups A and B (t=1.026,P=0.471), and HIF-1α mRNA and protein expressions in group B were significantly higher than those in group A (t=22.672,P=0.015;t=18.396,P=0.013), but, there was no significant difference in protein expressions of Bax and Bcl-2 between groups A and B (t=0.594,P=0.781;t=1.251,P=0.342). The number of vacuolar necrosis cells in group D was significantly higher than that in group C, and HIF-1α positive cells were observed in group D. The apoptosis rate of group D was significantly higher than that of group C (t=27.143,P=0.002). The mRNA expressions of HIF-1α, COL II, Aggrecan, and SOX9 in group D were significantly lower than those in group C (t=21.097,P=0.015;t=34.829,P=0.002;t=18.673,P=0.022;t=31.949,P=0.007). The protein expressions of HIF-1α and Bcl-2 in group D were significantly lower than those in group C (t=37.648,P=0.006;t=16.729,P=0.036), but the protein expression of Bax in group D was significantly higher than that in group C (t=25.583,P=0.011). Conclusion HIF-1α mRNA expression is up-regulated under hypoxia condition, which will increase the hypoxia tolerance of endplate chondrocytes. Cell apoptosis is suppressed by the activation of HIF-1α in endplate chondrocytes under hypoxia condition.
Objective： To observe the effect of estrogen on the expr ession of pigment epithelium derived factor (PEDF) in cultured retinal Muuml;ller cells under the anoxic condition． Methods：After the anoxic retinal Muuml;ll er cells were tre ated with estrogen (E2) with the concentration of 10-6、10-5 and 10-7 mmol/L, t he level of expression of PEDF mRNA and the protein was detected by reverse tran scriptionpolymerase chain reaction and Western blotting analysis. Results：Th e expression of PEDF mRNA and protein decreased 24 hours after anoxia. E2 with t he concentration of 10-5 and 10-6 mmol/L inhibited the decrease of expression of PEDF mRNA and protein induced by anoxia, which related to the concentration of E2． Conclusion：strogen can regulate the expression of PEDF, which ma y play an important role in the regulation of retinal neovascularization．
ObjectiveTo investigate the effects of hypoxia inducible factor 1α (HIF-1α) overexpression on the differentiation of stem cells derived from human exfoliated deciduous teeth (SHED) into vascular endothelial cells.MethodsSHED was isolated from the retained primary teeth donated by healthy children by using collagenase digestion method. The third generation cells were identified by flow cytometry and alizarin red and alkaline phosphatase (ALP) staining after osteogenic differentiation culture. The SHED were divided into blank control group (SHED without any treatment), empty group (SHED infected with empty lentivirus), HIF-1α overexpression group (SHED infected with HIF-1α overexpression lentivirus), Wnt inhibitor group (SHED interfered by IWR-1), and combination group (HIF-1α overexpressed SHED interfered by IWR-1). Real-time fluorescence quantitative PCR (qRT-PCR) and Western blot were used to analyze the expressions of HIF-1α mRNA and protein in the SHED of blank control group, empty group, and HIF-1α overexpression group. Then the SHED in 5 groups were induced differentiation into vascular endothelial cells for 14 days. The expressions of cell surface marker molecule [von Willebrand factor (vWF) and CD31] were detected by flow cytometry. The mRNA expressions of vascular cell adhesion protein 1 (VCAM-1), KDR (Kinase-inserted domain containing receptor), and VE-cadherin (VE) were analyzed by qRT-PCR. The protein expressions of phosphate-glycogen synthasc kinase 3β (p-GSK3β) and β-catenin were analyzed by Western blot. The tube forming ability of induced cells was detected by Matrigel tube forming experiment. The ability of endothelial cells to phagocytic lipid after differentiation was detected by DiI-labeled acetylated low density lipoprotein (DiI-Ac-LDL) phagocytosis.ResultsAfter identification, the cells were SHED. After lentivirus transfection, compared with the blank control group and the empty group, the expressions of HIF-1α mRNA and protein in the HIF-1α overexpression group increased significantly (P<0.05). Compared with the blank control group and the empty group, the expressions of VCAM-1, KDR, and VE mRNA, the percentages of vWF positive cells and CD31 positive cells, and the relative expression of β-catenin protein were significantly higher (P<0.05), the relative expression of p-GSK3β protein was significantly lower (P<0.05), the number of tubules formed and the ability to phagocytic lipids significantly increased (P<0.05) in the HIF-1α overexpression group; while the indicators in the Wnt inhibitor group were opposite to those in the HIF-1α overexpression group (P<0.05). Compared with the HIF-1α overexpression group, the expressions of VCAM-1, KDR, and VE mRNA, the percentages of vWF positive cells and CD31 positive cells, and the relative expression of β-catenin protein were significantly lower (P<0.05), the relative expression of p-GSK3β protein was significantly higher, and the number of tubules formed and the ability of phagocytosis of lipids significantly reduced, showing significant differences between groups (P<0.05).ConclusionOverexpression of HIF-1α can promote SHED to differentiate into vascular endothelial cells by activating Wnt/β-catenin signaling pathway.
Objective To investigate the protection on the intrahepatic cholangiocyte mediated by hypoxic preconditioning (HP) after liver transplantation and the role of vascular endothelial growth factor (VEGF). Methods The model of autologous liver transplantation was established, and the rats were divided into 3 groups: autologous liver transplantation group, hypoxic preconditioning before operation group (HP group) and sham operation group. At 6, 12, 24, 48 h after operation, blood samples were collected for examination of the serum total bilirubin (TBIL), direct bilirubin (DBIL) and alkaline phosphatase (ALP), and the expression of VEGF was detected by immunohistochemical method. The pathological changes of cholangiocytes were observed by light microscope. Results As compared with autologous liver transplantation group, the levels of seurm TBIL, DBIL and ALP in HP group were lower (P＜0.05), while the expression of VEGF in HP group was higher at the whole process (P＜0.05). The degrees of billiary epithelium damage and inflammatory infiltration in autologous liver transplantation group were more severe than those in HP group. Conclusion HP has protective effect on cholangiocytes after liver transplantation, in which VEGF may play an important role.
ObjectivesTo systematically review the risk factors of postoperative hypoxemia in patients undergoing coronary artery bypass grafting.MethodsPubMed, EBCO, The Cochrane Library, CNKI, VIP and WanFang Data databases were electronically searched to collect case-control studies and cohort studies on the risk factors of postoperative hypoxemia in patients undergoing coronary artery bypass grafting from inception to December 2018. Two reviewers independently screened literature, extracted data and assessed risk of bias of included studies, then, meta-analysis was performed by using RevMan 5.3 software.ResultsA total of 20 articles were included, including 3 926 patients. The results of meta-analysis showed that: age (OR=2.94, 95%CI 0.81 to 5.07, P=0.007), body mass index (OR=1.94, 95%CI 0.77 to 3.12, P=0.001), smoking (OR=2.72, 95%CI 1.68 to 4.42, P<0.000 1), diabetes history (OR=1.63, 95%CI 1.37 to 1.93, P<0.000 01), preoperative lung diseases (OR=4.11, 95%CI 1.64 to 10.28, P=0.003), complicated ventricular aneurysm (OR=1.57, 95%CI 1.12 to 2.21, P=0.01), left ventricular end-diastolic diameter (OR=1.28, 95%CI 0.12 to 2.44, P=0.03), aortic occlusion time (OR=13.25, 95%CI 4.93 to 21.57, P=0.002), operation time (OR=9.33, 95%CI 5.36 to 13.30, P<0.000 01), number of bypass branches (OR=0.19, 95%CI 0.02 to 0.36, P=0.03), intraoperative infusion volume (OR=383.46, 95%CI 282.16 to 484.76, P<0.000 01) and postoperative pulmonary infection (OR=6.00, 95%CI 3.83 to 9.42, P<0.000 01) were the risk factors for postoperative hypoxemia in patients undergoing coronary artery bypass grafting. Preoperative ejection fraction (OR=−2.60, 95%CI −4.56 to −0.64, P=0.009) and preoperative partial oxygen pressure (OR=−3.14, 95%CI −4.72 to −1.56, P=0.000 1) were the protective factors for postoperative hypoxemia.ConclusionsCurrent evidence shows that age, body mass index, smoking, diabetes history, preoperative lung diseases, complicated ventricular aneurysm, left ventricular end-diastolic diameter, aortic occlusion time, operation time, number of bypass branches, intraoperative infusion volume and postoperative pulmonary infection are risk factors for postoperative hypoxemia in patients undergoing coronary artery bypass grafting. Due to limited quality and quantity of included studies, the above conclusion is required to be assessed by further studies.
Objective To observe the effects of peritoneal ventilation with pure oxygen in the rabbits with hypoxaemia and hypercapnia induced by mechanical controlled hypoventilation. Methods Sixteen rabbits were invasively ventilated after trachea incision. Hypoxaemia and hypercapnia were induced by hypoventilation which was implemented both by degrading ventilation parameters and respiratory depression induced by intravenous infusion of muscle relaxant. Then pure oxygen was insufflated into the peritoneal cavity and arterial blood gases were measured every 30 minutes for two hours. Results The PaO2 was ( 52. 50 ±3. 46) mmHg at baseline and increased to ( 76. 46 ±7. 79) mm Hg, ( 79. 62 ±9. 53) mm Hg,( 78. 54 ±7. 18) mmHg, and ( 81. 1 ±8. 3) mm Hg, respectively at 30, 60, 90, and 120 minutes after the peritoneal ventilation with pure oxgen( all P lt; 0. 05) . Meanwhile PaCO2 was ( 63. 84 ±9. 09) mm Hg at baseline and ( 59. 84 ±14. 22) mmHg, ( 59. 16 ±15. 5) mmHg, ( 60. 02 ±7. 07) mmHg, and ( 61. 38 ±6. 56) mm Hg, respectively at 30, 60, 90, and 120 minutes after the peritoneal ventilation with pure oxgen with no significant change( P gt;0. 05) . Conclusion Peritoneal ventilation can obviously improve hypoxaemia induced by mechanical controlled hypoventilation, whereas hypercapnia remains unchanged.
ObjectiveTo investigate the expression and significance of CD73 in rats with intermittent hypoxia and high fat diet.MethodsThe rat model of chronic intermittent hypoxia combined with high fat diet was established. Twenty-four healthy male Wistar rats in the SPF level were randomly divided into 4 group, with 6 rats in each group, namely group A (normoxia and normal diet), group B (normoxia and high fat diet), group C (intermittent hypoxia and normal diet)and group D (intermittent hypoxia and high-fat diet). After 6 weeks of experiment, the serum lipid levels, myocardial morphological changes under microscope, the expression level of CD73 protein detected byimmunohistochemistry and Western blot in myocardial cells in rats were compared among these groups.ResultsThe serum lipid levels were significantly different among these groups (P<0.05). HE results showed that the myocardial cells of group A had no obvious abnormalities; disorganized visible myocardial fibers with focal necrosis in groups B and C; myocardial cell injury was most obvious in group D, in which visible muscle fibers arranged in disorder, and grain was not clear, part of the muscle fibers were dissolved predominantly. Compared with group A, CD73 protein expression levels in myocardial cells in groups B, C, and D were significantly elevated (P<0.01). Furthermore, CD73 protein expression level in myocardial cells in group D was significantly higher than those in groups B and C (P<0.01). Western blot showed consistent results as immunohistochemistry: compared with group A, CD73 protein expression levels in groups B, C, and D were significantly elevated (P<0.05), and CD73 protein expression level in myocardial cells in group D was significantly higher than those in groups B and C (P<0.01).ConclusionChronic intermittent hypoxia and high fat diet can cause myocardial cell damage and upregulate CD73 expression in the cardiomyocytes.