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1.
AIM To investigate the effect of sinomenine (SN) on the damage of human neuroblastoma SK-N-SH cells induced by 1-methyl-4-4 phenylpyridine (MPP+) and its mechanism for exploring the pathogenesis of Parkinson disease. METHODS SN was used to treat MPP+-induced SK-N-SH cells. The levels of malondialdehyde (MDA) and glutathione (GSH) in cell culture supernatants were measured by ELISA. The apoptosis was analyzed by flow cytometry. The protein expression levels of Bcl-2 and Bax were determined by Western blot. The expression levels of long noncoding RNA ANRIL and microRNA-626 (miR-626) were detected by RT-qPCR. Dual-luciferase reporter assay was used to evaluate the relationship between ANRIL and miR-626. After ANRIL small interfering RNA was transfected into SK-N-SH cells, the effects of ANRIL expression knock-down on MPP+-induced SK-N-SH cell apoptosis, the protein expression levels of Bcl-2 and Bax, and the levels of MDA and GSH in cell culture supernatants were examined. RESULTS After treatment with MPP+, the apoptotic rate, Bax protein level and ANRIL expression in SK-N-SH cells were increased (P<0.05), and the Bcl-2 protein level and miR-626 expression were decreased (P<0.05). The level of MDA in cell culture supernatants was increased (P<0.05), and the level of GSH was decreased (P<0.05). After SN treatment or ANRIL expression knock-down, decreased apoptotic rate, Bax protein level and ANRIL expression (P<0.05), and increased Bcl-2 protein level and miR-626 expression in MPP+-induced SK-N-SH cells were observed (P<0.05). The level of MDA in the cell culture supernatants was decreased (P<0.05), and the level of GSH was increased (P<0.05). CONCLUSION SN attenuates MPP+-induced damage in SK-N-SH cells by regulating ANRIL/miR-626 signaling pathway.  相似文献   

2.
AIM: To investigate the neuroprotective effect of progesterone against adenosine triphosphate (ATP)-injured human neuroblastoma SH-SY5Y cells.METHODS: The SH-SY5Y cells in the logarithmic phase were divided into different groups according to the progesterone and ATP concentrations. The cell viability was measured by CCK-8 assay. The membrane permeability was detected using fluorescent dye YO-PRO-1. Cytosolic Ca2+ concentration was measured with fluorescent dye Fluo-3/AM. The expression of purinergic P2X7 receptor was assessed by Western blot.RESULTS: The viability of the SH-SY5Y cells was significantly decreased (P<0.05) and YO-PRO-1 uptake was obviously increased (P<0.05) in a concentration-dependent manner compared with control group when SH-SY5Y cells were treated with ATP at 1, 3, 5 and 7 mmol/L for 2 h. The viability reduction of the SH-SY5Y cells induced by ATP was obviously counteracted by treatment with progesterone at 3, 10 and 30 nmol/L for 30 min (P<0.05) as compared with ATP group. YO-PRO-1 fluorescence enhancement induced by ATP in SH-SY5Y cells was significantly reduced (P<0.05) by progesterone (30 nmol/L) or P2X7 receptor antagonist KN-62 (500 nmol/L) pretreatment for 30 min, and no obvious difference between treatments with progesterone and KN-62 was observed. Cytosolic Ca2+ fluorescence intensity in normal group was a little, but that in ATP group was increased (P<0.05). Progesterone or KN-62 pretreatment significantly decreased the cytosolic fluorescence intensity of Ca2+ induced by ATP (P<0.05). However, no obvious difference between treatments with progesterone and KN-62 was found. The expression of P2X7 receptor in ATP group was significantly higher than that in control group (P<0.05), and progesterone inhibited ATP-induced P2X7 receptor expression (P<0.05).CONCLUSION: Progesterone inhibits P2X7 receptor expression, membrane pore formation, intracellular Ca2+ increase and cell death induced by ATP, so progesterone may protect SH-SY5Y cells against ATP-induced injuries.  相似文献   

3.
AIM: To investigate the protective effect of astragaloside IV (ASIV) on angiotensin II (Ang II)- induced apoptosis of H9c2 cardiomyocytes. METHODS: H9c2 cardiomyocytes were treated with different concentrations of Ang II and ASIV. The effects of Ang II and ASIV on the viability of H9c2 cells was measured by CCK-8 assay. The optimum concentration of Ang II was 1 μmol/L and the concentrations of ASIV were 25, 50 and 100 μmol/L. The H9c2 cells was divided into 6 groups:control group, ASIV group, Ang II group, Ang II+ASIV (25 μmol/L) group, Ang II+ASIV 50 (μmol/L) group and Ang II+ASIV (100 μmol/L) group. The morphological changes of the H9c2 cells were observed under inverted phase-contrast microscope. Apoptosis was detected by TUNEL assay. The generation of reactive oxygen species (ROS) was detected by DCFH-DA staining. The protein expression of Bax, Bcl-2, nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) was determined by Western blot. H9c2 cardiomyocytes were transfected with negative control shRNA (NC) or Nrf2-shRNA (shRNA), and the cells were divided into 8 groups:NC+control group, NC+AngⅡgroup, NC+ASIV group, NC+AngⅡ+ASIV group, shRNA+control group, shRNA+AngⅡgroup, shRNA+ASIV group and shRNA+AngⅡ+ASIV group. ROS level was detected by ROS detection kit. The protein expression of Nrf2 and HO-1 was determined by Western blot. RESULTS: Ang II decreased the viability of H9c2 cells in a concentration-dependent manner (P<0.05). ASIV reversed the effect of Ang II on the viability of H9c2 cells in a concentration-dependent manner (P<0.05). Compared with control group, the apoptotic rate, the level of ROS and the protein expression of Bax in Ang II group were increased significantly, while the protein expression of Bcl-2, Nrf2 and HO-1 was decreased significantly (P<0.05). Compared with Ang II group, ASIV reversed the increase in apoptotic rate of H9c2 cells induced by Ang II in a concentration-dependent manner, reduced ROS level, down-regulated the protein expression of Bax and up-regulated the protein expression of Bcl-2, Nrf2 and HO-1 (P<0.05). After shRNA transfection, the effects of ASIV decreasing ROS production induced by Ang II and up-regulating the expression of Nrf2 and HO-1 were eliminated. CONCLUSION: ASIV protects H9c2 cardiomyocytes from apoptosis induced by Ang II, which may be related to reducing ROS generation and mediating the Nrf2/HO-1 signaling pathway.  相似文献   

4.
AIM: To investigate the effect of triptolide on the inhibition of microglial activation in 1-methyl-4-phenyl pyridinium (MPP+)-induced hemiparkinson disease rats.METHODS: The rat model of Parkinson disease was established by intranigral injection of MPP+. The rats were randomly divided into sham group, MPP+ group, triptolide group and vehicle group. The survival of dopaminergic neurons was detected by the immunofluorescence of tyrosine hydroxylase (TH) in the substantia nigra (SN). The activation of microglia was determined by immunofluorescence of OX-42 (microglia marker) in the SN. The expression of chemokine receptor CX3CR1 in SN was measured by Western blotting.RESULTS: Intranigral injection of MPP+ increased the fluorescence intensity of the microglial marker, and promoted DA neuron degenerative death. Immunohistological analysis showed that the OX-42 density was decreased (P<0.01) and tyrosine hydroxylase (TH) positive neurons were increased in the triptolide group (P<0.01). The expression of CX3CR1 was lower in triptolide group than that in model group (P<0.05).CONCLUSION: Triptolide may improve PA neurons function in MPP+-induced rats through inhibiting CX3CR1 expression and microglial activation.  相似文献   

5.
AIM: To investigate the effects of aliskiren on the injury of SH-SY5Y cells induced by oxygen-glucose deprivation (OGD) and its possible mechanisms. METHODS: The SH-SY5Y cells were randomly divided into control group, OGD group and aliskiren (5.0, 10.0 and 20.0 μmol/L) groups. The cell viability was measured by CCK-8 assay. The levels of excitatory amino acid transporter 2 (EAAT2/GLT-1), EAAT3/EAAC1, EAAT4, endothelin-1 (ET-1) and S100 calcium-binding protein β subunit (S-100β) in the SH-SY5Y cells were detected by ELISA. The morphological changes of the cells were observed by Hoechst 33258 staining. Meanwhile, the content of lactic acid (LD) and activity of Na+-K+-ATPase were also analyzed. RESULTS: The viability of SH-SY5Y cells was not more than 60% after OGD injury for 4 h, so the appropriate time for OGD injury was 4 h. Compared with control group, the protein levels of GLT-1, EAAC1 and EAAT4 in the SH-SY5Y cells of OGD group were significantly decreased (P<0.05), but the protein levels of ET-1 and S-100β were significantly increased (P<0.05). Compared with OGD group, treatment with aliskiren dose-dependently increased the protein levels of GLT-1, EAAC1 and EAAT4 in the SH-SY5Y cells, but decreases in the levels of ET-1 and S-100β were observed (P<0.05). The results of Hochest 33258 staining showed that aliskiren significantly reduced the apoptosis of SH-SY5Y cells. Compared with control group, a significant increase in the content of LD (P<0.05) and a significant decrease in Na+-K+-ATPase activity (P<0.05) were found in the SH-SY5Y cells of OGD group. Compared with OGD group, aliskiren dose-dependently decreased the content of LD, but increased the Na+-K+-ATPase activity in the SH-SY5Y cells (P<0.05). CONCLUSION: Aliskiren has good neuroprotective effects on SH-SY5Y cells after OGD injury. The underlying mechanisms may be associated with the increases in the protein levels of GLT-1, EAAC1 and EAAT4, the enhancement of Na+-K+-ATPase activity, and the decreases in the levels of ET-1 and S-100β and the content of LD.  相似文献   

6.
AIM: To investigate the effects of salidroside on intracellular free calcium concentration [Ca2+]i, apoptosis, mitochondrial membrane potential (MMP) and activity during injury induced by hypoxia/hypoglycemia in cultured SH-SY5Y cells. METHODS: Mitochondrial activity was measured by methylthiazolyl tetrazolium test. MMP,[Ca2+]i and apoptosis were measured by flow cytometry. RESULTS: SH-SY5Y cells were cultured in a hypoxia/hypoglycemia condition for 2, 4, 6 and 12 h,[Ca2+]i and apoptosis rate significantly increased compared with control group (P<0.01). After hypoxia /hypoglycemia cultures, MMP and mitochondrial activity declined 29.17% (P<0.01) and 38.80% (P<0.01) at 2 h, 56.72% (P<0.01) and 63.58% (P<0.01) at 12 h, were lower than that in control group (P<0.01). Salidroside significantly decreased [Ca2+]i and apoptosis rate, and increased MMP and mitochondrial activity in hypoxia /hypoglycemia-treated SH-SY5Y cells. CONCLUSIONS: Salidroside might inhibit the decline in MMP and mitochondrial activity induced by hypoxia /hypoglycemia, and has an inhibitory effects on neuronal apoptosis. The mechanism might be related to inhibiting intracellular calcium overload.  相似文献   

7.
AIM:To investigate the inhibitory effect of thioredoxin 1 (Trx-1) over-expression on oxidative stress injury in 1-methyl-4-phenylpyridinium (MPP+)-induced rat pheochromocytoma PC12 cells by regulating NF-κB signaling pathway.METHODS:The PC12 cells were damaged by treatment with MPP+ at 1, 3 and 5 mmol/L, and the optimal concentration of 3 mmol/L was selected. The cell viability was measured by MTT assay. The oxidative stress indexes lactate dehydrogenase (LDH) activity, superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in the cell culture supernatant were detected, and the protein expression of Trx-1 was determined by Western blot. Lentiviral infection with Ad-Trx-1-GFP sequence was used to establish a model of MPP+-treated PC12 cells with Trx-1 over-expression. The effects of Trx-1 over-expression on the cell viability, oxidative stress responses and NF-κB signaling pathway were determined by MTT assay, commercial kits and Western blot. The effects of phorbol 12-myristate 13-acetate (PMA), an activator of NF-κB signaling pathway, on the viability and oxidative stress of PC12 cells were observed. The NF-κB signaling pathway inhibitor pyrrolidine dithiocarbamate (PDTC) was used in MPP+-treated PC12 cells with Trx-1 over-expression, and the cell viability and oxidative stress responses were measured. RESULTS:The viability of PC12 cells, SOD activity in the supernatant and protein expression of Trx-1 were decreased, while LDH activity and MDA content in the supernatant were increased significantly by treatment with MPP+ at 1, 3 and 5 mmol/L. The effect of MPP+ at 3 mmol/L and 5 mmol/L was significantly greater than that at 1 mmol/L (P<0.05), and no significant difference between 3 mmol/L and 5 mmol/L was observed (P>0.05). The inhibitory effect of MPP+ on the viability of PC12 cells, and the oxidative stress injury and activation of NF-κB signaling pathway induced by MPP+ were significantly attenuated by over-expression of Trx-1. The inhibitory effect of MPP+ on the viability of PC12 cells and the oxidative stress injury induced by MPP+ were promoted by the activation of NF-κB signaling pathway, while the protective effects of Trx-1 over-expression on the MPP+-treated PC12 cells were enhanced by the inhibition of NF-κB signaling pathway. CONCLUSION:Over-expression of Trx-1 protects MPP+-treated PC12 cells from oxidative stress injury by regulating NF-κB signaling pathway.  相似文献   

8.
AIM:To investigate the effect of erianin on the viability and apoptosis of human lung cancer A549 cells and its possible mechanism. METHODS:A549 cells and BEAS-2B cells were cultured in vitro and treated with erianin at 10, 20, 40, 80 and 160 nmol/L for 48 h. The cell viability was measured by CCK-8 assay. The apoptosis and reactive oxygen species (ROS) were analyzed by flow cytometry. The activity of superoxide dismutase (SOD) was detected by WST-8 method, and the content of malondialdehyde (MDA) was detected by barbituric acid method. The protein levels of nuclear factor E2-related factor 2 (Nrf2), NAD(P)H:quinone oxidoreductase-1 (NQO1), heme oxygenase-1 (HO-1), p38 MAPK, p-p38 MAPK, caspase-3 and cleaved caspase-3 were determined by Western blot.RESULTS:Erianin remarkably reduced the viability of A549 cells in a dose-dependent manner (P<0.05) with IC50 at 52.64 nmol/L. Erianin also induced apoptosis (P<0.05), increased ROS level and MDA content (P<0.05), diminished SOD activity (P<0.05), and down-regulated the protein levels of Nrf2, NQO1 and HO-1 (P<0.05), in a dose-dependent manner. Meanwhile, erianin up-regulated the levels of p-p38 MAPK and cleaved caspase-3 (P<0.05), and these effects were inhibited by N-acetyl-L-cysteine and SB203580 (P<0.05).CONCLUSION:Erianin may induce apoptosis of human lung cancer A549 cells most likely via inhibiting SOD activity and down-regulating the protein levels of Nrf2, NQO1 and HO-1, thus resulting in an increase in ROS and activation of p38 MAPK.  相似文献   

9.
AIM: To investigate the effect of nuclear factor E2-related factor 2 (Nrf2) activation by 18α-glycyrrhetinic acid (18α-GA) on the proliferation and self-renewal of adult neural stem cells (aNSCs), and to explore an effective way of maintaining the viability of aNSCs. METHODS: NSCs were dissociated from subventricular zone of the mice at postnatal days 0, 60, and 300. The expression levels of Nrf2 in the NSCs at various ages were compared. After treatment with 18α-GA, the expression of Nrf2 was examined by real-time PCR and Western blot. shRNA lentiviral vector (LV) carrying green fluorescent protein (GFP) gene was constructed to knock down Nrf2 expression. The knockdown efficiency in the aNSCs was detected by real-time PCR and Western blot. Subsequently, the aNSCs were divided into DMSO group, 18α-GA group, LV-GFP group and LV-Nrf2-shRNA group. BrdU incorporation assay, Tuj1 staining, CCK-8 assay, Hoechst 33342/PI staining and detection of reactive oxygen species (ROS) were performed to analyze the proliferation, differentiation, viability, apoptosis and oxidative stress levels of the NSCs. RESULTS: The mRNA expression level of Nrf2 in adult and aged NSCs was significantly lower than that in newborn NSCs (P<0.01), while the ROS level of aNSCs was significantly higher (P<0.05). After treatment with 18α-GA, the expression level of Nrf2 in the aNSCs was significantly up-regulated as compared with DMSO group (P<0.01). Increased number of BrdU+ and Tuj1+ cells was observed in 18α-GA group, indicating that 18α-GA-treated cells had higher viability (P<0.05). Meanwhile, there were fewer apoptotic cells and lower ROS level in 18α-GA group than those in DMSO group (P<0.05). After knockdown of Nrf2 in aNSCs and then treated with 18α-GA, there were less BrdU+ and Tuj1+ cells, as well as the aNSCs with lower viability in LV-Nrf2-shRNA group (P<0.05). Moreover, the ROS level was increased in LV-Nrf2-shRNA group as compared with LV-GFP group (P<0.05). CONCLUSION: Activation of Nrf2 by 18α-GA elevates the antioxidant capacity of aNSCs, thus ameliorating the cell proliferation and differentiation potentials.  相似文献   

10.
AIM: To observe the effect of rapamycin (Rapa) on human neuroblastoma SH-SY5Y cell injury induced by oxygen-glucose deprivation (OGD), and to explore the role of autophagy in this process. METHODS: The SH-SY5Y cells were randomly divided into 4 groups:normal control group:the cells were cultured without OGD treatment; Rapa group:the cells were pretreated with Rapa for 1 h; OGD group:the culture medium was replaced by glucose-free medium and the cells were transferred to a humidified incubation chamber flushed by a gas mixture of 1% O2, 94% N2 and 5% CO2 for 12 h; Rapa+OGD group:the cultured cells were treated with Rapa for 1 h, and then were given the same treatments as those in OGD group. The cell viability was assessed by MTT assay. The degree of the cell damage was evaluated by determining the leakage of lactate dehydrogenase (LDH). The enzyme activity of caspase-3 was detected. TUNEL staining were used to detect the variation of cell apoptosis. The protein levels of apoptosis-related proteins Bax and Bcl-2, autophagy-related protein beclin-1 and autophagy marker protein LC3B were determined by Western blot. RESULTS: Compared with OGD group, the viability of the SH-SY5Y cells was significantly increased, and the activity of caspase-3 was significantly reduced in Rapa+OGD group (P<0.05). The SH-SY5Y cell injury was apparent after OGD with a great increase in the apoptotic rate (P<0.05). Compared with OGD group, the apoptotic rate significantly decreased in Rapa+OGD group (P<0.05). Compared with control group, the protein level of Bcl-2 was significantly decreased (P<0.05) and the protein level of Bax was significantly increased in OGD group. Compared with OGD group, the levels of Bcl-2, beclin-1 and LC3B-Ⅱ were significantly increased and the protein level of Bax was significantly increased in Rapa+OGD group (P<0.05). CONCLUSION: Rapamycin has a protective effect on in vitro cultured SH-SY5Y cells injured by OGD. The mechanism may be related to the promotion of autophagy.  相似文献   

11.
AIM: To observe the effect of docosahexaenoic acid(DHA) on H2O2-induced apoptosis in human retinal pigment epithelium cells and its molecular mechanism. METHODS: Human retinal pigment epithelium cell line ARPE-19 was cultured in vitro, and 12.5 mmol/L H2O2 was used to mimic the oxidative stress condition. The cells were treated with 30~100μmol/L DHA for 4~24 h. The expression of heme oxygenase-1(HO-1) at mRNA and protein levels was detected by real-time PCR and Western blot, respectively. The enzymic activity of HO-1 was measured by colorimetry. Production of reactive oxygen species(ROS) was determined by fluorescent probe. Activation of NF-E2-related factor 2(Nrf2) was examined by immunofluorescence method. Apoptosis of ARPE-19 cells was analyzed by flow cytometry. RESULTS: The mRNA and protein expression and the enzymic activity of HO-1 were significantly increased in the ARPE-19 cells after DHA treatment. Meanwhile, nuclear translocation of Nrf2 was also observed. Apoptosis appeared and ROS was produced upon H2O2 incubation. In contrast, DHA at 100μmol/L significantly abrogated H2O2-induced apoptosis and ROS production. Furthermore, silencing of HO-1 by specific siRNA, or treatment with ZnPP, an inhibitor of HO-1, partly counteracted the protective effect against H2O2-induced apoptosis and ROS production. CONCLUSION: DHA protects retinal pigment epithelial cells against oxidative stress via induction of heme oxygenase-1 expression after Nrf2 activation.  相似文献   

12.
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13.
AIM: To investigate the protective effects of glucagan-like peptide-1 (GLP-1) on cardiac microvascular injury in diabetic rats and the underlying mechanism. METHODS: Diabetic rats were induced by intraperitoneal injection of streptozocin, and then randomized to 3 months of vehicle or exenatide (a GLP-1 analogue) treatment. Before and after the treatment, body weight, blood glucose and blood pressure were measured. Cardiac microvascular permeability was detected by transmission electron microscopy. Cardiac microvascular endothelial cells (CMECs) were isolated and cultured in normal glucose (5.5 mmol/L), high glucose (25 mmol/L), and high glucose plus GLP-1 (10-8 mol/L). The production of reactive oxygen species (ROS) was examined by superoxide assay kit and dihydroethidium staining. The protein expression of GLP-1 receptor (GLP-1R), Kelch-like epichlorohydrin-associated protein 1 (Keap1), nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) was determined by Western blot. RESULTS: Exenatide treatment for 3 months improved the cardiac microvascular permeability in the diabetic rats. The GLP-1R was expressed in CMECs. GLP-1 inhibited high glucose-induced ROS production (P<0.05). Compared with high glucose group, the protein expression of Keap1 was decreased, and Nrf2 and HO-1 were increased significantly (P<0.05). CONCLUSION: GLP-1 inhibits oxidative stress in high glucose-induced CMECs, and improves the cardiac microvascular permeability in diabetic rats. The protective effects of GLP-1 may be related to Keap1-Nrf2 signaling pathway.  相似文献   

14.
AIM: To study the relaxation effect of isoliensinine on high K+-induced isolated mouse airway smooth muscle (ASM) and the underlying mechanism. METHODS: The muscle tension transducer was used to detect the effects of isoliensinine on high K+-induced precontraction and Ca2+ influx in ASM. The technique of patch-clamp and calcium imaging system were respectively used to examine the effects of isoliensinine on LVDCC currents and[Ca2+]i of the ASM cells (ASMCs). RESULTS: Isoliensinine significantly relaxed precontracted ASM induced by high K+ in a concentration-dependent manner. The maximum relaxation ratio was(95.3±3.9)% by isoliensinine at 100 μmol/L. In addition, LVDCC currents were measured using the whole-cell patch-clamp technique, which were abolished by isoliensinine. High K+-induced 340/380 nm fluorescence ratio of Fura-2 was 0.63±0.10 in ASMCs, while it decreased to 0.36±0.05 after the addition of isoliensinine (P<0.01). When isoliensinine was added at the peak point of[Ca2+]i, the ratio rapidly decreased from 0.74±0.02 to 0.42±0.05 (P<0.01). Moreover, isoliensinine inhibited high K+-induced Ca2+ influx-mediated contraction of ASM. CONCLUSION: Isoliensinine inhibits LVDCC currents, terminates Ca2+ influx and reduces[Ca2+]i, eventually resulting in relaxation of the ASM, indicating isoliensinine might be a potential bronchodilator.  相似文献   

15.
AIM: To explore the effect of Yiqi-Yangyin recipe on myocardial ischemia-reperfusion injury (MIRI) in rats with diabetes mellitus (DM) and the possible mechanism. METHODS: The rats were divided into normal group (control group), DM sham operation (DM-S) group, DM+MIRI group, low-, medium-and high-dose Yiqi-Yang-yin recipe (TL, TM and TH) groups (7.5, 15 and 30 g/kg decoction of Yiqi-Yangyin recipe by gavage), and Nrf2 inhibitor (bardoxolone methyl) group (30 mg/kg bardoxolone methyl by intragastric administration). The gavage volume was 1 mL/kg. There were 15 rats in each group, and they were administered continuously for 7 d. The tail vein blood was collec-ted after the last administration to detect the blood sugar and lipid levels in the rats. The serum levels of cardiac troponin I (cTnI), tumor necrosis factor-α (TNF-α), interleukin (IL)-1β and IL-10 were measured by ELISA. Echocardiography was used to detect the changes of cardiac function in the rats after blood collection. After cardiac function test, the rats were sacrificed to obtain cardiac tissues, and the volume changes of myocardial infarction were assessed by triphenylte-trazole chloride staining. The histopathological changes of myocardium was observed by HE staining. The cardiomyocyte apoptosis was determined by TUNEL assay. The protein levels of phosphorylated extracellular signal-regulated kinase (p-ERK), nuclear factor E2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) in the myocardium were determined by Western blot. The myocardial activity of superoxide dismutase (SOD) was measured by nitro blue tetrazolium method, the content of malondialdehyde (MDA) was tested by thiobarbituric acid method, and the production of reactive oxygen species (ROS) was analyzed by iron ion reduction method. RESULTS: Compared with control group, the levels of fasting blood glucose (FBG), total cholesterol (TC) and triglyceride (TG) in DM-S group and DM+MIRI group were significantly elevated, while the level of high-density lipoprotein cholesterol (HDL-C) was significantly lowered (P<0.05). Compared with DM-S group and DM+MIRI group, the levels of FBG, TC, TG in TL, TM, TH and bardoxolone methyl groups were significantly decreased, while HDL-C level was significantly increased (P<0.05). Compared with control group and DM-S group, heart rate (HR) and left ventricular end-diastolic pressure (LVEDP) were increased in DM+MIRI group, mean arterial pressure (MAP), left ventricular systolic pressure (LVSP) and left ventricular ejection fraction (LVEF) were decreased, serum levels of cTnI, TNF-α, IL-1β and IL-10 were increased, the myocardial infarction volume percentage was increased, the myocardial cell breakage and necrosis were increased, the myocardial cell apoptotic rate was increased, the protein levels of p-ERK1/2, Nrf2 and HO-1 were decreased, MDA and ROS levels were increased, and the activity of SOD was decreased (P<0.05). Compared with DM+MIRI group, HR and LVEDP were decreased in TL, TM, TH and bardoxolone methyl groups, MAP, LVSP and LVEF were increased, the serum levels of cTnI, TNF-α, IL-1β and IL-10 were decreased, the myocardial infarction volume percentage was decreased, myocardial cell breakage and necrosis were decreased, myocardial cell apoptotic rate was decreased, the protein levels of p-ERK1/2, Nrf2 and HO-1 were increased, the MDA and ROS levels were decreased, and the activity of SOD was increased (P<0.05). CONCLUSION: Yiqi-Yangyin recipe protects the myocardial tissue of DM+MIRI rats from injury and reduces the oxidative stress level, which may be achieved by activating ERK/Nrf2/HO-1 pathway.  相似文献   

16.
AIM: To investigate the role of hydrogen molecule on apoptosis-related proteins in glomerular mesangial cells cultured with high glucose and to explore its possible mechanism. METHODS: Mouse glomerular mesangial cells cultured in vitro were divided into 4 groups:normal control group (C group, 5.5 mmol/L glucose), mannitol group (G group, 5.5 mmol/L glucose+19.5 mmol/L mannitol), high glucose group (H group, 25 mmol/L glucose), high glucose+hydrogen-rich water group (HH group, 25 mmol/L glucose+hydrogen-rich water), and cultured for 48 h. The protein levels of Bax, Bcl-2, cleaved caspase-3, nuclear factor E2-related factor-2 (Nrf2), heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase-1 (NQO-1) were determined by Western blot, and the mRNA expression of HO-1 and NQO-1 was determined by RT-PCR. The level of intracellular reactive oxygen species (ROS) was detected by dihydroethidium method, and the activity of superoxide dismutase (SOD) was measured by WST-8 assay. RESULTS: Compared with C group, the protein levels of Bax and cleaved caspase-3 were up-regulated, and Bcl-2 was down-regulated in H group (P <0.05). No significantly difference of the protein levels mentioned above between C and HH group was observed. Compared with H group, the protein levels of Bax and cleaved caspase-3 were down-regulated, and Bcl-2 was up-regulated in HH group (P <0.05). The level of intracellular ROS was higher and the activity of SOD was lower in H group than those in C group (P<0.05). However, there was no difference of the SOD activity between C group and HH group. The level of intracellular ROS decreased and the activity of SOD increased in HH group as compared with H group (P<0.05). Compared with C group, clearly reduced protein expression of Nrf2, HO-1 and NQO-1, and decreased mRNA expression of HO-1 and NQO-1 in H group were observed (P<0.05). Compared with H group, the protein levels of Nrf2, HO-1 and NQO-1 as well as the mRNA levels of HO-1 and NQO-1 were obviously increased in HH group (P<0.05).CONCLUSION: Hydrogen molecule inhibits the expression of pro-apoptotic proteins and induces the expression of anti-apoptotic proteins in glomerular mesangial cells cultured with high glucose. The mechanism may be related to activation of Nrf2 signaling pathway.  相似文献   

17.
AIM: To investigate the protective effect of mesenchymal stem cell (MSC)-conditioned medium (MSCCM) on myocardial cell line H9c2 and its mechanism. METHODS: Verification of MSC was performed by flow cytometry analysis, followed by MTT assay to determine the optimal incubation time of MSCCM with myocardial cells. The cells were divided into 4 groups: normal (N) group, model (M) group, M+MSCCM group and MSCCM group. The cells in M+MSCCM group and MSCCM group were pre-incubated with MSCCM for 24 h. The cells in M group and M+MSCCM group were treated with 300 μmol/L H2O2 for 4 h to imitate oxidative injury of myocardial cells. Mitochondrial membrane potential and apoptotic rate of injured myocardial cells were detected by flow cytometry. The ROS production was measured by fluorescence microscopy. The nuclear translocation of Nrf2 and expression of HO-1 was examined by Western blot. RESULTS: No difference of mitochondrial membrane potential, apoptotic rate or ROS production between MSCCM group and N group was observed (P>0.05). The mitochondrial membrane potential depolarization, apoptotic rate and ROS production in M+MSCCM group were significantly lower than those in M group (P<0.01). The nuclear translocation of Nrf2 and expression of HO-1 in the myocardial cells were increased with MSCCM incubation time prolonged. CONCLUSION: MSCCM protects the myocardial cells against oxidative injury induced by H2O2. The anti-oxidative mechanism would be associated with the activation of Nrf2/ARE pathway.  相似文献   

18.
AIM: To investigate the effects of naringenin (NAR) on the myocardium as well as its effects on adenosine monophosphate-activated protein kinase (AMPK)/nuclear factor (NF)-E2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) signaling pathways in diabetic mice. METHODS: C57BL/6 mice (n=50) were randomly divided into normal group (N group) and model group. The mouse model of type 1 diabetes mellitus was established by intraperitoneal injection of streptozotocin (STZ), then the mice were divided into diabetes group (D group), diabetes+low dose of NAR intervention group (D+L-NAR group), diabetes+middle dose of NAR intervention group (D+M-NAR group) and diabetes+high dose of NAR intervention group (D+H-NAR group). The mice in intervention groups were received NAR at low, middle and high doses respectively by gavage, and the mice in N group and D group were received equal volume of normal saline. After 6 weeks, the mice were sacrificed to observe the effects of NAR at different doses on the body weight and blood glucose. The histopathological changes of the cardiac tissues were observed with HE staining. The myocardial collagen volume fraction (CVF) was calculated by Masson staining. Immumohistochemical staining was used to test the protein levels of interleukin-6 (IL-6) and IL-10, and the TUNEL was used to observe the apoptosis of myocardial tissues. The production of reactive oxygen species (ROS) in the myocardial cells was analyzed by fluorescence probe of DHE, and superoxide dismutase (SOD) activity and malondiodehyde (MDA) content in the myocardial cells were measured by SOD and MDA kits. Western blot was applied to determine the protein levels of p-AMPKα, AMPKα, Nrf2, HO-1, NAD(P)H:quinone oxidoreductase 1 (NQO1) and cleaved caspase-3 in the myocardial tissues. RESULTS: Compared with N group, the blood glucose of the mice in D groups was increased and the body weight was decreased significantly. Compared with D group, the blood glucose of the mice in NAR intervention groups was decreased and the body weight was increased. Compared with N group, the CVF, apoptotic rate and the protein levels of IL-6, cleaved caspase-3 were increased, while the protein levels of IL-10, p-AMPKα, Nrf2, HO-1, NQO1 and SOD activity were decreased, the ROS production rate and MDA content was increased significantly in D group (P < 0.05). Compared with D group, the CVF, apoptotic rate and the protein levels of IL-6, cleaved caspase-3 were relatively decreased, conversely the protein levels of IL-10, p-AMPKα, Nrf2, HO-1, NQO1 were increased in NAR intervention groups(P < 0.05). No significantly difference of the ROS production rate, SOD activity and MDA content between D group and D+L-NAR group was observed. However, the ROS production rate and MDA content was decreased,SOD activity were increased in D+M-NAR group and D+H-NAR group as compared with D group. CONCLUSIONS: NAR attenuates myocardial injury in diabetic mice, and its mechanism may be related to regulation of AMPK/Nrf2/HO-1 signaling pathway, enhancement of the antioxidant reaction, reduction of myocardial fibrosis, apoptosis and inflammation.  相似文献   

19.
AIM:To investigate the effect of shikonin on the apoptosis and oxidative stress induced by high concentration of glucose in vascular endothelial cells. METHODS:Rat thoracic aortic endothelial cells were randomly divided into 5 groups:normal control group (with glucose at concentration of 5.5 mmol/L in cell culture medium), high glucose group (with glucose at concentration of 33 mmol/L in cell culture medium), high glucose+low shikonin group (with glucose at concentration of 33 mmol/L and shikonin at concentration of 0.1 μmol/L in cell culture medium), high glucose+medium shikonin group (with glucose at concentration of 33 mmol/L and shikonin at concentration of 1 μmol/L in cell culture medium), and high glucose+high shikonin group (with glucose at concentration of 33 mmol/L and shikonin at concentration of 10 μmol/L in cell culture medium). After treatments, the cell viability was measured by CCK-8 assay and cell apoptotic rate was analyzed by flow cytometry. In addition, the status of oxidative stress was evaluated by determining the levels of malondialdehyde (MDA), reactive oxygen species (ROS), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). The activation of Nrf2/HO-1 signaling pathway was determined by Western blot. RESULTS:Compared with high glucose group, shikonin reversed high glucose-induced decrease in cell viability and increase in apoptosis in a concentration-dependent manner. High concentration of glucose induced high levels of MDA and ROS, while decreased the levels of SOD and GSH-Px. However, after treatment with shikonin, the contents of MDA and ROS were decreased, while the activities of SOD and GSH-Px were increased as compared with high glucose group. Furthermore, the high concentration of glucose up-regulated the protein levels of cleaved caspase-3, HO-1 and Nrf2 (nuclear). Compared with high glucose group, the protein levels of cleaved caspase-3, HO-1 and Nrf2 (nuclear) were partly decreased after treatment with shikonin. CONCLUSION:Shikonin alleviates high glucose-induced vascular endothelial cell apoptosis. Its mechanism may be related to activation of Nrf2/HO-1 signaling pathway and down-regulation of oxidative stress in vascular endothelial cells.  相似文献   

20.
AIM: To investigate the role of mitochondrial calcium uniporter (MCU) in high glucose(HG)-induced apoptosis of cardiac myocytes. METHODS: Cardiac myocytes were exposed to normal glucose (5.5 mmol/L glucose+ 19.5 mmol/L mannitol), HG (25 mmol/L glucose), or HG combined with 5 μmol/L spermine for 72 h. Mitochondrial free Ca2+ concentration ([Ca2+]m), MCU at mRNA and protein levels, pyruvate dehydrogenase (PDH) activity, mitochondrial membrane potential (Δψm), the levels of ATP and reactive oxygen species (ROS), and apoptosis were determined. RESULTS: The [Ca2+]m, the mRNA and protein levels of MCU, PDH activity, ATP levels, and Δψm were reduced (P<0.05), while ROS content and the protein levels of caspase-9 and caspase-3 were increased in HG group (P<0.05). Adding 5 μmol/L spermine returned these parameters toward control levels (P<0.05). Moreover, apoptosis was reduced by adding spermine and HG treatment (P<0.05). CONCLUSION: HG-induced cardiac myocyte apoptosis may be associated with the decreased MCU expression and activity, abnormal mitochondrial Ca2+ handling, deviant mitochon-drial respiratory chain, and mitochondrial dysfunction.  相似文献   

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