Preparation of glycine liposomes and it's effect on cardiomyocyte injury induced by hypoxia/reoxygenation

AIM: To prepare glycine liposome microparticle and observe the effect of glycine liposomes on cardiomyocyte injury induced by hypoxia/reoxygenation. METHODS: （1） Reverse-phase evaporation method was used to produce glycine liposomes, the effects of different organic solvents: aether, chloroform and two mixtures of aether/chloroform on entrapment efficiency were evaluated, transmission electron microscope was used to detect the particle diameter of glycine liposomes. （2） A cardiomyocyte injury model was established by using hypoxia/reoxygenation, the activities of lactate dehydrogenase (LDH), creatine kinase (CK) and creatine kinase isoenzyme (CK-MB) of each group were detected. RESULTS: The entrapment efficiency of glycine liposomes prepared with the mixtures of aether/chloroform is highest compared with other organic solvents （64.8％, P<0.01）; all of glycine, glycine liposomes and blank liposomes inhibited the release of LDH, CK and CK-MB induced by hypoxia/reoxygenation in cultured cardiomyocytes, and the inhibition by glycine liposomes was the most obvious （P<0.05，P<0.01）. CONCLUSIONS: The mixtures of aether/chloroform can be used to prepare glycine liposomes, which can get higher entrapment efficiency; glycine liposomes protects cultured cardiomyocytes against hypoxia/reoxygenation-induced injury, and the protective effect of glycine liposomes is better than that of glycine.     

Liver X receptors attenuate myocardial ischemia-reperfusion injury through modulating GLUT-4

AIM：To investigate whether liver X receptors (LXRs) attenuate myocardial ischemia-reperfusion (I/R) injury in isolated rat heart through modulating glucose transporter 4 (GLUT-4). METHODS：Isolated rat hearts were used to establish the model of ischemia-reperfusion injury using Langendorff apparatus. The hearts were divided into 7 groups: LXR agonist T0901317 (0.1, 0.5 and 10 μmol/L) pretreatment groups, ischemic preconditioning group, control group, control+DMSO group, and I/R group. The releases of lactate dehydrogenase (LDH) and creatine kinase (CK), the infarct size, the hemodynamic parameters (left ventricle developed pressure,left ventricle end-diastolic pressure, coronary flow and ±dp/dt max), the relative mRNA level of GLUT-4 and the protein content of GLUT-4 in the myocardial cell membrane were compared between these groups. RESULTS：Besides producing hemodynamic disorders, I/R increased the activities of LDH and CK, and the infarct size in model groups. Treatment with T0901317 significantly suppressed ischemia-reperfusion injury-induced increases in LDH and CK, and reduced the infarct size. T0901317 also significantly ameliorated the parameters of haemodynamics. Treatment with T0901317 significantly increased GLUT-4 expression at mRNA and protein levels in the myocardial cell membrane. CONCLUSION：Liver X receptors may attenuate myocardial ischemia-reperfusion injury in isolated rat heart by modulating the expression of GLUT-4.     

Effect of L-carnitine on apoptosis and oxidant damage of cardiomyocytes induced by hypoxia/reoxygenation in vitro

AIM: To examine the effects of L-carnitine on apoptosis and oxidant injury in cultured neonatal rat cardiomyocytes induced by hypoxia/reoxygenation and its possible mechanism. METHODS: The cultured cardiomyocytes were divided into three groups， control， A/R group （anoxia for 120 min, reoxygenation for 240 min） and L-carnitine treatment group, in which cells were exposed to 20 mg/L, 50 mg/L, 100 mg/L, 200 mg/L L-carnitine respectively at 2 h before anoxia. The superoxide dismutase (SOD), succinate dehydrogenase (SDH) activities and malondialdehyde (MDA) content were examined, and the apoptosis was determined by flow of cytometry (FCM). In addition, the ultrastructure was observed by transmission electron microscopy. RESULTS: In A/R group, SOD and SDH activities were lower, the apoptosis rate and MDA content were higher than those in control group (P<0.05). In L-carnitine treatment group, SOD and SDH activities were higher, the apoptosis rate and MDA content were lower than those in A/R group, a L-carnitine concentration-dependent effect was found. Moreover, impairment of myocardial ultrastructure was more severe in A/R group than L-carnitine treatment group. CONCLUSION: L-carnitine can protect cardiomyocytes against hypoxia/reoxygenation-induced injury in a dose-dependent manner.     

Role of heme oxygenase-1 in cardioprotection against anoxia/re oxygenation induced injury and itsmechanisms

AIM:To investigate the role of HO-1 in pro tection of rat hearts against anoxia/reoxygenation-induced injury and its under lying mechanism.METHODS:Cardiac contractility,lactate dehydrogenase (LDH) and infarct area were analyzed by the Langendorff method in isolated rat hearts.RESULTS:After intraperitoneal injection of HO-1 inducer hemin,CO concentration in rat blood enhanced (P<0.01 vs control group).Pretreatm ent with hemin prevented the increase in LVEDP and decrease in LVDP,±dp/d tmax during the anoxia and reoxygenation period in hearts.Hemin had n o effect on changes of coronary flow,but it really inhibited the release of LDH from anoxia/reoxygenation hearts.Hemin also reduced the infarct area in anoxia heart after 2 h reoxygenation (P<0.01).CO concentration in rat blood redu ced after intraperitoneal injection of HO-1 inhibitor ZnPP (P<0.01 vs contr ol group).ZnPP aggravated the decrease in LVDP and ±dp/dtmax.Co mpared with anoxia/reoxygenation heart,pretreatment of ZnPP enhanced the LDH re lease and enlarged the infarct area (P<0.05).GC inhibitor methylene blue a nd cyclooxygenase-2 (COX-2) inhibitor celecoxib both partly abolished the protec tion effect of hemin on LVEDP,LVDP and ±dp/dtmax.Pretreatment o f methylene blue or celecoxib also cancelled the inhibition of LDH release and r eduction of infarct area caused by hemin (P<0.05).CONCLUSION:HO-1 inducer hemin protects heart from anoxia/reoxy genation-induced injury.The cardiac protection of HO/CO might be through GC pathway,and the activation of COX-2 might be also involved in this process.     

Protective effect of quercetin on rat cardiomyocytes against anoxia/reoxygenation injury is mediated by PKCε signaling pathway

AIM：To investigate whether quercetin (Que) protects cardiomyocytes from anoxia/reoxygenation (A/R) injury through protein kinase C epsilon (PKCε) pathway. METHODS：Primary cardiomyocytes were isolated from neonatal SD rats and exposed to A/R (3 h of anoxia followed by 2 h of reoxygenation) as well as Que and/or εV1-2 (a selective PKCε inhibitor) preconditioning. The expression of PKCε in the cells was detected by Western blotting. The levels of lactate dehydrogenase (LDH) and creatine kinase (CK) in cell culture supernatants, the reactive oxygen species (ROS) and mitochondrial membrane potential in the cells, the opening of mitochondrial permeability transition pore (mPTP) and the cell viability and apoptosis were also measured. RESULTS：The expression of PKCε protein was significantly increased in the cardiomyocytes pretreated with 40 μmol/L Que 72 h before A/R (P<0.01 vs A/R group). Meanwhile, Que preconditioning could increase cell survival rate, decrease ROS production and cell apoptosis, alleviate the loss of mitochondrial membrane potential and inhibit the opening of mPTP induced by A/R injury (P<0.01 vs A/R group). However, pretreatment with Que and εV1-2 attenuated these protective effects of Que (P<0.01 vs Que+A/R group). CONCLUSION：One of the mechanisms underlying the cardioprotective effect of Que might be the increase in PKCε protein expression and the activation of its downstream pathway.     

Effects of PDCD5 on hypoxia/reoxygenation-induced autophagy and apoptosis of cardiomyocytes

AIM: To investigate the influence of programmed cell death 5 (PDCD5) on apoptosis and autophagy in the cardiomyocytes exposed to hypoxia/reoxygenation (H/R) and its potential mechanism.METHODS: H9c2 cells were exposed to H/R. PDCD5 was downregulated by RNA interference. The cell viability was measured by MTT assay. TUNEL assay was used to detect cell apoptosis. The mRNA and protein levels were determined by RT-qPCR and Western blot.RESULTS: The expression of PDCD5 was upregulated in the cardiomyocytes after H/R injury. Furthermore, H/R injury obviously reduced the cell viability and enhanced the apoptosis and autophagy of the cardiomyocytes. However, knockdown of PDCD5 increased the cell viability, and attenuated H/R-induced apoptosis, accompany with reduction of Bax and augment of Bcl-2 expression. Additionally, silencing PDCD5 markedly inhibited H/R-induced autophagy by regulating the expression of LC3-II/LC3-I and beclin-1. Moreover, downregulation of PDCD5 suppressed NF-κB signaling by redu-cing the protein level of p-P65.CONCLUSION: Silencing PDCD5 suppresses H/R-induced H9c2 cells apoptosis and autophagy by blocking NF-κB signaling pathway. The result indicates a new strategy for the prevention and treatment of myocardial I/R injury.     

Role of glycine receptor in protection of glycine against anoxia/reoxygenation injury in cardiomyocytes

AIM:To investigate whether glycine receptor is involved in the protection of glycine against anoxia/reoxygenation injury in cardiomyocytes by detecting oxygen free radical metabolism, apoptosis and intracellular calcium overload. METHODS:The neonatal rat cardiomyocytes were cultured and exposed to anoxia and reoxygenation (A/R) in the presence of glycine receptor antagonist, glycine or in free chloride buffer. The superoxide dismutase (SOD) activity, the contents of malondialdehyde (MDA) and nitric oxide (NO), the intracellular free calcium concentration and the apoptotic rate in the cardiomyocytes were determined. RESULTS:SOD activity and NO content in cardiomyocytes were lower, but MDA content, intracellular free calcium concentration and apoptotic rate in cardiomyocytes were higher in A/R group than those in control. Pretreatment with glycine inhibited the above changes caused by A/R, which was reversed by strychnine treatment and in the free chloride medium. CONCLUSIONS:Glycine inhibits free radical production, attenuates calcium overload, decreases apoptotic rate and increases SOD activity and NO release in cardiomyocytes exposed to A/R. These findings suggest that glycine exerts a protective effect against A/R injury via glycine receptor and glycine protects the neonatal rat cardiomycytes from A/R-induced injury in a chloride-dependent manner.     

p38 MAPK-Pim-3 signal pathway may be involved in cardiomyocyte anoxia preconditioning against anoxia/reoxygenation injury

AIM: To investigate the role of mitogen-activated protein kinases (MAPKs) pathways and the molecular mechanism by which the proto-oncogene Pim-3 protects cardiomyocyte against anoxia/reoxygenation (A/R) injury. METHODS: The primarily cultured neonatal rat ventricular cardiomyocytes were randomly divided into 4 groups: control group; A/R group; APC+A/R group; SB203850, U0126 or SP600125+APC+A/R group. The cells were pre-incubated with U0126 (ERK1/2 inhibitor), SP600125 (SAPK/JNK inhibitor), or SB203850 (p38 MAPK inhibitor) at concentration of 10 μmol/L for 30 min before the APC. The activities of p38 MAPK, JNK and ERK1/2 were detected by Western blotting. The viability of cardiomyocytes was assayed by MTT and the apoptosis of cardiomyocyte was detected by TUNEL. RESULTS: U0126, SB203850, and SP600125 abolished the increased expression of ERK1/2, p38-MAPK, and JNK proteins induced by APC+A/R or A/R, respectively. The expression level of Pim-3 protein significantly decreased when the p38 MAPK signal pathway was inhibited. Meanwhile, the activity of LDH and the apoptosis index increased, and the viability of cardiomyocytes decreased. CONCLUSION: Pim-3 expression through a p38 MAPK signaling pathway may protect cardiomyocytes from A/R injury.     

Anti-aging Klotho protein reduce the hypoxia/reoxygenation injury of neonatal rat myocardial cells

AIM: To study the effects of anti-aging Klotho protein on neonatal rat myocardial cells with hypo-xia/reoxygenation (H/R) injury. METHODS: The cardiomyocytes of neonatal SD rats were cultured to establish hypoxia/reoxygenation model. The myocardial cells were divided into normal control group, H/R model group, different concentrations of Klotho protein (0.1 μmol/L, 1 μmol/L and 10 μmol/L) pretreatment groups. The myocardial cells pulse frequency was observed before and after H/R. The cell viability was measured by MTT assay. The leakages of LDH, CK and AST, the content of MDA and the activity of SOD were detected. The apoptotic rate of the myocardial cells was analyzed by flow cytometry. The mRNA expression of endoplasmic reticulum stress markers and apoptosis-related molecules GRP78, CRT, CHOP and caspase-12 was measured by real-time PCR. The protein levels of CHOP, caspase-12 and phosphorylated Akt in the myocardial cells were determined by Western blot. RESULTS: Compared with normal control group, the pulse frequency, cell viability rate and SOD activity of myocardial cells were significantly decreased, the cell apoptotic rate as well as the contents of LDH, CK, AST and MDA were increased in H/R model group. The mRNA expressions of GRP78, CRT, CHOP and caspase-12 as well as the protein levels of CHOP and caspase-12 were increased, whereas p-Akt level was decreased obviously. Compared with H/R model group, the pulse frequency, cell viability rate and SOD activity were increased significantly, the cell apoptotic rate as well as the contents of LDH, CK, AST and MDA were decreased in Klotho pretreated group. The mRNA expression of GRP78, CRT, CHOP and caspase-12 as well as the protein levels of CHOP and caspase-12 were decreased, while p-Akt level increased significantly. CONCLUSION: Anti-aging Klotho protein improves the myocardial cell survival and inhibits the apoptosis by increasing the resistance of the cells to oxidative stress and excessive endoplasmic reticulum stress response, which is related with the activation of Akt phosphorylation in H/R-injured mycardial cells.     

第27届国际园艺大会将于2006年8月13—19日在韩国汉城召开

由国际园艺学会和韩国园艺学会共同主办的第27届国际园艺大会将于2006年8月13～19日在韩国汉城召开(同期举办展览)。会议主题是“全球园艺:多样性与和谐”。会议组织委员会主席为韩国庆熙大学李政明(Jung MyungLee)教授,副主席为韩国汉城市立大学JeongSikLee教授和韩国汉城大学     

Role of voltage-dependent anion channel in cardiomyocytes subjected to anoxia/reoxygenation

AIM: To investigate the role of voltage-dependent anion channel (VDAC) in H9c2 cardiomyocytes subjected to anoxia/reoxygenation (A/R).METHODS: The shRNAs targeting VDAC1 mRNA were inserted into pSUPER plasmid.H9c2 cells were randomly divided into 5 groups as follows: control group, A/R group, anoxia preconditioning (APC) group, pSUPER-VDAC1-A/R group and pSUPER-A/R group. The expression of VDAC1 was detected by Western blotting. Cellular injury was evaluated by measuring the cell viability and the release of lactate dehydrogenase (LDH) and creatine phosphokinase (CPK). The mitochondria membrane potential was determined by flow cytometry.RESULTS: VDAC1 expression was up-regulated in A/R group and was inhibited in APC group. Similarly, down-regulation of VDAC1 expression by shRNA protected H9c2 cells from A/R injury. Moreover, we found that, with silencing VDAC1 expression, mitochondrial membrane potential was well preserved in H9c2 cells subjected to A/R.CONCLUSION: Down-regulation of VDAC1 protects H9c2 cells against A/R injury and its possible mechanism appears to be related to the regulation of mitochondial permeability transition pore opening.     

Effect of polyamines on intracellular calcium of the rat cardiomyocytes with anoxia and reoxygenation

AIM: To study the effect of extrogenous low concentration polyamine on cardiomyocyte calcium overload caused by anoxia and reoxygenation. METHODS: Enzymatically isolated rat ventricular myocytes were perfused with normal Tyrode solution for 8 min, then change to anoxia solution for 32 min, at last back to normal Tyrode solution perfusion for 8 min to establish the cardiomyocyte model of anoxia and reoxygenation. Spermine was added extracellularly to the bath before anoxia and spermine, spermidine or putrescine was added extracellularly after reoxygenation. Intracellular calcium fluorescence intensity (FI) was measured continuously by laser scan confocal microscope (LSCM). RESULTS: In the unstimulated state, exogenous spermine (1 mol/L) did not change resting ［Ca2+］i in the rat cardiomyocytes. Adding spermine before anoxia antagonized the ［Ca2+］i elevating caused by anoxia/reoxygenation. Adding spermine after reoxygenation also lowed the enhanced ［Ca2+］i caused by reoxygenation. Considering the potency of two conditions, the former was more efficacious than the later. Spermidine and putrescine also lowed the enhanced ［Ca2+］i caused by reoxygenation, but they were less efficacious than spermine. CONCLUSIONS: The results indicate that spermine given before anoxia or after reoxygenation, antagonized or lowed the cardiomyocytes calcium overload caused by anoxia/reoxygenation, but the later was weaker than the former. The order of potency of the polyamine lighten cardiomyocytes calcium overload caused by anoxia/reoxygenation was spermine>spermindine>putrescine.     

Effects of glycine on intracellular of free calcium and tumor necrosis factor-α of cardiomyocytes during hypoxia/reoxygenation

AIM: To observe the influence of glycine on intracellular free calcium, the concentration of tumor necrosis factor-α and the survival rate of myocardial cells during hypoxia/reoxygenation (H/R). METHODS: The simulated model of myocardial ischemia-reperfusion with the primary cultured cardiomyocytes of neonatal rats was established, and the cultured cardiomyocytes were divided into seven groups, control group, hypoxia/reoxygenation group, glycine (0.5 mmol/L) plus hypoxia/reoxygenation group, glycine (1.0 mmol/L) plus hypoxia/reoxygenation group, glycine (2.0 mmol/L) plus hypoxia/reoxygenation group, glycine (4.0 mmol/L) plus hypoxia/reoxygenation group, 4.0 mmol/L glycine group. RESULTS: Within certain concentration (0.5-2.0 mmol/L), the glycine could inhibit the calcium overload resulting from hypoxia/reoxygenation injury in cells in a dose-dependent manner with the optimal inhibitory effect at 2.0 mmol/L. Glycine inhibited the secretion of tumor necrosis factor-α from myocardial cells and increased the survival rate of myocardial cells. CONCLUSION: Glycine has a protective effect on hypoxia/reoxygenation myocardial cells, which may be related to inhibiting calcium overload and decreasing the production of tumor necrosis factor-α.     

Effect of AMPKα2 gene silencing by shRNA recombinant plasmid on chloride-mediated anoxia/reoxygenation injury in rat cardiomyocytes

AIM: To explore the role of AMP-activated protein kinase α2 subunit (AMPKα2) gene in chloride-mediated anoxia/reoxygenation (A/R) injury by transfection of short-hairpin RNA (shRNA) expression vector targeting to AMPKα2 gene into H9c2 cardiomyocytes. METHODS: Recombinant shRNA expression vector pSuper-AMPKα2 targeting to AMPKα2 gene was constructed and transfected into H9c2 cardiomyocytes. The protein expression of AMPKα2 was determined by Western blotting. The cells were divided into 5 groups: control group, A/R group, Cl^--free A/R group, pSuper+Cl^--free A/R group and pSuper-AMPKα2 shRNA+Cl^--free A/R group. After treatment, the cell viability was detected by MTT assay. LDH activity was analyzed with an automatic biochemical analyzer. The apoptotic rate and the level of intracellular ROS was measured by flow cytometry. The activity of SOD and GSH-Px was analyzed by a colorimetric method. RESULTS: The result of sequencing proved that the recombinant plasmid pSuper-AMPKα2 shRNA was correctly constructed. The protein level of AMPKα2 significantly decreased after the plasmid was transfected into the cardiomyocytes. Compared with A/R group, the cell viability and the activity of SOD and GSH-Px were significantly increased, while the activity of LDH, apoptotic rate and ROS production were significantly decreased in Cl^--free A/R group. The protective effect of Cl^--free solution on the A/R-induced injury of cardiomyocytes was abolished, and the ROS production was increased and the activity of SOD and GSH-Px was decreased after the cells were transfected with pSuper-AMPKα2 shRNA. CONCLUSION: Recombinant plasmid pSuper-AMPKα2 shRNA is successfully constructed, and silencing of AMPKα2 gene abolishes the protective effect of Cl^--free solution on A/R injury.     

Protection of calcium antagonists against cardiomyocyte injury caused by anoxia and reoxygenation

AIM: To investigate the protective effect of calcium antagonists on anoxia/reoxygenation (A/R) injury of cardiomyocytes. METHODS: Primary-cultured cardiomyocytes were divided into four groups, namely A/R, A/R+nifedipine (Nif), A/R+ruthenium red (Ru)+heparin (Hep) and control groups. The following parameters were measured in all groups: intracellular calcium concentration (i), cardiac cell viability, ATP content, lactate dehydrogenase (LDH) activity in the medium, PKC and MAPK activity and ³[H]-Leucine (³[H]-Leu) incorporation. RESULTS: In comparison with A/R group,A/R+nifedipine (Nif) and A/R+ruthenium red (Ru)+heparin (Hep) groups showed a marked decrease in[Ca²⁺]i and LDH content,and a significant increase in cell viability, ATP content, activity of PKC and MAPK and [³H]-Leu incorporation (P<0.05 or P<0.01). CONCLUSION: A/R mediated Ca²⁺ overload resulted in cardiomyocyte injury, which could be attenuated by blocking Ca²⁺ entry and release.     

Role of ROS/PKC/p38 MAPK pathway in protective effects of polysaccharide from Fructus cornion rat cardiomyocytes against hypoxia-reoxygenation injury

AIM: To investigate the effect of polysaccharide from Fructus corni(PFC) on cardiomyocytes against hypoxia/reoxygenation (H/R) injury and its possible relationship with ROS/PKC/p38 MAPK pathway.METHODS: Primary cardiomyocytes were isolated from neonatal SD rats and randomly divided into normal group, H/R group, PFC (20 mg/L, 100 mg/L and 200 mg/L) preconditioning+H/R groups, chelerythrine+PFC (100 mg/L)+H/R group and SB203580+PFC (100 mg/L)+H/R group. The cell viability was measured by inverted microscopic observation. Apoptosis in the cardiomyocytes was detected by Hoechst 33258 staining and fluorescence microscopy. The levels of lactate dehydrogenase (LDH) and superoxide dismutase (SOD) in the cell culture supernatants, and the reactive oxygen species (ROS) in the cells were also measured by microplate reader. The protein levels of PKC, p-p38 MAPK and HSP70 in the cells were detected by Western blotting.RESULTS: Compared with normal group, the cell viability and beating frequency were decreased in H/R group. LDH and ROS contents, apoptotic rate and p-p38 MAPK level increased significantly (P<0.01). Compared with H/R group, PFC preconditioning increased beating frequency, SOD activity and the protein level of PKC and HSP70, and decreased ROS production, the protein level of p-p38 MAPK and cell apoptotic rate. However, the effect of PFC was inhibited by chelerythrine or SB203580.CONCLUSION: PFC may protect cardiomyocytes from hypoxia/reoxygenation injury. Its mechanism is possibly involved in the inhibition of ROS via increasing the activity of SOD and the activation of PKC, and suppression of excessive activation of p38 MAPK.     

Effect of glycine liposomes on mitochondrial membrane potential and apoptosis in cultured cardiomyocytes

AIM: To observe the effect of glycine liposomes on the mitochondrial membrane potential and the apoptosis rate in cardiomyocytes induced by hypoxia/reoxygenation injury. METHODS: A cardiomyocyte injury model was established by using hypoxia/reoxygenation. DiOC6(3) as fluorescence molecular probe was used to detect the mitochondrial membrane potential in each group. The method of Annexin V associated with PI was used to detect the apoptosis ratio in each group. RESULTS: (1) The result of flow cytometry showed that the mitochondrial membrane potential of cardiomyocytes in H/R group was obviously lower than that in control group (P<0.01). The decrease in mitochondrial membrane potential in Gly-liposome group was the lowest, the percentage of cells about the part of hypofluorescence was (9.61±0.76)%, which was lower than that in glycine group (P<0.01). (2) The apoptosis rate of cardiomyocytes in H/R group was higher than that in control group (20.78±1.58)%,P<0.01. After the treatment of Gly-liposome, the apoptosis rate of cardiomyocytes was lower than that in glycine group (P<0.01). No difference in the apoptosis ratios between blank-liposome group and H/R group was observed(P>0.05).CONCLUSION: Glycine liposomes protect cultured cardiomyocytes against hypoxia/reoxygenation injury. Glycine liposomes produce the better protective effects than glycine.     

Effect of rosuvastatin on cerebral microvascular endothelial cell injury induced by oxygen-glucose deprivation/reoxygenation

AIM: To explore the effect of rosuvastatin on the oxygen-glucose deprivation (OGD)/reoxygenation induced injury of cerebral microvascular endothelial cells (BMECs). METHODS: BMECs derived from BALB/c mice were isolated and cultured. BMECs were pretreated with rosuvastatin, followed by OGD for 3 h or 6 h and reoxygenation for 24 h. The morphological changes of BMECs were observed under light microscope. MTT assay was used to measured the cell viability, and carboxyfluorescein diacetate succinimidyl ester (CFDA-SE) staining was used to assess the proliferation of BMECs. The protein levels of cleaved caspase-3 was observed by immunofluorescence staining. The protein levels of Bcl-2, Bax, matrix metalloproteinase (MMP) 2, MMP9, phosphorylated nuclear factor kappa B (p-NF-κB), phosphorylated P38 mitogen-activated protein kinase (p-P38) and phosphorylated c-Jun N-terminal kinase (p-JNK) were determined by Western blot. RESULTS: Rosuvastatin at 10 μmol/L improved the viability of the BMECs with OGD/reoxygenation-induced damage, and maintained the structure of BMECs. Moreover, rosuvastatin significantly prohibited the protein levels of cleaved caspase-3, MMP2, MMP9, p-NF-κB, p-P38 and p-JNK, and up-regulated the ratio of Bcl-2/Bax (P<0.05). CONCLUSION: Rosuvastatin reduces OGD/reoxygenation-induced injury of BMECs by inhibiting the expression of apoptosis-related proteins and MMPs, suggesting that rosuvastatin has potential value for the maintenance of blood-brain barrier.     

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《园艺学报》是中国园艺学会和中国农业科学院蔬菜花卉研究所主办的学术期刊,创刊于1962年,刊载有关果树、蔬菜、观赏植物、茶及药用植物等方面的学术论文、研究报告、专题文献综述、问题与讨论、新技术新品种以及园艺研究动态与信息等,适合园艺科研人员、大专院校师生及农业技术推广部门专业技术人员阅读参考。《园艺学报》是中文核心期刊,被英国《CAB文摘数据库》、美国CA化学文摘、日本CBST科学技术文献速     

Effects of adiponectin on reducing endoplasmic reticulum stress injury induced by hypoxia-reoxygenation in cultured rat neonatal cardiomyocytes

AIM: To investigate the effects of adiponectin (APN) on hypoxia-reoxygenation (H/R) induced endoplasmic reticulum stress injury in cultured cardiomyocytes. METHODS: Primary cultured cardiomyocytes were obtained from neonatal rats by enzymatic digestion method. The α-actin expression as molecular marker of the cardiomyocytes was observed by immunocytochemistry. The cells cultured for 72 h were used in the experiment and divided into groups randomly: control group, H/R group, APN+H/R (3 mg/L, 10 mg/L, 20 mg/L, 30 mg/L) groups. The morphological changes of the cardiomyocytes were observed under phase contracted microscope. The content of LDH was measured. The cardiomycocyte apoptosis was detected by flow cytometry. The expression levels of GRP78 and caspase-12 were examined by RT-PCR and Western blotting. RESULTS: The apoptotic rate was significantly increased in H/R group as compared to that in control group (68.20%±1.73% vs 0.73%±0.21%, P<0.05). The levels of LDH in H/R group were also significantly increased. Compared to untreated cells, the protein and mRNA levels of GRP78 and caspase-12 increased significantly in H/R cells. The APN preconditioning significantly reversed these changes. The indexes above improved obviously as compared to H/R group (P<0.05) in a dose-dependent manner. CONCLUSION: Hypoxia/reperfusion induces endoplasmic reticulum stress in rat cardiomyocytes. Adiponectin decreases the endoplasmic reticulum stress injury and plays a protective role by extenuation of cadiomyocyte apoptosis.