Regulation of AMPK/PPARα/SCAD signal pathway on cardiac hypertrophy

AIM：To study the effect of short-chain acyl-coenzyme A dehydrogenase (SCAD）on cardiac hypertrophy and to explore the role of adenosine monophosphate-activated protein kinase (AMPK)/peroxisome proliferator-activated receptor α (PPARα） signal pathway in the regulation of SCAD during the development of cardiac hypertrophy. METHODS：The optimal sequence of SCAD interference was chosen by Western blotting and real-time PCR. The cardiomyocytes were treated with fenofibrate (10 μmol/L) for 24 h and subsequently stimulated with the optimal sequence of SCAD interference. The changes of SCAD expression at mRNA and protein levels, the enzyme activity of SCAD, the cardiomyocyte surface area and free fatty acids were determined. Using real-time PCR for analyzing the markers of cardiac hypertrophy, the mRNA expression of atrial natriuretic factor (ANF） and brain natriuretic peptide (BNP) was detected to judge the development of cardiac hypertrophy. The cardiomyocytes were treated with fenofibrate (10 μmol/L) or AMPK activator 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR, 0.5 mmol/L) for 30 min and subsequently stimulated with phenylephrine (PE, 20 μmol/L) for 24 h. The changes of cardiomyocyte surface area, free fatty acids, and the expression of SCAD, PPARα and p-AMPKα (T172) at mRNA and protein levels were observed. RESULTS：The effect of optimal sequence siRNA-1186 and PE on the cardiomyocytes was the same. Compared with control group, the expression of ANF and BNP at mRNA level, the cardiomyocyte surface area and free fatty acids were increased obviously in siRNA-1186 group. After pretreated with fenofibrate (10 μmol/L), the expression of PPARα and SCAD, and the enzyme activity of SCAD were significantly increased, while the free fatty acids were decreased, indicating that fenofibrate prevented the development of cardiac hypertrophy induced by knockdown of SCAD. Compared with control group, the expression of SCAD, PPARα and p-AMPKα (T172) at mRNA and protein levels was significantly down-regulated, and the enzyme activity of SCAD was obviously decreased in PE group. Compared with PE group, the expression of SCAD, PPARα and p-AMPKα (T172) was significantly up-regulated, and the cardiomyocyte surface area and the content of free fatty acids were obviously decreased in the cardiomyocytes pretreated with fenofibrate or AICAR for 30 min. CONCLUSION：Down-regulation of SCAD is related to the cardiac hypertrophy and energy metabolism. AMPK/PPARα/SCAD signaling pathway may regulate cardiac hypertrophy directly.     

Effects of short-chain acyl-CoA dehydrogenase on collagen expression and proliferation of rat cardiac fibroblasts

AIM: To investigate the effect of short-chain acyl-CoA dehydrogenase (SCAD) on collagen expression and proliferation of rat cardiac fibroblasts and to explore the relationship between SCAD and cardiac fibrosis. METHODS: The model of proliferation and collagen expression of rat cardiac fibroblasts induced by angiotensin II was established. After treatment with siRNA-1186, the expression of SCAD at mRNA and protein levels, fatty acids beta oxidation rate, ATP, the enzyme activity of SCAD and free fatty acids in the rat cardiac fibroblasts were determined. RESULTS: The mRNA and protein expression of SCAD was decreased in the rat cardiac fibroblasts induced by angiotensin II compared with the control cells, and the expression of collagen I and collagen III was significantly upregulated. Compared with negative control group, SCAD expression and activity, fatty acid beta-oxidation rate and ATP significantly decreased in siRNA-1186 group, but the content of free fatty acids were obviously increased in the rat cardiac fibroblasts, and the expression of collagen I and collagen III was significantly up-regulated. CONCLUSION: The expression and synthesis disorder of collagen may be triggered by down-regulation of SCAD. SCAD may be a promising therapeutic target for myocardial fibrosis.     

Effects of ERK1/2/PPARα/SCAD signal pathways on physiological cardiac hypertrophy and pathological cardiac hypertrophy

AIM：To investigate the different effects of ERK1/2/PPARα/SCAD (short-chain acyl-CoA dehydrogenase) signal pathways on the cardiac hypertrophy induced by insulin-like growth factors 1 (IGF-1) or phenylephrine (PE). METHODS：The neonatal rat cardiomyocytes induced by IGF-1 were used as the model of physiological cardiac hypertrophy, and those induced by PE were used as the model of pathological cardiac hypertrophy. The surface area of the cardiomyocytes, the expression of p-ERK1/2, PPARα and SCAD, the activity of SCAD and the content of free fatty acid in the cardiomyocytes were measured. RESULTS：Compared with the control cells, the surface area of the cardiomyocytes induced by IGF-1 and PE were both increased. Compared with the controls, the expression of SCAD and PPARα, and the activity of SCAD in the cardiomyocytes induced by IGF-1 were increased, while the expression of p-ERK1/2 was decreased. However, the cardiomyocytes treated with PE showed decreased expression of SCAD and PPARα, decreased activity of SCAD and increased expression of p-ERK1/2. Meanwhile, the decrease in free fatty acid in IGF-1-induced cardiomyocytes and the increase in PE-induced cardiomyocytes indicated that the fatty acid utilization was increased in the cardiomyocytes induced by IGF-1, but decreased in the cardiomyocytes induced by PE. CONCLUSION：The changes of p-ERK1/2, PPARα and SCAD in the cardiac hypertrophy induced by IGF-1 or PE indicate that the effects of ERK1/2/PPARα/SCAD signal pathways are different between physiological cardiac hypertrophy and pathological cardiac hypertrophy, and that SCAD may be a molecular marker of these 2 different cardiac hypertrophies and a potential therapeutic target for pathological cardiac hypertrophy.     

Effects of short-chain acyl-CoA dehydrogenase on hypertensive vascular remodeling

AIM: To investigate the changes of short-chain acyl-CoA dehydrogenase (SCAD) in hypertensive vascular remodeling and to explore the relationship between SCAD and vascular remodeling in hypertension.METHODS: The spontaneously hypertensive rats (SHR; 24 weeks old) and Wistar rats (24 weeks old) were used as experimental control groups. The SHR and Wistar rats of 16 weeks old were trained by swimming as experimental groups. The systolic pressure was measured periodically. The thickness of vascular wall and the diameter of the vascular lumen were measured. The contents of ROS and ATP, the enzyme activity of SCAD, and the expression of SCAD at mRNA and protein levels in the aorta were determined. The free fatty acid in the serum and aorta was also measured.RESULTS: Compared with Wistar group, the diameter of vascular lumen decreased in SHR group. The thickness of vascular wall, the ratio of vascular wall and the diameter of vascular lumen, and the blood pressure in SHR group were increased significantly (P<0.05). Compared with SHR group, the diameter of vascular lumen increased in SHR+swim group. The thickness of vascular wall, the ratio of vascular wall and the diameter of vascular lumen, and the blood pressure in SHR+swim group were decreased significantly. Compared with control group, the expression of SCAD at mRNA and protein levels, the enzyme activity of SCAD, and the content of ATP were decreased in SHR group. However, the free fatty acid in the serum and aorta, and the content of ROS in the aorta were increased in SHR group. The expression of SCAD at mRNA and protein levels, the enzyme activity of SCAD, the content of ATP were increased in Wistar+swim group and SHR+swim group. However, the free fatty acid in serum and aorta, and the content of ROS in the aorta were decreased in Wistar+swim group and SHR+swim group.CONCLUSION: Decrease in SCAD expression may be associated with hypertensive vascular remodeling. Swimming training can reverse hypertensive vascular remodeling by increasing the expression of SCAD in the aorta.     

Effect of 11,12-EET on cardiomyocyte apoptosis and bcl-2 mRNA gene expression in donor hearts of immature rabbit underwent prolonged protection

AIM: To study apoptosis and bcl-2 mRNA gene expression of cardiomyocytes in donor hearts of immature rabbits underwent prolonged protection by 11, 12-epoxyeicosatrienoic acid (11, 12-EET), and further probe into the possible mechanisms. METHODS: 24 isolated immature rabbit hearts were performed to the model in a Langendorff perfusion apparatus and randomly assigned to normal control group,ST control group and EET group. The isolated rabbit hearts in ST control group and EET group were stored for 24 hours with 4 ℃ hypothermia, and underwent 30 minutes of reperfusion (37 ℃). TUNEL and in situ hybridization (ISH) methods were applied in the present study and apoptotic cells and bcl-2 mRNA gene expression were observed. RESULTS: The numbers of apoptotic cardiomyocytes in ST group and EET group were higher than that in normal control group, and the numbers of apoptotic cardiomyocytes were significantly decreased in EET group and bcl-2 mRNA positive expression were higher than that in ST control group, respectively. CONCLUSIONS: There were apoptosis during the prolonged protection of donor heart in our study, and we proved that: ①11,12-EET could decrease cardiomyocyte apoptosis significantly. ②Up-regulation of the bcl-2 mRNA expression in cardiomyocytes may be one of the mechanism responsible for inhibition of cardiomyocyte apoptosis by 11, 12-EET.     

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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     

《新编拉汉英植物名称》王宗训主编

AIM: To study the protective effects of non-mitogenic human fibroblast growth factor (nm-haFGF) on cardiomyocytes injured by reactive oxygen free radicals. METHODS: The cardiomyocytes were isolated from neonatal SD mouse by trypsin digestion. The cardiomyocytes injury model was established by exposing the cells to hydrogen peroxide (H2O2), and the injury status in cardiomyocytes were evaluated by examining the cellular viability, measuring cell apoptosis and observing the change of cellular morphology. nm-haFGF was added to the culture medium, and the changes of cellular viability, superoxide dismutase (SOD), malondialdehyde (MDA) and celluar apoptosis were observed. RESULTS: A dose-dependence relation between the concentration of H2O2 and the cardiomyocytes injury was observed. 10-80 μg/L nm-haFGF dose-dependently increased cardiomyocyte viability and the general SOD activity, as well as decreased the content of MDA and the quantity of cardiomyocyte apoptosis. CONCLUSION: The higher the concentration of H2O2, the more serious the cardiomyocyte injury. nm-haFGF may have a good protective effects on cardiomyocytes treated with H2O2.     

Effect of PKC activition on cardiac myocyte apoptosis and Bcl- 2 expression during myocardial ischemia/reperfusion in rats

AIM: To explore the effect of PKC activition on cardiac myocyte apoptosis and expression of bcl- 2 during myocardial ischemia/reperfusion(I/R) in rats. METHODS: TUNEL,immunohistochemistry and in situ hybridization were used. RESULTS: The TUNEL data showed that the numbers of positive cardiac myocyte nucleus and the percentage of positive cardiac myocyte nucleus in PMA+IR_{3 h} group decreased significantly(P＜0.05,P＜0.01), compared to those in IR_3h group. The number of Bcl-2 protein positive cardiomyocytes and the percentage of Bcl-2 protein positive cardiomyocytes in PMA+IR_3h group were higher than those in IR_3h group (P＜0.01) bcl- 2 mRNA expression showed the same changes in PMA+IR_0h group compared to IR_1h group.CONCL USIONS:Activation of PKC decreased cardiomyocyte death during I/R.Upregulation of bcl-2 gene expression in cardiomyocytes during I/R may be one of the mechanisms of decreasing cardiomyocyte death by PCK activating during I/R.     

Effects of ischemic preconditioning on cardiac myocyte apoptosis and expression of bcl-2 during myocardial ischemia/reperfusion in rats

AIM:To explore the effect of ischemic preconditioning on cardiac myocyte apoptosis and the expression of bcl-2 during myocardial ischemia/reperfusion in rats. METHODS:We use TUNEL,immunohistochemical and in situ hybridization(ISH) methods to detect the cardiac myocyte apoptosis and the expression of bcl-2 during myocardial ischemia/reperfusion in rats. RESULTS:①The numbers of positive cardiac myocyte nuclear and the percentage of positive cardiac myocyte nuclear in IP+I/R_3h group decreased significantly(P<0.05,P<0.01)compared with I/R_3h group,respectively.②The numbers of bcl-2 protein positive cardiomyocyte and the percentage of bcl-2 protein positive cardiomyocyte in IP+I/R_3h group were higher(P<0.01)than that of I/R_3h group,respectively.The numbers of positive bcl-2 mRNA cardiomyocyte and the percentage of positive bcl-2 mRNA cardiomyocyte in IP+I/R_1h group were higher(P<0.01)than that of I/R_1h group,respectively.CONCLUSION:① The first window of IP's protection could reduce cardiomyocyte apoptosis significantly.② Up-regulating the protein expression of bcl-2 in cardiomyocytes during I/R may be one of the mechanisms of first window of IP's protection.     

Effect of short-chain acyl-CoA dehydrogenase on cardiac hypertrophy induced by hypertension or exercise training

AIM: To investigate the differential expression of short-chain acyl-CoA dehydrogenase (SCAD) in cardiac hypertrophy induced by hypertension or exercise training. METHODS: Spontaneously hypertensive rats (SHR) were used as the model of pathological cardiac hypertrophy. The swim-trained rats were used as the model of physiological cardiac hypertrophy. The systolic pressure, cardiac hypertrophy parameters, echocardiogram parameters, free fatty acid in serum and cardiac muscle, and the expression and activity of SCAD in the left ventricle were measured. RESULTS: Compared with the control rats, trained rats developed an athletic heart, of which cardiac function was enhanced, whereas SHR developed hypertensive cardiac hypertrophy, of which cardiac function was deteriorated. Compared with the control rats, the ratios of left ventricular weight to body weight were both increased in trained rats and SHR, showing that the degrees of cardiac hypertrophy were similar in the 2 models. Compared with the control rats, the decrease of free fatty acid both in serum and myocardium indicated that the fatty acid utilization was increased in the left ventricle of trained rats. Meanwhile, the expression and activity of SCAD in the left ventricle of trained rats were increased. However, free fatty acid both in serum and myocardium were increased, indicating that the fatty acid utilization was decreased in the left ventricle of SHR. Furthermore, SHR had the decreased expression and activity of SCAD in the left ventricle. CONCLUSION: The changes of SCAD are different in cardiac hypertrophy induced by hypertension and exercise training, indicating that SCAD may be used as a molecular marker of physiological and pathological cardiac hypertrophy, and a potential therapeutic target of pathological cardiac hypertrophy.     

Effects of PPARβ and NO on high glucose and insulin-induced cardiomyocyte hypertrophy

AIM: To investigate the role of peroxisome proliferator-activated receptor β(PPARβ)-nitric oxide(NO) signal pathway in cardiomyocyte hypertrophy induced by high glucose(25.5 mmol/L) and insulin(0.1 μmol/L)(HGI). METHODS: The cardiomyocyte hypertrophy was characterized in rat primary cardiomyocytes by measuring the cell surface area, protein content, and the mRNA expression of atrial natriuretic factor(ANF). The mRNA and protein expression were measured by real-time PCR and Western blotting, respectively. The activity of NO synthase(NOS) and NO content were measured by a reagent kit through ultraviolet spectroscopy. RESULTS: HGI induced profound change of hypertrophic morphology, and significantly increased the cell surface area, protein content and mRNA expression of ANF(P<0.01), but decreased the expression of PPARβ at mRNA and protein levels(P<0.05). At the same time, the expression of inducible NOS(iNOS) was obviously elevated(P<0.01), which occurred in parallel with the rising NOS activity and NO concentration(P<0.01). GW0742(1 μmol/L), a selective PPARβ agonist, inhibited the cardiomyocyte hypertrophy induced by HGI(P<0.01), and up-regulated the expression of PPARβ at both mRNA and protein levels. Meanwhile, GW0742 also inhibited the increases in iNOS expression, NOS activity, and NO content induced by HGI, which were abolished by GSK0660(1 μmol/L), a selective PPARβ antagonist(P<0.01). CONCLUSION: PPARβ down-regulation and the following iNOS-NO activation are involved in the cardiomyocyte hypertrophy induced by HGI.     

Inhibitory effect of fructose-1, 6-diphosphate on adriamycin-induced cardiomyocyte apoptosis in rats

AIM:To study the effect of fructose-1, 6-diphosphate (FDP) on adriamycin(ADM)-induced cardiomyocyte apoptosis in rats. METHODS:Twenty-four Wistar rats were randomly divided into three groups: control group, ADM treated group and FDP intervention group. The contents of malondialdehyde (MDA) and NO₂^-/NO₃^-, the activities of glutathione peroxidase (GPx) and superoxide dismutase (SOD) were determined by colorimetric method in myocardial tissue, and the cardiomyocyte apoptosis was detected by TUNEL method in myocardial tissue, and the expression of inducible nitric oxide synthase (iNOS) mRNA, Bcl-2 mRNA and Bax mRNA in myocardial tissue were detected by in situ hybridization. RESULTS:The contents of NO₂^-/NO₃^- and MDA in myocardial tissue, the expressive levels of iNOS mRNA and Bax mRNA in cardiomyocyes and its apoptotic amounts in FDP intervention group were significantly lower than those in ADM treated group (P＜0.01). However, the activities of SOD and GPx in myocardial tissue, the expressive level of Bcl-2 mRNA of cardiomyocytes in FDP intervention group were significantly higher than those in ADM treated group (P＜0.01). CONCLUSION:FDP antagonized the reduced expression of Bcl-2 mRNA and increased expression of Bax mRNA in myocardial tissue induced by ADM, and in turn inhibited ADM-induced cardiomyocyte apoptosis.     

PQS attenuates cardiomyocyte apoptosis induced by thapsigargin through inhibiting endoplasmic reticulum stress

AIM:To study the effect of Panax quinquefoliumsaponin (PQS) on cardiomyocyte apoptosis induced by thapsigargin (TG). METHODS:Primary cultured cardiomyocytes from neonatal SD rats were divided into control group, TG group, PQS (40 mg/L, 80 mg/L and 160 mg/L)+TG group, si-PERK+TG group, and mock+TG group. The cells were treated with 1 μmol/L TG for 24 h to induce apoptosis. The PERKgene in the cardiomyocytes was knocked down by RNAi. The cell viability was detected by CCK-8 assay. Apoptosis was analyzed by flow cytometry. Wes-tern blotting was used to determine the expression of ERS molecules GRP78, CRT, ATF4 and CHOP, anti-apoptosis protein Bcl-2 and pro-apoptosis protein Bax. RESULTS:Compared with control group, TG significantly and the apoptosis, reduced the cell viability (P<0.05), increased the phosphorylation of PERK and eIF2α, increased the expression of GRP78, CRT, ATF4, CHOP and pro-apoptosis protein Bax, and decreased the expression of anti-apoptosis protein Bcl-2 (P<0.05). Compared with TG group, PQS treatment (160 mg/L) significantly reduced the apoptosis and increased the cell viability (P<0.05). All the 3 different concentrations of PQS significantly increased the expression of anti-apoptosis protein Bcl-2 and reduced the expression of pro-apoptosis protein Bax (P<0.05) in a dose-dependent manner. PQS pretreatment and knockdown of PERK both reduced the protein levels of GRP78, CRT, PERK, p-PERK, eIF2α, p-eIF2α, ATF4, CHOP and pro-apoptosis protein Bax, and increased the expression of anti-apoptosis protein Bcl-2 (P<0.05). CONCLUSION: PQS at concentration of 160 mg/L attenuated cardiomyocyte apoptosis induced by TG. PQS had the similar effect as PERKknockdown on cardiomyocyte apoptosis. The mechanism may be associated with inhibiting PERK-eIF2α-ATF4-CHOP pathway of ERS-related apoptosis.     

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.     

Changes of aryl hydrocarbon receptor in cardiac hypertrophy induced by high glucose in vitro

AIM: To investigate the changes of aryl hydrocarbon receptor (AhR) in the process of cardiomyocyte hypertrophy induced by high glucose, and to explore its potential mechanisms. METHODS: The rat cardiomyocytes (H9c2 cells) were divided into normal glucose group, high glucose group, DMSO group and resveratrol (an AhR antagonist) group. The content and distribution of AhR were observed with immunofluorescence staining. The myocardial cells were stained with rhodamine-labeled phalloidin to visualize cytoskeleton, and the cell surface area were determined after imaging by fluorescence microscopy. The generation of reactive oxygen species (ROS) in the cardiomyocytes was measured using a fluorescent probe DCFH-DA. The mRNA expression of AhR, CYP1A1, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) were evaluated by real-time quantitative PCR (RT-qPCR). The protein levels of AhR, CYP1A1, ANP and BNP were assessed by Western blot. RESULTS: AhR was constitutively presented in the cytosol under normal-glucose condition and was translocated to the nuclei under high-glucose condition. High glucose induced cardiac hypertrophy, and increased ROS generation. Significant reductions in the cell size and ROS generation were observed after treated with resveratrol. The expression of AhR, CYP1A1, ANP and BNP at mRNA and protein levels in high glucose group was increased as compared with normal glucose group and resveratrol group, and the above-mentioned indexes significantly decreased in resveratrol group as compared with DMSO group. CONCLUSION: High glucose-induced cardiac hypertrophy increases AhR expression, which may be involved in the maintenance of glucose homeostasis in the cardiomyocytes. AhR translocation to the nucleus induced by high glucose results in the increases in CYP1A1 expression and ROS generation, which may be an important mechanism of high glucose-induced cardiomyocyte hypertrophy.     

Role of TRIM8 in mouse cardiomyocyte apoptosis induced by high glucose and high free fatty acid

AIM: To investigate the effects of tripartite motif-containing protein 8 (TRIM8) on the apoptosis of mouse cardiomyocytes (MCMs) induced by high glucose and high free fatty acid (HGHF) and the underlying mechanism. METHODS: The MCMs were divided into normal glucose (NG) group (glucose at 5.5 mmol/L), high glucose (HG) group (glucose at 33 mmol/L), high free fatty acid (HF) group (sodium palmitate at 300 μmol/L) and HGHF group (glucose at 33 mmol/L and sodium palmitate at 300 μmol/L). The expression of TRIM8 in the MCMs was knocked down by siRNA, and the MCMs was further divided into control group, scrambled siRNA (Scra-siRNA)/PBS group, TRIM8-siRNA/PBS group, Scra-siRNA/HGHF group and TRIM8-siRNA/HGHF group. To further confirm the specific mechanism of TRIM8 in the MCM injury induced by HGHF, the MCMs were subgrouped into HGHF/DMSO group, HGHF+TRIM8-siRNA+DMSO (HGHF+Ts/DMSO) group, HGHF/ML385 group and HGHF+Ts/ML385 group. Accordingly, apoptosis was analyzed by flow cytometry, and the levels of reactive oxygen species (ROS) were measured by flow cytometry and DHE staining. The expression of TRIM8, nuclear factor E2-related factor 2 (Nrf2), glutamate-cysteine ligase catalytic subunit (GCLC), heme oxygenase-1 (HO-1) and NAD(P)H:quinone oxidoreductase 1 (NQO-1) at mRNA and protein levels was determined by qPCR and Western blot. RESULTS: HGHF increased the expression of TRIM8, and suppressed the expression of Nrf2, GCLC, HO-1 and NQO-1 in the MCMs (P < 0.05). Compared with Scra-siRNA/HGHF group, the intracellular ROS content and apoptotic rate were decreased in TRIM8-siRNA/HGHF group (P < 0.05). Correspondingly, the expression of the antioxidant molecule Nrf2 and its downstream genes GCLC, HO-1 and NQO-1 was increased (P < 0.05). In contrast, the addition of Nrf2 inhibitor ML385 partially reversed the inhibitory effect of TRIM8 expression knock-down on HGHF-induced apoptosis of MCMs. CONCLUSION: TRIM8 exacerbates the HGHF-induced cardiomyocyte apoptosis by modulating Nrf2 antioxidative pathway.     

Panax quinquefolium saponins attenuate hypoxia/reoxygenation-induced injury in rat cardiomyocytes

AIM： To investigate the effects of Panax quinquefolium saponins (PQS) and calcineurin (CaN) signal pathway on cardiomyocyte injury induced by myocardial hypoxia/reoxygenation(H/R). METHODS：Cultured cardiomyocytes isolated from neonatal Sprague-Dawley rats were used to establish the H/R model. The cells were transfected with pCDB-CaN plasmid to overexpress CaN, or exposed to the CaN inhibitor FK506 to interfere the CaN expression. The cardiomyocytes were divided into control group, H/R group, PQS+H/R group, CaN+PQS+H/R group, pCDB+PQS+H/R group and FK506+PQS+H/R group. The apoptosis was analyzed by flow cytometry. The activity of CaN in the cardiomyocytes was detected. The protein expression of CaN was determined by Western blotting. RESULTS：Compared with control group, the apoptosis of the cardiomyocytes in CaN group was significantly increased. Compared with PQS+H/R group, the cell apoptosis, the expression of Bcl-2 and Bax, the activity of CaN and its protein expression in FK506 group were not significantly different. CONCLUSION：Inhibition of CaN activity reduces the H/R injury in cardiomyocytes. However, the mechanism of PQS protecting cardiomyocytes from H/R injury may not be associated with the CaN signaling pathway.     

Proctective effect of microRNA-30a on regulating beclin-1 expression in hypoxia-reoxygenated neonatal rat cardiomyocytes

AIM: To explore the potential mechanism of microRNA-30a (miR-30a) overexpression in neonatal rat cardiomyocytes during hypoxia/reoxygenation (H/R). METHODS: The miR-30a overexpression was induced in primary neonatal rat cardiomyocytes by lentivirus transfection. The cardiomyocytes were divided into 5 groups: normal group, H/R group, LV-GFP+H/R group, LV-GFP-miR-30a+H/R group and 3-methyladenine(3-MA)+H/R group. The expression level of miR-30a after lentivirus transfection and H/R was determined by real-time PCR, while the protein levels of LC3 and Beclin-1 after H/R and lentivirus transfection were detected by Western blotting. The cardiomyocyte death after H/R were measured by TUNEL and PI staining. RESULTS: Compared with LV-GFP group, significant down-regulation of Beclin-1 protein level was observed in cardiomyocytes with miR-30a overexpression, while the protein levels of Beclin-1 and LC3 in the cardiomyocytes with miR-30a overexpression were down-regulated after H/R, and apoptosis of these cells were significantly decreased after H/R. CONCLUSION: The protein level of Beclin-1 is down-regulated in cardiomyocytes with miR-30a overexpression. Inhibition of autophagy decreases the cardiomyocyte death after H/R.     

矮化中间砧‘宫藤富士’苹果栽植密度对树体生长、冠层光照和果实产量的影响

以2011年春季定植的矮化中间砧苹果成品苗(3年根1年干的‘宫藤富士’/SH6/平邑甜茶)为试材,设置7种不同的栽植密度(株行距分别为1 m×3 m、1.5 m×3 m、2 m×3 m、0.75 m×4 m、1 m×4 m、1.25 m×4 m和1.5 m×4 m),细纺锤形整枝修剪,自栽植第2年,连续7年调查7种栽植密度对树体生长、冠层光照分布、果实产量和品质的影响。随着树龄的增长,不同栽植密度下树干粗度和总枝量逐年增加,不同处理间树干粗度无显著差异,第7年1 m×3 m和0.75 m×4 m两个栽植密度下树体总枝量超过140万条·hm-2,第8年均超过140万条·hm^-2。栽植前期(第2～4年)各栽植密度树体短枝比例不断增加,长枝比例不断减少,第5年各栽植密度枝类组成趋于稳定;综合稳产3年(第6～8年)树体的枝类组成数据,4 m行距的短枝比例明显高于3 m行距,长枝比例略低。树体冠层平均相对光照强度由高到低的株行距处理依次为1.5 m×4 m(63.87%)、1.25 m×4 m(61.44%)、2 m×3 m(61.27%)、1 m×4 m(59.19%)、...     

Effects of T-cadherin on hypoxia/reoxygenation-induced apoptosis of cardiomyocytes from neonatal rats

AIM: To investigate the mechanism of cardiomyocyte apoptosis induced by hypoxia/reoxygenation (H/R) by silencing a new adiponectin receptor T-cadherin through adenovirus-mediated RNA interference. METHODS: The primary cardiomyocytes were isolated from neonatal rats and cultured for 72 h. The cardiomyocytes were randomly divided into control group, H/R group, APN+H/R group, Ad-T-cadherin-siRNA+APN+H/R group and Ad-HK (adenovirus negative control)+APN+H/R group. The transfection ability and efficiency were examined. The expression of T-cadherin at mRNA and protein levels was detected by RT-PCR and Western blotting. The apoptotic rate was analyzed by flow cytometry and TUNEL. RESULTS: High purity of neonatal rat cardiomyocytes was obtained by primary culture. After 48 h, over 90% of myocardiocytes were infected at MOI=100. The transfected myocardiocytes showed a low expression level of T-cadherin under normal physiological condition. Compared with APN+H/R group, the cell apoptotic rate significantly increased in Ad-T-cadherin-siRNA+APN+H/R group (P<0.05). Compared with H/R group, the difference was not statistically significant (P>0.05). CONCLUSION: Ad-T-cadherin-siRNA effectively infects myocardial cells in vitro and successfully reduces the expression of T-cadherin in myocardial cells. The inhibitory effect of adiponectin on H/R-induced cardiomyocyte apoptosis is attenuated by decreasing the expression of T-cadherin.