苹果miR396家族鉴定及在不定根发育过程中的表达分析

分析了苹果miR396家族进化特性及其在苹果不定根发育过程中的表达模式。结果表明：苹果miR396家族有4条成熟体和7条前体序列（pre-miRNA）。Mfold预测显示Pre-miR396家族7个成员序列均可形成典型稳定的茎环二级结构,最小折叠自由能介于–62.9 kal ? mol-1（pre-miR396b）~–51.9 kal ? mol-1（pre-miR396g）之间。系统发育进化树分析显示,pre-miR396家族亲缘关系可分为3个亚组（G1、G2、G3）,每个亚组内基因数量不同,分别含有11、9、19个。靶基因预测显示,苹果miR396靶基因包括MdGRF1、MdGRF2和MdGRF5等,降解组测序进一步验证了miR396对其候选靶基因MdGRF1、MdGRF2和MdGRF5的剪切关系。苹果miR396家族成员在侧根和果实中的表达量显著高于其他组织,其候选靶基因表达量则在花芽和腋芽中显著高于其他组织;不定根发育过程中,miR396家族不同成员表达模式存在显著差异,整体上呈上调表达趋势,其候选靶基因呈下调表达趋势;外源IBA处理显著诱导miR396家族成员的表达,尤其是在不定根诱导期和根系生长期更为显著。     

Prostaglandin E1 protects heart after myocardial infarction by inhibiting endoplasmic reticulum stress in rats

AIM To investigate the effect of prostaglandin E1 (PGE1) on heart after myocardial infarction (MI) in rats and its related molecular mechanism. METHODS Fifty male SD rats were divided into sham group, model group and model+PGE1 group. The MI rat model was established by ligation of left anterior descending coronary artery. Cardiac function in the rats was detected by echocardiogaphy. The myocardial histomorphologic changes were evaluated by HE and Masson staining. The MI area was measured by TTC staining. The cardiomyocyte death was detected by TUNEL staining. The protein levels of endoplasmic reticulum stress (ERS)-related factors glucose-regulated protein 78 (GRP78), C/EBP homologous protein (CHOP) and caspase-12, and apoptosis-related factors Bcl-2, Bax and cleaved caspase-3 were determined by immunofluorescence staining and Western blot. RESULTS Compared with sham group, the cardiac function in model group was decreased, with significant myocardial pathological changes. The MI area was enlarged, and the death of cardiomyocytes was promoted. The protein levels of GRP78, CHOP, caspase-12, Bax and cleaved caspase-3 in the myocardial tissues were significantly increased, while Bcl-2 was decreased (P<0.01). Compared with model group, the cardiac function in model+PGE1 group was significantly improved, and the myocardial pathological damage was significantlty attenuated. The MI area and myocardial cell death were significantly reduced. The protein levels of GRP78, CHOP, caspase-12, Bax and cleaved caspase-3 in the myocardial tissues were significantly decreased, while Bcl-2 was increased (P<0.01). CONCLUSION PGE1 reduces collagen deposition and inflammation, and improves cardiac function by reducing ERS level, thus protecting cardiomyocytes from MI damage.     

Protective effect of curcumin on ERS-induced NASH rat hepatocytes

AIM: To investigate whether curcumin reduces hepatocyte apoptosis in the rats with non-alcoholic steatohepatitis (NASH) by inhibiting endoplasmic reticulum stress (ERS) and thus exerting a protective effect on the liver. METHODS: Male SD rats (n=30) were randomly divided into normal control group (n=10), model group (n=10) and curcumin group (n=10). NASH model was established by feeding the rats with high-fat diet for 4 weeks. The rats in curcumin group was given curcumin (200 mg/kg) daily by gavage, while the rats in model group and normal control group were given the same volume of saline. Four weeks later, the rats were killed, and their blood and liver tissues were collected. The serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were detected, liver histopathological changes were observed by HE staining, the expression of glucose-regulated protein 78 (GRP78) and C/EBP homologous protein (CHOP) was determined by Western blot, and apoptosis was detected by TUNEL method. RESULTS: Compared with normal control group, the levels of serum ALT and AST in model group were significantly increased, and the levels of serum ALT and AST in curcumin group were significantly lower than those in model group (P<0.05). At the same time, the steatosis and inflammation of hepatocytes in curcumin group were less than those in model group, and no obvious necrosis was observed. Compared with normal control group, the protein expression levels of GRP78 and CHOP in model group were increased, while the protein expression levels of GRP78 and CHOP in curcumin group were decreased compared with model group (P<0.01). TUNEL results showed that apoptotic hepatocytes in model group were significantly more than those in normal control group, while those in curcumin group were significantly fewer than those in model group. CONCLUSION: Hyperlipidemia induces excessive ERS in the hepatocytes, thus triggering apoptosis and leading to NASH. The mechanism of curcumin reducing hepatocyte apoptosis may be related to its inhibition of ERS.     

Protective effect of morinda officinalis oligosaccharides monomer HexB on hypoxia/reoxygenation-induced injury in HUVECs

AIM:To explore the protective effect of morinda officinalis oligosaccharides monomer HexB on hypoxia/reoxygenation (H/R)-induced injury in human umbilical vein endothelia cells (HUVECs). METHODS:HUVECs were treated with HexB, 4-phenylbutyric acid (4-PBA) and thapsigargin (TG), respectively. The cells were divided into control group, HexB group, H/R group, HexB+H/R group, 4-PBA+H/R group, TG group and HexB+TG group. The cell viability was measured by CCK-8 assay. The apoptotic rate was detected by flow cytometry. Western blot was used to determine the protein levels of endoplasmic reticulum stress (ERS) related molecules chaperone protein glucose-regulated protein 78 (GRP78), C/EBP homologous protein (CHOP), apoptosis-related protein caspase-12 and phosphorylated c-Jun NH₂-terminal kinase (p-JNK). RESULTS:The viability of HUVECs was reduced in H/R group and TG group (P<0.05), increased in HexB+H/R, 4-PBA+H/R and HexB+TG group (P<0.05). The apoptotic rate, the protein levels of GRP78, CHOP, caspase-12 and p-JNK were increased in H/R group and TG group (P<0.05), weakened in the HexB+H/R group (P<0.05), 4-PBA+H/R group and HexB+TG group (P<0.05). No significant change in the apoptotic rate, cell viability, protein levels of GRP78, CHOP, caspase-12, p-JNK between HexB+H/R group and 4-PBA+H/R group was observed. CONCLUSION:HexB attenuates HUVECs injury caused by H/R through suppressing ERS and apoptosis. The possible mechanism may be involved in the apoptotic pathways related to GRP78, CHOP, caspase-12 and p-JNK.     

Effects of bisoprolol plus peridopril on myocardial endoplasmic reticulum stress in rats with doxorubicin-induced heart failure

ATM: To investigate the effects of bisoprolol (Bis) plus peridopril (Per) on myocardial endoplasmic reticulum stress (ERS) in rats with heart failure.METHODS: Male SD rats were randomly divided into normal control group, sham group, doxorubicin (DOX) group, bisoprolol treatment group (DOX+Bis group), peridopril treatment group (DOX+Per group) and bisoprolol plus peridopril treatment group (DOX+Bis+Per group). A rat model of heart failure was induced by intraperitoneal injection of DOX. Distilled water, bisoprolol, peridopril, and bisoprolol plus peridopril were administrated by gastric gavage for 35 d, respectively. The indexes of cardiac functions and plasma levels of brain natriuretic peptide (BNP) were measured, myocardial apoptosis was analyzed by TUNEL assay and myocardial protein expression of GRP78, CHOP, JNK, caspase-12 and SERCA2a was detected by Western blot.RESULTS: Compared with normal control group and sham group, cardiac output (CO), left ventricular fractional shortening (FS), and left ventricular ejection fraction (EF) of the rats in DOX group decreased significantly (P<0.01), the cardiomyocyte apoptotic index increased significantly (P<0.01), the myocardial protein levels of SERCA2a decreased significantly, and GRP78, CHOP, JNK and caspase-12 increased significantly (P<0.01). Compared with DOX group, CO, FS and EF of the rats in DOX+Bis group, DOX+Per group and DOX+Bis+Per group increased significantly (P<0.01), cardiomyocytes apoptotic indexes in DOX+Bis group, DOX+Per group and DOX+Bis+Per group decreased significantly (P<0.01), myocardial protein levels of SERCA2a in DOX+Bis group, DOX+Per group and DOX+Bis+Per group increased significantly, while GRP78, CHOP, SERCA2a, JNK and caspase-12 decreased significantly (P<0.05). Indicators except JNK in DOX+Bis+Per group were changed more significantly than those in DOX+Bis group or DOX+Per group (P<0.05).CONCLUSION: Bisoprolol plus peridopril therapy improves cardiac functions in a rat model of doxorubicin-induced heart failure with more significant effectiveness than using bisoprolol or peridopril alone, which may be related to inhibition of myocardial ERS and apoptosis.     

Protective effect of Yiqi-Wenyang-Huoxue-Huatan formula on HHPH rat brain by suppressing excessive endoplasmic reticulum stress

AIM: To investigate whether Yiqi-Wenyang-Huoxue-Huatan formula (YWHHF) attenuats brain injury induced by hypoxia-hypercapnia pulmonary hypertension (HHPH) in the rats by inhibiting excessive endoplasmic reticulum stress response. METHODS: Healthy SPF male SD rats (n=50) were randomly divided into 5 groups: control group, hypoxia-hypercapnia group, low-dose YWHHF group, middle-dose YWHHF group and high-dose YWHHF group. The rats in control group lived in normal environment, while the rats in other 4 groups were raised for 4 weeks in oxygen tank with low oxygen concentration and high CO₂ concentration. YWHHF was perfused in the rats of low-, middle-and high-dose groups at 0.15, 0.3 and 0.6 g/kg daily, respectively. The rats in hypoxia-hypercapnia group were given isometric distilled water. The surgery was performed on the rats after 4 weeks, and the brain and lung tissues were quickly collected to detect brain water content and observe the morphological changes after mean pulmonary artery pressure recording and heart perfusion. The caspase-3 activity and the apoptotic index of the brain cells were determined. The expression of c-Jun N-terminal kinase (JNK), caspase-12, C/EBP homologous protein (CHOP) and glucose-regulated protein 78 (GRP78) at protein and mRNA levels in brain tissues was detected by Western blot and RT-PCR. RESULTS: Compared with control group, mean pulmonary artery pressure, brain water content, brain apoptotic index, caspase-3 activity, and the protein and mRNA levels of JNK, caspase-12, CHOP and GRP78 in the rest 4 groups were increased, and the brain and lung tissues had obvious damage under light microscope. Compared with hypoxia-hypercapnia group, mean pulmonary artery pressure, brain water content, brain apoptotic index, caspase-3 activity, and the protein and mRNA expression of JNK, caspase-12, CHOP and GRP78 in low-, middle-and high-dose YWHHF groups were decreased, and the pathological damage of the brain and lung tissues was obviously reduced under light microscope. These changes in middle-dose YWHHF group were the most significant. CONCLUSION: YWHHF effectively relieves the brain injury induced by HHPH in rats, which may be associated with inhibiting excessive endoplasmic reticulum stress response.     

Role of glucose-regulated protein 78 in pulmonary microvascular remodeling during development of rat hepatopulmonary syndrome

AIM：To explore the mechanisms of pulmonary microvascular remodeling induced by glucose-regulated protein 78 (GRP78) in the development of hepatopulmonary syndrome (HPS) in rats. METHODS：The Wistar rats were randomly divided into HPS groups at the 4th, 6th and 8th weeks, and normal control groups at the corresponding time points. The rat model of HPS produced in the process of liver cirrhosis was induced by employing multiple pathogenic factors to the animals. The rats in normal control group were designed by feeding with standard diet and mineral water. The expression of GRP78, factor Ⅷ- related antigen (FⅧ-RAg), C/EBP homologous protein (CHOP, also called growth arrest and DNA damage-inducible protein 153, GADD153), caspase-12, Bcl-2 and nuclear factor (NF)-κB in the lung tissues were measured by immunohistochemistry. The expression of VEGF at mRNA and protein levels in the lungs was measured by the methods of RT-PCR and Western blotting, respectively. RESULTS：The expression of GRP78, FⅧ-RAg,VEGF and Bcl-2 proteins was gradually increased with the HPS development. The protein expression of NF-κB was also gradually increased, especially in nucleus. GRP78 protein in the lung was positively correlated with the expression of FⅧ-RAg and VEGF, but negatively correlated with the expression of CHOP/GADD153 and caspase-12. The protein levels of GRP78 and FⅧ-RAg, and VEGF at both mRNA and protein levels were higher, and the protein levels of CHOP/GADD153 and caspase-12 were lower in the rats with HPS at every time point than those in normal control rats (P<0.05). CONCLUSION：Overexpression of GRP78 in the lung may be the critical pathogenesis of HPS by promoting cell survival and proliferation, and inhibiting cell apoptosis, thus leading to pulmonary microvascular remodeling.     

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.     

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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     

Fuzi polysaccharide protects neonatal rat cardiomyocytes with hypoxia-reoxygenation by inhibiting endoplasmic reticulum stress

AIM: To investigate whether the protection mechanism of Fuzi polysaccharide (FPS) is related to inhibition of endoplasmic reticulum stress in cultured neonatal rat cardiomyocytes with hypoxia/reoxygenation (H/R). METHODS: Cultured rat myocardial cells were divided into control group, H/R group (hypoxia for 3 h and reoxygenation for 6 h) and different concentrations of FPS (0.1 g/L, 1 g/L, 10 g/L or 20 g/L) +H/R groups. The cell survival was detected by MTT assay and cell apoptosis of cardiomyocytes was measured by flow cytometry using Annexin V-FITC staining. The expression of glucose-regulated protein 78 (GRP78), CCAAT/enhancer-binding protein homologous protein (CHOP) and caspase-12 were determined by Western blotting. The mRNA expression of CHOP and caspase-12 was detected by quantitative PCR. RESULTS: After reoxygenation, the expression of GRP78, CHOP and caspase-12 in cardiomyocytes was increased. Compared with H/R group, the expression of GRP78, CHOP and caspase-12 in FPS+H/R groups was significantly inhibited, the survival rate of cardiomyocytes was increased and the apoptosis of cardiomyocytes was inhibited. This protective effect of FPS was in a dose-dependent manner and reached its peak at 10 g/L. CONCLUSION: Fuzi polysaccharide protects cardiomyocytes from H/R injury. The mechanism is related to inhibiting endoplasmic reticulum stress.     

Effects of naringenin on PI3K/AKT signaling pathway and endoplasmic reticulum stress and its related apoptotic pathways in rats with myocardial ischemia/reperfusion injury

AIM To observe the effect of naringenin on cardiac injury in ischemia/reperfusion （I/R） rats, and to explore whether the role of naringenin is involved in PI3K/AKT signaling pathway and endoplasmic reticulum stress and its related apoptotic pathways. METHODS SD rats （n=48） were randomly divided into sham operation （sham） group, model （I/R） group, naringenin treatment （NAR） group and naringenin+LY294002 （NL） group. Myocardial I/R injury model was prepared by ligation of left anterior descending coronary artery of rats for 30 min followed by reperfusion for 120 min. After reperfusion, the serum levels of cardiac troponin I （cTnI） was measured by ELISA. HE staining, TTC staining and TUNEL staining were used to detect the myocardial histopathological changes, myocardial infarction area and myocardial cell apoptotic rate. The mRNA levels of endoplasmic reticulum stress-related indicators glucose-regulated protein 78 （GRP78）, C/EBP homologous protein （CHOP） and caspase-12 were detected by RT-qPCR. The protein levels of cleaved caspase-3, GRP78, CHOP, caspase-12, p-PI3K and p-AKT were determined by Western blot. RESULTS Compared with I/R group, the serum content of cTnI, myocardial pathological damage, myocardial infarction area and myocardial cell apoptotic rate were significantly reduced （P<0.05）, the protein levels of cleaved caspase-3, GRP78, CHOP and caspase-12 were decreased （P<0.05）, and the protein levels of p-PI3K and p-AKT were increased in NAR group （P<0.05）. LY294002 attenuated the protective effect of naringenin to some extent. CONCLUSION Naringenin reduces myocardial I/R injury in rats possibly by activating PI3K/AKT signaling pathway and subsequently regulating endoplasmic reticulum stress and its related apoptotic pathways.     

Dexmedetomidine reduces renal injury induced by lung ischemia/reperfusion in mice through inhibiting endoplasmic reticulum stress response

AIM:To investigate the effect of dexmedetomidine (DEX) on renal injury induced by lung ischemia/reperfusion (I/R) in mice and its relationship with endoplasmic reticulum stress response.METHODS:Healthy SPF male C57BL/6J mice,weighing 20~24 g,aged 8~10 weeks,were randomly divided into 5 groups (n=10 each):sham operation group (sham group),I/R group,atipamezole (Atip) group,DEX group,and DEX+Atip group.In vivo lung I/R model was established by occlusion of the left pulmonary artery for 30 min followed by 180 min of reperfusion in the mice.The Atip (250 μg/kg),DEX (20 μg/kg) and DEX+Atip were intraperitoneally infused into the mice before left pulmonary hilus was blocked in Atip group,DEX group and DEX+Atip group,and other operations were the same as I/R group.After experiment,the mice were killed,and the renal tissues were harvested to observe the morphological changes.The enzymatic activity of caspase-3,serum creatinine and blood urea nitrogen,and cell apoptotic index of the renal cells were also analyzed.The expression of c-Jun N-terminal kinase (JNK),caspase-12,CCAAT/enhancer-binding protein homdogous protein (CHOP) and glucose-regulated protein 78(GRP78) at mRNA and protein levels in the renal tissues was determined by RT-PCR and Western blot.RESULTS:Compared with sham group,the enzymatic activity of caspase-3,serum creatinine and blood urea nitrogen,renal cell apoptotic index,and the mRNA and protein levels of JNK,caspase-12,CHOP and GRP78 in I/R group were significantly increased (P<0.01),and the renal tissues had obvious damage under light microscope.Compared with I/R group,Atip group and DEX+Atip group,the enzymatic activity of caspase-3,serum creatinine and blood urea nitrogen,renal cell apoptotic index,and the mRNA and protein levels of JNK,caspase-12 and CHOP in DEX group were significantly decreased,and the expression level of GRP78 significantly increased (P<0.01).Furthermore,the renal tissue damage was obvious reduced.CONCLUSION:DEX effectively relieves the renal injury induced by lung I/R in mice,which may be associated with exciting α₂-adrenergic receptor and inhibiting endoplasmic reticulum stress response.     

Panax quinquefolium saponins protect rat myocardium from ischemia/reperfusion injury through attenuating excessive endoplasmic reticulum stress

AIM: To investigate whether excessive endoplasmic reticulum stress (ERS) is involved in the protective mechanism of Panax quinquefolium saponins (PQS) against ischemia/reperfusion (I/R) injury in rat myocardium. METHODS: The model of myocardial I/R injury in vivo was made by ligating the left anterior descending artery for 45 min followed by 24 h of reperfusion in SD rats. The hemodynamics and serum content of cardiac troponin T (cTnT) were measured. The myocardial infarct size was measured by Evans blue and 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. Cardiomyocyte apoptosis was detected using in situ TDT-mediated dUTP nick end labeling (TUNEL). The protein levels of glucose-regulated protein 78 (GRP78), calreticulin (CRT), C/EBP homologous protein (CHOP), caspase-12, apoptosis-associated proteins Bax and Bcl-2 were determined by Western blotting. RESULTS: Compared with I/R group, the mean arterial pressure in PQR+IR group was decreased by 32.0%, and left ventricular±dp/dtmax was increased by 64.0% and 35.0%, respectively.The serum content of cTnT was decreased by 53.3%, the percentage of area of necrosis (AN)/area at risk (AAR) was reduced by 65.5% and the apoptosis rate was decreased by 54.9%.The myocardial pathological changes were improved. Bcl-2 protein expression was increased by 110.0% and that of Bax was decreased by 47.8%. CRT protein expression was decreased by 43.4 %, CHOP protein expression and the protein level of cleaved caspase-12 were decreased by 38.6% and 23.7% in PQS+I/R group. CONCLUSION: PQS alleviates I/R injury in myocardium by inhibition of excessive ERS.     

Protective effect of Huangqi injection combined with puerarin injection on myocardium of type 2 diabetes mice

AIM:To investigate the effect of Huangqi injection combined with puerarin injection on the myocardium of the mice with type 2 diabetes. METHODS:Diabetic KKAy mice were randomly divided into model group and treatment group (Huangqi injection combined with puerarin injection). The male KKAy mice of the same age were used as control group. All mice were sacrificed at 21, 24 and 28 weeks. Morphological changes of the myocardium were observed by HE staining. Apoptosis of the cardiomyocytes was measured by TUNEL staining. The mRNA levels of glucose-regulated protein 78(GRP78), C/EBP hoinologous protein (CHOP) and p53 up-regulated modulator of apoptosis (PUMA) were detected by real-time PCR, and the protein levels of GRP78, CHOP, PUMA, caspase-3, cleaved caspase-3, caspase-9, cleaved caspase-9, poly(ADP-ribose) polymerase (PARP) and cleaved PARP were determined by Western blot. RESULTS:Cardiomyocyte hypertrophy, partly dissolved sarcoplasm and necrosis were observed in model group, and these lesion were alleviated in treatment group. Obvious increased apoptosis in model group and significantly decreased apoptosis of cardiomyocytes in treatment group was observed (P<0.05). At 21, 24 and 28 weeks, the mRNA and protein levels of GRP78, CHOP and PUMA and the protein levels of cleaved caspase-3, cleaved caspase-9 and cleaved PARP in model group were increased significantly as compared with control group (P<0.01), and these in treated group were decreased compared with model group. CONCLUSION:Huangqi injection combined with puerarin injection has cardioprotective effects on type 2 diabetes mice and its mechanism of the action was implemented via inhibiting the activation of endoplasmic reticulum stress and caspase pathway, thus resulting in suppressed 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.     

PERK-mediated endoplasmic reticulum stress is involved in angiotensin II-induced myocardial hypertrophy

AIM: To investigate the role of protein kinase R-like endoplasmic reticulum kinase (PERK)-mediated endoplasmic reticulum stress (ERS) in angiotensin II (AngⅡ) -induced myocardial hypertrophy. METHODS: In the hypertrophy model of AngⅡ-induced cardiomyocytes isolated from neonatal Sprague-Dawley rats, the methods of morphological observation, [³H]-leucine incorporation and surface area measurement were employed to assess the cardiomyocyte hypertrophy. Real-time PCR, RT-PCR and Western blotting were used to detected the expression of glucose-regulated protein 78 (GRP78), calreticulin (CRT), PERK, eukaryotic initiation factor 2α (eIF2α) and C/EBP homologous protein (CHOP) at mRNA and protein levels. RESULTS: Compared with control group, Ang II-treated cardiomyocytes showed that the mRNA and protein expression of CRT increased by 146.4% and 125.3%, respectively (P<0.05). The mRNA and protein expression of GRP78 increased by 84.0% and 77.6%, respectively (P<0.05). The mRNA and protein expression of PERK increased by 165.4% and 132.1%, respectively (P<0.05).The mRNA and protein expression of eIF2α was increased by 110.9% and 46.5%, respectively (P<0.05). The mRNA and protein expression of CHOP also increased by 117.7% and 63.3%, respectively (P<0.05). CONCLUSION: PERK-mediated ERS response is involved in AngⅡ-induced cardiomyocyte hypertrophy.     

Glucose-regulated protein 78 promote cardiomyocyte apoptosis in liver cirrhotic rats

AIM: To explore the cardiomyocyte apoptosis induced by glucose-regulated protein 78/immunoglobulin heavy chain-binding protein (GRP78/BiP) in liver cirrhotic rats and its mechanism. METHODS: The liver cirrhotic rat model was established with multiple pathogenic factors, and sampled at the time points of 4 weeks, 6 weeks and 8 weeks. In experiment 1, the heart was collected and weighed, the thickness of the left ventricular wall was measured, and the ratios of the left ventricular wall thickness to the heart weight, and the heart weight to the body weight were calculated. In experiment 2, TUNEL was used to detect the apoptotic cardiomyocytes,and the protein levels of GRP78/BiP, CCAAT/enhancer-binding protein homologous protein/growth arrest and DNA damage-inducible protein 153 (CHOP/GADD153), caspase-12, nuclear factor-κB p65 (NF-κB p65) and B-cell lymphoma/leukemia-2 (Bcl-2) in the myocar-dium were detected by the method of immunohistochemistry. RESULTS: Compared with the normal myocardium, significant larger ratios of the left ventricular wall thickness to the heart weight and the heart weight to the body weight, a significant increase in the cardiomyocyte apoptosis, and a significant larger positive expression index of GRP78/BiP in the hearts 8 weeks after modeling were observed. The protein levels of CHOP/GADD153 and caspase-12 were gradually increased during the development of liver cirrhosis and were significantly increased at 8 weeks. The positive expression of NF-κB p65 and Bcl-2 showed consistent changes, and were markedly higher at 4 weeks than those at the other time points. The positive expression index of GRP78/BiP was positively correlated with the apoptotic index, and the levels of CHOP/GADD153 and caspase-12. The positive expression index of CHOP/GADD153 was negatively correlated with NF-κB p65 and Bcl-2. CONCLUSION: Elevated expression of GRP78/BiP may play a crucial role in the pathogenesis of liver cirrhotic cardio-myopathy mediated by endoplasmic reticulum stress.     

Effect of endoplasmic reticulum stress on cardiac myocyte apoptosis in mouse congestive heart failure induced by myocardial infraction

AIM: To explore the effect of endoplasmic reticulum stress on cardiac myocyte apoptosis in mouse congestive heart failure induced by myocardial infraction.METHODS: The mouse model of heart failure was established by ligating the left anterior descending coronary to produce acute myocardial infarction. Thirty-two mice were divided into 4 groups: sham group and groups of post-operation at time points of 2, 4 or 6 weeks, respectively. The ventricular dilatation and left ventricular functions were assessed by echocardiography. The expression of GRP78, CHOP, caspase-12, cleaved caspase-12, JNK and phosphorylated-JNK was detected by Western blotting. The cardiac myocyte apoptosis was determined by terminal deoxynucleotidyl transferase mediated dUTP-biotin nick end labeling (TUNEL).RESULTS: The cardiac expression of endoplasmic reticulum chaperones GRP78 was significantly increased in the hearts with functional failure. The upregulated expression of CHOP, phosphorylated-JNK and cleaved caspase-12 illuminated that the CHOP-JNK- caspase-12 dependent pathways for endoplasmic reticulum-initiated apoptosis were activated in the heart with functional failure by myocardial infraction. CONCLUSION: These findings suggest that the congestive heart failure induced by myocardial infraction is associated with endoplasmic reticulum stress and activation of CHOP, JNK, caspase-12 dependent pathways for endoplasmic reticulum-initiated apoptosis.     

Effect of grape seed proanthocyanidin on cisplatin-induced nephrotoxicity in mice

AIM:To study the protective effect of grape seed proanthocyanidin (GSP) on nephrotoxicity induced by cisplatin (CP) in mice. METHODS:Male C57 mice were randomly divided into 4 groups: control group (N group, n=10), CP group (intraperitoneal injection of CP at dose of 20 mg/kg, n=20), GSP group (intragastric administration of GSP at dose of 500 mg/kg, n=15) and CP+GSP group (intragastric administration of GSP 30 min prior to intraperitoneal injection of CP and intragastric administration of the same dose of GSP 72 h later, n=20). On the 5th day after CP treatment, blood and kidney samples were collected. The renal pathological changes were examined by HE staining. The protein expression of glucose-regulated protein 78 (GRP78) and phosphorylated extracellular signal-regulated kinase (p-ERK) was examined by Western blotting and immunohistochemical staining. RESULTS:Renal index, blood urea nitrogen and serum creatinine were significantly higher in CP group than those in N group. The renal tissues were heavily damaged in CP group. The protein expression of GRP78 and p-ERK increased in CP group compared with N group. GSP treatment alleviated the increase in the renal index, blood urea nitrogen and serum creatinine. The damages of the renal tissues were attenuated. The protein expression of GRP78 and p-ERK was obviously reduced in CP+GSP group compared with CP group. CONCLUSION:GSP prevents kidney from CP-induced damage by suppression of endoplasmic reticulum stress.     

Effect of hyperlipidemia and influence of simvastatin on endoplasmic reticulum stress in rat kidney

AIM: To investigate the role of endoplasmic reticulum stress in renal injury caused by hyperlipidemia and the influence effect of simvastatin. METHODS: Thirty male Wistar rats were randomly divided into three groups: rats in control group (n=10) were fed with normal diet; rats in high fat group (n=10) were fed with high fat diet; animals in simvastatin+high fat group (n=10) were fed with high fat diet and were received simvastatin 10 mg·kg-1·d-1 by gastric irrigation. After 18 weeks, the quantitative urine protein in 24 h, the serum cholesterol and triglycerides levels were tested. The pathological changes of renal tissue were observed under optic microscope. The expressions of GRP78 and p-JNK in renal tissues were examined by immunohistochemistry. The apoptotic cells in the kidney were detected by TUNEL staining. The mRNA expressions of GRP78 and CHOP were examined by RT-PCR. RESULTS: The quantitative urine protein in 24 h, the serum lipid, the expressions of GRP78 and p-JNK proteins, the mRNA expressions of GRP78 and CHOP as well as the apoptotic cells in renal tissues were increased in high fat group (P<0.01).The quantitative urine protein in 24 h, the serum lipid, the expression of GRP78 and p-JNK proteins, the mRNA expressions of GRP78 and CHOP as well as the apoptotic cells in renal tissues were remarkably reduced in simvastatin+high fat group than those in high fat group (P<0.05). CONCLUSION: The endoplasmic reticulum stress is engaged in the renal injury caused by hyperlipidemia. The simvastatin play a role in renal protection by inhibiting the endoplasmic reticulum stress in the kidney.