Effects of orlistat on the viability of human gallbladder cancer cells / 中华消化外科杂志

Objective:To investigate the effects of orlistat on the viability of human gall-bladder cancer (GBC) cells.Methods:The experimental study was conducted. The human GBC NOZ cells with high expression of FSAN was screened out through in vitro cultivating human GBC-SD, SGC-996 and NOZ cells. The cell proliferation assay, clone formation assay and protein detection experiment were used to analysis of the effects of orlistat on the viability of human GBC cells. Cell grouping: NOZ cells cultured with medium were set as the control group, cultured with medium + 10 μmol/L orlistat were set as the low-dose orlistat group, cultured with medium + 100 μmol/L orlistat were set as the high-dose orlistat group, respectively. Observation indicators: (1) expression of FASN protein in human GBC cells; (2) effects of orlistat on the proliferation of human GBC NOZ cells; (3) effects of orlistat on apoptosis of human GBC NOZ cells. Measurement data with normal distribution were represented as Mean± SD, the ANOVA test was used for comparison between groups and the least significant difference method was used for pairwise comparison. Results:(1) Expression of FASN protein in human GBC cells. Results of western blot showed that the relative expression of FASN protein in human GBC NOZ, GBC-SD and SGC-996 cells was 0.57±0.06, 0.12±0.04 and 0.10±0.02, respectively, showing a significant difference among them ( F=115.67, P<0.05). There were significant differences between the NOZ cells and the GBC-SD or the SGC-996 cells ( P<0.05), and there was no significant difference between the GBC-SD cells and the SGC-996 cells ( P>0.05). (2) Effects of orlistat on the proliferation of human GBC NOZ cells. ① Results of cell proliferation assay showed that the absorbance value of NOZ cells was 2.34±0.12, 1.57±0.08 and 1.07±0.13 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=205.88, P<0.05). ② Results of clone formation assay showed that the number of NOZ cells clones was 257±23, 153±11 and 83±11 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=92.64, P<0.05). ③Results of western blot showed that the relative expression of Cyclin-D1 protein of NOZ cells was 2.31±0.10, 1.52±0.05 and 1.23±0.11 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=120.73, P<0.05). The relative expression of CDK-4 protein of NOZ cells was 1.58±0.04, 1.21±0.02 and 1.19±0.04 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a signifi-cant difference among them ( F=110.45, P<0.05). (3) Effects of orlistat on apoptosis of human GBC NOZ cells. Results of western blot showed that the relative expression of Bcl-2 protein of NOZ cells was 1.07±0.03, 0.36±0.03 and 0.15±0.02 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a significant difference among them ( F=1 242.93, P<0.05). The relative expression of Bax protein of NOZ cells was 0.51±0.03, 0.38±0.05 and 1.38±0.04 in the control group, low-dose orlistat group and high-dose orlistat group, respectively, showing a signifi-cant difference among them ( F=583.51, P<0.05). Conclusion:Orlistat can inhibit the growth of human GBC NOZ cells and promote their apoptosis.

Role of PNPT1 in cardiomyocyte apoptosis induced by oxygen-glucose deprivation / 南方医科大学学报

OBJECTIVE@#To explore the effect of inhibiting polyribonucleotide nucleotidyl-transferase 1 (PNPT1) on oxygen-glucose deprivation (OGD)-induced apoptosis of mouse atrial myocytes.@*METHODS@#Cultured mouse atrial myocytes (HL-1 cells) with or without OGD were transfected with PNPT1-siRNA or a negative control siRNA (NC-siRNA group), and the cell survival rate was detected using CCK-8 assay. The expression levels of ACTB and TUBA mRNA were detected with qPCR, and the protein expression of PNPT1 was detected with Western blotting. The apoptosis rate of the treated cells was determined with flow cytometry, the mitochondrial membrane potential was detected using JC-1 kit, and the mitochondrial morphology was observed using transmission electron microscope.@*RESULTS@#With the extension of OGD time, the protein expression levels of PNPT1 increased progressively in the cytoplasm of HL-1 cells (P < 0.05). Transfection with PNPT1-siRNA significantly reduced PNPT1 expression in HL-1 cells (P < 0.05). Exposure to OGD significantly enhanced degradation of ACTB and TUBA mRNA (P < 0.05) and markedly increased the apoptosis rate of HL-1 cells (P < 0.05), and these changes were significantly inhibited by transfection with PNPT1-siRNA (P < 0.05), which obviously increased mitochondrial membrane potential and improved mitochondrial morphology of HL-1 cells exposed to OGD.@*CONCLUSION@#Inhibition of PNPT1 improves mitochondrial damage and reduces degradation of apoptotic-associated mRNAs to alleviate OGD-induced apoptosis of mouse atrial myocyte.

Anti-tumor effect and its related mechanisms of cinobufotalin combined with cisplatin on H22 liver cancer mice / 中国中药杂志

In order to observe the anti-tumor effect of cinobufotalin on H22 liver cancer mice and to explore its regulatory mechanism, 50 Kunming mice were subcutaneously inoculated with H22 intraperitoneal passage cells under the armpit to establish H22 hepatocellular carcinoma model. They were then randomly divided into model group, cinobufotalin low dose group, cinobufotalin high dose group, cisplatin group and cisplatin+cinobufotalin group, which received 0.01% ethanol solution, 1 mg·kg~(-1) cinobufotalin, 5 mg·kg~(-1) cinobufotalin, 5 mg·kg~(-1) cisplatin, 5 mg·kg~(-1)cisplatin + 5 mg·kg~(-1) cinobufotalin respectively for 10 days. The general condition of mice during the intervention was observed, and the inhibition rate, tumor mass, thymus index, histopathological changes of the tumors, apoptotic rate of the tumors, the expressions of phosphatidylinositol 3-kinase(PI3 K), protein kinase B(Akt), apoptosis related gene(Fas), Fas ligand(FasL) mRNA and protein phosphorylated Akt(pAkt) protein in the tumors of each group were compared. The results showed that during the modeling period, the mice showed a decline in food intake, dark fur, poor mental status, and gradually worsened over time. The mental status of mice in each intervention group was improved gradually, especially in the cisplatin+cinobufotalin group. As compared with the model group, the tumor mass of each intervention group was lower(P<0.05). As compared with the cinobufotalin low dose group, the tumor mass was lower and inhibition rate was higher in the cinobufotalin high dose group, cisplatin group and cisplatin+cinobufotalin group(P<0.05). As compared with the cinobufotalin high dose group and the cisplatin group, the tumor mass was lower and the inhibition rate was higher in cisplatin+cinobufotalin group(P<0.05). As compared with the model group, the thymus index was higher in cinobufotalin high dose group and cisplatin + cinobufotalin group, while was lower in cisplatin group(P<0.05). As compared with the cinobufotalin low dose group, the thymus index was higher in the cinobufotalin high dose group and lower in the cisplatin group(P<0.05). As compared with the cinobufotalin high dose group, the thymus index was lower in cisplatin group(P<0.05). As compared with cisplatin group, the thymus index was higher in cisplatin+cinobufotalin group(P<0.05). Pathological staining showed that a large number of heterogeneous cells and mitotic phenomena were observed in the model group. Cell fragments and neutrophils were observed in the tumor tissues of the intervention groups, showing diffuse necrosis, and the diffuse necrosis was more obvious in the cisplatin+cinobufotalin group. As compared with the model group, the apoptotic rate of the tumors and the relative expressions of Fas mRNA and protein were higher in the intervention groups, while the relative expressions of PI3 K, FasL mRNA and protein and the relative expression of pAkt protein were lower in the intervention groups(P<0.05). As compared with the cinobufotalin low dose group, the apoptotic rate of the tumors and relative expression of Fas and protein were higher in the cinobufotalin high dose group, cisplatin group and cisplatin+cinobufotalin group, while the relative expressions of PI3 K, FasL mRNA and protein and pAkt protein were lower(P<0.05). As compared with the cinobufotalin high dose group and the cisplatin group, apoptotic rate of the tumors and the relative expression of Fas mRNA and protein were higher in the cisplatin+cinobufotalin group, while the relative expressions of PI3 K, FasL mRNA and protein and pAkt protein were lower in the cisplatin+cinobufotalin group(P<0.05). In summary, cinobufotalin has significant anti-tumor effect on H22 liver cancer mice, and can enhance the immune function of mice and synergistically enhance the effect of chemotherapy. Its mechanism may be associated with regulating PI3 K/Akt/Fas/FasL signaling pathway related genes and protein expression.

Study on Improvement Effects of Total Ginsenosides on D-galactose Induced PC 12 Cell Senescence and Its Me- chanism / 中国药房

OBJECTIVE：To study the improvement effects of total ginsenosides on the senescence of PC 12 cells induced by D-galactose and its mechanism. METHODS ：Rat pheochromocytoma （PC12）cells were treated with D-galactose to establish cell senescence model. CCK- 8 method was used to screen the D-galactose modeling concentration and total ginsenosides concentration. Normal control group ，model group ，total ginsenosides low and high concentration groups were set up. Cell senescence ，cell apoptosis rate ，apoptotic cycle and mitochondrial membrane potential （MMP），cell adenosine triphosphate （ATP）and reactive oxygen species （ROS）levels in each group were detected. The expression of apoptosis related proteins [B lymphoma 2（Bcl-2）and its related egg X protein （Bax），cytochrome C （Cyt-C）] and oxidative damage related proteins [nuclear factor 2 related factor 2 （Nrf2），heme oxygenase 1（HO-1）] were detected. In addition ，positive drug group [ 5 mmol/L N-acetyl-L-cysteine（NAC）] and positive control group [ D-galactose+5 mmol/L NAC] were set up to compare the levels of oxidative damage related proteins. RESULTS：D-galactose could significantly inhibit the survival rate of PC 12 cells，with a critical concentration of 20 mg/mL. The total ginsenosides could significantly increase the survival rate of D-galactose induced senescent cells with a median effective concentration（EC50）of 65 μg/mL，and then the low and high concentrations of total ginsenosides were set at 55 and 65 μg/mL. Compared with normal control group ，the number of aging cells increased ，the apoptotic rate and percentage of G 1 phase were significantly increased i n model group. the percentage of S phase ，MMP and ATP contents ，the protein expression of Bcl- 2 and Cyt-C in mitochondria were decreased significantly ，whileROS content ，the protein expression of Bax ，Nrf2 and Cyt-C protein in endochylema were increased significantly （P＜0.05 or P＜0.01）. Compared with model group ，the number of E-mail：sunqiao150509@163.com aging cells reduced ，the apoptosis rates and percentage of G 1 phase were significantly decreased in total ginsenosides low and high concentration groups ，the percentage of S phase ，the contents of MMP and ATP （except for low concentration group ），protein expression of Bcl- 2，Nrf2 and HO- 1 as well as protein expression of Cyt-C in mitochondria were increased significantly ；ROS level （except for low concentration group ）and Bax protein as well as protein expression of Cyt-C were decreased significantly. The protein expression of Nrf 2 and HO- 1 were increased significantly in positive control group （P＜0.05 or P＜0.01）， but it was lower than that of total ginsenosides groups . CONCLUSIONS：Total ginsenosides can improve D-galactose induced senescence of P 12 cells，the mechanism of which may be related to activating Nrf 2 antioxidant signal pathway to antagonize D-galactose induced oxidative stress and alleviating mitochondrial dysfunction.

Effects of PD 98059 Combined with Paclitaxel on Cell Proliferation and Apoptosis of Human Gastric Signet Ring Cell Carcinoma Cells / 中国药房

OBJECTIVE：To investiga te the effects of MEK/ERK pathway specific inhibitor PD 98059 combined with paclitaxel on the proliferation and apoptosis of human gastric signet ring cell carcinoma （SRCC）cells. METHODS ：Using human SRCC KATO Ⅲ cells as object ，CCK-8 assay was used to detect cell proliferation after treated with paclitaxel ，PD98059 and two drug combination for 48 h，and the proliferation rate was calculated. Flow cytometry ，Western blotting and Transwell assay were used to detect the cell proliferation ，the expression of apoptosis related protein （Cleaved-caspase-3）and cell migration after treated with paclitaxel，PD98059 and two drug combination for 48 or 24 h. RESULTS ：After treated with paclitaxel （1 μg/mL），PD98059（5， 20，40 μmol/L）and two drug combination （1 μg/mL+5，20，40 μmol/L），the proliferation rate of cells was increased significantly in administration groups ，and the combination groups were significantly higher than paclitaxel and PD 98059 alone groups （P＜ 0.05）. After treated with paclitaxel （1 μg/mL），PD98059（5，20，40 μmol/L）and two drug combination （1 μg/mL+40 μmol/L）， early and late apoptosis rate ，the protein expression of Cleaved-caspase- 3 were significantly increased in paclitaxel group and combination group ；combination group was significantly higher than paclitaxel and PD 98059 alone group （P＜0.05）. The number of migrated cells in administration groups were reduced significantly ，and the combination group was significantly lower than paclitaxel and PD 98059 alone group （P＜0.05）. CONCLUSIONS ：Paclitaxel and PD 98059 can inhibit the proliferation and migration of human SRCC KATO Ⅲ cells，paclitaxel can also promote the apoptosis and the expression of apoptosis related protein，which may be related to the inhibition of MEK/ERK pathway. The effect of the combination of the two drugs is better than paclitaxel or PD 98059 alone.

Experimental Research on the Effects of Zhuangtongyin Decoction on Arrhythmia after Myocardial Ischemia-re- perfusion / 中国药房

OBJECTIVE：To investigate the e ffects of Zhuangtongyin water extract （ZTYWE）on hemorheology and blood lipid level of myocardial ischemia-reperfusion model rats ，and preliminarily explore its molecular biology foundation of reducing arrhythmia after myocardial ischemia-reperfusion. METHODS ：Totally 75 SD rats were divided into control group ，model group and ZTYWE low-dose ，middle-dose and high-dose groups. Myocardial ischemia-reperfusion model was established by ligating the left ventricle cyclotron endings for 30 min then reperfusing for 60 min. Since the first day after surgery ，control group and model group were intraperitoneally administrated with same volume normal saline ；ZTYWE low-dose ，middle-dose and high-dose groups were intraperitoneally administrated with medicine solution （6.8，13.6，27.2 g/kg），once a day ，for continuous 28 d. At the 7th， 14th，21st，28th day after surgery ，the incidence rate of arrhythmia of rats were detected. The hemorheology indicators and serum contents of TC ，TG，LDL-C，HDL-C were determined after the last administration. The morphology of myocardial tissue was observed by HE staining. The protein expressions of apoptosis-related factor FAS and FAS-L were detected by Western blotting assay. RESULTS ：Compared with control group ，the incidence rates of arrhythmia of rats in model group were significantly increased；the whole blood contrast viscosity ，hematocrit（HCT），in vitro forming length and quality of thrombosis ，platelet aggregation rate ，erythrocyte filtration index （IF），as well as the serum contents of TC ，TG and LDL-C were significantly increased；in vivo thrombus formation time as well as the serum content of HDL-C were significantly decreased ；the protein expression of FAS and FAS-L were significantly increased （P＜0.05 or P＜0.01）；obvious pathological changes were observed in myocardial tissue. Compared with model group ， the incidence rates of arrhythmia of rats in ZTYWE groups were in obvious reducing trend ； except for the HCT in 163.com low-dose ZTYWE group without significance ， the blood contrast viscosity ，HCT，in vitro forming length and quality of thrombosis ，platelet aggregation rate ，IF as well as the serum contents of TC ，TG and LDL-C were significantly decreased ；in vivo thrombus formation time as well as the serum contents of HDL-C were significantly increased ；the protein expressions of FAS and FAS-L were significantly decreased （P＜0.05 or P＜ 0.01）；the pathological changes of myocardial tissue were improved. CONCLUSIONS ：Through the promoting blood circulation and removing blood stasis effect ，Zhuangtongyin decoction can improve indexes of hemorheology index of ischemia-reperfusion model rats ，reduce the contents of TC ，TG and LDL-C ，enhace the contents of HDL-C ，down-regulate the protein expression of FAS and FAS-L ，so as to inhibit myocardial cells apoptosis and reduce arrhythmia.

Effect of Ferula ferulaeoides on growth and apoptosis of human gastric cancer MGC-803 transplantation tumor in nude mice / 中国中药杂志

In this paper,the effects of active fractions of Ferula ferulaeoides on the growth and apoptosis of human gastric cancer cell MGC-803 transplantation tumor were systematically studied. The subcutaneous ectopic transplantation tumor model was established in human gastric cancer MGC-803 nude mice by cell suspension implantation method. The anti-tumor rate and organ index were used to evaluate the anti-tumor effect of the active fractions of F. ferulaeoides on the tumor-bearing nude mice. HE staining,TUNEL staining,RT-PCR,Western-blot and ELISA were used for pathological examination,apoptosis observation,and detection of apoptosis-related genes,proteins and cytokines expression. The results showed that as compared with the model group,the low,medium and high doses of the active fraction of F. ferulaeoides had inhibitory effects on xenografts in nude mice,respectively,in a dose-dependent manner; the apoptotic ratio was increased with the increase of drug concentration. As compared with the model group,F. ferulaeoides could down-regulate the expression of survivin mRNA in nude mice,and the protein expression levels of Bax,Bcl-2,caspase-3 and caspase-9 in tumor tissues of nude mice could be increased to different degrees in F. ferulaeoides groups. The contents of IL-10 and TGF-β1 in plasma of nude mice were decreased in high dose group of F. ferulaeoides active fractions. The results indicated that F. ferulaeoides can significantly inhibit the growth of human gastric cancer MGC-803 subcutaneously transplanted tumor,and its mechanism may be related with down-regulating the expression of survivin mRNA,and up-regulating the expression of apoptosis-related proteins Bax,caspase-3 and caspase-9.

Effects of Cimetidine on L ow Dose Rate Irradiation-induced Liver Cell Apoptosis in Beagle Dogs and Its Mechanism / 中国药房

OBJECTIVE： To study the effects of cimetidine on low dose rate irradiation-induced liver cell apoptosis in Beagle dogs. METHODS： Healthy male Beagle dogs were randomly divided into normal control group， model control group， positive drug group （lentinan， 21.33 mg/kg） and cimetidine low-dose， medium-dose and high-dose groups （5.33， 10.67， 21.33 mg/kg）， with 4 Beagle dogs each. Except for normal control group， other groups were given 60Co-γ accumulative irradiation （dosage rate： 0.040 8 mGy/min） for 23 d； the medication groups were given relevant medicine orally before irradiation， once a day. Twenty-four hours after stopping irradiation， TUNEL method was used to detect the apoptosis of liver cells in Beagle dogs. The percentage of apoptotic cells was calculated. The expression level of apoptosis-related proteins （Bax， Bcl-2， Caspase-3， p53） in liver tissue was detected by immunohistochemistry. RESULTS： Compared with normal control group， apoptotic cells and Bax， Caspase-3， p53 positive cells were increased significantly in liver tissue of Beagle dogs in model control group； the percentage of apoptotic cells， protein expression levels of Bax， Caspase-3 and p53 were increased significantly； Bcl-2 positive cells were decreased significantly， and its protein expression level was decreased significantly （P＜0.05 or P＜0.01）. Compared with model control group， above positive cells of liver tissue in Beagle dogs were changed to different extents in medication groups； the percentage of apoptotic cells and protein expression levels of p53 in medication groups， protein expression levels of Bax in positive drug group， cimetidine low-dose and high-dose groups as well as protein expression levels of Caspase-3 in cimetidine groups were decreased significantly； protein expression levels of Bcl-2 were increased significantly in cimetidine groups. The percentage of apoptotic cells in cimetidine medium-dose and high-dose groups as well as protein expression levels of Caspase-3 in cimetidine groups were all lower than positive control group. Protein expression level of p53 in cimetidine low-dose group was significantly higher than positive drug group （P＜0.05 or P＜0.01）. CONCLUSIONS： Cimetidine can inhibit the low dose rate irradiation-induced apoptosis of liver cells in Beagle dogs， and certainly protect liver cells against irradiation. The mechanism of it may be associated with up-regulating the protein expression of Bcl-2 and down-regulating the protein expression of Bax， Caspase-3 and p53 in liver cells.

Study on the Inhibitory Effects and Mechanism of New Small Molecular Kinase Inhibitors Ibr- 7 on Human Pancreatic Cancer Capan-2 Cells / 中国药房

OBJECTIVE： To observe the inhibitory effects and possible mechanism of new small molecular kinase inhibitors Ibr-7 [Irutinil（Ibr） derivatives] on human pancreatic cancer Capan-2 cells. METHODS： Taking Capan-2 cells as objects， CCK-8 method was used to determine the proliferation of cells after treated with 1， 2， 4， 8 μmol/L Ibr/Ibr-7 for 48 h. The survival rates of cells were calculated. Sensitization effects of 1 μmol/L Ibr/Ibr-7 on different doses of gemcitabine/paclitaxel （0.062 5， 0.125， 0.25， 0.5， 1 μmol/L） were detected. Clone formation test was used to detect the situation of cell clone formation after treated with 1， 2， 4 μmol/L Ibr/Ibr-7 for 48 h. The number of cell colony formation was recorded. Flow cytometry or JC-1 method was used to detect the apoptosis of cells after treated with 2， 4， 8 μmol/L Ibr-7 for 24 or 16 h and the changes of mitochondrial transmembrane potential； total apoptotic rate and the percentage of mitochondrial membrane potential decrease were calculated. Western blotting was used to detect the expression of related apoptotic protein （PARP， Noxa， Bcl-2， Bax， Mcl-1， Bcl-xL）. RESULTS： After treated with 1， 2， 4， 8 μmol/L Ibr/Ibr-7 for 48 h， the survival rates of cells were decreased significantly； those of Ibr-7 groups were significantly lower than those of same-dose Ibr groups； IC50 of Ibr-7 was significantly lower than that of Ibr （P＜0.05 or P＜0.01）. After combined with Ibr/Ibr-7， the survival rate of cells was significantly lower than that of same-dose gemcitabine/paclitaxel alone group， and the Ibr-7 combination group was significantly lower than same-dose Ibr combination group （P＜0.05 or P＜0.01）. After treated with 2， 4 μmol/L Ibr and 1， 2， 4 μmol/L Ibr-7 for 48 h， the number of cell clone formation was decreased significantly， while Ibr-7 groups were significantly lower than same-dose Ibr groups （P＜0.01）. After treated with different doses of Ibr-7 for 24 or 16 h， total apoptosis rate of cells （2， 4， 8 μmol/L）， the proportion of cell mitochondrial membrane potential decrease （8 μmol/L）， the relative protein expression of Noxa （2， 4， 8 μmol/L） and Bax （8 μmol/L） were increased significantly， while the protein expression of PARP （8 μmol/L）， Bcl-2 （4 μmol/L）， Mcl-1 （2， 4， 8 μmol/L） were decreased significantly； above indexes （except for relative expression of PARP and Bcl-2） of 8 μmol/L Ibr-7 group were significantly better than same-dose Ibr group （P＜0.05 or P＜0.01）. There was no statistical significance in protein expression of Bcl-xL among those groups （P＞0.05）. CONCLUSIONS： Compared with Ibr， Ibr-7 has better inhibitory and apoptotic effects on human pancreatic cancer Capan-2 cells in vitro， and has stronger chemotherapeutic drug sensitization activity， the mechanism of which may be associated with reducing mitochondrial transmembrane potential， down-regulating the protein expression of PARP， Bcl-2 and Mcl-1 and up-regulating the protein expression of Noxa and Bax.

Hawthorn Extract Alleviates Atherosclerosis through Regulating Inflammation and Apoptosis Related Factors: An Experimental Study / 中国结合医学杂志

OBJECTIVE@#To determine the effects of hawthorn extract on serum lipid levels, pathological changes in aortic atherosclerosis plaque, inflammatory factors, and apoptosis-related protein and mRNA expression in apolipoprotein E gene knockout (ApoE) mice.@*METHODS@#Thirty-six ApoE mice were fed with a high-fat diet starting at the age of 8 weeks. Mice were randomly divided into 3 groups by a random number table including model group, hawthorn extract group, and simvastatin group, 12 mice in each group. Twelve 8-week-old C57BL/6 mice were fed a basic diet and served as control. The mice in the control and model groups were administered 0.2 mL saline daily, the mice in the hawthorn extract and simvastatin groups were administered with 50 mg/kg hawthorn extract or 5 mg/kg simvastatin daily for 16 weeks. After 16 weeks, plasma lipids including total cholesterol (TC), triglyceride (TG), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) were determined by an enzymatic assay. Aortic atherosclerotic lesions were observed by light microscopy, scanning and transmission electron microscopy, respectively. Plasma levels of monocyte chemoattractant protein-1 (MCP-1), interleukin-1β (IL-1β), adiponectin (APN), and hypersensitive C-reactive protein (hs-CRP) were measured by enzyme-linked immunosorbent assay (ELISA). Protein and mRNA expressions of Bax and Bcl-2 in the aorta were assessed by Western blotting and quantitative real-time polymerase chain reaction (qRT-PCR), respectively.@*RESULTS@#Compared to the control group, the plasma levels of TC, TG and LDL-C were significantly increased and HDL-C were significantly decreased in the model group (P<0.01). Compared to the model group, treatment with hawthorn extract significantly decreased the plasma levels of TC, TG, and LDL-C and increased the plasma level of HDL-C in ApoE mice (P<0.01). The levels of MCP-1, IL-1ß, and hs-CRP in the model group were significantly increased and APN was significantly decreased compared with the control group (P<0.01). Compared to the model group, treatment with hawthorn extract decreased the levels of MCP-1, IL-1ß, and hs-CRP and increased the APN level (P<0.01). Compared to the control group, the protein and mRNA expression of Bax in the model group were significantly increased and the expression of Bcl-2 was significantly decreased (P<0.01). Hawthorn extract also reduced the protein and mRNA expression of Bax and increased the Bcl-2 expression in the aorta (P<0.01).@*CONCLUSION@#Hawthorn extract has anti-atherosclerosis and stabilizing unstable plaque effects. The mechanism may be related to the inflflammation and apoptosis signaling pathways.

Effect of 3 neuroprotective measures on expressions of Fas/FasL in brain tissue of neonatal rats with hypoxicischemic brain damage / 中国中西医结合急救杂志

Objective To observe the effects of 3 neuroprotective measures on the expressions of apoptosis-related factors and their ligands (Fas and FasL) in brain tissue of neonatal rats with hypoxic ischemic brain injury. Methods One hundred and twenty Wistar rats 7 days old were selected as experimental subjects, the rats were divided into four groups: neural stem cell, erythropoietin (EPO), ω-3 unsaturated fatty acid treatment groups and hypoxic ischemic brain damage model group according to random number table method, with 30 rats in each group. Neural stem cell group, EPO group and ω-3 unsaturated fatty acid group were respectively injected with neural stem cells, EPO and ω-3 unsaturated fatty acid, each 5 mL via tail vein after modeling; the hypoxic ischemic brain damage model group was given equal volume of normal saline. At 6, 12, 24, 48 and 72 hours after administration of drug, 6 rats were sacrificed in each group, brain tissue was taken, the mRNA expression levels of Fas/FasL, protein expression levels of Toll-like receptor 4 (TLR4), nuclear transcription factor-κB (NF-κB), tumor necrosis factor-α (TNF-α), interleukin (IL-1β, IL-6) and cell apoptotic rate in hippocampus tissue were measured. Results ① mRNA expressions: the mRNA expressions of Fas and FasL of the 3 experimental groups were significantly lower than those of the hypoxic ischemic brain damage model group, the degrees of descent after administration for 24 hours were the most significant, neural stem cell treatment group < EPO treatment group < ω-3 unsaturated fatty acid treatment group < hypoxic ischemic brain damage model group [Fas mRNA expression (2-ΔΔCt): 140.5±2.9, 156.4±2.5, 165.2±2.7 vs. 173.7±2.8, FasL mRNA expression (2-ΔΔCt): 143.1±4.3, 154.6±1.5, 160.7±1.4 vs. 174.7±2.8], the differences were statistically significant (all P < 0.05). ② Protein expressions: the protein expressions of TLR4, NF-κB, TNF-α, IL-1β, IL-6 of the 3 experimental groups were significantly lower than those of the hypoxic ischemic brain damage model group (TLR4/GAPDH: 0.7±0.2, 0.6±0.1, 0.2±0.1 vs. 1.4±0.1; NF-κB/GAPDH: 6.7±0.4, 5.3±0.1, 1.1±0.2 vs. 11.2±0.3; TNF-α/GAPDH: 14.3±1.4, 11.2±1.2, 3.2±2.1 vs. 23.2±0.5; IL-1β/GAPDH: 9.4±0.2, 7.4±0.3, 2.2±0.3 vs. 13.4±0.1; IL-6/GAPDH: 36.2±4.4, 39.3±1.5, 26.2±2.1 vs. 51.4±1.4, all P < 0.05), the protein expression levels of above indexes in neural stem cell treatment group < those of EPO treatment group < those of ω-3 unsaturated fatty acid treatment group < those of hypoxic ischemic brain damage model group. ③ Apoptotic rates:after drug administration, the apoptotic rates of the ω-3 unsaturated fatty acid group, EPO treatment group, neural stem cell treatment group were obviously lower than the rate of model group [(3.7±0.3)%, (3.4±0.2)%, (2.5±0.1)% vs. (5.5±0.4)%, all P < 0.05]. Conclusion The mRNA expressions of Fas/FasL in the brain of neonatal rats with hypoxic-ischemic brain damage are high, and the treatment with each of the following agents; neural stem cells, EPO and ω-3 unsaturated fatty acid can reduce the mRNA expressions of Fas/FasL in such rats' brain tissues.

Apoptosis induction and growth inhibition effects of Schisandra chinensis polysaccharides on brain tumor stem cells / 吉林大学学报(医学版)

Objective:To investigate the effect of Schisandra chinensis polysaccharide(SCP)on the growth of brain tumor stem cells(BTSCs),and to clarify the mechanism of inhibiting the growth of BTSCs of SCP. Methods:The primary human glioma cells were cultured,then the BTSCs were isolated by CD133 immunomagnetic sorting.The neural stem cell surface markers CD133 and Nestin were detected by immunofluorescence assay.The proliferation rate of BTSCs was examined by MTT assay.Annexin V-PI analysis was used to analyze the apoptotic rate of BTSCs.The expression levels of Bax,Bcl-2 and Caspase-3 proteins in BTSCs in various groups were detected by ELISA assay.Results:The results of immunofluorescence staining showed that the expressions of CD133 and Nestin were positive in BTSCs.Compared with control group,the proliferation rates of BTSCs in 200,400 and 800 mg·L-1SCP groups were decreased,especially in 400 and 800 mg·L-1SCP groups(P<0.05).The results of Annexin V-PI analysis showed that the apoptotic rate of BTSCs in 800 mg·L-1SCP group was increased compared with control group(P<0.05).The ELISA results showed that the expression levels of Bax in 200,400 and 800 mg·L-1SCP groups were significantly increased(P<0.05),and the values of Bax/Bcl-2 were significantly increased(P<0.05);compared with control group,the Bcl-2 expression level in the BTSCs in 800 mg·L-1SCP group was decreased(P<0.05).The expression level of Caspase-3 protein in 800 mg·L-1SCP group was also significantly increased compared with control group(P<0.01).Conclusion:SCP could inhibit the growth of BTSCs,and the induction of apoptosis may be one of mechanisms.

Apoptosis induction and growth inhibtion effects of Schisandra chinenis polysaccharides on brain tumor stem cells / 吉林大学学报(医学版)

Objective: To investigate the effect of Schisandra chinensis polysaccharide (SCP) on the growth of brain tumor stem cells (BTSCs), and to clarify the mechanism of inhibiting the growth of BTSCs of SCP. Methods: The primary human glioma cells were cultured, then the BTSCs were isolated by CD133 immunomagnetic sorting. The neural stem cell surface markers CD133 and Nestin were detected by immunofluorescence assay. The proliferation rate of BTSCs was examined by MTT assay. Annexin V-PI analysis was used to analyze the apoptotic rate of BTSCs. The expression levels of Bax, Bcl-2 and Caspase-3 proteins in BTSCs in various groups were detected by ELISA assay. Results: The results of immunofluorescence staining showed that the expressions of CD133 and Nestin were positive in BTSCs. Compared with control group, the proliferation rates of BTSCs in 200, 400 and 800 mg · L-1 SCP groups were decreased, especially in 400 and 800 mg · L-1 SCP groups (P<0.05). The results of Annexin V-PI analysis showed that the apoptotic rate of BTSCs in 800 mg · L-1 SCP group was increased compared with control group (P<0.05). The ELISA results showed that the expression levels of Bax in 200, 400 and 800 mg · L-1 SCP groups were significantly increased (P<0.05), and the values of Bax/Bcl-2 were significantly increased (P<0.05); compared with control group, the Bcl-2 expression level in the BTSCs in 800 mg · L-1 SCP group was decreased (P<0.05). The expression level of Caspase-3 protein in 800 mg · L-1 SCP group was also significantly increased compared with control group (P<0.01). Conclusion: SCP could inhibit the growth of BTSCs, and the induction of apoptosis may be one of mechanisms.

Influence of genipin in abilities of proliferation and invasion and apoptosis of gastric cancer SGC 7901 cells and their mechanisms in vitro / 吉林大学学报(医学版)

Objective: To investigate the regulatory effect of genipin (GP) on the gastric cancer SGC 7901 cells, and to clarify its mechanism. Methods: The SGC 7901 cells in logarithmic growth phase were selected and divided into control group and different concentrations (5. 0, 10. 0, and 20. 0 mg · L-1) of GP groups. MTT was used to detect the inhibitory rates of proliferation of SGC 7901 cells at different time points (24, 48, and 72 h). Transwell chamber cell invasion assay was used to detect the invasion ability of SGC 7901 cells in vitro. The expression levels of Bcl-2, Bax and caspase-3 proteins in the SGC 7901 cells were detected by Western blotting method. Results: The results of MTT assay showed that the inhibitory rates of proliferation of the cells in different concentrations of GP groups after treated with GP for different time were increased significantly compared with control group (P< 0. 01). The Transwell cell invasion assay results showed that compared with control group, the number of transmembrane cells in 10. 0 and 20. 0 mg · L-1 GP groups was decreased significantly after treated for 72 h (P< 0. 01). The results of Western blotting method showed that compared with control group, the expression level of Bcl-2 protein in 20. 0 mg · L-1 GP group was decreased after treated for 72 h (P<0. 01), and the expression levels of caspase-3 and Bax proteins were increased (P<0. 05). Conclusion; GP can inhibit the abilities of proliferation and invasion in vitro of SGC 7901 cells, and induce the apoptosis; its mechanism may be related to the regulation of Bcl-2, caspase-3, and Bax protein expressions.

Effect of Huangjiao Granule on Inflammatory Factors and Apoptosis-related Proteins in Rats with Cerebral Ischemia-reperfusion Injury / 中国康复理论与实践

@#Objective To investigate the effect of Huangjiao granule on inflammatory factors and apoptosis-related proteins in rats with cerebral ischemia-reperfusion injury.Methods A total of 40 Sprague-Dawley rats were divided into sham operation group, model group and Huangjiao granule group, with ten rats in each group, and ten rats standby. The cerebral ischemia for two hours and reperfusion model was established by suture method. The sham operation group and the model group were given saline 10 ml/kg intragastrically 30 minutes before operation. The Huangjiao granule group was given Huangjiao granule solution 10 ml/kg (the content of crude drug was 1 g/ml) intragastrically 30 minutes before ischemia-reperfusion. Longa scoring method was used to evaluate the neurological function score 24 hours after reperfusion, while the percentage of cerebral infarction volume was detected by TTC staining, the pathological morphology of brain tissue was observed by HE staining, the cell apoptosis of brain tissue was detected with TUNEL, the levels of interleukin (IL)-1β, IL-6 and tumor necrosis factor α (TNF-α) in serum were detected by ELISA, the expression of cleaved-caspase 3, cleaved-caspase 9, Bax and Bcl-2 proteins of brain tissue was evaluated by Western blotting.Results Compared with the model group, the neurological score decreased, the percentage of cerebral infarction volume and the apoptosis rate of the brain tissue decreased in Huangjiao granule group (P<0.05). The levels of IL-1β, IL-6 and TNF-α in serum and the expression of cleaved-caspase 3, cleaved-caspase 9 and Bax proteins of brain tissues significantly decreased (P<0.05), and the expression of Bcl-2 protein of brain tissues inreased in Huangjiao granule group (P<0.05).Conclusion The protective effect of Huangjiao granule on rats with cerebral ischemia-reperfusion injury may be related to the inhibition of inflammatory response and the reduction of apoptosis.

Effects of High Thoracic Epidural Anesthesia on Cerebral Blood Flow and Hippocampal Apoptosis during Global Cerebral Ischemia and Reperfusion in Rats / 现代生物医学进展

Objective:To investigate the effects of high thoracic epidural anesthesia (HTEA) on the cerebral blood flow (CBF) and hippocampal apoptosis-related proteins Bcl-2 and Bax during global cerebral ischemia and reperfusion (GCI) in rats.Methods:Fifteen-minute global ischemia was established by 4-vessel occlusion and epidural catheterization was performed through T4-5 intervertebral spaces in adult male Wistar rats.According to the different drugs infused into the epidural space,the rats were randomly divided into four groups:Sham group (0.9 ％ NaC1),Sham-HTEA group (0.25 ％ bupivacaine),GCI group (global cerebral ischemia,0.9 ％ NaC1) and HTEA group (global cerebral ischemia,0.25 ％ bupivacaine).And 0.25 ％bupivacaine or 0.9 ％ saline (20 μL·h-1) was infused continuously to the thoracic epidural space from 15 minutes before ischemia to 24 hours after reperfusion.Mean arterial pressure (MAP),heart rate (HR) and cerebral blood flow (CBF) were determined until 2 hours after reperfusion,and the hippocampal Bcl-2 and Bax proteins at 24 hours after reperfusion were examined by Western-blot.Results:Compared with the GCI group,HTEA group has no significant difference on MAP and HR during ischemia and 2 hours after reperfusion,andcompared with the Sham group,MAP in GCI group increased in ischemia 0 min and decreased in reperfusion 0 min.The CBF in HTEA group was significantly lower than that in GCI group (123.1％± 35.2％ vs 177.5％± 32.4％,P＜0.01) in reperfusion 10 min,and higher than that in GCI group during the hypoperfusion of 60 to 120 minutes after reperfusion (P＜0.05),and the ratio of Bax/Bcl-2 in hippocampus was significantly decreased in HTEA group 24 hours after reperfusion (P＜0.01).Conclusions:Continuous HTEA infusion of 0.25 ％ bupivacaine 20 μL ·h-1 could maintain the hemodynamic stability,and improve the CBF of hypoperfusion period in rats,as well as reduce the ratio of Bax/Bcl-2 at 24 hours after reperfusion.

Podophyllotoxin derivative LN-13 induced multidrug resistant cell K562/A02 apoptosis and its molecular mechanism / 中国药理学通报

Aim To study the mechanism of action of the new derivative of podophyllotoxin(LN-13)in indu-cing the apoptosis of K562/A02 cells.Methods The MTT method was taken to detect the inhibition of LN-13 and VP-16 on K562/A02 proliferation and inhibi-tion rate and IC50 values were obtained 48 hours later. The K562/A02 cell morphological change induced by LN-13 were observed through Hochest33342 and PI staining after 48 hours later.Flow cytometry was taken to detect the apoptosis of K562/A02 cells induced by LN-13.The reverse transcription-polymerase chain re-action was taken to detect the Bcl-2,Bax,caspase-3 and mdr-1 mRNA expression.The expression of P-gp was detected by Western blot.Results The growth of K562 /A02 cells was obviously inhibited by LN-13 when IC50 value was 3.32 μmol · L-1 .LN-13 could obviously induced cell apoptosis observed by Ho-chest33342 and PI staining.Flow cytometry detection showed that LN-13(2,4,8 μmol·L-1 )could induce cell apoptosis and apoptosis ratio reached 15.0%, 48.0%,68.96%,respectively.The reverse transcrip-tion-polymerase chain reaction showed that LN-13 in-creased the Bax and Caspase-3 mRNA expression,and meanwhile the expression of mdr-1 mRNA decreased. Western blot showed that P-gp expression was de-creased as the LN-13 dose increased.The data were significantly different from those of control group.Con-clusion Podophyllotoxin derivative LN-13 can induce the apoptosis of K562 /A02 cells,which may be close-ly-related to regulating P-gp expression and apoptosis related gene mRNA expression.

Effect of hedyotisdiffusa on expressions of apoptosis related protein Fas, caspase3 and caspase7 in Renca renal cell carcinoma of model mice / 中国生化药物杂志

Objective To analyse the effect of hedyotisdiffusa on expressions of apoptosis related protein Fas , caspase3 and caspase7 in Renca renal cell carcinoma of model mice.Methods One hundred and twenty BALB/C male mice were randomly divided into normal control group, model control group, interleukin group, hedyotisdiffusa group, 30 mice in each group, half male and female.Excepted for normal control group , renal cell carcinoma models were established in the other groups , and were given corresponding drug treatment.The tumor size and weight , and the tumor inhibition rate of all mice were observed, and the expression levels of Fas,FasL,caspase3 and caspase7 were detected.Results Compared with model control group, the tumor size and weight were lower , the tumor inhibition rate were higher , the positive expression rates and protein levels of Fas were higher , the positive expression rates and protein levels of FasL were lower, the positive expression rates and expression levels of Caspase3,Caspase7 protein were higher in the mice of interleukin group and hedyotisdiffusa group, all with significant differences ( P<0.05 ).The above indicators in hedyotisdiffusa group were significant improved compared with interleukin group (P<0.05).Conclusion The hedyotisdiffusa can effectively promote the expressions of apoptosis related protein Fas,caspase3 and caspase7 in renal cell carcinoma of model mice, inhibit the expression of FasL protein, improve the tumor suppression rate.

Study on Time-effect and Dose-effect of Paeonol on the Apoptosis of Knee Osteoarthritis Articular Chon-drocyte in Rabbits and the mRNA Expression of Its Related Protein Bcl-2 and Bax / 中国药房

OBJECTIVE:To study the time-effect and dose-effect of paeonol on the apoptosis of knee osteoarthritis(OA)artic-ular chondrocyte in rabbits and the mRNA expression of its related protein Bcl-2 and Bax. METHODS:60 big-ear rabbits were ran-domly divided into normal (normal saline) group,model (normal saline) group,paeonol high-dose,medium-dose and low-dose groups and triamcinolone acetonide(positive drug)group,with 10 rabbits in each group. Except for normal group,OA model was induced by right knee anterior cruciate ligament (ACLT) and the medial meniscus 1/3 resection in those groups. After modeling, different doses of paeonol(0.8,0.4,0.2 mg/kg),triamcinolone acetonide 0.2 mg/kg were injected into right articular cavity twice a week. 4 weeks and 8 weeks after administration,articular cartilage specimens were collected. Ultrapathological structure changes of articular chondrocytes were observed by electron microscope. Apoptosis of cartilage cell was observed by TUNEL and apoptotic index was calculated. mRNA expression of apoptosis related genes of Bcl-2 and Bax in articular cartilage tissue of rabbits were de-tected by in situ hybridization technique. RESULTS:Compared with normal group,articular chondrocyte of model group showed early and middle stage apoptosis morphology change after 4 and 8 weeks,and apoptosis index increased significantly and the mRNA expression of Bcl-2 and Bax was up-regulated (P<0.01);4 and 8 weeks later after administration,compared with model group,apoptosis index decreased and mRNA expression of Bax was down-regulated in paeonol groups,while mRNA expression of Bcl-2 was up-regulated(P<0.05 or P<0.01). Electron microscopy ultrastructural observation showed articular chondrocyte of pae-onol high-dose and middle-dose groups were in early stage of apoptosis.CONCLUSIONS:Paeonol can slow down articular chondro-cyte degeneration and destroy in OA model rabbits in time and dose dependently. Its mechanism may be associated with expression up-regulation of Bcl-2 and expression down-regulation of Bax.

Effects of Cornus officinalis Polysaccharide on Learning and Memory Ability of Vascular Dementia Rats and Its Mechanism / 中国药房

OBJECTIVE:To study the effects of Cornus officinalis polysaccharide on learning and memory ability of vascular dementia (VD) model rats and its mechanism. METHODS:30 rats were randomly divided into sham operation (normal saline) group,model(normal saline)group and C. officinalis polysaccharide(0.28 g/kg)group. Except sham operation group,VD model was induced in other groups and given relevant medicine intragastrically once a day. After 4 weeks,the frequency of leaping over original platform position,water maze mean escape latencies,platform retention time and average swimming distance were detect-ed,and the mRNA and protein expression of brain-derived neurotrophic factor(BDNF)and apoptosis related gene Bcl-2 in hippo-campal tissue of rats were determined. RESULTS:Compared with sham operation group,learning and memory ability of rats in model group weakened,manifesting as the frequency of leaping over original platform position decreased,water maze mean escape latencies prolonged,platform retention time shortened,average swimming distance extended(P<0.05);mRNA and protein expres-sion of BDNF and Bcl-2 in hippocampus decreased significantly (P<0.05). Compared with model group,learning and memory ability of rats in C. officinalis polysaccharide group increased,manifesting as the frequency of leaping over original platform posi-tion increased,water maze mean escape latencies shortened,platform retention time prolonged,average swimming distance short-ened(P<0.05);mRNA and protein expression of BDNF and Bcl-2 in hippocampus was enhanced significantly(P<0.05). CON-CLUSIONS:C. officinalis polysaccharide can improve the learning and memory ability of VD rats,and its mechanism may be asso-ciated with the expression up-regulation of BDNF and Bcl-2 in hippocampus.