益气活血方对胃癌前病变大鼠胃黏膜Sonic Hedgehog信号通路的影响

目的:研究益气活血方对1-甲基-3-硝基-1-亚硝基胍(MNNG)诱导的新生大鼠胃癌前病变胃黏膜组织Sonic Hedgehog(Shh)信号通路的影响。方法:48只雄性乳鼠随机分为正常组8只,造模组40只。造模组灌服800mg/L的MNNG 0.1mL/只,正常组灌服等量0.9%氯化钠溶液,共10d。23周后将造模组随机均分为模型组,中药高、中、低剂量组,环耙明组。正常组、模型组灌服0.9%氯化钠溶液10mL/kg,中药高、中、低剂量组分别灌服38.4、19.2、9.6g/kg的益气活血方流浸膏,环耙明组腹腔注射0.2mg/kg环靶明,均每日1次,12周后HE染色观察胃黏膜组织病理学变化,免疫组化检测胃黏膜PCNA的表达,qRT-PCR法检测胃黏膜组织Shh、Ptch1、Smo、Gli1、SuFu、CyclinD1、CyclinE1、C-Myc mRNA的表达,Western Blot检测胃黏膜组织Shh、Ptch1、Smo、Gli1、SuFu、CyclinD1、CyclinE1、C-Myc、p-c-Myc蛋白表达。结果:与模型组比较,益气活血方高、中、低剂量组和环耙明组大鼠胃黏膜炎症及异型增生程度均显著减轻,PCNA表达水平显著降低(P<0.05,P<0.01),Ptch1及SUFU mRNA表达显著升高(P<0.05,P<0.01),Shh、Smo、Gli-1、CyclinD1及p-c-Myc蛋白的相对表达量显著降低而Ptch1及SUFU蛋白的表达显著升高(P<0.05,P<0.01)。结论:益气活血方可能通过下调Shh信号通路的表达起到对胃癌前病变的防治作用。     

Effects of Panax notoginseng saponins on proliferation and differentiation of rat hippocampal neural stem cells

We aimed to investigate the effects of Panax notoginseng saponins (PNS) on proliferation, differentiation and self-renewal of rat hippocampal neural stem cells (NSCs) in vitro. Rat hippocampal NSCs were isolated from post-natal day 1 (P1) rats and cultured in a serum-free medium. The neurospheres were identified by the expressions of nestin, class III β-tublin (Tuj-1) and glial fibrillary acid protein (GFAP). The cells were given PNS and subjected to oxygen glucose deprivation (OGD) as an in vitro model of brain ischemia reperfusion. The proliferation of NSCs was determined by MTT colorimetry, nestin/BrdU immunofluorescent double-labeling and RT-PCR. Differentiation of NSCs was assessed by immunofluorescent double-labeling of nestin/BrdU, nestin/vimentin, and nestin/Tuj-1. The primary cells and the first two passages of cells formed certain amount of neurospheres, the cells derived from a single cell clone also formed neurospheres. Nestin, BrdU, GFAP and Tuj-1-positive cells appeared in those neurospheres. Compared to the control group, PNS significantly promoted NSC proliferation and the expression of nestin/BrdU, and also enhanced Tuj-1, vimentin, and nestin mRNA expressions in hippocampal NSCs. PNS significantly increased area density, optical density and numbers of nestin/BrdU, nestin/vimentin, and nestin/Tuj-1 positive cells following OGD. These results indicate that PNS can promote proliferation and differentiation of hippocampus NCSs in vitro after OGD, suggesting its potential benefits on neurogenesis and neuroregeneration in brain ischemic injury.     

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??OBJECTIVE To investigate the effect of esculin (ES) against lens injury induced by D-galactose in rats. METHODS Male SD rats were randomly divided into five groups normal control group (control), D-galactose model group (model), ES 30 mg??kg^-1 administered group (ES30), ES 100 mg??kg^-1 administered group (ES100) and aminoguanidine 100 mg??kg^-1 administered group (AG). After treatment, blood glucose and glycosylated serum protein (GSP) content in serum were measured. The content of malondialdehyde (MDA), the level of glutathione (GSH), and the activity of aldose reductase (AR) in lens were also assayed. RESULTS Compared with control group, the level of blood glucose was significantly increased (P<0.01) in D-galactose model rats. In addition, in the lens tissue of D-galactose-induced rats, the content of MDA was significantly decreased (P<0.05),the level of GSH was significantly increased (P<0.05), and the activity of AR was significantly decreased (P<0.05). However, co-treatment with ES 30 mg??kg^-1 significantly decreased the level of GSP in serum(P<0.05), decreased the content of MDA (P<0.05), increased the level of GSH (P<0.05), and inhibited the activity of AR in lens (P<0.05). CONCLUSION Esculin could improve lens injury induced by D-galactose in rats. The mechanism may be related with increasing GSH level, decreasing MDA production, and inhibiting AR activity.     

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??OBJECTIVE To investigate the effect of genipin on hypoxia/reoxygenation induced oxidative stress injury in H9c2 cell line. METHODS For mimicking myocardial ischemia/reperfusion (I/R) injury, H9c2 cells were cultured in vitro and established hypoxia/reoxygenation induced oxidative stress injury model. H9c2 cells were divided into six groups: control group, H/R group, H/R+0.5 ??mol??L^-1genipin group, H/R+1.25 ??mol??L^-1genipin group, H/R+2.5 ??mol??L^-1genipin group and H/R+10 ??mol??L^-1genipin group. Cell viability was detected by cell counting kit-8 assay (CCK-8). The levels of lactate dehydrogenase (LDH), intracellular total superoxide dismutase (T-SOD) and malondialdehyde (MDA) were assessed using microplate reader. Confocal laser scanning microscope was performed to examine the production of reactive oxygen species (ROS) and the level of mitochondrial calcium (m) as well as the depolarization ratio of mitochondrial membranes potential(????m). The protein expression of cytochrome-C was detected by Western-blot. RESULTS Comparing to control group, the cell viability of H/R group was significantly decreased in a time-dependent manner(r=-0.82,P<0.01). Low concentration(1.25-40 ??mol??L^-1)of genipin could improve cell viability exposed to H/R treatment (P<0.05), on the contrary, high concentration(80-320 ??mol??L^-1) of genipin remarkably reduced the cell viability (P<0.05). In compared with H/R group, the levels of LDH release, MDA production and ROS production, the level of m, depolarization ratio of ????m and the protein expression of cytochrome-c in H/R+(1.25-10 ??mol??L^-1) genipin groups were notably lessened, while the level of T-SOD was increased(P<0.01 or P<0.05). CONCLUSION Genipin could reduce H/R-induced oxidative stress injury, the mechanism might be connected with balancing the oxidative stress products and anti-oxidation enzyme system as well as improving mitochondrial dysfunction.     

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??OBJECTIVE To observe the effect and mechanism of Bauhinia championii flavones (BCF) on anti-hypoxia/reoxygenation injury in cardiomyocytes via inhibiting necroptosis. METHODS The cardiomyocytes hypoxia/reoxygenation injury model was developed and pretreated with Bauhinia championii flavones. ELISA was used to evaluate the contents of TNF-??, and the activities of T-AOC were determined by ultraviolet spectrophotometric method. The protein expression of RIPK3 was observed by Western-blotting, and the necroptosis rate was determined by using Annex v-FITC/PI double staining.RESULTS Compared with model group, Bauhinia championii flavones pretreatment alleviated cardiomyocytes injury, increased T-AOC level, decreased the activity of TNF-??, down-regulated the expression of RIPK3, and inhibited cardiomyocytes necroptosis(P<0.05). It had synergistic effect when combined BCF with necrostatin-1(P<0.05). CONCLUSION BCF can inhibit necroptosis and has protective effects on hypoxia/reoxygenation injury, which are associated with increasing the level of T-AOC, down-regulating TNF-?? and RIPK3, and decreasing the cardiomyocytes necroptosis rate.     

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??OBJECTIVE To investigate the protective effect and possible mechanism of Rhizoma Coptis(RC) on lipopolysaccharide(LPS)-induced inflammatory injury in rat hepatocytes(BRL). METHODS LPS-induced BRL cells injury model was established in vitro, then the damaged cells were given different interventions and treatment with 0.175, 0.1 mg?? mL^-1 RC aqueous extract as the test drug, and dexamethasone(Dex) as positive control drug. The optimal test doses of LPS and RC aqueous extract were selected and determined by cell counting kit-8(CCK-8), the cellular apoptosis rate was determined by flow cytometry, TLR4/NF-??B and TLR4/IRF3 signaling pathways and the mRNA level of related inflammatory mediators(TNF-??, IL-1??, IL-6) were detected by RT-PCR, the NF-??B p65 protein expression was analysed by Western blot and immunofluorescence techniques. RESULTS ??Compared with normal control group, 0.1 mg??mL^-1 LPS affected on BRL cells for 24 h, the cell survival rate was decreased significantly(P<0.01), the apoptotic rate increased significantly(P<0.01), the mRNA level of TLR4, NF-??B, IRF3, TNF-??, IL-1??, IL-6 were significantly increased(P<0.01), and the NF-??B p65 protein expression was increased. ??Compared with the model group, 0.1 and 0.175 mg??mL^-1 RC affected on LPS-induced BRL cells for 24 h, the survival rate of BRL cells was increased significantly(P<0.05), the apoptotic rate decreased significantly(P<0.01), the mRNA level of TLR4, NF-??B, IRF3, TNF-??, IL-1??, IL-6 and the NF-??B p65 protein expression were decreased significantly(P<0.01). CONCLUSION Rhizoma Coptis has obviously protective effect on LPS-induced inflammatory injury in rat hepatocytes(BRL), the mechanism of which may be related with inhibiting apoptosis, reducing the release of inflammatory factors such as TNF-????IL-1?? and IL-6, blocking NF-??B p65 protein nuclear translocation, interfering the R4/NF-??B and TLR4/IRF3 signaling pathway.     

香青兰有效部位对OGD/R损伤诱导的人脑微血管内皮细胞保护作用研究

目的 采用泛半胱氨酸天冬氨酸蛋白酶(cysteine aspartic acid protease,Caspase)抑制剂Z-VAD-FMK联合氧糖剥夺/复氧(oxygen-glucose deprivation/re-oxygenation, OGD/R)建立体外脑缺血再灌注损伤模型,探讨了香青兰有效部位(effective parts of Dracocephalum moldavica, EPDM)对人脑微血管内皮细胞(human brain microvascular endothelial cells,HBMECs)的保护作用。方法 CCK-8法检测细胞活力,倒置显微镜下观察细胞形态,NO测定试剂盒检测NO含量,transwell法检测细胞迁移情况,免疫荧光检测血管内皮-钙黏蛋白(VE-Cadherin)的表达情况,免疫印迹检测酪氨酸激酶(SRC)、内皮素-1(ET-1)、紧密连接蛋白Claudin 5、内皮性一氧化氮合酶(eNOS)、及磷酸化eNOS的蛋白表达。结果 与对照组比较,Z-VAD-FMK联合OGD/R可使HBMECs活力下降,细胞形态变差,细胞迁移减少,NO释放量降低,VE-cadherin表达减少,Src、Claudin 5、磷酸化eNOS表达减少,ET-1表达升高;而EPDM预处理可部分逆转Z-VAD-FMK联合OGD/R引起的上述因素的变化。结论 EPDM对体外脑缺血再灌注模型下的HBMECs具有保护作用,可促进细胞增殖、迁移,增强血管舒缩调节能力,并通过增强紧密连接和黏附连接维持屏障功能。     

鹅不食草介导Nrf2/HO-1信号通路在脑缺血再灌注中抗氧化抗炎作用机制研究

目的:探讨鹅不食草对脑缺血再灌注大鼠Nrf2/HO-1信号通路介导的氧化应激反应和神经炎症影响。方法:SD大鼠分为假手术组、脑缺血组、依达拉奉组、鹅不食草100 mg/kg剂量组、鹅不食草200 mg/kg剂量组、鹅不食草400 mg/kg剂量组。采用中动脉栓塞介导大鼠缺血损伤,连续给药后评价大鼠神经功能,脑组织脑梗死面积、氧化因子、炎症因子含量,Nrf2、HO-1 mRNA与蛋白含量。结果:与假手术组相比,脑缺血组大鼠神经功能显著损伤(P<0.05),脑梗死体积增加(P<0.05),氧化因子、炎症因子含量增加(P<0.05)。与脑缺血组相比,鹅不食草组与依达拉奉组改善了大鼠神经功能(P<0.05),减少脑梗死面积(P<0.05),抑制氧化应激与炎症反应(P<0.05),而Nrf2、HO-1 mRNA与蛋白表达增加(P<0.05)。结论:鹅不食草可以介导Nrf2/HO-1信号通路减少大鼠脑中氧化应激反应与炎症因子含量,改善大鼠神经功能。     

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??OBJECTIVE To observe the influence of safflower yellow(SY) on inflammatory injury in cortex of APP/PS1 double Alzheimer??s disease(AD) transgenic mice.METHODS Six-month-old APP/PS1 transgenic male mice were used in the study.The mice were randomly divided into five groupsmodel group, galanthamine group,high,middle and low dose groups of Safflower Yellow,and wild-type mice with same age were selected into normal control group. Each group mice were performed Morris water maze test after given different drugs by gavage for three months. The level of IL-1??, IL-4, IL-10, IFN-?? and iNOS in cortex were detected by ELISA. RESULTS Compared with wild-type controls, the ability of learning and memory were decreased in the model group. The level of IL-1??, IFN-?? and iNOS increased as well as the expression of IL-4, IL-10 decreased(P<0.01). After SY treatment, the learning and memory abilities of middle dose group were elevated (P<0.01); it could obviously down-regulate the expression of IL-1??, IFN-??, iNOS and up-regulate the expression of IL-4 and IL-10 (P<0.01). High and low dose groups could obviously down-regulate the expression of IFN-??, iNOS and up-regulate the expression of IL-10 (P<0.01). High dose group does had obvious effect of up-regulating the expression of IL-4 (P<0.01), Both of High and low dose groups didn??t have statistical significance on the expression of IL-1??.CONCLUSION Safflower Yellow could improve the ability of learning and memory and exert protective effects on inflammation damage in the cortex of APP/PS1 transgenic mice.