小檗碱对心肌缺血再灌注损伤大鼠线粒体自噬及对PINK1/Parkin通路的影响

目的探讨小檗碱对心肌缺血再灌注损伤大鼠线粒体自噬及PTEN诱导激酶1(PTENinducedputativekinase1,PINK1)/帕金森病蛋白(Parkin)通路的影响。方法建立心肌缺血再灌注损伤大鼠模型,随机分组为模型组、小檗碱低、高剂量(75、150 mg/kg)组,自噬抑制剂三甲基腺嘌呤(3-MA,100 mmol/L)组、小檗碱+3-MA(150 mg/kg+100 mmol/L)组,每组12只,另取12只正常大鼠设为假手术组。分组处理后,超声检测大鼠左室功能,记录左室收缩末期内径(LVESD)、左室舒张末期内径(LVEDD)、左心射血分数(LVEF)、左室短轴缩短率(FS);三苯基氯化四氮唑(TTC)染色检测各组大鼠心肌梗死面积,酶联免疫吸附实验(ELISA)检测各组大鼠血清中磷酸肌酸激酶同工酶(CK-MB)、肌钙蛋白I(cTnI)水平;HE染色观察大鼠心肌组织病理变化;透射电镜观察心肌细胞超微结构及线粒体自噬并分析线粒体损伤评分;蛋白免疫印迹法检测各组大鼠心肌组织PINK1、Parkin蛋白及微管轻链蛋白3B(LC3B)、线粒体自噬受体p62(p62)、泛素特异性蛋白酶30(USP30)蛋白表达。结果与假手术组比较,模型组大鼠心肌组织病理损伤严重,线粒体肿胀及空泡化损伤较多,线粒体损伤评分、心肌梗死面积、LVEDD、LVESD、CK-MB、cTnI水平及PINK1、Parkin、LC3B、p62蛋白表达升高(P0.05),LVEF及FS、USP30蛋白表达降低(P0.05)。与模型组比较,3-MA组大鼠心肌组织及线粒体病理损伤加重,LVEF、FS、PINK1、Parkin、LC3B、p62蛋白表达降低(P0.05),线粒体损伤评分、心肌梗死面积、LVEDD、LVESD、CK-MB、cTnI水平、USP30蛋白表达升高(P0.05);小檗碱低、高剂量大鼠心肌组织及线粒体病理损伤减轻,LVEF、FS、PINK1、Parkin、LC3B、p62、USP30蛋白表达升高(P0.05),线粒体损伤评分、心肌梗死面积、LVEDD、LVESD、CK-MB、c TnI水平降低(P0.05)。小檗碱+3-MA组大鼠上述各项指标均与小檗碱高剂量组变化趋势相反,且有统计学差异(P0.05)。结论小檗碱可能通过激活PINK1/Parkin/P62/LC3B通路促进线粒体自噬,升高USP30表达,减少异常自噬,缓解心肌缺血再灌注损伤。     

黄芪甲苷调控线粒体自噬减轻5-Fu诱导老龄大鼠心肌毒性的实验研究#br#

目的　探索黄芪甲苷（AS-Ⅳ）通过PINK1/Parkin信号通路调控自噬减轻5-氟尿嘧啶（5-Fu）诱导老龄大鼠心肌毒性的疗效及机制。方法　18月龄雄性SD大鼠36只,按随机数字表法分为对照组、模型组、AS-Ⅳ组,每组12只。以5-Fu注射液30 mg/kg,腹腔注射,隔日1次,连续7次建立模型组;AS-Ⅳ稀释液,20 mg/kg,灌胃,每日1次,连续14次,建立AS-Ⅳ组。检测血清肌钙蛋白I（cTnI）、肌酸激酶同工酶（CK-MB）、心肌线粒体腺苷三磷酸（ATP）及膜电位水平、行HE及Masson染色观察心肌病理改变、免疫荧光染色检测微管相关蛋白LC3Ⅱ表达、电镜观察线粒体结构并采用Western blot检测PINK1/Parkin通路关键分子PINK1、Parkin、Beclin1、LAMP、LC3蛋白表达。结果　与模型组相比,AS-Ⅳ可减缓大鼠体质量下降,降低心肌cTnI及CK-MB水平,抑制线粒体ATP水平下降（P＜0.05）。与对照组相比,模型组心肌纤维排列紊乱,胶原纤维沉积,胶原容积百分比升高;经AS-Ⅳ干预后,胶原容积百分比减低（P＜0.05）。电镜及LC3Ⅱ荧光染色显示,经AS-Ⅳ干预后,线粒体损伤程度减轻,LC3Ⅱ表达量减低（P＜0.05）。Western blot结果提示AS-Ⅳ苷干预后Beclin1、PINK1、Parkin、LAMP和LC3Ⅱ/Ⅰ表达较模型组降低（P＜0.05）。结论　5-Fu可诱导心肌线粒体损伤,线粒体自噬过度,AS-Ⅳ可通过调节自噬减轻5-Fu心肌毒性。     

不同生境人参茎叶总皂苷对心律失常小鼠的作用比较

目的 探讨园参茎叶总皂苷(GSLS)和林下山参茎叶总皂苷(MSLS)对心律失常小鼠的改善作用.方法 将80只SPF级BALB/c小鼠随机分为正常组、模型组、园参茎叶总皂苷低(GSLS-L)、中(GSLS-M)、高(GSLS-H)剂量组、林下山参茎叶总皂苷低(MSLS-L)、中(MSLS-M)、高(MSLS-H)剂量组....     

Cadmium induces mitophagy through ROS-mediated PINK1/Parkin pathway

Context and objective: Recent reports have highlighted the relationship between cadmium (Cd) and autophagy, however, whether Cd can activate mitophagy remains enigmatic. This study aims to investigate the effects of Cd on mitophagy and its potential mechanism.

Methods: Mice were intraperitoneally injected with Cd for 3?d. Mitochondrial membrane potential (MMP), mitophagosomes, LC3-II/LC3-I ratio, PINK1 level and mitochondrial mass were evaluated to indicate the effects of Cd on mitophagy. To elucidate the mechanism, reactive oxygen species (ROS) scavenger N-acetyl-l-cysteine (NAC) or acetyl-l-carnitine (ALC) as well as the mitophagy inhibitor cyclosporine A (CsA) were introduced to verify the role of ROS in mitophagy.

Results and conclusions: The results showed that Cd significantly induced MMP collapse and typical mitophagosomes formation, increased LC3-II/LC3-I ratio and PINK1 level, and decreased mitochondrial mass, revealing that Cd could induce mitophagy. However, NAC or ALC pretreatment markedly decreased Cd-induced ROS and simultaneously rescued MMP and mitochondrial mass, suggesting ROS played a crucial role in regulating mitophagy. NAC or ALC also dramatically lessened PINK1 level and mitochondrial accumulation of Parkin, indicating that ROS were related to PINK1/Parkin pathway. Notably, CsA compromised Cd-induced mitophagy, PINK1 accumulation and Parkin translocation while failed to block ROS increase, suggesting ROS functioned as an upstream signal for PINK1/Parkin pathway. Taken together, the results indicated that Cd induced ROS-mediated mitophagy through PINK1/Parkin pathway in kidneys of mice. The present study proposes a new perspective to evaluate the nephrotoxicity and its molecular mechanism under Cd exposure in vivo.     

积雪草苷调控SIRT1-FOXO3-PINK1-Parkin通路介导的线粒体自噬保护肾缺血再灌注损伤的机制研究

目的　探讨积雪草苷（AC）调控沉默信息调节因子1（SIRT1）-叉头盒转录因子O3（FOXO3）-PTEN诱导性激酶蛋白1（PINK1）-E3泛素连接酶（Parkin）通路介导线粒体自噬对肾缺血再灌注损伤（RIRI）的保护作用及机制。方法　采用随机数字表法将50只雄性SD大鼠分为假手术组（Sham组）、模型组（Model组）、AC组、AC+SIRT1抑制剂（EX-527）组、EX-527组,每组10只。构建RIRI模型;AC组造模前予以AC混悬液80 mg/（kg·d）连续灌胃4周;AC+EX-527组造模前3 d予以含EX-527（5 mg/kg）的1% DMSO溶液腹腔注射,术中再灌注前20 min腹腔注射1次,余处理同AC组;EX-527组仅予以等量含EX-527的1%DMSO溶液腹腔注射。造模24 h后取材,检测血肌酐（Scr）、尿素氮（BUN）水平,HE染色观察肾组织病理改变及评分,Western blot检测组织SIRT1、FOXO3、PINK1、Parkin通路蛋白和自噬相关蛋白Beclin1、微管相关蛋白轻链3（LC3）A/B-Ⅰ、LC3A/B-Ⅱ表达水平,并计算（LC3A/B-Ⅱ）/（LC3A/B-Ⅰ）;紫外分光光度法检测组织ATP含量;JC染色法检测线粒体膜电位变化。结果　与Sham组比较,Model组Scr、BUN水平升高,肾组织发生病理损伤,通路及自噬相关蛋白表达量出现不同程度升高,ATP含量减少,线粒体膜电位水平下降（P＜0.05）;相比Model组,AC组Scr、BUN水平明显降低,肾组织病理损伤减轻,通路及自噬相关蛋白表达水平升高,ATP含量增高,线粒体膜电位升高（P＜0.05）;与AC组比较,经EX-527干预的AC+EX-527、EX-527组Scr、BUN水平出现不同程度升高,组织病理损伤加重现象,通路及自噬相关蛋白表达量均减少,ATP含量减少,线粒体膜电位水平下降（P＜0.05）,EX-527组程度较为明显。结论　AC通过上调SIRT1-FOXO3-PINK1-Parkin信号通路蛋白表达,促进线粒体自噬来改善肾组织细胞线粒体功能,抑制细胞凋亡,对RIRI起到保护作用。     

Baicalin mitigated IL-1β-Induced osteoarthritis chondrocytes damage through activating mitophagy

Mitophagy is related to chondrocyte homeostasis and plays a key role in the progress of osteoarthritis (OA). Baicalin has a protective effect on OA chondrocytes, the aim of this study was to explore whether the effect of Baicalin on IL-1β-induced chondrocyte injury is related to the regulation of mitophagy. The expression of collagen II in chondrocytes was detected to identify chondrocytes. The effects of different concentrations of Baicalin (10, 20 and 40 μM), autophagy inhibitor (3-Methyladenine), autophagy activator (rapamycin) and Baicalin combined with PI3K agonist (740Y-P) on the viability (cell counting kit 8), apoptosis (flow cytometry), autophagy activation (Monodansylcadaverine staining) and mitochondrial membrane potential (JC-1 kit) of IL-1β-induced chondrocytes were evaluated. The co-localization of autophagosome and mitochondria was determined by immunofluorescence. Apoptosis-, autophagy-, PI3K/AKT/mTOR pathway- and mitophagy-related proteins were detected by western blot. Our result revealed that Baicalin and rapamycin facilitated cell viability, autophagy and mitophagy, elevated mitochondrial membrane potential and suppressed apoptosis of IL-1β-induced rat chondrocytes. In addition, Baicalin and rapamycin upregulated the levels of Bcl-2, Beclin 1, LC3-II/LC3-I, p-Drp1, PINK1 and Parkin as well as downregulated the levels of Bax, cleaved caspase-3, P62, p-PI3K/PI3K, p-mTOR/mTOR and Drp1 in IL-1β-induced rat chondrocytes. However, 3-Methyladenine did the opposite effects of Baicalin and 740Y-P reversed the effects of Baicalin on IL-1β-induced rat chondrocytes. In conclusion, Baicalin activated mitophagy in IL-1β-induced chondrocytes by inhibiting PI3K/AKT/mTOR pathway and activating PINK1/Parkin and PINK1/Drp-1 pathway, thereby reducing the chondrocyte injury.     

黄柏及其酒和盐炙品改善热证大鼠能量代谢及其机制的研究

目的：探索黄柏、酒黄柏及盐黄柏对热证大鼠能量代谢的作用规律。方法黄柏及其酒和盐炙品水煎液9.519、1.058 g/kg ig大鼠7 d,测定生制黄柏对2,4-二硝基苯酚所致热证大鼠肛温,以及血浆中三碘甲腺原氨酸（T3）、四碘甲腺原氨酸（T4）、促甲状腺激素（TSH）、促甲状腺激素释放激素（TRH）、肝组织中乳酸脱氢酶（LDH）、琥珀酸脱氢酶（SDH）、肝糖原、Na+-K+-ATP 酶和 Ca2+-Mg2+-ATP 酶的含量。结果生黄柏、盐黄柏均可以降低热证大鼠肛温,而酒黄柏的作用不明显。生黄柏、盐黄柏可以有效降低热证大鼠的血浆中甲状腺功能轴（T3、T4、TSH、TRH）和肝组织中 LDH、SDH、Na+-K+-ATP酶、Ca2+-Mg2+-ATP酶的含量,并能增加热证大鼠肝组织中糖原含量,而酒炙后对能量代谢指标含量作用减弱。结论生黄柏能够改善热证大鼠的能量代谢,且与甲状腺途径有关。盐炙后寒性增强,对热证大鼠的能量代谢有进一步的改善作用,而酒炙后,缓和黄柏的寒性,对能量代谢改善效果不显著。     

丹酚酸B预处理对心肌缺血/再灌注损伤能量代谢的影响

目的 观察丹酚酸B预处理对大鼠心肌缺血/再灌注损伤（MI/RI）能量代谢的作用。方法 通过结扎冠状动脉30min再灌注2 h建立大鼠MI/RI模型,随机分为4组：假手术组、模型组及丹酚酸B高、低（20、10 mg/kg）组,于建立模型前7 d开始ip给药,每天1次;再灌注结束后,采用比色法测定血清乳酸脱氢酶（LDH）、肌酸激酶（CK）活力,染色法测定心肌梗死面积（MIA）,定磷法测定心肌组织Na⁺-K⁺-ATP酶、Ca²⁺-Mg²⁺-ATP酶活性。结果 与模型组（42.60%）比较,丹酚酸B高、低剂量组的MIA分别缩小至35.93%和37.21%,差异显著（P<0.05）;与模型组比较,丹酚酸B高、低剂量组血清CK、LDH活力均显著降低（P<0.05、0.01）;与模型组比较,丹酚酸B高、低剂量组心肌组织Na⁺-K⁺-ATP酶、Ca²⁺-Mg²⁺-ATP酶活性均显著升高（P<0.05、0.01）。结论 丹酚酸B预处理可保护MI/RI所致心肌损伤,作用途径可能与改善心肌组织的能量代谢相关。     

Blocking Parkin/PINK1-mediated mitophagy sensitizes hepatocellular carcinoma cells to sanguinarine-induced mitochondrial apoptosis

Hepatocellular carcinoma (HCC) remains a major clinical challenge. Although mitophagy is implicated in hepatocarcinogenesis, novel therapeutic options targeting mitophagy for HCC treatment still await further studies. Here, we demonstrate that sanguinarine induces cell death in HCC cell line MHCC-97H through the mitochondrial apoptosis pathway. Sanguinarine triggers mitochondrial dysfunction and PTEN-induced putative kinase 1 (PINK1)/Parkin upregulation and recruitment to mitochondria. Elevated levels of p62 and LC3-II/I ratios suggest that sanguinarine is both an inducer of autophagy and a blocker of autolysosome formation, which is further confirmed by LC3-II conversion levels in presence of autophagy and mitophagy inhibitors, as well as an autophagy activator. In addition, blocking autophagy promotes sanguinarine-induced cell death, indicating mitophagy plays a cytoprotective role in sanguinarine-treated cells. Our findings suggest that blocking mitophagy may contribute to sanguinarine-induced mitochondrial apoptosis through the prevention of damaged mitochondrial clearance.     

The Action of Thioridazine and Promazine on Biological Membranes: Relationship between ATPase Inhibition and Membrane Stabilization

The effects of promazine and thioridazine on hypotonic haemolysis of human erythrocytes are compared with their effect on Na⁺-K⁺-ATPase of washed human erythrocyte ghosts. Promazine (5 × 10^-5 - 5 × 10^-4 M) and thioridazine (10^-5 - 10^-4 M) stabilize erythrocytes against hypotonic haemolysis, but have lytic effects at higher concentrations. Both drugs inhibit Na⁺-K⁺-ATPase of erythrocyte membranes. This inhibition is slight at drug concentrations which have a membrane-stabilizing action, but is complete and irreversible at lytic concentrations of the drugs. Promazine and thioridazine also inhibit Na⁺-K⁺-ATPase in a membrane fraction prepared from rat hearts. The relative order of potency of the two drugs in this respect does not reflect their relative potency as general cardiac depressants. It is concluded that Na⁺-K⁺-ATPase is not a primary target for the action of promazine and thioridazine in the rat heart. It is suggested that inhibition of Na⁺-K⁺-ATPase by these agents is secondary to more general alterations of the physical properties of the cell membrane.     

Twitch responses dependent on external calcium ions in the mouse diaphragm in a potassium-free bathing solution

We examined the effect of external Ca²⁺ on twitches of mouse diaphragm under the inhibition of Na⁺-K⁺-ATPase in vitro. The muscle was directly stimulated in presence of d-tubocurarine (dTc). K⁺ removal potentiated the amplitude of twitches with a transient prior reduction. This potentiation depended on external Ca²⁺. Membrane potentials decreased after removal of both external K⁺ and Ca²⁺ and were restored after an addition of Ca²⁺. The removal of both ions increased the content of Na⁺ in tissues and decreased K⁺. These changes were restored to the levels in the K⁺-free bathing solution by the addition of Ca²⁺. These results imply that Na⁺/Ca²⁺ exchange can support twitch contraction under the inhibition of Na⁺-K⁺-ATPase activity.     

苦参碱对阿霉素诱导大鼠心肌损伤的保护作用及其机制研究

目的研究苦参碱对阿霉素诱导大鼠心肌损伤的保护作用及其机制。方法 SD大鼠随机分为对照组、模型组和苦参碱25、50、100 mg/kg组,每组各20只。模型组大鼠ip注射用阿霉素2.5 mg/kg,1次/周,连续给药6周,累积剂量15 mg/kg,建立心肌损伤模型。对照组ip等量生理盐水。苦参碱组造模前2 d ip注射用苦参碱25、50、100 mg/kg,连续给药5 d。观察大鼠心肌细胞病理学,采用酶联免疫吸附法检测大鼠血清线粒体偶联因子CF6水平,应用分光光度法测定Na~+-K~+-ATP酶、Ca2~+-ATP酶活力,采用试剂盒检测谷胱甘肽过氧化物酶(GSH-px)、总超氧化物歧化酶(SOD)活性及丙二醛(MDA)含量。结果苦参碱各组心肌组织肿胀,肌束间、间质有灶性出血现象显著减轻。与模型组比较,苦参碱各组血清CF6水平显著降低(P0.05);线粒体Na~+-K~+-ATP酶、Ca2~+-ATP酶活性显著升高(P0.05);心肌组织GSH-px活性及SOD活力升高,MDA含量显著降低(P0.05)。结论苦参碱能保护阿霉素引起的大鼠心肌损伤,其作用机制与改善线粒体ATP酶活性、降低线粒体偶联因子6水平、减轻氧化应激水平有关。     

Propentofylline and postischemic brain edema: Relation to Na+-K+-ATPase activity

Following the bilateral occlusion of common carotid arteries in gerbil, an increase in water content and sodium/potassium ratio as well as the inhibition of Na⁺-K⁺-ATPase was found. The xanthine derivative propentofylline (HWA 285) [3-methyl-1-(5-oxohexyl)-7-propylxanthine] given either before or after cerebral ischemia attenuated the development or postischemic brain swelling and the increase in sodium/potassium ratio and prevented the postischemic reduction of Na⁺-K⁺-ATPase activity. It is concluded that the action of propentofylline on brain edema during ischemia is mediated aside from other possible mechanism(s), by the influence of the drug on Na⁺-K⁺-ATPase activity.     

内皮-单核细胞激活多肽对人U118胶质瘤细胞自噬的影响及机制

目的研究内皮-单核细胞激活多肽(EMAP-Ⅱ)对人U118脑胶质瘤细胞自噬的影响及可能机制。方法 EMAP-Ⅱ处理人U118胶质瘤细胞后,采用MTT检测细胞活力;用免疫荧光法检测自噬标记物微管相关蛋白轻链-3(LC3)在U118细胞的分布;Western blot方法检测LC3-Ⅱ、自噬降解底物p62/SQSTM1和自噬相关蛋白Beclin1的蛋白表达水平。结果与EMAP-Ⅱ0 h组相比,0.05 nmol.L-1剂量以上的EMAP-Ⅱ可明显抑制人U118胶质瘤细胞的活力,降低线粒体膜电位(MMP),在EMAP-Ⅱ作用0.5 h时效果最明显;自噬抑制剂3-甲基腺嘌呤(3-MA)能够阻断EMAP-Ⅱ的作用,使细胞活力和线粒体膜电位基本恢复;EMAP-Ⅱ作用0.5 h后自噬标记物LC3在胞质内呈点状分布,荧光表达明显增强,并且LC3-Ⅱ的蛋白表达水平明显上调;同时伴有自噬降解底物p62/SQSTM1的表达下降;此外,自噬相关蛋白Beclin1的蛋白表达水平明显上调。结论 EMAP-Ⅱ能明显抑制人U118胶质瘤细胞活力,诱导U118细胞发生自噬,其机制可能与自噬相关蛋白Beclin1上调有关。     

灯盏花素通过激活线粒体自噬途径减轻糖尿病大鼠心肌缺血再灌注损伤

目的 探讨灯盏花素对糖尿病大鼠心肌缺血再灌注损伤(MIRI)及线粒体自噬途径的影响.方法 采用高糖高脂饮食联合链脲佐菌素制备糖尿病模型,糖尿病模型成功后再制备MIRI模型,具体分组为对照组(不做任何处理)、模型组(糖尿病+MIRI)、假手术组(糖尿病),均ip等体积生理盐水;灯盏花素低、高剂量组(糖尿病+MIRI),分...     

Reversible inhibition of mammalian brain acetylcholinesterase and sodium potassium-stimulated adenosine triphosphatase by cyclopropane.

Membrane-bound and purified forms of acetylcholinesterase (AChE) derived from beef brain caudate nucleus tissue were inhibited reversibly by cyclopropane at low gas pressures (0.025 to 0.25 atm). Inhibition followed mixed kinetics which suggested interactions of the anesthetic gas with both active sites(s) and other sites on the enzyme molecule. At gas pressures of 1 atm and higher, cyclopropane inhibited membrane-bound and solubilized preparations of brain Na⁺-K⁺-ATPase without affecting Mg²⁺-ATPase activity. This inhibition was reversible, followed uncompetitive kinetics and was not due to pressure per se. AT³²P-labeling experiments suggested that cyclopropane inhibited Na⁺-K⁺ATPase at or before the phosphorylation step in the enzyme reaction cycle.     

增龄对大鼠睾丸自噬水平和血睾屏障完整性的影响

目的探究增龄过程中大鼠睾丸自噬水平的变化及其对血睾屏障的影响。方法HE染色法观察6、12、18和24月龄SD♂大鼠睾丸组织形态学的变化。Western blot法检测各组大鼠睾丸自噬相关蛋白Beclin1、ATG5、ATG7和LC3Ⅱ,以及血睾屏障相关蛋白Occludin和β-catenin的表达水平变化。免疫荧光法检测睾丸自噬相关蛋白Beclin1和LC3,以及血睾屏障相关蛋白β-catenin的蛋白表达及定位。结果HE染色观察发现,增龄过程中大鼠睾丸组织形态结构发生明显变化,生精小管萎缩,生精细胞层数减少,细胞间间隙增大,且发生部分脱落现象;Western blot结果显示,增龄过程中大鼠睾丸自噬相关蛋白Beclin1、ATG5、ATG7和LC3Ⅱ,以及Occludin和β-catenin表达水平均逐渐下降;免疫荧光结果显示,增龄过程中大鼠睾丸生精小管上皮细胞中Beclin1和LC3蛋白表达逐渐下调,并且血睾屏障标志蛋白β-catenin表达也逐渐下降。结论增龄过程中大鼠睾丸生精功能减退,其机制与睾丸生精上皮细胞自噬水平下降,进而破坏血睾屏障的完整性有关。     

Synergistic effects of autophagy/mitophagy inhibitors and magnolol promote apoptosis and antitumor efficacy

Mitochondria as a signaling platform play crucial roles in deciding cell fate. Many classic anticancer agents are known to trigger cell death through induction of mitochondrial damage. Mitophagy, one selective autophagy, is the key mitochondrial quality control that effectively removes damaged mitochondria. However, the precise roles of mitophagy in tumorigenesis and anticancer agent treatment remain largely unclear. Here, we examined the functional implication of mitophagy in the anticancer properties of magnolol, a natural product isolated from herbal Magnolia officinalis. First, we found that magnolol induces mitochondrial depolarization, causes excessive mitochondrial fragmentation, and increases mitochondrial reactive oxygen species (mtROS). Second, magnolol induces PTEN-induced putative kinase protein 1 (PINK1)‒Parkin-mediated mitophagy through regulating two positive feedforward amplification loops. Third, magnolol triggers cancer cell death and inhibits neuroblastoma tumor growth via the intrinsic apoptosis pathway. Moreover, magnolol prolongs the survival time of tumor-bearing mice. Finally, inhibition of mitophagy by PINK1/Parkin knockdown or using inhibitors targeting different autophagy/mitophagy stages significantly promotes magnolol-induced cell death and enhances magnolol's anticancer efficacy, both in vitro and in vivo. Altogether, our study demonstrates that magnolol can induce autophagy/mitophagy and apoptosis, whereas blockage of autophagy/mitophagy remarkably enhances the anticancer efficacy of magnolol, suggesting that targeting mitophagy may be a promising strategy to overcome chemoresistance and improve anticancer therapy.     

RNA interference-mediated downregulation of Beclin1 attenuates cerebral ischemic injury in rats

Aim:

To test the role of the Beclin 1-dependent autophagy pathway in brain damage during cerebral ischemia.

Methods:

Focal cerebral ischemia was established in rats using a middle cerebral artery occlusion (MCAO) model. A lentiviral vector-associated RNA interference (RNAi) system was stereotaxically injected into the ipsilateral lateral ventricle to reduce Beclin1 expression. We measured the ipsilateral infarct volume, autophagosome formation, neurogenesis and apoptosis, all of which could be modulated by Beclin1 RNAi.

Results:

On the 14th day after MCAO, Beclin1 downregulation by RNAi increased the population of neural progenitor cells (BrdU⁺-DCX⁺), newborn immature cells (BrdU⁺-Tuj-1⁺) and mature neurons (BrdU⁺-MAP-2⁺), and reduced the apoptosis of immature neurons (caspase-3⁺-DCX⁺ and caspase-3⁺-Tuj-1⁺) surrounding the ischemic core of the ipsilateral hemisphere. Furthermore, RNAi-mediated downregulation of Beclin1 decreased infarct volume and inhibited histological injury and neurological deficits.

Conclusion:

RNAi-mediated downregulation of Beclin1 improves outcomes after transient MCAO.     

(Na+-K+)-ATPase inhibitors: Implications for new drug discovery

The (Na⁺-K⁺)-ATPase is a ubiquitous membrane-bound enzyme that actively transports Na⁺ out of the cell in exchange for a smaller ratio of extracellular K⁺. The current report focuses on the role of modifiers of (Na⁺-K⁺)-ATPase activity in the development of new pharmacological agents. More versatile biological test systems are proposed. Possible use of Na⁺-K⁺-ATPase activity modulators in treatment of hypertension and other diseases are discussed. It is concluded that the ubiquitous distribution and crucial role of the enzyme in normal and diseased cell function merits a critical biological and chemical reappraisal of the enzyme beyond the current narrow viewpoint dealing with the application of glycosides in the treatment of congestive heart failure.