Knockdown of human Oxa1l impairs the biogenesis of F1Fo-ATP synthase and NADH:ubiquinone oxidoreductase

The Oxa1 protein is a founding member of the evolutionarily conserved Oxa1/Alb3/YidC protein family, which is involved in the biogenesis of membrane proteins in mitochondria, chloroplasts and bacteria. The predicted human homologue, Oxa1l, was originally identified by partial functional complementation of the respiratory growth defect of the yeast oxa1 mutant. Here we demonstrate that both the endogenous human Oxa1l, with an apparent molecular mass of 42 kDa, and the Oxa1l-FLAG chimeric protein localize exclusively to mitochondria in HEK293 cells. Furthermore, human Oxa1l was found to be an integral membrane protein, and, using two-dimensional blue native/denaturing PAGE, the majority of the protein was identified as part of a 600-700 kDa complex. The stable short hairpin (sh)RNA-mediated knockdown of Oxa1l in HEK293 cells resulted in markedly decreased steady-state levels and ATP hydrolytic activity of the F₁F_o-ATP synthase and moderately reduced levels and activity of NADH:ubiquinone oxidoreductase (complex I). However, no significant accumulation of corresponding sub-complexes could be detected on blue native immunoblots. Intriguingly, the achieved depletion of Oxa1l protein did not adversely affect the assembly or activity of cytochrome c oxidase or the cytochrome bc₁ complex. Taken together, our results indicate that human Oxa1l represents a mitochondrial integral membrane protein required for the correct biogenesis of F₁F_o-ATP synthase and NADH:ubiquinone oxidoreductase.     

Selective toxicity of chrysin on mitochondria isolated from liver of a HCC rat model

Flavonoids are natural compounds that show various biological effects, such as the anti-cancer effect. Chrysin is a flavonoid compound found in honey and propolis. Studies have shown that chrysin has anti-cancer activity due to induction of apoptosis signaling. In the present study, we examined the cytotoxic effect of chrysin against liver mitochondria obtained from the hepatocellular carcinoma (HCC) rat model. Diethylnitrosamine (DEN) and 2-acetylaminofluorene (2-AAF) was used for induction of HCC. Mitochondria were isolated from liver hepatocytes using differential centrifugation. Then, hepatocytes and mitochondria markers related to apoptosis signaling were investigated. Our finding indicated an increase in mitochondrial reactive oxygen species (ROS) generation, collapse in the mitochondrial membrane potential (MMP), swelling in mitochondria, and cytochrome c release (about 1.6 fold) after exposure of mitochondria obtained from the HCC rats group with chrysin (10, 20, and 40 µM) compared to the normal rats group. Furthermore, Chrysin was able to increase caspase-3 activity in the HCC rats group (about 2.4 fold) compared to the normal rats group. According to the results, we proposed that chrysin could be considered as a promising complementary therapeutic candidate for the treatment of HCC, but it requires a further in vivo and clinical studies.     

紫红链霉菌对钉螺酶组织化学的影响

为研究紫红链霉菌灭螺作用的机理,将钉螺分别浸泡于紫红链霉菌培养液(含菌4×106/ml)及去氯水、培养基中36h后,用酶组织化学方法显示各组钉螺肝脏、中枢神经节、头足部及鳃的Mg2 激活的三磷酸腺苷酶(Mg2 ATPase)、胆碱脂酶(ChE)、一氧化氮合酶(NOS)、乳酸脱氢酶(LDH)、琥珀酸脱氢酶(SDH)并观察其变化。结果显示:菌液浸泡组钉螺的Mg2 ATPase活性在肝脏、中枢神经节、头足部及鳃部均明显减弱或完全失活,LDH在中枢神经节、头足部也有一定程度减弱,ChE、NOS、SDH在肝脏、中枢神经节、头足部及鳃部与去氯水组无明显差异;培养基组与去氯水组钉螺相应部位的Mg2 ATPase、ChE、NOS、LDH、SDH活性一致。结果提示:紫红链霉菌的灭螺作用机理主要在于破坏钉螺体内Mg2 ATPase和LDH活性,使ATP生成和利用障碍,最终因能量缺乏而丧失生命功能直至死亡     

老龄大鼠海马CA_3区神经元的酶细胞化学改变及人参茎叶皂甙的抗衰老作用

本实验应用酶细胞化学方法观察了老龄大鼠海马ＣＡ３区神经元的琥珀酸脱氢酶（ＳＤＨ）,酸性磷酸酶（ＡＣＰａｓｅ）的衰老变化,同时对比观察了人参茎叶皂甙的抗衰老作用。实验数据由彩色显微图像分析系统进行定量分析。实验结果提示老龄时海马ＣＡ３区神经元ＳＤＨ酶活性减弱,ＡＣＰａｓｅ活性增强。而人参茎叶皂甙具有促进神经元ＳＤＨ酶活性,降低ＡＣＰａｓｅ活性的作用。本结果为老龄时学习记忆能力下降及人参茎叶皂甙延缓衰老提供了一定的酶细胞化学变化的依据。     

丛枝菌根真菌与根瘤菌互作及类黄酮对互作效果的影响

研究了中华根瘤菌与AM真菌互作或加入适量的类黄酮(150nmol·L^-1和1．5μmol·L^-1)对紫云英的生物量、结瘤数、AM真菌的侵染率、菌丝ALP和SDH酶活性的影响．结果表明,与对照组[0、AMF、Rh]相比,Rh+AMF组紫云英的生物量、结瘤数、AM真菌侵染率和菌丝酶活都有显著差异;与对照组及Rh+AMF组相比,类黄酮处理组紫云英的生物量、结瘤数、AM真菌侵染率和菌丝酶活差异更加显著,但不同类黄酮间(apigenin和hesperitin),或同一种类黄酮不同浓度处理问(150nmol·L^-1和1．5umol·L^-1)差异不明显,说明AM真菌与根瘤菌互作对紫云英的结瘤固氮、AM真菌的侵染和菌丝的生长都有促进作用．当Rh+AMF组加入适量的类黄酮时。这种促进作用更加显著．     

温度对虎斑颈槽蛇3种组织SDH和LDH活性的影响

研究了虎斑颈槽蛇骨骼肌、肝脏、心肌组织中琥珀酸脱氢酶（SDH）、乳酸脱氢酶（LDH）活性对温度的依赖关系。结果表明：虎斑颈槽蛇骨骼肌、肝脏、心肌SDH在5-15℃范围内表现出较低活性,在25℃左右表现出较高活性,该温度可能为SDH的最适温度;除心肌KLDH活性在35℃以上开始下降,其余两种组织的LDH活性随温度的上升而升高。SDH、LDH活性呈现组织问的特异性,3种组织中,SDH活性心肌中最高,LDH活性骨骼肌中最高。     

肿瘤SDH基因突变的研究进展

线粒体琥珀酸脱氢酶(succinate dehydrogenase,SDH)是三羧酸循环和有氧电子传递呼吸链中的关键酶之一,包含A、B、C、D 4个亚基.4个亚基分别由4个基因编码,即SDHA、SDHB、SDHC和SDHD,4个基因突变可以诱发癌症,包括副神经节瘤(paraganglioma,PGL)、嗜铬细胞瘤(pheochromocytoma,PHEO)、肾细胞癌(renal cell carcinoma,RCC)、胃肠道间质瘤(gastrointestinal stromal tumors,GIST)、Leigh综合症等.近年来,突变的SDH已经被证实是一种重要的诊断与预后的生物标志物和治疗分子标靶.本文就SDH存在的各种突变以及在肿瘤发生、发展与转移的作用机理研究的进展进行全面的论述.     

Neuronal apoptosis by prolyl hydroxylation: implication in nervous system tumours and the Warburg conundrum

Oxygen-sensing mechanisms are often dysfunctional in tumours. Oxygen sensing is mediated partly via prolyl hydroxylation. The EglN prolyl hydroxylases are well characterized in regulating the hypoxia inducible factor α (HIF-α) hypoxic response, but also are implicated in HIF-independent processes. EglN3 executes apoptosis in neural precursors during development and failure of EglN3 developmental apoptosis can lead to certain forms of sympathetic nervous system tumours. Mutations in metabolic/mitochondrial enzymes (SDH, FH, IDH) impair EglN activity and predisposes to certain cancers. This is because the EglNs not only require molecular oxygen to execute hydroxylation, but also equally require the electron donor α-ketoglutarate, a metabolite from the Krebs cycle. Therefore EglN enzymes are considered oxygen, and also, metabolic sensors. α-Ketoglutarate is crucial for EglN hydroxylation activity, whereas the metabolites succinate and fumarate are inhibitors of the EglN enzymes. Since EglN activity is dependent upon metabolites that take part in the Krebs cycle, these enzymes are directly tied into the cellular metabolic network. Cancer cells tend to convert most glucose to lactate regardless of whether oxygen is present (aerobic glycolysis), an observation that was first made by Otto Warburg in 1924. Despite the striking difference in ATP production, cancer cells might favour aerobic glycolysis to escape from EglN hydroxylation, resulting in the accumulation of oncogenic HIFα and/or resistance to EglN3-mediated apoptosis.