线粒体与肝脏缺血再灌注损伤的关系

肝脏是体内最大的代谢器官 ,线粒体是细胞代谢的中心场所 ,线粒体的正常呼吸功能和 ATP生成对组织器官功能及细胞结构的完整性至关重要。但线粒体对缺血、缺氧非常敏感 ,缺血、缺氧的肝组织在血流恢复或复氧后其损伤度不但没有改善 ,反而进一步加重 ,这与细胞钙超载、氧自由基等因素有直接的关系 [1 ]。线粒体是联系钙超载、氧自由基和细胞死亡的中心环节。近年来线粒体在肝脏缺血再灌注损伤中的作用研究较多 ,进展较快 ,本文就这方面的内容综述如下。1　线粒体对钙稳态的调节及缺血再灌注钙超载1.1　细胞内钙稳态线粒体和内质网是细胞内…     

出血性休克或缺血所致线粒体损伤及可能机理

线粒体对出血性休克或缺血等病理因素是非常敏感的,线粒体损伤主要表现为线粒体呼吸功能下降,其功能的结构基础——线粒体内膜的结构和组成等发生了明显变化。同时对线粒体的损伤机理如缺氧、细胞酸中毒、线粒体内膜结构改变、抑制剂以及 Ca~(++)对线粒体损伤等作了概述。     

脓毒症与线粒体损伤

线粒体是细胞的能量供应站、钙离子浓度的调节器、细胞死亡的执行者。脓毒症中细菌毒素的直接损害、免疫损害、氧自由基损害使线粒体的结构和功能发生改变,进而引起线粒体钙超载、呼吸功能障碍、凋亡、DNA损伤,最终使ATP合成受阻,细胞能量供应不足;凋亡调控系统激活,发生细胞凋亡。     

线粒体融合蛋白2与缺血-再灌注损伤

线粒体是重要的细胞器之一,由线粒体外膜和线粒体内膜组成,其结构和功能受线粒体动力学调控.线粒体融合相关蛋白和线粒体分裂相关蛋白可参与线粒体融合和分裂过程,调控线粒体动力学,进而调节细胞结构、功能及能量代谢.其中线粒体融合蛋白(MFN)2是一种位于哺乳动物线粒体外膜上的蛋白,具有三磷酸鸟苷酶活性,可介导线粒体融合,参与线...     

线粒体调控成骨细胞功能的研究进展

成骨细胞是骨修复重建的关键细胞，其增殖、分化、矿化的异常是骨质疏松症等骨代谢疾病的主要发病原因。线粒体在成骨细胞成骨功能中的调控作用不容忽视。近年研究发现，线粒体调控成骨细胞的能力，主要和线粒体氧化磷酸化、线粒体生物发生、线粒体动力学、线粒体自噬介导线粒体数量和功能的维持、为细胞提供能量及信号传导密切相关。本文综述了近年来线粒体调控成骨细胞功能的相关研究进展。     

线粒体结构、功能和常用研究方法

线粒体是一切生物进行生命活动的动力之源,是所有真核生物进行能量代谢、产生ATP的重要场所。不同种类细胞和不同的生理状态,线粒体的形态和结构可能不同,随之它的功能也发生变化。近年来的研究表明线粒体新膜式结构“Crista Junction Model”已取代经典的“Baffle Model”。通过线粒体完整性与膜电位检测、ATP含量与呼吸链效率测定,氧化应激水平的观察等一系列方法,深化了对其功能及调控机制的认识。线粒体正成为当前生命科学和分子医学的新前沿。现就其结构、功能和常用研究方法作一简要总结。     

小鼠肺组织内质网线粒体结构偶联的分离与鉴定

目的:分离并鉴定C57BL/6小鼠肺组织中内质网线粒体结构偶联(ERMC)。方法:选取8~10周C57BL/6雄性小鼠10只,处死小鼠后,取1 g肺组织匀浆后,采用梯度离心法分别分离出肺组织细胞内的内质网、粗提线粒体、ERMC和纯化线粒体,然后采用Western blotting方法对上述结构进行鉴定。结果:通过梯度离心在离心管中间一层白色条带中含有ERMC,底层白色沉淀物为纯化的线粒体,随后将分离纯化的ERMC和其他组分进行鉴别,结果发现蛋白质二硫键异构酶(PDI)主要表达在内质网和ERMC,Cyto C主要分布在线粒体,而钙连蛋白主要表达在内质网,三磷酸肌醇受体-Ⅰ(IP3R-1)作为内质网上的钙离子通道,在内质网和ERMC均有表达,此外,Sigma-1受体分子伴侣(Sig-1R)多表达在ERMC和线粒体中。结论:上述特征性蛋白分子表达分布特点说明本次研究分离提纯肺组织内ERMC的方法有效,可将其应用于后续的实验。     

吸入麻醉药对鼠肝线粒体内膜损伤机制的研究

目的与方法：实验研究了４种吸入麻醉药对鼠肝线粒体细胞色素氧化酶活性及细胞色素含量的影响。结果：氟烷在高浓度可引起线粒体细胞色素氧化酶活性及细胞色素Ｃ含量明显降低；安氟醚、异氟醚高浓度对细胞色素氧化酶活性及细胞色素Ｃ含量也有降低作用；七氟醚对肝线粒体基本无影响。结论：提示氟烷肝毒性引起的线粒体呼吸功能障碍除了与低流性缺氧有关外，还与线粒体内膜结构改变导致对氧利用能力降低直接有关     

线粒体跨细胞转运参与中枢神经系统疾病研究进展

线粒体是细胞氧化磷酸化及合成ATP的主要场所,在维持钙稳态、调节活性氧自由基产生、能量平衡与代谢、诱导程序性细胞死亡中均发挥重要作用。越来越多的证据提示线粒体可在细胞间发生转移,对机体产生保护或有害的影响。多种转运方式介导线粒体的跨细胞转运,然而深入的转运机制仍未完全阐明。目前发现线粒体跨细胞转运在多种中枢神经系统疾病损伤修复中具有重要意义。文章主要就线粒体功能、线粒体跨细胞转运及其转运机制、线粒体跨细胞转运与中枢神经系统疾病进行概述。     

软骨细胞线粒体稳态失衡在骨关节炎发病机制中的作用研究进展

目的 总结软骨细胞线粒体稳态失衡在骨关节炎（osteoarthritis,OA）发病机制中的作用及其临床应用前景。方法 综述国内外相关文献,总结线粒体稳态失衡的相关机制、线粒体稳态失衡与OA发病机制的关系,以及在OA治疗中的应用前景。结果 研究表明,软骨细胞线粒体生物发生异常、氧化还原失衡、动力学失衡、自噬功能受损引起的线粒体稳态失衡,在OA发病机制中发挥重要作用。其中,线粒体生物发生异常会加速OA软骨细胞分解代谢反应,加剧软骨损伤。线粒体氧化还原失衡会导致活性氧（reactive oxygen species,ROS）大量堆积,抑制细胞外基质合成,诱导细胞铁死亡,最终导致软骨退化。线粒体动力学失衡则会导致线粒体DNA突变,腺嘌呤核苷三磷酸生成减少,ROS堆积,加速软骨细胞的凋亡。线粒体自噬功能受损时,功能失调的线粒体无法被及时清除,导致ROS大量堆积,进而使软骨细胞凋亡。研究发现如葛根素、红花黄、虾青素等药物可通过调节线粒体稳态抑制OA的发展,为OA的治疗提供了新思路。结论 软骨细胞线粒体稳态失衡是OA的重要发病机制之一,进一步探索线粒体稳态失衡相关机制对于OA的预防和治疗具有重要意...     

卵母细胞代谢及其调节

卵母细胞发育与成熟过程依赖大量ATP,因此物质代谢旺盛。卵母细胞以葡萄糖和/或其中间产物丙酮酸为底物,在线粒体内氧化磷酸化,产生大量ATP,是卵母细胞最主要的供能形式。但是,卵母细胞糖代谢模式有种属特异性差异。同时,卵母细胞也存在脂肪酸和氨基酸的代谢。除提供能量外,这些代谢产物在细胞信号传导、渗透压调节等方面也有重要作用。卵母细胞代谢的精确调控受多方面因素的影响如卵母细胞胞内、胞间、胞外的调控等。研究卵母细胞代谢及其机制,对探寻改善卵母细胞质量、提高卵母细胞体外成熟效能具有重要的意义。     

Diabetes and inflammatory diseases: An overview from the perspective of Ca2+/3'-5'-cyclic adenosine monophosphate signaling

A large amount of evidence has supported a clinical link between diabetes and inflammatory diseases, e.g., cancer, dementia, and hypertension. In addition, it is also suggested that dysregulations related to Ca²⁺ signaling could link these diseases, in addition to 3'-5'-cyclic adenosine monophosphate (cAMP) signaling pathways. Thus, revealing this interplay between diabetes and inflammatory diseases may provide novel insights into the pathogenesis of these diseases. Publications involving signaling pathways related to Ca²⁺ and cAMP, inflammation, diabetes, dementia, cancer, and hypertension (alone or combined) were collected by searching PubMed and EMBASE. Both signaling pathways, Ca²⁺ and cAMP signaling, control the release of neurotransmitters and hormones, in addition to neurodegeneration, and tumor growth. Furthermore, there is a clear relationship between Ca²⁺ signaling, e.g., increased Ca²⁺ signals, and inflammatory responses. cAMP also regulates pro- and anti-inflammatory responses. Due to the experience of our group in this field, this article discusses the role of Ca²⁺ and cAMP signaling in the correlation between diabetes and inflammatory diseases, including its pharmacological implications. As a novelty, this article also includes: (1) A timeline of the major events in Ca²⁺/cAMP signaling; and (2) As coronavirus disease 2019 (COVID-19) is an emerging and rapidly evolving situation, this article also discusses recent reports on the role of Ca²⁺ channel blockers for preventing Ca²⁺ signaling disruption due to COVID-19, including the correlation between COVID-19 and diabetes.     

全身麻醉药引发的内质网应激和线粒体功能障碍对阿尔茨海默病影响的研究

背景 阿尔茨海默病(Alzheimer's disease,AD)是一种常见的神经退行性病变,以大脑特定区域蛋白蓄积以及神经元缺失为特点.有研究证实全身麻醉药物可引发内质网应激(endoplasmic reticulum stress,ERS)和线粒体功能障碍,进而引起细胞凋亡,而AD的发病及进展与ERS和线粒体功能障碍密切相关. 目的 探究全身麻醉药引发的ERS和线粒体功能障碍对AD的影响. 内容 介绍ERS及其与线粒体功能障碍之间的联系以及全身麻醉药对AD发病及病程影响. 趋向 ERS、线粒体功能障碍及ERS-线粒体之间的联系近年来成为AD临床治疗的重要靶点,也为全身麻醉药物合理选择及配伍提供新思路.     

Effect of low [CaCl2] and high [MgCl2] cardioplegia and moderate hypothermic ischemia on myoplasmic [Ca] and cardiac function in intact hearts

Objective: Cold cardioplegia (CP) protects against ischemic damage in part by reducing [Ca²⁺]_i overload on reperfusion. Hyperkalemic cardioplegic solutions are widely used in coronary artery bypass procedures, and the specific ionic composition of these solutions may contribute to their variable myocardial protective effects secondary to reduced Ca²⁺_i loading. We reported previously that CP decreased the rise in cardiac diastolic (dia) [Ca²⁺]_i observed during 4 h cold storage at 3 °C in Krebs–Ringer's (KR) solution and decreased dia[Ca²⁺]_i and increased systolic (sys) [Ca²⁺]_i and function on reperfusion after cold storage. Our aim here was to determine if low Ca²⁺_o and high Mg²⁺_o adds to the protective effects of high K⁺_o by decreasing [Ca²⁺]_i during ischemia and reperfusion. Methods: We compared effects of 4.5 mM K⁺_o, 2.5 mM Ca²⁺_o and 2.4 mM Mg²⁺_o KR solution with a higher K⁺_o (18 mM), a lower Ca²⁺_o (1.25 mM) and/or higher Mg²⁺_o (7.2 mM) CP solutions on cardiac mechanic function and sys and dia[Ca²⁺]_i during and after moderate hypothermic global ischemia (17 °C for 4 h) in guinea pig intact hearts isolated by the Langendorff technique. Isovolumetric left ventricular pressure (LVP) was measured with a transducer connected to a fluid-filled balloon placed in the LV and [Ca²⁺]_i was measured using indo-1 fluorescence and a fiberoptic cable placed on the LV free wall. Results: For all CP groups compared to the KR control group after 60 min reperfusion, we observed significant lowering of dia[Ca²⁺]_i by 47%, left ventricular diastolic pressure (diaLVP) by 55%, and infarct size by 43%. We also found significant elevation of sys[Ca²⁺]_i by 25%, d[Ca²⁺]_i/dt_max and d[Ca²⁺]_i/dt_min by 33 and 34%, sys–diaLVP by 55%, dLVP/dt_max and dLVP/dt_min by 34 and 40%, coronary flow by 31%, cardiac efficiency by 21%, and MVO₂ by 25%. These results indicate that CP reduces myoplasmic Ca²⁺ loading and improves mechanical and metabolic function on warm reperfusion compared to KR. However, there were no differences in these indices of Ca²⁺_i cardiac function or metabolism among any CP group after warm reperfusion with KR solution. Conclusion: Increasing K⁺_o to produce cardiac arrest was the most cardioprotective effect of CP against ischemia reperfusion injury; lowering Ca²⁺_o or raising Mg²⁺_o did not add to this protective effect or additionally alter [Ca²⁺]_i.     

Endothelin- and sarafotoxin-induced receptor-mediated calcium mobilization in a clonal murine osteoblast-like cell line, MC3T3-E1/B

Previous studies have demonstrated that, in osteoblast-like MC3T3-E1 cells, various endothelin peptides and their homologous sarafotoxins generate prostaglandin E₂ (PGE₂) release through an ET_A receptor subtype. In this study, biphasic Ca²⁺ signals elicited with endothelin (ET)-1, ET-2, ET-3, β-ET, S6a1, and S6b (ET/S6) were measured by microspectrofluorimetric methods in cell suspensions loaded with Fura-2 acetoxymethylester (Fura-2 AM). Phospholipase C (PLC)-dependent calcium activation mechanisms seem to be involved. We found evidence of Ca²⁺ release from thapsigargin-sensitive and non-thapsigargin-sensitive intracellular Ca²⁺ stores as well as Ca²⁺ transmembrane inflow through multiple voltage-independent and Ni²⁺-sensitive cation channels. Using an ET_A receptor antagonist, BQ-123, we showed that this receptor was coupled to Ca²⁺ mobilization. All agonists tested, except S6c (an ET_B-receptor-specific agonist) induced receptor desensitization. Our results demonstrate that the ET/S6-induced Ca²⁺ signaling pathway is mediated via an ET_A-receptor subtype in MC3T3-E1/B cells.     

磨损微粒引起的成骨细胞内质网应激反应在骨溶解中的作用及机制研究

目的：分析内质网应激反应在骨溶解骨组织中成骨细胞凋亡和骨溶解发生发展中的作用,探讨人工关节松动的原因,为人工关节松动的防治提供新的思路和理论依据。方法：采用小鼠颅骨建立磨损微粒诱导骨溶解的动物模型,随机分成4组,每组7只：组1,空白对照组;组2,磨损微粒TiAl6V4纳米合金粉末（TiNPs）组;组3,内质网应激反应阳性对照（TiNPs+Tg）组;组4,内质网应激反应抑制剂（TiNPs+4-PBA）组。通过甲苯胺蓝染色、HE染色和ALP染色观察骨溶解的病理变化;Western Blotting方法检测骨溶解颅骨组织中内质网应激反应标志蛋白的表达变化;TUNEL和Caspase-3免疫组化方法检测骨溶解颅骨组织内成骨细胞的凋亡情况。结果：磨损微粒TiNPs能够在体外诱导骨溶解的发生、加重炎症细胞的浸润以及抑制成骨细胞分化成熟,同时磨损微粒还可以上调成骨细胞内质网应激反应标志蛋白以及促进骨溶解骨组织中成骨细胞的凋亡。在磨损微粒TiNPs的基础上加入内质网应激的抑制剂（4-PBA）后,骨溶解症状明显缓解,骨侵蚀和炎症浸润显著降低,成骨细胞的分化成熟得到改善,凋亡的成骨细胞急剧减少,内质网应激标志蛋白的表达也逐渐减弱。结论：内质网应激参与骨溶解的形成并在骨溶解的发生发展中发挥重要作用。同时,内质网应激可作为一种新的治疗靶点,为临床逆转或治疗骨溶解和无菌性松动提供新的思路和方法。     

Nucleobindin is produced by bone cells and secreted into the osteoid, with a potential role as a modulator of matrix maturation

Nucleobindin (Nuc), also known as CALNUC, is a Ca²⁺-binding protein, located in the nucleus, the Golgi apparatus and the endoplasmic reticulum (ER). The presence of a signal sequence in Nuc suggests secretion from the cell and it has been found in bone extracellular matrix. Within the present study, molecular biological and morphological methods were combined to evaluate the synthesis and distribution of Nuc in and around cells of rat metaphyseal and calvarial bone. Northern blot analysis and in situ hybridization of bone tissues confirmed that the protein was a product of bone cells. By electron microscopy, immunolabeling for Nuc was seen in osteoid of newly formed bone, on all surfaces facing the various bone cells and also in compact bone. Intracellularly, the gold particles were found in the rough ER of osteoblasts, which suggested synthesis of the protein by these cells. Compared to bone sialoprotein and osteopontin, Nuc demonstrated different localization pattern in bone trabeculae, with the majority of labeling restricted to nonmineralized osteoid. Moreover, the role of Nuc during the mineralization process was investigated in rat calvaria-derived primary osteoblasts grown under osteogenic conditions. Semiquantitative RT-PCR and Northern blot analysis showed Nuc expression to be low during cell proliferation, upregulated during differentiation and matrix maturation, but subsequently downregulated during mineralization. In summary, our data show that Nuc was synthesized by osteoblasts and osteocytes, and secreted into the osteoid, suggesting a role as a modulator of matrix maturation in the mineralization process in bone.     

卵泡刺激素、黄体生成素对卵母细胞成熟的调节作用

遵循两细胞两促性腺激素的理论,卵泡刺激素(FSH)、黄体生成素(LH)对于卵泡成熟和卵巢性腺类固醇的产生都是必需的。在体内FSH启动卵泡生长,LH促进卵母细胞成熟。FSH及LH的应用显著缩短了卵母细胞体外成熟(IVM)的时间,提高卵母细胞成熟及发育的潜能,提高妊娠率。因此揭示卵母细胞核质成熟的影响因素,对提高IVM卵母细胞的质量至关重要。     

褪黑素对老化卵母细胞体外发育能力的改善效果评价

目的探讨体外培养阶段(IVC)添加褪黑素(MT)对卵母细胞老化引起发育受损的改善效果。方法采用不同浓度的过氧化氢(H_2O_2)处理小鼠MII期卵母细胞诱导老化,浓度分别为0(对照组)、10、50、100、150μmol/L,体外受精(IVF)后统计各组二细胞率、囊胚形成率,检测老化卵母细胞的线粒体活性及丰度(Mitotracker Red、JC-1)、活性氧(ROS)水平、线粒体拷贝数等指标;在100μmol/L H_2O_2处理条件下,老化卵母细胞在IVF后的培养阶段分别添加不同浓度褪黑素(10-5、10-7、10-9 mol/L),统计二细胞率、囊胚率,并且分别检测各组获得囊胚的细胞数及凋亡率。结果不同浓度H_2O_2诱导卵母细胞老化后,囊胚发育率随H_2O_2浓度的升高而降低,50μmol/L和100μmol/L H_2O_2组囊胚形成率分别为(26.27±0.06)%和(28.46±3.45)%,相比对照组的(34.90±1.77)%显著降低,差异有统计学意义(P0.05);在H_2O_2诱导老化卵母细胞中,100μmol/L、150μmol/L H_2O_2浓度时,ROS水平相比对照组显著升高,差异具有统计学意义(P0.05);而活性线粒体的丰度及拷贝数呈现下降趋势,膜电位呈上升趋势,但相比对照组无显著性差异(P0.05);100μmol/L H_2O_2处理诱导卵母细胞老化后,在培养液中添加10-9 mol/L褪黑素组与老化对照组相比,囊胚率[(29.42±2.39)%vs.(20.87±4.12)%]、囊胚细胞数(39.36±9.78vs.37.91±4.25)均显著升高,凋亡率显著下降(2.57%vs.3.18%),差异均有统计学意义(P0.05);并且这些指标均达到与未经老化处理组的相似发育水平。结论老化卵母细胞体外受精后发育率和发育质量偏低的现象,可以通过在受精后体外培养阶段添加褪黑素得到改善。