小鼠胚着床前线粒体的分布和超微结构变化

为了解小鼠着档前细胞中线粒体的分布和超微结构的变化规律，观察了２细胞胚，４细胞胚，８细胞胚，桑椹胚，早期囊胚和晚期囊胚，２细胞期和桑椹胚期，线粒体绕胞核集，在挤紧的８细胞胚中，线粒体在细胞接触面处的胞质边缘密集。囊胚期，滋养层细胞的线粒体在胞核周围较宽的区域中分布。     

雄激素缺乏对雄性小鼠主动脉增龄性变化的影响

目的探讨雄激素缺乏对C57BL/6J雄鼠主动脉血管壁组织增龄性变化的影响。结论 24只8周龄C57BL/6J小鼠随机分为正常组（n=8）和去势组（n=8）,另外8只C57BL/6J小鼠作为自然衰老组饲养18个月,测定各组血清睾酮浓度,并分离小鼠主动脉测定丙二醛（maleic dialdehyde,MDA）、超氧化物歧化酶（superoxide dismutase,SOD）的浓度,Western-blot测定去磷酸化Rb蛋白表达量以及提取主动脉线粒体DNA,用巢氏PCR技术分析线粒体DNA缺失突变率。结果与正常组相比,去势组血清睾酮浓度明显下降[（1.05±0.53）ng/mlvs（8.67±0.97）ng/ml,P〈0.01];MDA浓度升高[（1.55±0.43）nmol/mgprotvs（2.42±0.41）nmol/mgprot,P〈0.01];SOD浓度降低[（27.92±2.28）U/mlvs（17.09±1.71）U/ml,P〈0.01],线粒体DNA缺失突变的突变率增加[（18.1713±2.4317）%vs（36.8475±3.3365）%,P=0.029];去磷酸化Rb蛋白表达量增加[（0.12±0.06）vs（0.36±0.07）,P=0.001]。去势组与自然衰老组相比,以上衰老标志物差异均无统计学意义（P〉0.05）。结论雄激素缺乏的小鼠主动脉组织衰老进程加快,内源性生理剂量的睾酮浓度能延缓雄性小鼠主动脉的衰老。     

福尔马林急性痛小鼠小脑线粒体相关分子表达变化

目的:探索急性痛状态下小脑线粒体可塑性变化规律。方法:成年雄性C57BL/6J小鼠42只随机分为对照组(control,n=15)和福尔马林组(formalin,n=27),formalin组小鼠再以3个关键时间点(5 min,30 min,1h)分为3组。利用动物行为学方法观察各组小鼠舔足情况、旷场实验和6个诺达思(Noldus)行为学指标;利用商品化试剂盒检测各组小鼠小脑组织匀浆上清中谷胱甘肽(GSH)、超氧化物歧化酶(SOD)的水平变化;利用real time RT-PCR方法检测小脑ANXA10、Drp1、Mfn1、Mfn2、OPA1、SGK1、TFAM、UCP2和UCP4的mRNA的表达。结果:与control组比较,formalin组小鼠舔足时间及次数呈明显双峰模式增加(0～5 min和20～40 min,P <0.05)。旷场实验可见formalin组小鼠呈明显焦虑样情绪(中央活动路程、中央区域停留时间以及中央进入次数均明显减少,P <0.05)。Noldus行为学结果显示formalin组小鼠运动能力减弱(包括移动距离、移动速度及后肢支撑能力显著减小,P &...     

急性肝衰竭小鼠线粒体分裂蛋白DRP1在肝脏和大脑皮层的表达变化

目的:观察C57小鼠急性肝衰竭(acute liver failure,ALF)过程中线粒体分裂蛋白(dynamic-related protein 1,DRP1)在肝脏和大脑皮层中的变化及与肝衰竭的相关性。方法:C57/BL小鼠,随机分为对照组、ALF1d、4 d、7 d组。腹膜腔内注射硫代乙酰胺(TAA)建立ALF模型,利用高架十字和旷场实验测定行为学,取血清检测谷氨酸转氨酶(ALT)、天冬氨酸转氨酶(AST)水平;之后取肝组织,HE染色观察肝组织的病理变化;用Western Blot和RT-q PCR检测DRP1在肝脏和大脑皮层中蛋白及mRNA水平的表达变化。结果:(1)行为学结果显示:ALF各组小鼠自发活动及探索行为均明显降低(P0.05);(2)肝功检测ALF组ALT、AST表达水平明显升高(P0.05);(3)肝组织病理学检查ALF组1 d时肝细胞出现大面积坏死,7 d时肝细胞坏死较1 d、4 d缓解;(4)Western Blot检测DRP1在ALF全肝组织和肝线粒体中明显降低(P0.05);而在大脑皮层全组织蛋白中明显升高(P0.05),但在提取的线粒体中则无明显改变(P0.05);(5)RT-q PCR检测DRP1在肝组织中ALF各组中表达明显降低(P0.05);而在大脑皮层组织中1 d时表达增多,持续到4 d,在7 d时恢复(P0.05)。结论:DRP1在急性肝衰竭小鼠的肝脏和大脑皮层中对线粒体形态学发挥着重要的作用。     

线粒体核糖体蛋白与人类线粒体疾病

哺乳动物线粒体核糖体(mitochondrial ribosome,mitoribosome)在漫长的进化阶段经过一系列的结构重组,rRNA比例降低,新增了部分线粒体核糖体蛋白(mitochondrial ribosomal proteins,MRPs),成为蛋白含量最丰富的核糖体.所有MRPs均为核基因编码,在细胞质中合成,再转运到线粒体,与线粒体基因(mitochondrial DNA,mtDNA)编码的两种rRNA结合.mtDNA除编码tRNA和rRNA外,还编码组成线粒体呼吸链复合体的13种蛋白质.由于线粒体核糖体负责翻译这13种蛋白,MRPs和其他翻译工具的突变和缺陷可造成线粒体的相关疾病.     

解偶联蛋白在肝性脑病小鼠黑质和前额叶皮层中表达变化

目的:观察不同时间内肝性脑病(Hepatic encephalopathy,HE)小鼠黑质和大脑前额叶皮层中线粒体解偶联蛋白2/4(Uncoupling Protein 2/4,UCP2/4)的变化及可能的关系。方法:取C57/BL小鼠,随机分为sham组、HE 1 d、4 d、7 d四组。HE组小鼠腹膜腔内注射硫代乙酰胺(TAA),sham组注射同等剂量的Na Cl。用Elisa检测小鼠血清中血清氨水平;随后分别运用Western Blot和RT-q PCR测定UCP2/4在黑质和前额叶皮层全组织蛋白和线粒体蛋白水平变化以及mRNA水平中的表达差异。结果:(1)肝性脑病组小鼠血氨水平较对照组小鼠明显升高(P0.01);在蛋白水平检测发现,(2)肝性脑病黑质中UCP2在总组织和线粒体水平表达均明显增加(P0.001),而UCP4却无明显变化(P0.05);(3)在肝性脑病前额叶皮层中UCP2和UCP4在总蛋白水平和线粒体提纯蛋白水平中均明显增加(P0.05);在RT-q PCR水平检测,(4)肝性脑病小鼠黑质中UCP2水平显著增加(P0.01),而UCP4未见明显变化(P0.05);(5)肝性脑病小鼠前额叶皮层发现UCP2(P0.001)和(P0.01)UCP4在mRNA水平均显著升高。结论:肝性脑病小鼠黑质和前额叶皮层中线粒体解偶联蛋白2和4发挥着重要的作用。     

线粒体与阿尔茨海默病

随着人类社会老龄化步伐的加快 ,老年人口所占比例逐渐增加 ,老年性痴呆的发病人数随年龄的增高而增多。阿尔茨海默病 (Ａｌｚｈｅｉｍｅｒ’ｓｄｉｓｅａｓｅ ,ＡＤ)作为老年性痴呆的一种重要类型 ,是中枢神经系统的一种渐进性退行性疾病 ,临床上以认知能力下降、记忆损害和痴呆为主要特征 ,神经病理上以脑细胞内神经纤维缠结 (ＮＦＴｓ)和细胞外老年斑 (ＳＰｓ)为主要特征 ,其中ＮＦＴ的主要成分为高度磷酸化的ｔａｕ蛋白 ,ＳＰｓ的主要成分为淀粉样蛋白前体 (ＡＰＰ)产生的β淀粉样肽 (Ａβ)。目前 ,ＡＤ的病因研究较多 ,其中线粒体因在…     

线粒体动力相关蛋白Drp1在小鼠脑出血后炎症反应中的作用

目的：探讨线粒体动力相关蛋白1(Drp1)在小鼠脑出血（ICH）后继发性炎症损伤中的作用及机制。方法：Western blot检测小鼠脑出血后Drp1和磷酸化Drp1(p-Drp1)的表达时间窗,并筛选出选择性Drp1抑制剂（Mdivi-1）在小鼠ICH模型中的最适药物浓度。将小鼠随机分为sham组、ICH组、ICH+溶剂对照组（ICH+Vehicle组）、ICH+Mdivi-1组。采用mNSS评估小鼠神经功能,干湿重法检测脑组织含水量,HE、Nissl染色观察小鼠脑组织形态学改变,MPO染色观察中性粒细胞浸润情况,Western blot检测炎症因子IL-6、TNF-α以及线粒体膜标志蛋白TOM20、COX4Ⅰ1蛋白表达水平。结果：与sham组相比,p-Drp1表达在ICH后12 h显著升高（P<0.01）。与ICH+Vehicle组相比,ICH+Mdivi-1组神经功能缺损评分升高（P<0.01）,脑组织含水量增多（P<0.05）,脑组织损伤程度加重,血肿周围炎症因子IL-6、TNF-α蛋白表达明显升高（P<0.05）,MPO阳性细胞数量明显增加,线粒体膜蛋...     

谷胱甘肽改善线粒体功能障碍减轻小鼠酒精性肝损伤

目的制备小鼠酒精性肝病动物模型,给予还原型谷胱甘肽,观察模型小鼠肝线粒体功能及肝损伤恢复情况。方法C57BL/6J雄性小鼠30只随机分为对照组(Control)、模型组(ALD)、干预组(Glutathione),每组10只。实验结束后处死小鼠进行取材,对小鼠及肝脏进行称重,对肝脏进行病理分析,TUNEL染色检测模型组和干预组肝细胞凋亡情况。检测血液生化指标ALT、AST、TG,比较各组肝组织谷胱甘肽(GSH)与氧化型谷胱甘肽(GSSG)的比值(GSH/GSSG)。结果与对照组相比,模型组小鼠肝重比升高,与模型组相比,干预组小鼠肝重比降低(P<0.05)。模型组小鼠肝脏有气球样变性和脂质沉积,细胞凋亡数量明显多于对照组,干预组小鼠肝脏空泡样病变和脂质沉积减轻,细胞凋亡数量少于模型组(P<0.05)。血清ALT、AST和TG水平模型组比对照组升高(P<0.05),干预组比模型组下降(P<0.05)。肝组织GSH/GSSG水平模型组比对照组显著下降,干预组比模型组显著上升(P<0.05)。结论谷胱甘肽减轻小鼠酒精性肝病模型肝损伤,与改善线粒体功能障碍相关。     

线粒体DNA片段诱导小鼠胚胎成纤维细胞恶性转化

目的 探讨线粒体ＤＮＡ（ｍｔＤＮＡ）片段的恶性转化效应和转化机制。方法 采用基因转入技术，观察经ｍｔＤＮＡ片段转染后小鼠胚胎成纤维细胞（ＮＩＨ３Ｔ３）在裸小鼠体内的成瘤能力，并对形成的肿瘤进行病理观察；同时应用荧光原位杂交（ＦＩＳＨ）方法，观察ｍｔＤＮＡ片段在ＮＩＨ３Ｔ３细胞核内的整合情况。结果 转染后１周，１８％～２０％的ＭＩＨ３Ｔ３细胞核中发现有目的基因探针的阳性杂交信号；转染后２周的培养细胞     

Different effect of methylazoxymethanol on mouse cerebellar development depending on the age of injection

Summary Methylazoxymethanol (MAM), a powerful antimitotic, has been extensively used to affect rodent CNS development. Here we show that MAM causes different effects on mouse cerebellum depending on the age of the injected pup. Sublethal doses were determined for each age. A single injection at birth permanently reduces the number of cells. In addition, the cytoarchitecture was greatly perturbed: Purkinje cells retained an immature aspect and were dispersed through the cerebellar cortex. A single dose of MAM injected into 5 day old mice also affected the number of cells but, at the level of light microscopy, the cytoarchitecture of the cerebellar cortex appeared not to be altered. Purkinje cells, however, showed some immaturity and degenerated around the 22nd postnatal day. This modulation of MAM effect appears to provide a good model for studying cerebellar ontogeny and neuronal plasticity.     

小鼠胚胎心脏发育期T型钙离子通道的表达

目的 检测哺乳动物心脏中T型钙离子通道α1亚基在小鼠胚胎心脏发育过程中表达部位和时间的分布特征。 方法 对小鼠胚胎心脏发育关键时期（Ｅ9.5至胚胎E18.5共9d）的胚胎进行连续切片,用RT-PCR和免疫组织化学的方法对α1G和α1H蛋白进行阳性检测。 结果 α1H从胚胎期10.5d左右开始表达并一直延续到发育晚期（E18.5）,而α1G则从E11.5d延续到E18.5;免疫组织化学的结果显示,T型钙离子通道主要在心肌层以及左右传导束支稳定表达,而在心内膜垫以及心室肌小梁的心内膜内皮和间充质细胞中不表达。 结论 T型钙离子通道对于心肌细胞以及心脏传导组织的形成起到了一定的作用。     

Bcl-2在小鼠肾脏发生发育中的表达

目的 探讨小鼠肾脏发育过程中抗凋亡蛋白Bcl-2的表达. 方法 选取胚龄16(E16d)、18d和出生后1d(P1d)、3、5、7d、14和21d的昆明小白鼠共54只,应用免疫组织化学技术并结合体视学分析方法,观察小鼠肾脏不同发育时期树脂切片上Bcl-2的表达. 结果 在小鼠肾脏发育过程中,Bcl-2的阳性表达主要出现在近端小管,从E18 d到P1 d,Bcl-2阳性表达肾小管面数密度的增长速度最快,在P7d之后,Bcl-2阳性表达肾小管面数密度的增长速度的变化不大. 结论 在肾的早期发育过程中,Bcl-2主要影响了近端小管的存活或死亡.     

Expression of neural cell adhesion molecule (NCAM) during the first molar development in the mouse

NCAM, the neural cell adhesion molecule, was immunolocalized in the mandibular first molar tooth germ of the mouse. NCAM was first detected in the tooth germ of the late bud stage, where only the cells in the outer part of the condensed mesenchyme (primitive dental follicle) exhibited faint immunoreactivity. The entire dental follicle was intensely immunostained for NCAM from cap stage to the stage when root formation started. During root formation, NCAM disappeared from the follicular tissue surrounding the cervical root as well as from the part covering the crown top. This loss of NCAM proceeded in the direction of the root apex, but even after the tooth had achieved functional occlusion, NCAM was still expressed by the mesenchymal cells adjacent to the root apex. On the other hand, NCAM was negative in the dental papilla until birth. After birth, NCAM-immunoreactivity appeared in the basal portion of the dental papilla, but this NCAM-positive area gradually diminished in width during the root elongation. Instead, another NCAM-positive zone appeared in the core of the pulp during root formation. Even in the tooth that had already erupted, the pulp core contained cells that were strongly positive for NCAM immunostaining. In addition to its expression in the above two mesenchymal cell lineages, NCAM was transiently expressed by epithelial components of the tooth germ, some of the cells of the dental lamina and the enamel organ. The results suggest that NCAM participates in several processes of tooth development.     

Expression of the neural cell adhesion molecule (NCAM) during second- and third-molar development in the mouse

Distribution of the neural cell adhesion molecule (NCAM) during the development of the mandibular second- and third-molars of the mouse was studied by indirect immunofluorescence techniques. At the initial stage, NCAM was intensely expressed by the mesenchymal cells surrounding the dental lamina, and by the cap stage NCAM expression by the mesenchymal cells became restricted to the dental follicle. After that, in addition to the follicular mesenchyme, some cells in the basal part of the dental papilla showed NCAM-immunoreactivity for a while after the hard tissue formation had started. During root formation, the follicular cells lost NCAM first from the level of the cervical root and later from the coronal part, while an additional NCAM positive area appeared deep in the dental papilla. Even after the teeth had erupted, NCAM was expressed in the tissue surrounding the apical root and in the pulp core. During the initial and bud stages, the pattern of NCAM expression in the second and third molars was different from that in the first molar, where NCAM was found only after the late bud stage; while from the cap stage onward, it changed in the same sequence as in the first molar. The different pattern of NCAM expression implies that there is a difference in developmental events between the early stages of the first and the other two molars. On the other hand, the common sequence of NCAM expression in the tooth germs later than the cap stage suggests that NCAM plays an essential role in the formation of the basic structure of the teeth and periodontal tissues.     

地塞米松介导胸腺细胞凋亡过程中线粒体和细胞结构蛋白的变化

目的：研究地塞米松(DEX)介导的小鼠胸腺细胞凋亡过程中线粒体质量和结构蛋白变化特点。 方法： 以地塞米松（DEX）诱导小鼠胸腺细胞凋亡为模型，利用Annexin V-FITC/PI双染流式细胞术研究细胞凋亡和坏死，JC-1染色流式细胞术检测线粒体膜电势（△Ψm）和线粒体质量，利用CFDA-SE染色流式细胞术检测细胞结构蛋白变化。 结果： 在1×10-6mol/L DEX诱导下，小鼠胸腺细胞在6 h凋亡比率为(51.25±5.51)%，对照组为(12.03±2.00)%，差异显著（P<0.01）； DEX组坏死比率为(30.25±3.67)%，对照组为(10.11±1.11)%，差异显著（P<0.01）。DEX组在6h时点的线粒体质量显著低于对照组（P<0.01），FL1平均荧光强度分别为（561.62±54.27）和（900.25±38.80）。DEX同时引起线粒体膜电势的显著下降（P<0.01），对照组FL2平均荧光强度为（267.51±26.48），DEX组为（133.17±12.29）。成熟T细胞培养48 h，CFDA-SE法仅检测到亲代单一细胞峰；而在Con A刺激条件下出现3个子代峰。对照组小鼠胸腺细胞在CFDA-SE染色培养6 h条件下，存在(5.25±1.15)％的低荧光强度细胞群，而在DEX刺激下，该群细胞占（47.39±9.76）％，并且在直方图结果上形成明显的细胞峰。 结论： DEX诱导小鼠胸腺细胞凋亡过程中，线粒体质量和细胞结构蛋白均有所下降；CFDA-SE染色流式细胞术可以作为基于细胞结构蛋白变化的凋亡定量检测方法。     

On the development of the sympathetic chain and the adrenal medulla in the mouse

Summary The first visible primordia of the sympathetic chain appear in the anterior thoracic region and run caudally through 8 segments in an embryo of 11 gestation days. At 12 gestation days the trunk reaches from the base of the skull into the sacral region. During the following days the trunk develops into ganglia with interganglionic fibers.At 16 gestation days there is a well developed trunk with a cranial ganglion cervicale superior and a ganglion stellatum in the anterior part of the thoracic region. In the remaining sympathetic chain there are segmentally arranged ganglia but in this common pattern large differences are noticed.At the 13th gestation day the first signs of the adrenal medulla and the splanchnic plexus appear in the form of sympathoblasts ventral to the sympathetic chain. The migration of sympathoblasts into the primordia of the adrenal cortex goes on for 3 days while the migration to the splancnic plexus in the mesenchym ventral and lateral to the aorta goes on at least 2 more days.     

A study of neurogenesis in the forebrain of opossum pouch young

Summary Morphological changes of growing axons and dendrites are traced in rapid Golgi preparations of the olfactory bulb and the striatum and the parietal, cingulate, and hippocampal cortex. In age-graded series of opossum pouch young changes in the form and position of the differentiating neurons are related to the genesis of the neuronal architecture of each region. In the olfactory bulb there is a specific sequence of changes, starting with the formation of the olfactory glomeruli and the apical dendrites of the mitral cells and ending with the formation of the granule cell processes and the lateral dendrites of the mitral cells. In the ganglionic eminence, the external processes of neuroblasts form axons as they pass from the matrix zone into the presumptive striatal neuropil. Subsequently the perikarya move through the external processes from the matrix zone to the origins of the axons, the external processes disappear, and the dendrites of the striatal neurons start to form. The region of neuronal differentiation is defined by the intersection of the primitive processes of the neuroblasts with the growth cones of specific afferent axons. In the cerebral cortex the neuroblasts resemble primitive epithelial cells (spongioblasts). Their perikarya are in the matrix zone; their primitive processes extend between the internal and external limiting membranes. When afferent growth cones appear in the mantle zone of the parietal area, the perikarya of the pyramidal neuroblasts move to the external limiting layer through their primitive processes. Next these processes disappear; then axons and dendrites differentiate. The order of these events differs in other cortical areas. There is no sign of free, ameboid migration of the neuroblasts. The classical problems of neurogenesis are redefined in the light of these findings.Supported by U.S. Public Health Service Research Grant NS 06115 and GRS Grant 5 S01 FR 05381-08 to Harvard University.With the technical aid of R. R. Morest and P. E. Palmer.     

新生儿小脑发育与胎龄的相关性研究

目的　通过颅脑超声检测25~41周新生儿出生时的小脑,分析小脑随胎龄的发育规律,利于临床早期评估小脑发育。 方法　提取318例25~41周适于胎龄儿颅脑超声图像,测量小脑横径,蚓部高度、前后径、周长和面积,作统计学分析。 结果　小脑横径与胎龄最适曲线为直线方程Y=0.1036X+0.1929,R2=0.727(P<0.001)。蚓部高度与胎龄最适曲线为直线方程Y=0.069X-0.3156,R2=0.823(P<0.001);蚓部前后径与胎龄最适曲线为直线方程Y=0.0509X-0.5798,R2=0.735(P<0.001);蚓部周长与胎龄最适曲线为直线方程Y=0.2811X-1.5932,R2=0.782(P<0.001);蚓部面积与胎龄最适曲线为直线方程Y=0.2283X-4.4559,R2=0.817(P<0.001)。其中蚓部高度与胎龄的正相关性最强,相关系数为0.907(P<0.001)。 结论　在适于胎龄儿中,建立出生时小脑横径,蚓部高度、前后径、周长和面积与胎龄之间的宫外参考范围,以此为依据可为临床宫外诊断和评估小脑发育提供参考。