YAP对胶质瘤细胞系U87MG细胞迁移的促进作用

目的评价Hippo信号传导通路下游转录辅助因子YAP对胶质瘤细胞系U87MG细胞迁移的作用。方法利用Westernblot法验证YAP在U87MG中的内源性表达,通过过表达YAP以及siRNA干扰技术分别外源性表达YAP及干扰内源性YAP表达,进而利用细胞划痕实验评价YAP在u87MG细胞迁移中的作用。结果Westernblot结果证实YAP在u87MG细胞中内源性表达;细胞划痕实验结果显示YAP过表达能够促进细胞的迁移,而干扰表达则阻碍细胞的迁移,与对照组比较差异均有统计学意义（P〈0．01）。结论YAP在胶质瘤细胞中表达而且在细胞迁移中发挥促进作用。     

槐定碱通过Wnt/β-catenin信号通路抑制人胶质瘤U87细胞迁移、侵袭

目的 探讨槐定碱对人胶质瘤U87细胞迁移、侵袭能力的影响及其作用机制。方法 体外培养人胶质瘤U87细胞,加入槐定碱[0 mg/ml（对照组）,1.0 mg/ml,2.0 mg/ml,3.0 mg/ml]共培养24 h,划痕实验检测细胞迁移能力,Transwell小室实验检测细胞侵袭能力,免疫印迹法检测细胞β-catenin、E-cadherin、Vimentin和MMP-9蛋白表达,RT-PCR检测细胞Vimentin、MMP-9 mRNA表达。结果 槐定碱明显抑制U87细胞迁移、侵袭能力（P<0.001）,而且随浓度增加抑制作用明显增强（P<0.001）。槐定碱明显抑制U87细胞β-catenin蛋白、Vimentin和MMP-9 mRNA和蛋白表达（P<0.01）,明显增强E-cadherin蛋白表达（P<0.01）,而且随浓度增加抑制作用明显增强（P<0.01）。结论 槐定碱能抑制人胶质瘤U87细胞的迁移和侵袭,机制可能与抑制Wnt/β-catenin信号通路的活性,阻断细胞上皮间质转化过程。     

面包海星皂苷-1对恶性胶质瘤细胞系U87MG增殖抑制的体外研究

目的:初步探讨面包海星皂苷-1(asterosaponins 1)对体外恶性胶质细胞瘤细胞生长的影响及其作用机理。方法:我们使用MTT、细胞周期分析、Hoechst 33342细胞核染色荧光显微镜观察、透射电子显微镜观察等方法了解面包海星皂苷-1对U87MG细胞的作用。结果:实验结果显示,面包海星皂苷-1在很低的浓度(IC50=4.3μg/mL)就够引起人胶质母细胞瘤细胞系U87MG细胞的增殖抑制。当面包海星皂苷-1作用浓度为3.4μg/mL和4.3μg/mL时,U87MG细胞出现S期阻滞,而其浓度为10.0μg/mL时,U87MG细胞出现G0/G1期阻滞。同时,出现的亚二倍体峰(sub-G1)随浓度和时间而增加。Hoechst 33342细胞核染色荧光显微镜观察以及透射电子显微镜下细胞形态观察显示出典型的凋亡细胞形态学特征。结论:体外实验初步显示,面包海星皂苷-1可明显抑制U87MG细胞的增殖,并促其凋亡,具有显著的抗恶性胶质细胞瘤的药理作用。     

靶向沉默c-fos基因表达对胶质瘤U87MG细胞增殖与侵袭的影响

目的探讨靶向沉默c-fos基因表达对胶质瘤U87MG细胞体外增殖与侵袭的影响。方法构建慢病毒载体c-fos-sh RNA,将其转染人脑胶质瘤U87MG细胞,同时将空载体转染细胞作为空载对照,未转染细胞作为空白对照;观察U87MG细胞形态、c-fos m RNA和蛋白表达变化、细胞侵袭和迁移能力、细胞生存率和细胞凋亡率。结果 c-fos-sh RNA慢病毒转染U87MG细胞48 h后,荧光显微镜下观察发现空白对照组U87MG细胞无绿色荧光,而空载体组和转染组U87MG细胞可见绿色强荧光和清晰的细胞轮廓。与空白对照组、空载体组相比,转染组U87MG细胞c-fos m RNA和c-fos蛋白表达显著降低(P0.05);慢病毒转染U87MG细胞24 h后,转染组U87MG细胞存活率显著降低(P0.05),细胞活力显著减弱(P0.05);细胞迁移和侵袭能力明显减弱(P0.05);慢病毒转染72 h后,转染组U87MG细胞凋亡数量显著上升(P0.05)。结论沉默c-fos基因表达可显著抑制脑胶质瘤U87MG细胞增殖和侵袭能力。     

人参皂甙Rh2对人胶质瘤细胞U87MG凋亡的影响

目的 观察人参皂甙Rh2对胶质瘤细胞凋亡的影响并初步探讨其可能机制。方法 将培养的人胶质瘤细胞U87MG随机分为人参皂甙Rh2组、人参皂甙Rh2+尼莫地平组和对照组。人参皂甙Rh2组在常规培养细胞时加入20 μg/ml的人参皂甙Rh2，人参皂甙Rh2+尼莫地平组在人参皂甙Rh2组培养细胞时加入浓度为10 μmol/L的尼莫地平。利用流式细胞仪检测U87MG细胞凋亡，利用激光共聚焦显微镜和流式细胞仪检测U87MG细胞内钙离子浓度。结果 与对照组相比，人参皂甙Rh2促进U87MG细胞凋亡（P<0.05），且增加细胞内游离钙离子浓度（P<0.05）；尼莫地平显著减少人参皂甙Rh2引起的U87MG细胞凋亡（P<0.05）。结论 人参皂甙Rh2可以通过增加细胞内游离钙离子浓度促进U87MG细胞凋亡。     

下调CRNDE基因表达对MAPK/ERK通路中相关基因的影响

目的探讨下调LncRNA CRNDE基因表达对MAPK/ERK通路及转录因子中相关基因的影响。方法将CRNDE基因干扰siRNA转染人胶质瘤U251细胞系,并同时设置空白组及对照组,利用实时PCR检测细胞系中MAPK/ERK通路及转录因子中相关基因表达量的变化。结果与空白组及对照组相比,转染组(si783、si809)中相关基因NF-κB、MEK2、ERK1、ERK2、c-Myc、Raf-1表达量均明显下降(均P 0. 05),具有统计学意义。结论 CRNDE可能通过调节MAPK/ERK通路中及转录因子中一系列相关基因表达,增加胶质瘤细胞的增殖和迁移能力,同时抑制细胞凋亡。     

ArdipusillosideⅠ诱导胶质母细胞瘤细胞凋亡实验研究

目的研究开发诱导胶质母细胞瘤细胞凋亡的新型抗肿瘤药物并探讨其机制。方法以U87MG、原代培养的人源性胶质母细胞瘤株和SVGp12细胞为研究对象,采用甲基噻唑基四唑（MTF）法检测ardipusilloside Ⅰ作用后细胞的增殖活性,通过流式细胞仪分析细胞周期,Hoechst33342细胞核染色,透射电子显微镜观察ardipusillosideⅠ作用后细胞形态学的改变。Western blot检测细胞内FasL、Fas、caspase-8和caspase-3的蛋白表达情况。结果ArdipusillosideⅠ以时间和浓度依赖的方式显著抑制了人胶质母细胞瘤U87MG细胞和原代培养人源性胶质母细胞瘤细胞的增殖活性。随着时间的延长和浓度梯度的增加,处理组逐渐显示出典型的凋亡细胞形态学特征。ArdipusillosideⅠ明显地改变了胶质母细胞瘤的细胞周期。结论ArdipusillosideⅠ成功诱导了人脑胶质母细胞瘤U87MG细胞和原代培养的人源性胶质母细胞瘤细胞发生凋亡。     

MAPK信号通路在创伤性脑损伤中的作用

目的通过建立小鼠创伤性脑损伤(TBI)模型,研究丝裂原活化蛋白激酶(MAPKs)通路中的细胞外调节蛋白激酶1/2(ERK1/2)通路、JNK通路和p38通路的激活及在TBI中的作用及机制。方法建立小鼠TBI模型,通过Western blot检测ERK1/2、JNK和p38的相对磷酸化水平,确定TBI后MAPK通路的激活情况;分别加入ERK1/2通路抑制剂(PD98059,500μmol/L)、JNK通路抑制剂(SP600125,500μmol/L)和p38通路抑制剂(SB203580,500μmol/L),通过脑干湿重检测、神经功能学评分和TUNEL染色评估不同抑制剂对TBI的作用,并通过Western blot检测ERK1/2、JNK和p38的相对磷酸化水平,明确ERK1/2通路、JNK通路和p38通路之间的相互调节作用。结果 TBI可分别引起ERK1/2通路、JNK通路和p38通路的激活;抑制ERK通路和JNK通路可减轻TBI引起的脑水肿、神经功能损伤和细胞凋亡,而抑制p38通路则加重TBI引起的脑水肿、神经功能损伤和细胞凋亡;抑制JNK通路可减少ERK1/2的相对磷酸化水平,而抑制p38通路可增加ERK1/2的相对磷酸化水平。结论 TBI后,ERK1/2通路和JNK通路的激活发挥促进损伤形成的作用,而p38通路的激活则起到神经保护的作用;ERK1/2通路的激活受到JNK通路的促进和p38通路的抑制,表明MAPK通路之间存在相互调节。     

多肽T9sP对胶质瘤细胞抗肿瘤活性的研究

目的 研究来源于E3泛素连接酶TRIM9分子短亚型TRIM9s蛋白的多肽片段T9sP对胶质瘤细胞U87的增殖、迁移和凋亡的影响。方法 采用CCK-8法检测T9sP对U87细胞增殖的影响;划痕实验检测多肽对U87细胞迁移的影响; Trans-well实验检测多肽对U87细胞侵袭的影响; Annexin V-FITC/PI双染后使用流式细胞仪检测多肽对U87细胞凋亡的影响。Western Blot法检测多肽对关键信号通路蛋白表达的影响。结果 穿膜肽TAT可以促进多肽T9sP进入肿瘤细胞。与对照组相比,多肽T9sP可以显著抑制U87细胞的增殖、迁移和侵袭,促进U87细胞凋亡,T9sP可以显著激活p38信号通路。结论 多肽T9sP具有发展为新型抗肿瘤多肽的潜力。     

ERK/P38MAPK信号转导通路在人脐动脉血管平滑肌细胞表型转化中的作用

目的探讨血小板源生长因子-BB（PDGF-BB）及米诺环素是否可通过调节ERK／P38MAPK信号通路,从而影响人脐动脉血管平滑肌细胞（HASMC）的表型转化。方法建立HASMC体外培养模型,分为无血清的DMEM、PDGF-BB（20ng／ml）、PDGF-BB（20ng／ml）＋PD98059（30μmol／L）＋SB203580（20μmol／L）、PDGF-BB（20ng／ml）＋米诺环素（15μmol／L）、PDGF-BB（20ng／ml）＋米诺环素（30μmol／L）五组,Western Blot法检测ERK1／2、P-ERK1／2、P38、P—P38蛋白表达。结果体外培养的HASMC在PDGF-BB（20ng／ml）＋米诺环素（15μmol／L）培养液孵育24h后,P-P38蛋白表达较PDGF-BB组明显下降（P〈0．01）;在PDGF-BB（20ng／ml）＋米诺环素（30μmol／L）组,P—ERK1／2和P—P38蛋白表达均较PDGF-BB组明显下降（P〈0．01）,表明米诺环素显著抑制ERK/P38MAPK信号通路。结论（1）PDGF-BB诱导HASMC的去分化与ERK／P38MAPK信号通路有关,如抑制ERK／P38MAPK信号通路的活性,则HASMC保持分化表型;（2）米诺环素对PDGF-BB诱导HASMC去分化的抑制作用是通过抑制ERK／P38MAPK信号通路的活性,下调PDGF-BB诱导的ERK1／2和P38磷酸化水平而实现的,与其对HASMC的细胞毒性无关。     

Neuroimaging in epilepsy in tropics

Epilepsy is a major public health problem in many tropical countries. Also, some of the tropical diseases are major contributors to the higher prevalence of epilepsy in these countries. The etiologic factors responsible for epilepsy in these countries are quite different from those in the developed world. This article discusses the etiologic factors and neuroimaging of epilepsy in light of the conditions in these tropical countries.     

癫痫与抑郁关系的研究进展

抑郁是癫痫患者中常见的精神障碍,严重地影响了患者的生活质量。传统的观点认为癫痫患者因为存在着诸多社会学问题易出现抑郁倾向,癫痫和抑郁是单向的联系,但大量的研究已经证明癫痫和抑郁之间存在双向的联系,一种异常状态的存在可能易转化为另一种异常状态的发展。癫痫和抑郁存在着共同的发病机制。本文主要就癫痫和抑郁的双向联系以及抗抑郁药物在癫痫患者中的应用进行阐述。     

Sudanophilic lipid accumulation in astrocytes in periventricular leukomalacia in monkeys

Summary Sudanophilic lipid accumulation is a characteristic feature of periventricular leukomalacia (PVL) of infants. At least two types of lipid-containing cells have been identified, one being the macrophage, the other the pre-myelin glial cell. A third type of lipid-containing cell has been seen in two monkeys with spontaneous PVL. Electron microscopically this cell appears to be an astrocyte. This probably represents a reaction of the astrocyte to hypoxia and may be the equivalent of the hypertrophic astrocytes found in human infants.Supported in part by NIH grant HDO 8633 and the Regional Primate Research Center Grant RR-00166     

Increase in cathepsin D activity in rat brain in aging

Cathepsin D-like activity in homogenates of five brain areas of 3-month-old and 24-month-old Fischer 344 rats was measured. With hemoglobin as substrate at pH 3.2, more than 90% of the activity was inhibited by pepstatin. In each area studied, activity was more than twice as high in the old rat brain: 140-160% higher in the cortex, cerebellum, pons-medulla, and striatum and 90-100% higher in the hippocampus and spinal cord. The greatly increased metabolic capacity in the absence of an increase in protein turnover may have a role in age-related pathological degeneration in the brain.     

Characteristics of nociceptors in the periodontium--an in vitro study in rats

Recent studies have indicated that nociceptors can be classified into various types according to their physiological properties. These studies have clarified that the frequency distribution of various nociceptor types is different among body sites and animal species. In the present study, we investigated the physiological properties of rat's periodontal nociceptors in an in vitro jaw-nerve preparation. Responses were recorded from functional single filaments in the inferior alveolar nerve. To determine the nociceptor type, calibrated von Frey filaments, heat, and bradykinin (BK) stimuli were used. We found five subtypes of nociceptors in the periodontal ligaments of the lower incisor: Adelta-high threshold mechanonociceptors (Adelta-HTM, n=28), Adelta-mechanoheat nociceptors (Adelta-MH, n=6), Adelta-polymodal nociceptors (Adelta-POLY, n=26), C-high threshold mechanonociceptors (C-HTM, n=3) and C-polymodal nociceptors (C-POLY, n=4). Most nociceptors were Adelta-innervated, while only a small number of C-innervated nociceptors were found. The present results suggest that periodontal nociceptors transmit mainly fast pain, and may thus play a role in rapid detection of injure-related stimuli during mastication.     

Gender in Voice Perception in Autism

Deficits in the perception of social stimuli may contribute to the characteristic impairments in social interaction in high functioning autism (HFA). Although the cortical processing of voice is abnormal in HFA, it is unclear whether this gives rise to impairments in the perception of voice gender. About 20 children with HFA and 20 matched controls were presented with voice fragments that were parametrically morphed in gender. No differences were found in the perception of gender between the two groups of participants, but response times differed significantly. The results suggest that the perception of voice gender is not impaired in HFA, which is consistent with behavioral findings of an unimpaired voice-based identification of age and identity by individuals with autism. The differences in response times suggest that individuals with HFA use different perceptual approaches from those used by typically developing individuals.     

Defects in Bioenergetic Coupling in Schizophrenia

Synaptic neurotransmission relies on maintenance of the synapse and meeting the energy demands of neurons. Defects in excitatory and inhibitory synapses have been implicated in schizophrenia, likely contributing to positive and negative symptoms as well as impaired cognition. Recently, accumulating evidence has suggested that bioenergetic systems, important in both synaptic function and cognition, are abnormal in psychiatric illnesses such as schizophrenia. Animal models of synaptic dysfunction demonstrated endophenotypes of schizophrenia as well as bioenergetic abnormalities. We report findings on the bioenergetic interplay of astrocytes and neurons and discuss how dysregulation of these pathways may contribute to the pathogenesis of schizophrenia, highlighting metabolic systems as important therapeutic targets.     

Ethnic disparities in problem behaviour in adolescence contribute to ethnic disparities in social class in adulthood

BACKGROUND: It is important for prevention of social class disparities to know how ethnic disparities in social class arise among migrant children. We contribute to this understanding by examining the role of problem behaviour in adolescence. METHODS: Prospective observational study with 753 Dutch native and 217 Turkish migrant adolescents (11-18 year) followed for 10 years. Internalising and externalising problems were assessed in adolescence and employment status and occupational level were assessed in adulthood. The difference in odds ratios (OR) before and after adjustment for internalising and externalising problems was an indication of the predictive value of disparities in internalising and externalising problems for the development of social class disparities. RESULTS: A total of 135 (62%) of the Turkish and 602 (80%) of the Dutch adults were employed. Internalising and externalising problems were not associated with employment status. Of the employed, 65 (48%) Turkish and 179 (30%) Dutch adults worked in low-level occupations (p < 0.0001). Internalising and externalising problems were associated with both ethnicity and occupation. The OR for low-level occupation for Turkish adults was 1.78 (1.19-2.65), indicating ethnic disparities. Adjustment for internalising problems lowered the OR with 36% to 1.50 (0.97-2.31), and adjustment for externalising problems lowered it with 8% to 1.72 (1.15-2.57). Findings were similar for men and women and did not vary by age. CONCLUSIONS: Ethnic disparities in occupational level in adulthood could partly be attributed to disparities in mental health between Turkish migrants and Dutch natives in adolescence. Prevention of ethnic disparities in mental health at young age may therefore also contribute to the prevention of occupational differences in adulthood.     

Progress in neuroprotection in Parkinson's disease

Slowing or aborting the progress of neurodegeneration in Parkinson's disease (PD) remains the most important unmet need of this disorder. There are several recent developments in trial design and also in drugs under investigation for possible neuroprotective effect. Emphasis has been placed on clinical as opposed to imaging end-points and these include change in a clinical rating scale, e.g. United Parkinson's disease Rating Scale (UPDRS), or time to additional therapy. The introduction of the delayed-start, or wash-in, trial design adds an additional dimension to drug evaluation for neuroprotection. Compounds that have been recently tested in clinical trial include the monoamine oxidase-B inhibitor rasagiline, the anti-apoptotic agents TCH346 and CEP1347, and the promitochondrial agent creatine. The dopamine agonists have been evaluated for a neuroprotective effect using imaging end-points. Perhaps the most important and simplest concept for neuroprotection has been the theory that early dopaminergic support for the degenerating dopaminergic system per se provides significant long-term clinical benefit for PD patients.     

Changes in parasympathetic system in medulla oblongata in male pigs in the course of tachycardia-induced cardiomyopathy

Background

Autonomic imbalance constituting a fundamental feature of heart failure (HF) has been assessed mainly at the periphery. Changes in the functioning of autonomic centers in the brain remain unclear. We investigated the molecular elements of parasympathetic system, i.e. α7 nicotinic acetylcholine receptor (α7nAChR) and enzymes metabolizing acetylcholine (acetylcholinesterase, AChE, choline acetyltransferase, ChAT) in medulla oblongata (MO) of male pigs with chronic tachycardia-induced cardiomyopathy.

Methods

The mRNA levels of AChE, ChAT, α7nAChR and X-box binding protein 1 (spliced form, XBP1s) in MO were analyzed using qPCR, AChE and ChAT activities using spectrophotometry, proteasome activity using fluorometry, and the protein level of α7nAChR using Western blotting.

Results

The development of systolic HF was accompanied by an increase in circulating catecholamines, a decrease in the AChE and α7nAChR mRNA in MO, an increase in AChE activity (all p < 0.05), and no change in either the mRNA or activity of ChAT. Both circulating catecholamine levels and AChE activity were inversely related to systolic function of left myocardial ventricle (p < 0.05). The level of α7nAChR protein in MO and its cytoplasmatic fraction were higher in pigs with moderate and severe HF as compared to the other animals (p < 0.01). There was no difference in proteasome activity in MO between diseased and healthy animals, whereas the XBP1s mRNA decreased during HF progression (p < 0.05).

Conclusions

Molecular elements of parasympathetic system are changed within the medulla oblongata during the progression of systolic non-ischemic heart failure in male pigs, indicating a functional link between MO and heart in HF.