Preferential resistance of dopaminergic neurons to glutathione depletion in a reconstituted nigrostriatal system

Depletion of glutathione in the substantia nigra is one of the earliest changes observed in Parkinson's disease (PD), and could initiate dopaminergic neuronal degeneration. Nevertheless, we have previously demonstrated that mesencephalic dopaminergic neurons in primary monolayer cultures are more resistant to the toxicity of glutathione depletion than nondopaminergic neurons. To extend this finding to a system that more closely resembles the in vivo situation, we characterized the effects of glutathione depletion on reaggregate cultures derived from ventral mesencephalic and their striatal target neurons, as well as supporting elements including glia. Dopaminergic neurons were found to be more resistant to the toxicity of buthionine-(S,R)-sulfoximine, an inhibitor of glutathione synthesis, than other nigrostriatal neurons, while striatal target cells exhibited an intermediate susceptibility when examined after 48 h. Glutathione depletion, however, decreased the intracellular content of catecholamines after 48 h and eventually led to the loss of dopaminergic neurons after 7 days. Our data indicate that the intrinsic resistance of dopaminergic neurons to the toxicity of glutathione depletion occurs in a variety of experimental paradigms, and suggest that global glutathione depletion alone is unlikely to account for the selective loss of dopaminergic neurons in PD. Rather, it is more likely that either the selective loss of glutathione from dopaminergic neurons, or the combination of glutathione loss with other insults contributes to the preferential death of dopaminergic neurons in PD.     

Estrogen effects on tyrosine hydroxylase-immunoreactive cells in the ventral mesencephalon of the female rat: further evidence for the two cell hypothesis of dopamine function

The present study was undertaken to examine the differential effect of estrogen (E) on the expression of tyrosine hydroxylase (TH) in the substantia nigra compacta (SNc) and in two subdivisions of the ventral tegmental area in ovariectomized (ovx) and ovx plus estradiol benzoate (ovx+E)-treated female rats. Cell counting of TH-immunoreactive perikarya of the SNc, paranigral (PN) and interfascicular (IF) nucleus was performed and compared. Our findings demonstrate that E eliminated TH immunoreactivity from a number of midbrain neurons, while it seemingly did not affect it in others. This signifies a differential effect of E on ventral mesencephalic dopaminergic neurons.     

蛋白酶体功能障碍对多巴胺能PC12细胞的特异性损伤

目的探讨蛋白酶体抑制剂lactacystin对多巴胺能PC12细胞的特异性损伤。方法不同浓度的lactacystin(1、5、10、15和20μmol/L)分别处理多巴胺能PC12细胞和胶质瘤U251细胞24 h,MTT法检测细胞活力;50μmol/L的lactacystin处理U251细胞24 h,MTT法检测细胞活力;10、20μmol/L lactacystin处理PC12细胞,W estern B lot检测细胞内多泛素化蛋白含量;单胺氧化酶B抑制剂selegiline(500μmol/L)和特异性酪氨酸羟化酶抑制剂-αMT(1 mmol/L)提前4 h预处理PC12细胞,再与10μmol/Llactacystin共同作用24 h,MTT法检测细胞活力,W estern B lot检测多泛素化蛋白含量。结果Lactacystin呈剂量依赖性损伤多巴胺能PC12细胞,其对胶质瘤U251细胞无毒性作用,而且其毒性与细胞内多泛素化蛋白生成增加相关。用以增加细胞内多巴胺含量的selegiline和用以减少细胞内多巴胺含量的α-MT都导致lactacystin毒性增强。结论蛋白酶体功能障碍特异性损伤多巴胺能细胞,而其特征性的神经递质多巴胺在其易感性中的作用复杂。     

丘脑底核高频电刺激对大鼠黑质-纹状体多巴胺代谢的影响

目的研究丘脑底核(STN)高频电刺激(HFS)对大鼠黑质-纹状体系统的影响。方法给予正常大鼠一侧STN-HFS,应用微透析观察其对纹状体多巴胺(DA)及其代谢产物的影响,应用逆转录-聚合酶链反应(RT-PCR)和W estern b lot观察其对黑质、纹状体酪氨酸羟化酶(TH)的影响。结果刺激侧纹状体DA代谢产物明显增多(P<0.05),DA水平无变化;RT-PCR检测发现刺激侧黑质、纹状体TH mRNA水平升高;W estern b lot检测发现刺激侧黑质TH表达无变化,而纹状体TH表达明显升高(P<0.01)。结论STN-HFS可能通过影响黑质-纹状体DA代谢及其限速酶的水平发挥作用。     

内蒙古经典型苯丙酮尿症PAH基因外显子11突变的检测

目的：研究内蒙古地区经典型苯丙酮尿症（PKU）苯丙氨酸羟化酶（PAH）基因突变的特点和频率，以提高该地区PKU的基因诊断率。方法：应用PCR—SSCP-银染法和DNA直接测序的方法，对22例内蒙古地区PKU病人PAH基因外显子11进行了检测。站果：检出一种已知突变Y356X（TAC→TAA），突变频率为13．6％。结论：内蒙古地区PKU病人PAH基因外显子11的突变频率均高于我国的其它北方人群，也高于云南PKU病人的突变频率，这为内蒙古地区PKU病人PAH基因分析和产前诊断提供了科学依据。     

小分子EGFR酪氨酸激酶抑制剂盐酸埃罗替尼

盐酸埃罗替尼是一种小分子表皮生长因子酪氨酸激酶可逆抑制剂,通过抑制酪氨酸激酶的磷酸化,阻断信号传导,抑制肿瘤生长.临床前研究表明其对表皮生长因子酪氨酸激酶有抑制作用;临床研究显示该药对多种肿瘤有抗肿瘤活性,不良反应较轻,与化疗药物合用不增加毒性.2004年经FDA批准上市,用于一线化疗失败的局部晚期或转移性非小细胞肺癌的治疗.     

C-KIT突变在伴t(8;21)或inv(16)的急性髓系白血病中的研究进展

近来,在急性髓系白血病(AML)中Ⅲ型受体酪氨酸激酶家族中的成员C-KIT基因的异常引起了人们的重视。在C-KIT突变中,8号外显子突变(mutKIT8)和17号外显子突变(mutKIT17)成为了研究的热点,其在伴有t(8;21)或inv(16)的AML中较常见,与临床预后密切相关。国内外的大量研究表明:C-KIT突变预示着有较高的复发率,预后不良,特别是mutKIT17;伊马替尼通过抑制C-KIT受体酪氨酸激酶活性,对伴有C-KIT突变的患者有良好的疗效,而那些伴有C-KIT激酶结构域D816密码子突变的恶性细胞对其不敏感,但对达沙替尼、PKC412、AP23464、AP23848等敏感。可见,筛查C-KIT突变对预后判断和指导治疗具有重要意义。     

Identification of Inhibitors and Drug Targets for Human Adenovirus Infections

Adenoviruses can cause infections in people of all ages at all seasons of the year. Adenovirus infections cause mild to severe illnesses. Children, immunocompromised patients, or those with existing respiratory or cardiac disease are at higher risk. Unfortunately, there are no commercial drugs or vaccines available on the market for adenovirus infections. Therefore, there is an urgent need to discover new antiviral drugs or drug targets for adenovirus infections. To identify potential antiviral agents for adenovirus infections, we screened a drug library containing 2138 compounds, most of which are drugs with known targets and past phase I clinical trials. On a cell-based assay, we identified 131 hits that inhibit adenoviruses type 3 and 5. A secondary screen confirmed the antiviral effects of 59 inhibitors that inhibit the replication of adenoviruses type 3 or 5. Most of the inhibitors target heat shock protein, protein tyrosine kinase, the mTOR signaling pathway, and other host factors, suggesting that these host factors may be essential for replicating adenoviruses. Through this study, the newly identified adenovirus inhibitors may provide a start point for developing new antiviral drugs to treat adenovirus infections. Further validation of the identified drug targets can help the development of new therapeutics against adenovirus infections.     

General anesthetics selectively modulate glutamatergic and dopaminergic signaling via site-specific phosphorylation in vivo

Isoflurane, propofol and ketamine are representative general anesthetics with distinct molecular mechanisms of action that have neuroprotective properties in models of excitotoxic ischemic damage. We characterized the effects of these agents on neuronal glutamate and dopamine signaling by profiling drug-induced changes in brain intracellular protein phosphorylation in vivo to test the hypothesis that they affect common downstream effectors. Anesthetic-treated and control mice were killed instantly by focused microwave irradiation, frontal cortex and striatum were removed, and the phosphorylation profile of specific neuronal signaling proteins was analyzed by immunoblotting with a panel of phospho-specific antibodies. At anesthetic doses that produced loss of righting reflex, isoflurane, propofol, and ketamine all reduced phosphorylation of the activating residue T183 of ERK2 (but not of ERK1); S897 of the NR1 NMDA receptor subunit; and S831 (but not S845) of the GluR1 AMPA receptor subunit in cerebral cortex. At sub-anesthetic doses, these drugs only reduced phosphorylation of ERK2. Isoflurane and ketamine also reduced phosphorylation of spinophilin at S94, but oppositely regulated phosphorylation of presynaptic (tyrosine hydroxylase) and postsynaptic (DARPP-32) markers of dopaminergic neurotransmission in striatum. These data reveal both shared and agent-specific actions of CNS depressant drugs on critical intracellular protein phosphorylation signaling pathways that integrate multiple second messenger systems. Reduced phosphorylation of ionotropic glutamate receptors by all three anesthetics indicates depression of normal glutamatergic synaptic transmission and reduced potential excitotoxicity. This novel approach indicates a role for phosphorylation-mediated down-regulation of glutamatergic synaptic transmission by general anesthetics and identifies specific in vivo targets for focused evaluation of anesthetic mechanisms.     

五味子乙素对慢性应激抑郁大鼠海马BDNF/TrkB/CREB通路的影响

目的 研究五味子乙素对慢性应激抑郁大鼠海马脑源性神经营养因子（BDNF）/酪氨酸激酶B（TrkB）/环磷腺苷效应元件结合蛋白（CREB）信号通路的影响。方法 40只SD大鼠随机选择10只作为对照组,其余大鼠采用慢性不可预知温和应激（chronic unpredictable mild stress,CUMS）结合孤养制备抑郁症模型,造模结束后随机分为3组：模型组、盐酸氟西汀（3 mg·kg^-1）组、五味子乙素（5 mg·kg^-1）组,每天ig给药1次,连续8周。分别于造模前、造模后及给药后进行旷场、悬尾、强迫游泳行为学实验;通过苏木素-伊红（HE）染色观察大鼠海马形态学改变;免疫组织化学染色（IHC）法观察大鼠海马BDNF蛋白表达;实时荧光定量PCR（qRT-PCR）法检测大鼠海马BDNF、TrkB、CREB mRNA相对表达量;Westernblotting检测大鼠海马BDNF、TrkB、CREB蛋白相对表达量。结果 与对照组比较,模型组大鼠旷场实验水平、垂直得分显著降低（P＜0.05）,悬尾不动时间和强迫游泳漂浮时间显著增加（P＜0.05）;HE染色结果显示海马神经元结构损伤,IHC结果显示海马BDNF表达明显降低;海马BDNF、TrkB、CREB mRNA及蛋白相对表达显著降低（P＜0.05）。与模型组比较,盐酸氟西汀及五味子乙素组大鼠水平、垂直得分显著增加（P＜0.05）,不动时间和漂浮时间显著减少（P＜0.05）;海马神经元结构明显复原,海马组织中BDNF染色明显增加;BDNF、TrkB、CREB mRNA和蛋白相对表达量显著增加（P＜0.05）。结论 五味子乙素可以改善慢性应激抑郁大鼠抑郁样行为、海马区神经元数量及形态,其机制可能与上调BDNF/TrkB/CREB信号通路有关。