Mes42细胞及细胞因子对中脑和纹状体神经前体细胞的分化作用

了解中脑神经前体细胞和纹状体神经前体细胞在Mes42细胞及细胞因子作用下的体外诱导分化情况。采用含细胞因子／和Mes42细胞条件培养基和细胞膜碎片的分化培养基培养中脑神经前体细胞和纹状体神经前体细胞，孵育7d后行免疫细胞化学染色了解其体外诱导分化情况。IL－1α（100pg/ml),IL-11(1ng/ml),GDNF(1μg/ml）和LIF(1ng/ml)可诱导中脑神经前体细胞分化成TH阳性神经元，增加分化的TH阳性细胞数目。Mes42细胞条件培养基和其细胞膜裂解碎片以及细胞因子IL－11，LIF，GDNF，IL－1α共同培养神经前体细胞，诱导分化的TH阳性细胞数目无明显增加，但TH阳性细胞的形态更趋成熟。而纹状体神经前体细胞在上述分化培养基内鲜见TH阳性细胞。中脑神经前体细胞可在体外被诱导分化成TH阳性神经元，可能成为帕金森病移植治疗的供体细胞。     

大鼠胚胎中脑神经前体细胞体外诱导多巴胺能神经元的实验研究

目的 在体外多种条件下诱导大鼠胚胎中脑神经前体细胞，比较各自分化出多巴胺能神经元的比例。方法 体外分离、培养出大鼠胚胎中脑腹侧神经前体细胞并传代，通过白介素-1α（IL-1α)、白细胞抑制因子(LIF)、胶质细胞衍生物神经营养因子(GDNF)、纹状体条件培养液和中脑膜裂解碎片诱导分化后，比较子代中酪胺酸羟化酶(TH)免疫阳性细胞的数量和形态变化。结果 IL-1α能增加中脑神经前体细胞分化出的TH细胞数量，但形态不成熟；其他条件不能增加TH细胞数，但与IL-1α共同作用时可使TH免疫阳性细胞形态变得成熟．结论 IL-1α可以诱导中脑神经前体细胞分化多巴胺能神经元，LIF、GDNF、纹状体条件培养液和中脑膜裂解碎片存多巴胺能神绎元的形态发育中起重要作用．     

体外中脑微环境下骨髓基质干细胞向多巴胺能神经元分化的实验研究

目的 应用中脑条件培养基与细胞因子联合诱导骨髓间质干细胞(MSCs)转化为多巴胺能(DA)神经元,探索体外诱导MSCs定向分化为DA能神经元的最佳条件.方法 取雄性Wistar大鼠股骨和胫骨骨髓,进行MSCs的体外培养和传代扩增.采用贴壁培养法使MSCs得到纯化.碱性成纤维生长因子(bFGF)预诱导后,依据加入的神经营养因子不同分为对照组及实验组[单唾液酸四己糖神经节苷脂(GM1)组、胶质源性神经营养因子(GDNF)组、GDNF+GM1组、GDNF+GM1+中脑条件培养基组].诱导过程中在倒置显微镜下观察细胞形态变化,分别在诱导第3、7天进行神经元特异烯醇化酶(NSE)、胶质纤维酸生蛋白(GFAP)、酪氨酸羟化酶(TH)免疫细胞化学检测.计数NSE和TH阳性细胞数,并计算阳性细胞百分比.结果 各实验组于诱导第3、7天见不同数量的NSE、TH阳性神经元样细胞,GFAP阴性.对照组及实验各组7 d时NSE阳性细胞数(个/视野,n=10)分别为:对照组2.214±0.779,GM1组22.014±3.624,GDNF组31.345±2.850,GDNF+GM1组40.314±4.203,GDNF+GM1+中脑条件培养基组45.257±5.999.实验各组以GDNF+GM1+中脑条件培养基组NSE阳性细胞率最高,依次为GDNF+GM1组、GDNF组、GM1组,组间比较差异有统计学意义(P＜0.05).单独应用GDNF和GM1不能诱导MSCs表达TH,二者联合应用可诱导MSCs表达TH,加入中脑条件培养基可使TH阳性细胞率明显增加,而且随着共培养时间的延长,TH阳性表达增加更显著.结论 中脑条件培养基与细胞因子联合可明显促进MSCs向神经元样细胞分化,并促进细胞表达TH.     

IL-1α对神经前体细胞Nurr1基因的诱导表达

目的:了解中脑和纹状体神经前体细胞Nurr1基因的表达以及IL-1α对中脑神经前体细胞和纹状体神经前体细胞Nurr1基因的诱导表达作用.方法:采用RT-PCR,Westem Blot和免疫荧光染色方法观察中脑和纹状体神经前体细胞内Nurr1 mRNA和蛋白质的表达,采用半定量RT-PCR方法观察IL-1α对中脑和纹状体神经前体细胞Nurr1 mRNA的诱导表达作用.结果:中脑和纹状体神经前体细胞均表达Nurr1mRNA和蛋白质,中脑神经前体细胞的Nurr1表达量多于纹状体神经前体细胞.IL-1 α可诱导中脑神经前体细胞Nurr1 mRNA的快速表达,IL-1α(100pg/mL)作用1 h后,Nurr 1mRNA表达增加,3 h后Nurr1 mRNA表达达到高峰,24 h后恢复至基线水平.而纹状体神经前体细胞的Nurr1mRNA在IL-1α诱导后无明显变化.结论:IL-1α可能通过Nurr1的过度表达促进中脑神经前体细胞向多巴胺能神经元方向分化.     

4000/人胚胎干细胞源TH 阳性细胞电压门控性通道的初步研究

目的：检测人胚胎干细胞源TH阳性细胞的神经元性电生理特性。方法：采用我们实验室改良后的“无血清四步法经拟胚体培养体系”的方法,体外诱导人胚胎干细胞源性TH阳性细胞,在对其细胞核型及特异性标志物进行检测的基础上,运用全细胞膜片钳记录的方法,检测其细胞膜上电压门控性离子通道的电生理特性。结果：分化前后细胞核型保持正常;诱导得到的细胞形态一致,大多数细胞（>90%）表达多巴胺能神经元的标志β-tubulion和TH,并仍表达神经前体细胞的标志nestin;膜片钳检测显示诱导分化的TH阳性细胞具有神经元性电压门控钠、钾离子通道。结论：人胚胎干细胞经体外定向诱导分化为TH阳性细胞后具有一定的DA能神经元特性,特别是神经元性电生理特性。     

IL-1α对神经前体细胞Nurr1基因的诱导表达

目的：了解中脑和纹状体神经前体细胞Nurr1基因的表达以及IL-1α对中脑神经前体细胞和纹状体神经前体细胞Nurr1基因的诱导表达作用。方法：采用RT-PCR，Western Blot和免疫荧光染色方法观察中脑和纹状体神经前体细胞内Nurr1 mRNA和蛋白质的表达，采用半定量RT-PCR方法观察IL-1α对中脑和纹状体神经前体细胞Nurr1 mRNA的诱导表达作用。结果：中脑和纹状体神经前体细胞均表达Nurr1 mRNA和蛋白质，中脑神经前体细胞的Nurr1表达量多于纹状体神经前体细胞。IL-1α可诱导中脑神经前体细胞Nurr1 mRNA的快速表达，IL-1α(100pg／mL)作用lh后，Nurr1 mRNA表达增加，3h后Nurr1 mRNA表达达到高峰，24h后恢复至基线水平。而纹状体神经前体细胞的Nurr1mRNA在IL-1α诱导后无明显变化。结论：IL—1α可能通过Nurr1的过度表达促进中脑神经前体细胞向多巴胺能神经元方向分化。     

GDNF和GM1体外联合诱导骨髓基质干细胞向多巴胺能神经元分化的实验研究

目的应用GDNF和GM1体外联合诱导MSCs转化为多巴胺能(DA)神经元。探索体外诱导MSCs定向分化为DA能神经元的最佳条件。方法取雄性Wistar大鼠股骨和胫骨骨髓,密度梯度离心法分离获取单个核细胞。进行MSCs的体外培养和传代扩增。采用贴壁培养法使MSCs得到纯化。依据加入的神经营养因子不同分为对照组及实验组(GM1组、GDNF组、GDNF GM1组)。诱导过程中在倒置显微镜下观察细胞形态变化,分别在诱导第3天、第7天进行NSE、GFAP、TH免疫细胞化学检测。计数NSE和TH阳性细胞数,并计算阳性细胞百分比。采用SPSS10.0软件进行统计学处理。结果对照组可见少量NSE阳性细胞。各实验组比较发现,GDNF GM1组NSE阳性细胞率最高,GDNF组次之,GM1组最低。单独应用GDNF和GM1不能诱导MSCs表达TH,联合应用GDNF和GM1可诱导MSCs表达TH。随着诱导时间的延长,TH阳性表达增加。结论GDNF能够单独诱导MSCs向神经元样细胞分化,GM1不能单独诱导MSCs分化为神经元样细胞,但与GDNF联合,不仅可明显促进MSCs向神经元样细胞分化,而且部分细胞能够表达TH。     

胶质细胞源性神经营养因子诱导骨髓基质细胞向多巴胺能神经元分化的观察

目的 探讨胶质细胞源性神经营养因子（GDNF）在体外能否诱导骨髓基质细胞（BMSCs）向多巴胺（DA）能神经元分化及可能机制。方法无菌条件下，抽取成年SD大鼠胫骨内骨髓组织，分离制备成单细胞悬液进行培养。将增殖传代至第5代的BMSCs随机分为GDNF诱导组和对照组。继续培养7d后，应用BrdU／GFAP、BrdU/NeuN和TH免疫荧光单标和双标技术检测BMSCs增殖和分化情况。结果两组BMSCs继续培养7d后，增殖仍然活跃，有部分细胞向神经元和胶质样细胞分化，呈Brdu，GFAP、BrdU/NeuN和TH阳性表达，但GDNF组的增殖力更强，向神经元和TH神经元分化的数量明显多于对照组（P〈0．05）。结论GDNF能促进BMSCs的增殖和诱导BMSCs分化成神经元和胶质样细胞，其中少部分可分化为TH神经元（即DA能神经元）。     

胚胎大鼠脑纹状体和中脑神经干细胞的培养

目的研究大鼠脑不同部位胚胎神经干细胞的增殖分化特性。方法采用无血清培养基分离和培养大鼠脑的纹状体和中脑的神经干细胞,通过巢蛋白(nestin)表达和5-溴脱氧尿嘧啶(5-bromo-2'-deoxyuridine BrdU)染色,鉴定细胞的增殖能力;通过新生神经元、星形胶质细胞和少突胶质细胞的特异性免疫细胞化学染色,鉴定培养细胞的多潜能性。通过酪氨酸羟化酶的染色(tyrosine hydroxylase,TH)鉴定多巴胺神经元。结果二者在体外培养都可增殖成球,并能分化成神经系统3个谱系的细胞。纹状体增殖传代3个月,中脑培养细胞增殖维持3周。中脑干细胞分化TH阳性细胞比例高于纹状体。结论培养的胎鼠脑细胞是神经干细胞。纹状体干细胞增殖能力高于中脑干细胞,中脑干细胞更倾向于分化成TH阳性细胞。     

周围神经细胞悬液对体外培养的骨髓基质干细胞诱导分化的影响

目的应用周围神经细胞悬液、中脑条件培养基与细胞因子联合诱导骨髓基质干细胞(MSCs)转化为多巴胺(DA)能神经元。探索体外诱导MSCs定向分化为DA能神经元的最佳条件。方法取雄性SD大鼠股骨和胫骨骨髓,进行MSCs的体外培养和传代扩增。碱性成纤维细胞生长因子预诱导后,依据处理因素不同分为对照组及实验组(周围神经细胞悬液组、周围神经细胞悬液 中脑条件培养基组)。倒置显微镜下观察细胞形态变化,在诱导第7天进行神经元特异烯醇化酶(NSE),酪氨酸羟化酶(TH)免疫细胞化学检测。计数NSE和TH阳性细胞数,并计算阳性细胞百分比。结果对照组及实验各组7d NSE阳性细胞数分别为2.304±0.767,37.411±2.89.37.836±2.836(细胞数/每视野)。各组以周围神经细胞悬液 中脑条件培养基组NSE阳性细胞率最高,差异有统计学意义(P<0.01)。对照组及实验各组7d TH阳性细胞数分别为0,10.44±0.511,16.671±0.544(细胞数/每视野)。以周围神经细胞悬液 中脑条件培养基组TH阳性细胞数量较多,差异非常显著(P<0.01)。结论周围神经细胞悬液、中脑条件培养基与细胞因子联合可明显促进MSCs向神经元样细胞分化,并促进细胞表达TH。     

The neuroendocrinology of stress and the pathophysiology and therapy of depression and anxiety

Clinical and preclinical studies have gathered substantial evidence that stress response alterations play a major role in the development of major depression, panic disorder, and post-traumatic stress disorder. The stress response, the hypothalamic pituitary adrenocortical (HPA) system and its modulation by corticotropin-releasing hormones (CRH),corticosteroids,and their receptors, and the roles of natriuretic peptides and neuroactive steroids are described. We review the role of the HPA system in major depression, panic disorder, and post-traumatic stress disorder and its possible relevance for treatment. Impaired glucocorticoid receptor function in major depression is associated with an excessive release of neurohormones such as CRH, to which a number of signs and symptoms characteristic of depression can be ascribed. In panic disorder, a role of central CRH in panic attacks has been suggested. Atrial natriuretic peptide (ANP) is causally involved in sodium lactate-induced panic attacks. Furthermore, preclinical and clinical data on its anxiolytic activity suggest that nonpeptidergic ANP receptor ligands may be potentially useful in the treatment of anxiety disorders. Post-traumatic stress disorder is characterized by a peripheral hyporesponsive HPA system and elevated CRH concentrations in the CSF. This dissociation is probably related to an increased risk of this disorder. We further review recent data that describe an important role of GABA(A)-receptor modulatory,3 alpha-reduced neuroactive steroids in major depression, anxiety, and its treatment. Antidepressants are effective in both depression and anxiety disorders and have major effects on the HPA system,especially on glucocorticoid and mineralocorticoid receptors. Normalization of HPA system abnormalities is a strong predictor of the clinical course, at least in major depression and panic disorder. Currently,CRH-R1 or glucocorticoid receptor antagonists and ANP receptor agonists are being studied and may provide future treatment options more closely related to the pathophysiology of these disorders.     

Polymorphisms of DRD4 and DRD3 and risk of avoidant and obsessive personality traits and disorders

We investigated whether polymorphisms of the dopamine D4 receptor (DRD4) and polymorphisms of the dopamine D3 receptor (DRD3) were associated with personality disorder symptomatology rather than with personality traits such as novelty seeking. DNA was obtained from 145 depressed patients in a clinical trial. These patients were assessed for the presence of personality disorder symptoms and disorders. The 2-repeat allele of the DRD4 exon III polymorphism was associated with increased rates of avoidant and obsessive personality disorder symptomatology. The T,T genotype of the DRD4 -521 C>T polymorphism was also associated with increased rates of avoidant and obsessive personality disorder symptomatology. The Gly9,Gly9 genotype of the DRD3 Ser9Gly polymorphism was associated with increased rates of obsessive personality disorder symptomatology. None of these three polymorphisms were associated with novelty seeking or other temperament traits on the Temperament and Character Inventory. Our results suggest that genetic polymorphisms of DRD4 and DRD3 may well be associated with personality traits, and that conflicting findings to date may arise from the problem of phenotype definition.     

沙盘游戏疗法在地中海贫血患儿心理干预中的应用

本文目的是对沙盘游戏疗法在地中海贫血患儿心理干预中的应用进行综述,以期为地中海贫血患儿的心理康复提供参考。地中海贫血是以珠蛋白生成障碍为主要特征的遗传性疾病,由于长期输血治疗,患儿存在较多的心理和行为问题。沙盘游戏疗法作为一种有效、实用的儿童心理治疗方法,对提高地中海贫血患儿的康复效果、改善生存质量有重要的临床意义。     

癫痫共病抑郁的机制及临床诊疗

本文目的是探讨癫痫共病抑郁的可能机制及临床诊疗。癫痫是一种常见的、慢性的、致残性的神经疾病,癫痫患者生活质量下降,存在明显的负性情绪,常伴发各种精神疾病。癫痫与抑郁具有共同的神经生物学基础,可能存在共同的发病机制。本文从癫痫共病抑郁的发病机制、临床诊断及治疗方面予以总结归纳。     

Efficacy and Effectiveness of Child and Adolescent Psychotherapy and Pharmacotherapy

Decades of intervention research have produced a rich body of evidence on the effects of psychotherapies and pharmacotherapies with children and adolescents. Here we summarize and critique that evidence. We review findings bearing on the efficacy of psychosocial treatments and medications under controlled experimental conditions. We also report evidence, where available, on the effectiveness of both classes of treatment with clinically referred youth treated in real-world clinical contexts. In general, the large body of evidence on efficacy contrasts sharply with the small base of evidence on effectiveness. Addressing this gap through an enriched research agenda could contribute importantly to linking scientific inquiry and clinical practice—to the benefit of both ventures. This is one element of a multifaceted agenda for future research and for synthesis of research, which will require the interplay of multiple disciplines related to child and adolescent mental health.     

Seclusion and restraint and prediction of violence

The authors studied the use of seclusion and restraint on an inpatient unit in a state psychiatric hospital. Of 69 randomly selected inpatients, 51% experienced seclusion or restraint at least once. More psychotic than nonpsychotic patients required seclusion or restraint. However, neither psychosis/nonpsychosis nor voluntary/involuntary admission status predicted the likelihood of violent threats or actions. Patients experiencing seclusion and restraint showed a nonsignificant trend toward longer mean length of stay in the hospital. The frequency of patient behavior leading to seclusion or restraint appeared to be directly related to the stimulation caused by the presence of many staff members and other patients.     

Comparison of trigeminal and spinal modulation of pain and nociception

Modulation of pain and nociception by noxious counterstimulation, also called "diffuse noxious inhibitory controls" or DNIC-like effect, is often used in studies of pain disorders. It can be elicited in the trigeminal and spinal innervation areas, but no study has previously compared effects in both innervation areas. Therefore, we performed a study comparing DNIC-like effects on the nociceptive flexion reflex (NFR) and the nociceptive blink reflex as well as the respective pain sensations. In 50 healthy volunteers, the blink reflex elicited with a concentric electrode and the NFR were recorded before and after immersion of the contralateral hand in cold water. Responses were recorded as the subjective pain sensation and the reflex size. The cold water immersion of the contralateral hand elicited a reduction of both subjective pain sensation and reflex amplitude following the stimulation of both reflexes. However, there were no strong correlations between the individual reductions of both subjective pain sensation and reflex amplitude for both reflexes, and neither when results of the two reflexes were compared with each other. The dissociation between DNIC-like effects on pain and on nociception, which had been found previously already for the NFR, implies that both effects need to be studied separately.