黄芩苷对注意缺陷多动障碍大鼠突触体ATP酶和LDH的影响及对AC/cAMP/PKA信号通路的调控作用

目的研究黄芩苷对注意缺陷多动障碍(ADHD)模型大鼠脑突触体腺苷三磷酸(ATP)酶、乳酸脱氢酶(LDH)的影响及对腺苷酸环化酶(AC)/环磷酸腺苷(cAMP)/蛋白激酶A(PKA)信号通路的调控作用。方法将40只SHR大鼠随机分为模型组、盐酸哌甲酯组(0.07 mg/mL)、黄芩苷低(3.33 mg/mL)、中(6.67 mg/mL)、高剂量组(10 mg/mL),每组8只,另设WKY大鼠8只为正常对照组。Percoll密度梯度离心法制备脑突触体,电镜观察突触体结构;运用比色法检测突触体ATP酶、LDH活性;运用ELISA法检测突触体内AC、cAMP、PKA的含量。结果与正常对照组比较,模型组大鼠脑突触体存在ATP酶活性降低、LDH活性升高及AC、cAMP、PKA含量降低的改变(P0.05)。与模型组比较,盐酸哌甲酯及黄芩苷中、高剂量均能显著升高ADHD模型大鼠脑突触体内ATP酶活性(P0.05),降低LDH活性(P0.05),并显著提高AC、cAMP、PKA含量(P0.05)。与盐酸哌甲酯组比较,高剂量黄芩苷治疗ADHD模型大鼠对改善上述各指标水平的效果更为显著(P0.05)。与黄芩苷低剂量组比较,黄芩苷高剂量组ATP酶活性显著升高(P0.05);黄芩苷中、高剂量组LDH活性显著降低,AC、cAMP、PKA含量均显著升高(P0.05)。与黄芩苷中剂量组比较,黄芩苷高剂量组ATP酶活性显著升高(P0.05)。结论盐酸哌甲酯及黄芩苷均能改善ADHD模型大鼠脑突触体ATP酶、LDH活性;黄芩苷的作用与剂量存在相关性,且高剂量黄芩苷较盐酸哌甲酯效果更为显著;黄芩苷可能是通过上调AC/cAMP/PKA信号通路发挥治疗作用。     

激活Gs蛋白对胆红素诱导的大鼠小脑颗粒神经元凋亡影响的研究

目的　研究激活Gs蛋白对胆红素诱导的小脑颗粒神经元凋亡的影响及作用机制。方法　采用原代培养的新生大鼠小脑颗粒神经元 ,用霍乱毒素 (CT)激活Gs蛋白 ,用二乙酸荧光素(FDA)染色及四甲基偶氮唑蓝 (MTT)法测定胆红素诱导的小脑神经元存活率 ,12 5I放射免疫法测定细胞内环磷酸腺苷 (cAMP) ,Fura 2 /AM荧光比值成像技术测定细胞内钙浓度 ([Ca2 ]i)。结果　CT诱导细胞内cAMP升高并呈剂量依赖性地保护胆红素诱导的小脑颗粒神经元凋亡。当加入腺苷环化酶(AC)刺激剂 (Forskolin)及cAMP类似物 (8 溴 环磷酸腺苷 )后 ,虽导致细胞内cAMP升高 ,但不能阻断或减弱胆红素诱导的神经元凋亡。另发现胆红素浓度依赖性触发细胞内钙升高 ,而CT可降低胆红素导致的细胞内钙的升高。结论　Gs蛋白可能参与了小脑颗粒神经元凋亡的调控 ,激活Gs蛋白拮抗胆红素诱导的神经元凋亡的机制是通过降低细胞内钙而不依赖于cAMP的途径     

新生儿缺氧缺血性脑病T细胞活化受抑的机制探讨

目的：研究轻、中、重度缺氧缺血性脑病（HIE）新生儿T细胞功能增殖和活化功能状态及可能机制。方法：分HIE新生儿组（60例）及健康新生儿组，采用尼龙毛法分离两组外周血单个核细胞中的T细胞；用CTLL－2细胞株活性检测测定T细胞培养上清中IL－2活性；流式细胞仪间接免疫荧光法检测T细胞表面白细胞介素2受体α（IL－2Rα）表达，3H－TdR掺入法测定T淋巴细胞增殖反应。放射免疫分析法测定T细胞内环磷酸腺苷（cAMP)，环磷酸鸟苷（cGMP)。结果：中、重度HIE患儿T细胞功能明显受抑，T细胞转化率及T细胞IL－2及IL－2R的表达下降（P均＜0．05），其活化T细胞内cAMP含量升高，cGMP含量降低，此变化与T细胞IL－2及IL－2R的表达下降（P均＜0．05），其活化T细胞内cAMP含量升高，cGMP含量降低，此变化与T细胞功能受抑密切相关。结论：新生儿HIE是T细胞功能不同程度受抑，可能与胞内环核苷酸代谢系统信号传导途径有关。     

新生儿缺氢缺血性系列疾病的探讨

目的 分析窒息致新生儿缺氧缺血性系列疾病(HID)临床资料和预后，探讨改善预后的措施。方法对72例HID进行资料分析。结果 在72例HID中，新生儿缺氧缺血性脑病(HIE)64例，新生儿缺氧缺血性肾脏损害(HIR)40例，新生儿缺氧缺血性心脏损害(HIM)24例，经治疗，存活60例，死亡7例，放弃治疗5例；胎儿期缺氧缺血较出生后缺氧致脏器损害多而重(χ^2＝8．974，P＜0．01)，且预后差(χ^2＝4．816，P＜0．05)；保持患儿血糖在正常的高值，预后明显优于低值正常血糖的思儿(χ^2＝6．038，P＜0．051，而中值正常水平血糖想儿预后与低值正常水平血糖患儿无显著差异χ^2＝1．302，P＜0．05)。结论 胎儿期缺氧缺血性HID和血糖在正常的低值的HID预后均差，保持血糖在正常的高值预后佳。     

哮喘患儿血浆一氧化氮、环磷酸腺苷/环磷酸鸟苷比值变化的临床意义

目的　研究支气管哮喘患儿血浆一氧化氮 (NO)、环磷酸腺苷 (cAMP) /环磷酸鸟苷 (cGMP)比值变化及其临床意义。方法　采用硝酸还原酶法和放射免疫法测定哮喘患儿 4 0例急性期和缓解期血浆NO3 -/NO2 -、cAMP及cGMP水平与cAMP/cGMP比值变化 ,并设 2 3例健康儿童为对照组。结果　 1.哮喘患儿急性期血浆NO3 -/NO2 -水平显著高于缓解期和对照组 (P均 <0 .0 1)。 2 .哮喘患儿急性期血浆cGMP水平明显高于缓解期 (P <0 .0 5 ) ,显著高于对照组 (P <0 .0 1)。 3.哮喘患儿急性期血浆cAMP明显低于缓解期和对照组 (P均<0 .0 1)。 4 .哮喘患儿急性期cAMP/cGMP比值显著低于缓解期和对照组 (P均 <0 .0 1)。 5 .缓解期血浆NO3 -/NO2 -和cGMP水平下降 ,cAMP水平上升及其cAMP/cGMP比值与对照组相比 ,差异无显著性 (P均 >0 .0 5 )。6 .哮喘患儿急性期血浆NO3 -/NO2 -与cGMP水平呈正相关 (r =0 .4 0 1　P <0 .0 1)。结论　血浆内源性NO、cAMP、cGMP可能参与哮喘的发病机制 ,血浆NO、cAMP/cGMP比值变化可作为监测和指导哮喘患儿疗效和评价哮喘药物疗效的较好生化指标。     

MDM2mRNA 过度表达与儿童急性淋巴细胞白血病免疫分型、FAB 分型及高白细胞的关系

为了研究癌基因MDM2mRNA过度表达与儿童急性淋巴细胞白血病(ALL)的免疫分型、FAB分型及临床高白细胞性的关系,用RT-PCR法检测了32例急性淋巴细胞白血病患儿外周血单个核细胞内MDM2mRNA表达水平,并分析MDM2mRNA过度表达与不同型别的关系。高白细胞性白血病(HAL)MDM2表达明显高于非高白细胞性白血病(NHAL)组(P＝0．0099);T淋巴细胞白血病(T-ALL)与非T淋巴细胞白血病(Non-T-ALL)间(P＝0．7345),L     

热性惊厥儿血清干扰素α,白细胞介素2和肿瘤坏死因子α测定 …

目的探讨热性惊厥(FC)的免疫作用机制。 方法采用双抗体夹心酶联免疫吸附试验(ELISA)对30例FC儿进行血清干扰素α(IFN-α)、白细胞介素2(IL-2)和肿瘤坏死因子α(TNF-α)测定,并与20例正常儿童比较。 结果FC组血清IFN-α、IL-2、TNF-α水平均明显高于对照组(t＝5．29,2．19,5．22P分别＜0．001,＜0．05,＜0．001),相关分析发现FC患儿IFN-α与IL-α之间呈显著正相关(r＝0．527P＜0．005)。 结论FC儿存在细胞免疫功能异常,其参与了FC的发病过程,且与FC复发可能有一定的关系。     

肾病儿童外周血单个核细胞活化蛋白1和糖皮质激素受体的DNA结合活性的变化

目的 探讨肾病综合征（NS）儿童外周血单个核细胞（PBMC）中活化蛋白1（AP－1）和糖皮质激素受体（GR）的DNA结合活性的变化,体外实验加入地塞米松（DEX）以及泼尼权治疗后对二者的影响。方法 用凝胶电泳迁移率变化分析法（EMSA）和同位素放射性自显影法分析了NS儿童PMBC中AP－1、GR的DNA结合活性。结果 （1）NS组基础状态AP－1DNA活性为264．1（0．6－351．8）,对照组为0．8（0－1．1）,两组比较差异有非常显著意义（Z＝2．72,P＜0．01）;佛波酯刺激状态下AP－1DNA活性为514．8（1．1－616．7）,对照组为1．1（0－36．6）,两组比较差异有非常显著意义（Z＝2．64,P＜0．01）;经DEX作用后NS组AP－1的DNA活性与对照组比较,差异无显著意义（Z＝0．64,P＞0．05）。（2）NS组基础状态GR的DNA活性为7．6（0．2－11．7）,对照组为45．2（8．1－95．2）,两组比较差异有显著意义（Z＝2．56,P＜0．05）;TPA刺激状态GR的DNA活性为5．3（0．0－10．7）,对照组为78．1（7．8－97．4）,两组比较差异有显著意义（Z＝2．40,P＜0．05）,经DEX作用后GR的DNA活性与对照组比较,差异无显著意义（Z＝0．96,P＞0．05）。（3）经泼尼松治疗尿蛋白转阴1周后,AP－1 DNA活性为1．05（0．95－1．44）,与治疗前[2．87（1．00－4．20）]比较,差异有显著意义（Z＝2．24,P＜0．05）。治疗后GR的DNA活性为3．91（1．00－4．70）,与治疗前[1．04（0．73－1．24）]比较,差异有显著意义（Z＝2．0,P＜0．05）。结论 （1）NS儿童的PBMC中AP－1 DNA结合活性异常升高,而GRDNA结合活性降低;（2）糖皮质激素可抑制AP－1、增强GR的DNA结合活性。     

内皮素、一氧化氮在毛细支气管炎和哮喘息儿中的变化

目的 探讨血内皮素(ET)和一氧化氮(NO)在毛细支气管炎(毛支)和婴幼儿哮喘(哮喘)中的变化及意义。方法 用放射免疫分析法和分光光度比色法分别检测毛支、哮喘患儿急性期和缓解期血ET和NO，并与正常儿童进行比较。结果 血浆ET在毛支组和哮喘组，急性期均明显高于正常组(P均＜0．01)，缓解期均下降，与正常组比较无显著差别(P均＞0．05)；两组急性期比较无差别(P＞0．05)。与血浆ET一样，血清NO在毛支组和哮喘组，急性期均显著高于正常组(P＜0．01和P＜0．05)；缓解期则下降，与对照组比无显著差别，(P均＞0．05)；两组急性期相比，差异无显著性(P＞0．05)。两组急性期血浆ET与血清NO呈明显正相关(P＜0．05)。结论 在毛支和哮喘中，血ET和NO均升高，二者关系密切，均可能参与毛支及哮喘的发病。     

儿童神经系统疾病血清、脑脊液神经元特异性烯醇酶变化的意义

目的　探讨血清、脑脊液神经元特异性烯醇酶 (NSE)在神经系统疾病中的意义。方法　采用电化学发光法检测 85例神经系统疾病血清 ,脑脊液NSE活性。结果　病毒性脑炎 (病脑 )、癫、高热惊厥 (FC)患儿血清NSE含量均明显高于对照组 (P <0 .0 0 5 ) ,而相互间无显著性差异 (P >0 .0 5 )。病脑、癫患儿脑脊液NSE与对照组比较显著升高 (P <0 .0 0 1) ,而FC组与对照组无明显差别 (P >0 .0 5 ) ,且相互间有显著性差异 (P<0 .0 5 )。结论　病脑、癫均存在脑损伤 ,血清、脑脊液NSE含量均明显升高 ,脑脊液NSE变化更具特异性 ,可作为病脑、癫和FC的鉴别诊断指标     

Ontogeny of beta-adrenergic regulation of adenylate cyclase in intrapulmonary arteries from fetal and postnatal lambs.

The enzyme adenylate cyclase is critically involved in the regulation of vasodilatation in the developing pulmonary circulation in that it mediates the vascular smooth muscle effects of beta-adrenergic agonists and vasodilatory prostaglandins. These agonists activate receptors coupled to the catalytic subunit of the enzyme by stimulatory guanine nucleotide-dependent regulatory proteins. We examined the ontogeny of the function of the adenylate cyclase system in intrapulmonary arterial segments from fetal lambs at 110-115 (F1) and 125-135 d gestation (F2) and from postnatal lambs at 7-14 (P1) and 35-56 d of age (P2). The function of the intact enzyme system and its components was assessed in incubations measuring cAMP accumulation during phosphodiesterase inhibition. beta-Adrenergic mediation of adenylate cyclase was examined because the binding characteristics of the smooth muscle receptors can be readily quantified. Nonstimulated (basal) cAMP accumulation was similar in F1 and F2, it increased 8-fold from F2 to P1, and it was equivalent in P1 and P2. cAMP accumulation with isoproterenol increased 5.9-fold from F1 to F2 and was similar in F2, P1, and P2. Radioligand binding studies revealed that the greater response to isoproterenol in F2 versus F1 is not related to an increase in beta-adrenergic receptor density or affinity. cAMP accumulation with forskolin, which activates adenylate cyclase by actions that involve both the stimulatory guanine nucleotide-dependent regulatory protein and the catalytic subunit, was similar in the four age groups, indicating that there are no maturational changes in the funciton of these enzyme system components.(ABSTRACT TRUNCATED AT 250 WORDS)     

Lessons from fragile X regarding neurobiology, autism, and neurodegeneration

The fragile X mental retardation 1 gene (FMR1) mutation causes two disorders: fragile X syndrome (FXS) in those with the full mutation and the fragile X-associated tremor/ataxia syndrome (FXTAS) in some older individuals with the premutation. FXS is caused by a deficiency of the FMR1 protein (FMRP) leading to dysregulation of many genes that create a phenotype with ADHD, anxiety, and autism. FXTAS is caused by the elevation of FMR1-mRNA to levels 2 to 8 times normal in the premutation. This causes an RNA gain of function toxicity leading to brain atrophy, white matter disease, neuronal and astrocytic inclusion formation, and subsequent ataxia, intention tremor, peripheral neuropathy, and cognitive decline. The neurobiology and pathophysiology of FXS and FXTAS are described in detail.     

Two boys with fragile x syndrome and hepatic tumors

Hepatic tumors are rare childhood neoplasms with uncertain etiology. We report the cooccurrence of hepatic tumors in 2 boys with fragile X syndrome, one with hepatoblastoma and another with desmoplastic nested spindle cell tumor of liver. The pathogenesis of fragile X syndrome involves silencing of the fragile X mental retardation 1 gene and consequent loss of FMR1 protein. We speculate regarding molecular pathways that might explain the cooccurrence of the 2 conditions. Further examination of a possible functional link between hepatic neoplasia and loss of FMRP is warranted.     

Spermine and spermidine, modulators of the cell surface enzyme adenylate cyclase

Adenylate cyclase activity was measured in membrane preparations of cultured fibroblasts from controls and patients with cystic fibrosis. Enzyme activity increased as the transition from exponential growth to confluence occurred; sodium fluoride-stimulated activity more markedly displayed this relationship than basal cyclase activity. The in vitro addition of spermine (1 X 10(-6) to 2 X 10(-3) M) to membrane preparations caused inhibition of basal and sodium fluoride-stimulated enzyme activity, with 50% inhibition of basal activity occurring at 10(-6) M spermine. Spermidine (10(-4) M) caused 15--25% inhibition of adenylate cyclase activity. The increase in fibroblast adenylate cyclase activity during the transition from exponential growth was comparable in cells obtained from cystic fibrosis patients and control subjects. Basal and sodium-fluoride stimulated adenylate cyclase activity as well as inhibition of this enzyme activity by spermidine and spermine were undistinguishable between the different cell genotypes. A potential modulation of cellular proliferative activity through polyamine interaction with the adenylate cyclase system is postulated.     

Ontogeny of adenylate cyclase and phosphodiesterase activities in swine tissues.

The ontogenic patterns of the basal- and fluoride-stimulated adenylate cyclase and the cAMP phosphodiesterase activites as well as the tissue levels of cAMP were assessed in swine adipose tissue, heart, skeletal muscle, and liver. The developmental patterns were complex. However, there was a general tendency in all tissues toward lower enzyme activities in the oldest animals (150 days) for both enzymes. In adipose tissue, the decreased enzyme levels could partially be attributed to an increase in adipocyte size.     

Cyclic guanosine monophosphate metabolism in human amnion cells trisomic for chromosome 21

An extra copy of chromosome 21, a small chromosome or a specific segment of it, is the cause of the disorder known as Down's syndrome (DS). Genes mapped to this chromosome include superoxide dismutase-1 (SOD-1) along with other enzymes. Gene dosage effects have been shown for some of these enzymes, including SOD-1. Increased SOD-1 has been suggested to stimulate the cGMP-forming enzyme, guanylate cyclase (GC). In the present study we have used amnion cells from DS subjects and normal subjects in order to indirectly test the effects of SOD-1 on the cGMP metabolism. We have measured the cAMP and cGMP content, SOD-1 activity, GC activity and cGMP phosphodiesterase (G-PDE) activity in amnion cells from DS subjects and normal subjects, respectively. The levels of cGMP in DS amnion cells were lower than in normal cells, although the SOD-1 activity was higher in DS amnion cells. Furthermore, the GC activity and the G-PDE activity were found to be lower in the trisomic cells. Our results do not support the suggestion that SOD-1 has a stimulatory effect on the GC activity.     

Molecular pathogenesis of fragile X syndrome

The fragile X syndrome is an X-linked genetic disorder; manifesting primarily as intellectual disability. The disease is caused by the absence of functional FMRP, a protein encoded by the FMR1 gene. The expansion of trinucleotide repeats within the first exon of the gene contributes to most cases of the syndrome. This review summarizes the present knowledge of the relationship between the molecular defect in the FMR1 gene and the clinical phenotype associated with disease.     

智力低下和孤独症男童脆性X基因突变的研究

目的应用改良基因检测方法,探讨脆性X综合征致病基因(fragileXmentalretardation"1,FMR1)在中国人群智力低下和孤独症中的作用。方法收集2002～2006年小儿神经、遗传代谢门诊诊断的男性孤独症患儿44例、非家族性智力低下男性患儿40例,建立适用于男性的FMR1基因突变检查方法,对检查阳性者以pfxa3探针进行Southern杂交。结果在44例孤独症患儿中,发现1例pfxa3杂交片段约0.2kb,为FMR1前突变;40例智力低下患者中FMR1基因未见异常。结论在孤独症人群中发现的1例FMR1基因前突变,其致病意义有待进一步阐明。     

DNA testing for fragile X syndrome in schools for learning difficulties

Fragile X syndrome is the most common inherited cause of mental retardation. Early diagnosis is important not only for appropriate management of individuals but also to identify carriers who are unaware of their high risk of having an affected child. The disorder is associated with a cytogenetically visible fragile site (FRAXA) at Xq27.3, caused by amplification of a (CGG)n repeat sequence within the gene at this locus designated FMR1. Clinical and molecular studies have been undertaken to screen for fragile X syndrome in 154 children with moderate and severe learning difficulties of previously unknown origin. Southern blot analysis of peripheral blood showed the characteristic abnormally large (CGG)n repeat sequence associated with fragile X syndrome in four of the 154 children. The findings were confirmed by cytogenetic observation of the fragile site and by further molecular studies. The families of the affected children were offered genetic counselling and DNA tests to determine their carrier status. These findings show that there are still unrecognised cases of fragile X syndrome. Given the difficulty of making a clinical diagnosis and the implications for families when the diagnosis is missed, screening in high risk populations may be justified. The issues involved in screening all children in special schools for fragile X syndrome are discussed.     

DNA testing for fragile X syndrome in schools for learning difficulties.

Fragile X syndrome is the most common inherited cause of mental retardation. Early diagnosis is important not only for appropriate management of individuals but also to identify carriers who are unaware of their high risk of having an affected child. The disorder is associated with a cytogenetically visible fragile site (FRAXA) at Xq27.3, caused by amplification of a (CGG)n repeat sequence within the gene at this locus designated FMR1. Clinical and molecular studies have been undertaken to screen for fragile X syndrome in 154 children with moderate and severe learning difficulties of previously unknown origin. Southern blot analysis of peripheral blood showed the characteristic abnormally large (CGG)n repeat sequence associated with fragile X syndrome in four of the 154 children. The findings were confirmed by cytogenetic observation of the fragile site and by further molecular studies. The families of the affected children were offered genetic counselling and DNA tests to determine their carrier status. These findings show that there are still unrecognised cases of fragile X syndrome. Given the difficulty of making a clinical diagnosis and the implications for families when the diagnosis is missed, screening in high risk populations may be justified. The issues involved in screening all children in special schools for fragile X syndrome are discussed.