CITED2基因突变与先天性心脏病关系研究

目的 研究CITED2基因突变与先天性心脏病（CHD）的关系。方法 2013年1月至2015年1月山东大学齐鲁儿童医院收集368例散发型CHD患儿和200名健康儿童血液样本进行DNA抽提, PCR扩增CITED2外显子区, Sanger测序后, 进行GeneBank对比和氨基酸序列分析。结果 发现4例杂合突变。病例1室间隔缺损患儿为c.399C＞T同义突变（p.His133His）; 病例2室间隔缺损患儿为SGJ区c.574A＞G的错义突变（p.Ser192Gly）; 病例3房间隔缺损患儿为已知 SNP位点（rs191856368）; 病例4动脉导管未闭患儿为新发现的错义突变（p.Ser96Phe）。这些突变在对照组中均未检测到。结论 CITED2基因Ser192Gly和p.Ser96Phe突变可能与CHD发生有关。     

住院患儿细菌培养标本分离肺炎链球菌特征分析

目的 分析住院患儿细菌培养标本中所检出肺炎链球菌的分布、血清型及耐药性特征。方法　收集2013年3月至2014年2月在首都医科大学附属北京儿童医院呼吸、感染及重症监护病房住院的195例患儿送检的细菌培养标本,对其进行肺炎链球菌常规检测、血清分型和药物敏感性试验。按照2013年临床实验室标准化委员会（CLSI）推荐的抗菌药物敏感性试验标准进行结果判断。应用WHONET 5.6软件对药敏结果进行耐药性分析。结果　195例患儿分离到肺炎链球菌,其中婴幼儿占66.1%。195例中共检出20种血清型,其中19F最多,占32.3%,其次19A,占20.5%。肺炎链球菌对红霉素和克林霉素最不敏感（敏感率＜5%）,对青霉素不敏感率为46.7%。与对青霉素敏感的肺炎链球菌进行抗菌药物耐药性比较后显示,青霉素不敏感肺炎链球菌对头孢噻肟、头孢吡肟、美罗培南和复方新诺明不敏感率均高于敏感菌株,差异存在统计学意义（P＜0.05）。结论　在儿科,肺炎链球菌的检出、流行型别的确定以及耐药性检测均不容忽视,应进行疫苗预防接种和合理选择抗菌药物进行抗感染治疗。     

单纯性肥胖外周血单核/巨噬细胞趋化因子与胰岛素抵抗的相关性研究

了解单纯性肥胖儿童外周血单核/巨噬细胞趋化因子与胰岛素抵抗相关参数的关系。方法　2006年5月至2008年5月广西医科大学第一附属医院儿科70 例确诊为单纯性肥胖儿童作为研究对象,30名健康体检儿童作为健康对照组。流式细胞仪测定外周血单核细胞CD68阳性率;酶联免疫吸附（ELISA）法测定血浆巨 噬细胞炎性蛋白-1α（MIP-1α）、单核细胞趋化蛋白-1（MCP-1）水平;放射免疫分析（RIA）法测定血浆脂联素（ADPN）、血浆胰岛素（Ins）浓度,计算体质 指数（BMI）,按HOMA模型计算胰岛素抵抗指数（InRI）,并与相关变量进行Pearson相关分析或偏相关分析。结果　单纯性肥胖组血浆MIP-1α、MCP-1水平高于 健康对照组,两组比较差异有统计学意义（P < 0.05）。两组外周血CD68阳性率比较差异无统计学意义（P > 0.05）。两组InRI比较差异有统计学意义 （P < 0.05）。单纯性肥胖组血浆MIP-1α、MCP-1水平分别与BMI、腰围（WC）和InRI呈正相关 （P < 0.05）。血ADPN分别与BMI、WC和InRI呈负相关（P < 0.05）。外 周血单个核细胞CD68阳性率与BMI、WC和InRI无相关性（ P > 0.05）。结论　单纯性肥胖患儿体内存在低程度炎症反应,肥胖儿童外周血单核/巨噬细胞趋化因子 与胰岛素抵抗及中心性肥胖相关,可以作为肥胖相关疾病的早期预测指标之一。     

An innovative flow cytometric approach for small-size platelet microparticles: influence of calcium

Microparticles (MPs) are small submicron membrane-derived vesicles shed from a variety of cells and they have been implicated in different disorders. Accordingly, understanding of physiological characteristics of MPs and improvement of methods of their quantification are important for further advance in the field. Although flow cytometry is the most widely applied technique for MP analysis, it is limited by lack of adequate standardisation. Annexin V (AnV), which binds surface phosphatidylserine (PS) with high affinity, has been long regarded as a marker of MPs, but AnV binding is Ca2+-dependent and it is unclear how [Ca2+] concentrations could affect AnV binding to MPs and its enumeration. MPs from citrated and heparinised plasma were labelled with AnV, anti-CD42b and quantified using an Apogee A50 flow cytometer. The small-size MP gate was defined with the use of size beads (from 0.1 to 0.5 μm) and confirmed with an in vitro assessment of platelet stimulation. Biotinylated anti-CD42b antibodies were then bound to streptavidin conjugated with different fluorochromes, leading to an amplified signal of platelet MPs (PMPs). Moderate increase of [Ca2+] concentrations in the annexin V staining buffer allows initial plasma recalcification and more accurate MP quantification in citrated plasma. Thrombin stimulation of platelet-free plasma containing only MPs did not produce any changes in the concentration of AnV+ MPs, but decreased the anti-CD42b binding. The results also indicate that prolonged storage and thrombin induce the release of AnV+ MPs whereas PS exposure in pre-existent MPs is not affected by thrombin. In conclusion, we present a sensitive protocol for the analysis of circulating and in vitro induced small-size PMPs that might contribute to future cardiovascular and clinical research.     

Four novel and two recurrent NHLRC1 (EPM2B) and EPM2A gene mutations leading to Lafora disease in six Turkish families

Lafora disease (LD) is a type of autosomal recessive, progressive myoclonus epilepsy resulting mostly from mutations in the EPM2A and NHLRC1 genes. Mutational analysis in both genes was initiated with the aim of establishing LD DNA diagnosis in Turkey. Four novel NHLRC1 (p.G131X, p.P69S and p.D82H) and EPM2A (p.V7A) and two recurrent NHLRC1 (p.D146N) and EPM2A (p.R241X) mutations were identified in six families. The delineation of causative mutations in patients provided early disease diagnosis for other family members and contributed to the knowledge of LD pathogenesis.     

One-year psychosocial functioning in patients in the early vs. late stage of bipolar disorder

Rosa AR, González‐Ortega I, González‐Pinto A, Echeburúa E, Comes M, Martínez‐Àran A, Ugarte A, Fernández M, Vieta E. One‐year psychosocial functioning in patients in the early vs. late stage of bipolar disorder. Objective: The aim of this 1‐year follow‐up study was to compare functional outcome as well as clinical differences between patients with first‐ and multiple‐episode bipolar disorder. Method: Bipolar disorder patients with first (n = 60) and multiple episodes (n = 59) were recruited from two hospitals in Spain. The Functioning Assessment Short Test (FAST) was used to assess functioning. The Hamilton Depression Rating Scale (HAMD) and the Young Mania Rating Scale (YMRS) were administered to assess mood symptoms. Results: As expected, patients with first episode experienced a greater functioning compared to patients with multiple episodes (11.26 ± 10.94 vs. 26.91 ± 13.96; t = 6.436, P < 0.001). There were significant demographic and clinical differences between both groups. Baseline depressive symptoms (F = 9.553, df = 4, 102; P < 0.001) and age (F = 14.145, df = 4, 103; P < 0.001) were significantly associated with poor functional recovery at 6‐month and 12‐month assessment, respectively, in a group of patients with multiple episodes. Conclusion: Our data give support to the model of staging in bipolar disorder, showing that the enduring neurotoxicity of repeated episodes may contribute to sustained impairment in multiple areas of psychosocial functioning.     

Bioenergetic dysfunction in Huntington's disease human cybrids

In this work we studied the mitochondrial-associated metabolic pathways in Huntington's disease (HD) versus control (CTR) cybrids, a cell model in which the contribution of mitochondrial defects from patients is isolated. HD cybrids exhibited an interesting increase in ATP levels, when compared to CTR cybrids. Concomitantly, we observed increased glycolytic rate in HD cybrids, as revealed by increased lactate/pyruvate ratio, which was reverted after inhibition of glycolysis. A decrease in glucose-6-phosphate dehydrogenase activity in HD cybrids further indicated decreased rate of the pentose-phosphate pathway. ATP levels of HD cybrids were significantly decreased under glycolysis inhibition, which was accompanied by a decrease in phosphocreatine. Nevertheless, pyruvate supplementation could not recover HD cybrids' ATP or phosphocreatine levels, suggesting a dysfunction in mitochondrial use of that substrate. Oligomycin also caused a decrease in ATP levels, suggesting a partial support of ATP generation by the mitochondria. Nevertheless, mitochondrial NADH/NAD_t levels were decreased in HD cybrids, which was correlated with a decrease in pyruvate dehydrogenase activity and protein expression, suggesting decreased tricarboxylic acid cycle (TCA) input from glycolysis. Interestingly, the activity of alpha-ketoglutarate dehydrogenase, a critical enzyme complex that links the TCA to amino acid synthesis and degradation, was increased in HD cybrids. In accordance, mitochondrial levels of glutamate were increased and alanine was decreased, whereas aspartate and glutamine levels were unchanged in HD cybrids. Conversely, malate dehydrogenase activity from total cell extracts was unchanged in HD cybrids. Our results suggest that inherent dysfunction of mitochondria from HD patients affects cellular bioenergetics in an otherwise functional nuclear background.     

Daytime sleepiness and polysomnography in obstructive sleep apnea patients

BackgroundExcessive daytime sleepiness (EDS) is the major complaint in subjects with obstructive sleep apnea syndrome (OSAS). However, EDS is not universally present in all patients with OSAS. The mechanisms explaining why some patients with OSAS complain of EDS whereas others do not are unknown.ObjectiveTo investigate polysomnographic determinants of excessive daytime sleepiness (EDS) in a large multicenter cohort of patients with obstructive sleep apnea (OSAS).MethodsAll consecutive patients with an apnea–hypopnea index greater than 5 h⁻¹ who were evaluated between 2003 and 2005. EDS was assessed using the Epworth Sleepiness Scale (ESS), and patients were considered to have EDS if the ESS was >10.ResultsA total of 1649 patients with EDS ((mean [±SD] Epworth 15 ± 3) and 1233 without EDS (Epworth 7 ± 3) were studied. Patients with EDS were slightly younger than patients without EDS (51 ± 12 vs 54 ± 13 years, p < 0.0001), had longer total sleep time (p < 0.007), shorter sleep latency (p < 0001), greater sleep efficiency (p < 0.0001) and less NREM sleep in stages 1 and 2 (p < 0.007) than those without EDS. Furthermore, patients with EDS had slightly higher AHI (p < 0.005) and arousal index (p < 0.001) and lower nadir oxygen saturation (p < 0.01).ConclusionsPatients with OSAS and EDS are characterized by longer sleep duration and increased slow wave sleep compared to those without EDS. Although patients with EDS showed a mild worsening of respiratory disturbance and sleep fragmentation, these results suggest that sleep apnea and sleep disruption are not the primary determinants of EDS in all of these patients.     

Spinal cord transection modifies ileal fluid and electrolyte transport in rats

Spinal cord injury (SCI) is associated with severe autonomic changes, including inhibition of gastrointestinal (GI) motility. GI motility changes are known to affect electrolytes transport and these changes have not been adequately studied after SCI. We studied the ileal permeability to fluid and electrolytes in rats submitted to experimental spinal cord transection (SCT), between T4 and T5, throughout the first week after SCT. SCT increased ileal secretion of Na+ (P<0.05) and decreased the Cl(-) absorption during the first week post SCI (P<0.05). Water transport was also significantly altered, leading to increased water secretion following the Na+ gradient. Ileal secretion of K+ was significantly increased 1 and 7 days after spinal cord injury. To our knowledge, the present findings are the first direct evidence that SCT alters ileal electrolyte transport in rats. Further studies are necessary to evaluate the mechanisms involved in this phenomenon.     

Characterization of central and peripheral components of the hypothalamus-pituitary-adrenal axis in the inbred Roman rat strains

Several studies performed in outbred Roman high- and low-avoidance lines (RHA and RLA, respectively) have demonstrated that the more anxious line (RLA) is characterized by a higher hypothalamic-pituitary-adrenal (HPA) response to certain stressors than the less anxious one (RHA). However, inconsistent results have also been reported. Taking advantage of the generation of an inbred colony of RLA and RHA rats (RHA-I and RLA-I, respectively), we have characterized in the two strains not only resting and stress levels of peripheral HPA hormones but also central components of the HPA axis, including CRF gene expression in extra-hypothalamic areas. Whereas resting levels of ACTH and corticosterone did not differ between the strains, a greater response to a novel environment was found in RLA-I as compared to RHA-I rats. RLA-I rats showed enhanced CRF gene expression in the paraventricular nucleus (PVN) of the hypothalamus, with normal arginin-vasopressin gene expression in both parvocellular and magnocellular regions of the PVN. This enhanced CRF gene expression is not apparently related to altered negative corticosteroid feedback as similar levels of expression of brain glucorticoid and mineralocorticoid receptors were found in the two rat strains. CRF gene expression tended to be higher in the central amygdala and it was significantly higher in the dorsal region of the bed nucleus of stria terminalis (BNST) of RLA-I rats, while no differences appeared in the ventral region of BNST. Considering the involvement of CRF and the BNST in anxiety and stress-related behavioral alterations, the present data suggest that the CRF system may be a critical neurobiological substrate underlying differences between the two rat strains.