Evidence that the LRRK2 ROC domain Parkinson's disease‐associated mutants A1442P and R1441C exhibit increased intracellular degradation

Mutations in the leucine‐rich repeat kinase 2 (lrrk2) gene are the leading genetic cause of Parkinson's disease (PD). In characterizing the novel ROC domain mutant A1442P, we compared its steady‐state protein levels, propensity to aggregate, and toxicity with the pathogenic R1441C mutant and wild‐type (WT) LRRK2. Mutant (R1441C and A1442P) and WT LRRK2 fused to green fluorescent protein (GFP) and FLAG were transiently expressed in HEK293 cells using plasmid constructs. Western analysis and fluorescence microscopy consistently demonstrated lower mutant LRRK2 protein levels compared with WT. A time‐course expression study using flow cytometry showed that WT LRRK2 expression increased initially but then plateaued by 72 hr. Conversely, R1441C and A1442P mutant expression attained 85% and 74% of WT levels at 24 hr but fell to 68% and 55% of WT levels by 72 hr, respectively. We found that proteasome inhibition markedly increased mutant LRRK2 to levels approaching those of WT. Taken together, our findings reveal increased intracellular degradation for both mutants. Furthermore, the impact of mutant and WT LRRK2 expression on HEK293 cell viability was assessed under normative and oxidative (hydrogen peroxide) conditions and found not to differ. Expression of WT and mutant LRRK2 protein gave rise to intracellular aggregates of similar appearance and cellular localization. In summary, we provide evidence that the novel A1442P mutant and the previously investigated R1441C pathogenic mutant exhibit increased intracellular degradation, a property reportedly demonstrated for the pathogenic LRRK2 kinase domain mutant I2020T. © 2013 Wiley Periodicals, Inc.     

一例3-甲基戊烯二酸尿症Ⅶ型患儿的临床和遗传学分析

目的对一例全面发育落后的婴儿进行临床和遗传学分析,明确其病因。方法采集患儿及其家系成员的病史,应用实验室检查、遗传代谢病筛查和新一代测序技术对该家系进行临床和遗传学分析。结果先证者临床表现为对声音反应不灵敏,竖头不稳,不能翻身、逗笑,不认识母亲。实验室检查血乳酸、血糖等正常,尿有机酸中3-甲基戊烯二酸、3-甲基戊二酸水平增高,提示为"3-甲基戊烯二酸尿症可能"。头颅磁共振扫描显示胼胝体压部T1W信号偏低,髓鞘化落后于月龄。高通量测序发现CLPB基因存在复合杂合变异c.1085G>A和c.1700A>C,分别遗传自父亲和母亲,二者均为新变异。根据美国医学遗传学与基因组学学会标准,两个变异均预测为疑似致病变异。结论该患儿可能为CLPB基因变异引起的3-甲基戊烯二酸尿症Ⅶ型。高通量测序技术为分析该类疾病提供了有力的诊断工具。     

Variability in retinoblastoma genome stability is driven by age and not heritability

Retinoblastoma (RB) is a childhood intraocular cancer initiated by biallelic inactivation of the RB tumor suppressor gene (RB1^?/?). RB can be hereditary (germline RB1 pathogenic allele is present) or non‐hereditary. Somatic copy number alterations (SCNAs) contribute to subsequent tumorigenesis. Previous studies of only enucleated RB eyes have reported associations between heritability status and the prevalence of SCNAs. Herein, we use an aqueous humor (AH) liquid biopsy to investigate RB genomic profiles in the context of germline RB1 status, age, and International Intraocular Retinoblastoma Classification (IIRC) clinical grouping for both enucleated and salvaged eyes. Between 2014 and 2019, AH was sampled from a total of 54 eyes of 50 patients. Germline RB1 status was determined from clinical blood testing, and cell‐free DNA from AH was analyzed for SCNAs. Of the 50 patients, 23 (46.0%; 27 eyes) had hereditary RB, and 27 (54.0%, 27 eyes) had non‐hereditary RB. Median age at diagnosis was comparable between hereditary (13 ± 10 months) and non‐hereditary (13 ± 8 months) eyes (P = 0.818). There was no significant difference in the prevalence or number of SCNAs based on (1) hereditary status (P > 0.56) or (2) IIRC grouping (P > 0.47). There was, however, a significant correlation between patient age at diagnosis, and (1) number of total SCNAs (r[52] = 0.672, P < 0.00001) and (2) number of highly‐recurrent RB SCNAs (r[52] = 0.616, P < 0.00001). This evidence does not support the theory that specific molecular or genomic subtypes exist between hereditary and non‐hereditary RB; rather, the prevalence of genomic alterations in RB eyes is strongly related to patient age at diagnosis.     

Computational study on the mechanism and kinetics for the reaction between HO2 and n-propyl peroxy radical

The n-propyl peroxy radical (n-C₃H₇O₂) is the key intermediate during atmospheric oxidation of propane (C₃H₈) which plays an important role in the carbon and nitrogen cycles in the troposphere. In this paper, a comprehensive theoretical study on the reaction mechanism and kinetics of the reaction between HO₂ and n-C₃H₇O₂ was performed at the CCSD(T)/aug-cc-pVDZ//B3LYP/6-311G(d,p) level of theory. Computational results show that the HO₂ + n-C₃H₇O₂ reaction proceeds on both singlet and triplet potential energy surfaces (PESs). From an energetic point of view, the formation of C₃H₇O₂H and ³O₂via triplet hydrogen abstraction is the most favorable channel while other product channels are negligible. In addition, the calculated rate constants for the title reaction over the temperature range of 238–398 K were calculated by the multiconformer transition state theory (MC-TST), and the calculated rate constants show a negative temperature dependence. The contributions of the other four reaction channels to the total rate constant are negligible.

The negative temperature dependence for the HO₂ + n-C₃H₇O₂ reaction in lower temperature regime.     

归因训练对老年性抑郁症康复的疗效观察

目的探讨归因训练对老年性抑郁症的增效作用。方法将老年抑郁症患者58例,随机分为归因训练合并西酞普兰组（研究组）和单纯西酞普兰组（对照组）,进行对照研究,采用汉密顿抑郁量表（HAMD）和临床疗效总评量表（CGISI）于治疗前及治疗后6周末、半年末,进行评分比较。结果治疗6周末研究组同对照组评分比较有显著性差异（P〈0．01）。治疗半年末,研究组和治疗6周末同组比较有显著性差异（P〈0．01）,而对照组无显著性差异（P〉0．05）。两组半年未复发率比较有显著性差异（X^2=6．44,P〈0．05）。结论归因训练能提高老年性抑郁症的近期和远期疗效,降低复发率。     

Association of KIR Genotypes and Haplotypes with Susceptibility to Chronic Hepatitis B Virus Infection in Chinese Han Population

Killer immunoglobulin-like receptor （KIR） genes can regulate the activation of NK and T cells upon interaction with HLA class I molecules. Hepatitis B virus （HBV） infection has been regarded as a multi-factorial disorder disease. Previous studies revealed that KIRs were involved in HCV and HIV infection or clearance. The aim of this study was to explore the possibility of the inheritance of KIR genotypes and haplotypes as a candidate for susceptibility to persistent HBV infection or HBV clearance. The sequence specific primer polymerase chain reaction （SSP-PCR） was employed to identify the KIR genes and pseudogenes in 150 chronic hepatitis B （CHB） patients, 251 spontaneously recovered （SR） controls, and 412 healthy controls. The frequencies of genotype G7 M, FZ1 increased in CHB patients compared with healthy control subjects. The frequency of genotype AH was higher in SR controls than that in both CHB patients and healthy controls. The carriage frequencies of genotype G and AH were higher; while, the frequencies of AF and AJ were lower in SR controls than those in healthy control subjects. The frequency of A haplotype was lower, whereas, the frequency of B haplotype was higher in CHB patients and SR controls than those in healthy controls. In healthy controls, haplotype 4 was found lower compared with that in CHB patients and SR controls and the frequency of haplotype 5 was higher in SR controls than that in other two groups. Based on these findings, it seems that the genotypes M and FZ1 are HBV susceptive genotypes; AH, on the other hand, may be protective genotypes that facilitate the clearance of HBV. It appears that the haplotype 4 is HBV susceptive haplotype, whereas, haplotype 5 may be the protective haplotype that facilitates the clearance of HBV. Cellular ＆ Molecular Immunology. 2008;5（6）：457-463.     

颈动脉窦粥样硬化与冠状动脉粥样硬化程度的相关性

背景:颈动脉内中膜厚度可用作冠状动脉粥样硬化的替代指标,但它与冠状动脉事件的关系尚未得到充分研究。目的:探讨颈动脉内中膜厚度在预测冠状动脉粥样硬化性心脏病及其程度中的价值。设计:病例分析。单位:解放军总医院心内科。对象:实验于2000-01/2002-01在解放军总医院完成。将285例可疑冠状动脉粥样硬化性心脏病患者按造影结果分为0支组,1支组,2支组,3支组4组,年龄33～74岁,平均(54.48±9.44)岁,其中男164例,女121例。方法:①测量体质量、身高,计算体质量指数(BMI)=体质量(kg)/身高(m)2。②测量卧位左臂血压。③采静脉血检测血脂。④用高分辨率B型超声,10MHz线性探头检测颈总动脉,颈内动脉,颈外动脉和颈动脉窦。将颈动脉内中膜厚度≥1.0mm定义为颈动脉粥样硬化。用标准的Judkins技术作冠状动脉造影。根据管腔狭窄≥50%的病变血管支数将冠状动脉粥样硬化性心脏病的程度记录为0,1,2,3。左主干狭窄≥50%定义为2支病变,左主干合并右冠病变定义为3支病变。主要观察指标:①不同冠状动脉病变程度患者不同测量位点的内中膜厚度。②冠状动脉粥样硬化性心脏病严重程度的预测因子。③颈动脉不同位点内中膜厚度值对冠状动脉粥样硬化性心脏病的预测价值。④不同危险因子与内中膜厚度的关系。结果:285例患者全部完成实验进入结果分析。①不同冠状动脉病变程度患者不同测量位点的内中膜厚度经卡方检验,左、右颈动脉窦的F值最大,分别为65.64和63.24。左、右颈动脉窦的内中膜厚度值随冠状动脉病变程度的加重而增厚。②对冠状动脉粥样硬化性心脏病严重程度的预测因子进行回归分析表明,男性、高密度脂蛋白胆固醇、低密度脂蛋白胆固醇、左颈动脉窦、右颈动脉窦、右颈内动脉、右颈外动脉的内中膜厚度为冠状动脉粥样硬化性心脏病严重程度的独立预测因子。左颈动脉窦和右颈动脉窦是冠状动脉粥样硬化性心脏病的最重要的预测因子(t=3.61,P=0.391;t=2.58,P=0.247)。③颈动脉不同位点内中膜厚度值对冠状动脉粥样硬化性心脏病的预测价值中左右颈动脉窦的阳性预测值最高,分别为是89.9%,88.8%。④对不同危险因子与内中膜厚度的关系行多元回归分析,其结果是:年龄,体质量指数,胆固醇,三酰甘油,高密度脂蛋白胆固醇和低密度脂蛋白胆固醇是左颈动脉窦处动脉粥样硬化的危险因子。结论:颈动脉窦内中膜厚度与冠状动脉粥样硬化性心脏病程度明显相关。二者有共同的病理与血液动力学基础。颈动脉窦内中膜厚度可以作为筛选冠状动脉粥样硬化性心脏病的指标。