载脂蛋白基因与空间记忆力及胆固醇代谢的相关性分析

目的 研究载脂蛋白E(apoE)基因在早老痴呆疾病中的作用.方法 选取C57BL/6J鼠,通过显微注射法建立人突变apoE4近交系转基因鼠,PCR初筛,再将首建鼠与正常C57BL/6J鼠繁育,将获得的6只转基因鼠作为转基因组,经Southern杂交鉴定,同时出生的结果为阴性的6只小鼠作为对照组,用酶法测定2组小鼠2、9...     

Crystal structures of substrate-bound and substrate-free cytochrome P450 46A1, the principal cholesterol hydroxylase in the brain

By converting cholesterol to 24S-hydroxycholesterol, cytochrome P450 46A1 (CYP46A1) initiates the major pathway for cholesterol removal from the brain. Two crystal structures of CYP46A1 were determined. First is the 1.9-A structure of CYP46A1 complexed with a high-affinity substrate cholesterol 3-sulfate (CH-3S). The second structure is that of the substrate-free CYP46A1 at 2.4-A resolution. CH-3S is bound in the productive orientation and occupies the entire length of the banana-shaped hydrophobic active-site cavity. A unique helix B'-C loop insertion (residues 116-120) contributes to positioning cholesterol for oxygenation catalyzed by CYP46A1. A comparison with the substrate-free structure reveals substantial substrate-induced conformational changes in CYP46A1 and suggests that structurally distinct compounds could bind in the enzyme active site. In vitro assays were performed to characterize the effect of different therapeutic agents on cholesterol hydroxylase activity of purified full-length recombinant CYP46A1, and several strong inhibitors and modest coactivators of CYP46A1 were identified. Structural and biochemical data provide evidence that CYP46A1 activity could be altered by exposure to some therapeutic drugs and potentially other xenobiotics.     

去痴灵对AD模型小鼠学习记忆及脑内突触体素表达的影响

目的 观察去痴灵对阿尔茨海默病(AD)小鼠学习记忆以及脑内突触体素(synaptophysin,SYN)表达的影响.方法 选用昆明种小鼠,右侧脑室注射β-淀粉样蛋白25-35(Aβ25-35)制备AD动物模型.造模1 w后,治疗组分别灌胃低、中、高剂量(3.05、6.10、12.20 g·kg~(-1)·d~(-1))去痴灵,以石杉碱甲为阳性对照药,连续28 d.给药结束后,对各组小鼠进行Morris水迷宫测试、脑组织SYN含量测定以及海马CA1区病理结构观察.结果 与模型组比较,去痴灵各剂量组均明显缩短水迷宫逃避潜伏期(P<0.05),中、高剂量组跨越平台次数显著增加(P<0.05);各治疗组AchE阳性纤维密度明显增加(P<0.05),脑内SYN含量明显升高(P<0.05),突触结构改善明显;各治疗组上述指标组间比较差异不显著(P>0.05).结论 去痴灵可明显改善Aβ所致的小鼠记忆障碍及突触丧失,其脑内突触再生,突触体素含量增加可能是去痴灵改善AD记忆的重要途径之一.     

ACAT inhibitor pactimibe sulfate (CS-505) reduces and stabilizes atherosclerotic lesions by cholesterol-lowering and direct effects in apolipoprotein E-deficient mice

The objective of the present study was to determine whether a novel acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, pactimibe sulfate (CS-505), could reduce atherosclerotic lesions beyond and independent of the reduction achieved by cholesterol lowering alone from two different types of lesions. (1) Early lesion model. Twelve-week-old apolipoprotein E (apoE)^−/− mice were treated with 0.03 or 0.1% (w/w) CS-505, 0.1 or 0.3% avasimibe (CI-1011), or 3% cholestyramine for 12 weeks. Each treatment significantly reduced plasma cholesterol by a similar degree (43–48%). The antiatherosclerotic activity of 0.1% CS-505, however, was more efficacious than the effects of the other treatments (90% versus 40–50%). (2) Advanced lesion model. Twenty-four-week-old apoE^−/− mice were treated with 0.03 or 0.1% CS-505 or 0.1% CI-1011 for 12 weeks. CS-505 at 0.1% revealed enhanced lesion reduction compared with 0.1% CI-1011 (77% versus 54%), whereas the plasma cholesterol-lowering effect of 0.1% CS-505 was almost the same as that of 0.1% CI-1011. Furthermore, immunohistochemical analysis demonstrated that CS-505 significantly reduced the number of macrophages and expression of matrix metalloproteinase (MMP)-2, MMP-9, and MMP-13. These data indicate that CS-505 can reduce and stabilize atherosclerotic lesions. This antiatherosclerotic activity is exerted via both cholesterol lowering and direct ACAT inhibition in plaque macrophages.     

Lipid-Lowering Therapy in Patients with Average Cholesterol and High Cardiovascular Risk,Insights from the Heart Protection Study

Although the benefits of cholesterol-lowering with statins has been established for patients with high cholesterol levels as well as for patients with prior occlusive coronary heart disease, substantial uncertainty has existed about the long-term benefits of these agents in particular types of patient, including patients with moderate or low baseline cholesterol levels pre-treatment, women, the elderly, and those with prior occlusive non-coronary vascular disease. The Heart Protection Study was designed to resolve these uncertainties and to provide substantially more safety information of statins. In this report the study is presented with the significance and practical implications of the results briefly discussed.     

黑龙江省东部地区汉族人群PCSK9基因第1外显子多态性与脂代谢的相关性

目的 研究黑龙江省东部地区汉族人群前蛋白转化酶枯草溶菌素(PCSK)9基因第1外显子多态性与脂代谢的相关性.方法 因心绞痛和(或)运动试验阳性及有冠脉管腔狭窄证据而需入院行冠状动脉造影者220例中,110例冠心病(CHD)者为病例组,110例非CHD者为对照组,检测PCSK9基因第1外显子基因多态性及血脂水平,并对基因多态性与血脂水平进行相关性分析.结果 共发现c.121C>G、c.299C>T、c.427-428insGCT、A53V、P56S和c.556A>G 6个突变位点,其中在A53V突变位点上发现CC和TC两种基因型,但未发现TT基因型.病例组和对照组各基因型间血清三酰甘油(TG)、总胆固醇(TC)、高密度脂蛋白胆固醇(HDL-C)和低密度脂蛋白胆固醇(LDL-C)水平差异具有统计学意义(P<0.05).病例组中TC基因型血清TG、TC和LDL-C水平均显著高于CC基因型(P<0.05);对照组中TC基因型血清TC水平显著高于CC基因型(P<0.05).结论 PCSK9基因第1外显子在黑龙江省东部地区汉族人群中存在多态性,且与CHD患者脂代谢有关.     

Apolipoprotein(a) and cardiovascular disease in Type 2 (non-insulin-dependent) diabetic patients with and without diabetic nephropathy

Summary The relative mortality from cardiovascular disease is on average increased five-fold in Type 2 (non-insulin-dependent) diabetic patients with diabetic nephropathy compared to non-diabetic subjects. We assessed the possible contribution of dyslipidaemia in general and elevated serum apolipoprotein(a) (apo(a)) in particular. Type 2 diabetic patients with normo-, micro- and macroalbuminuria were compared with healthy subjects. Each group consisted of 37 subjects matched for age, sex and diabetes duration. Serum creatinine in the nephropathy group was 105 (54–740) mol/l. The prevalence of ischaemic heart disease (resting ECG, Minnesota, Rating Scale) was 57, 35, 19 and 2% in macro-, micro- and normoalbuminuric diabetic patients and healthy subjects, respectively. The prevalence of ischaemic heart disease was higher in all diabetic groups as compared to healthy subjects (p<0.05), and higher in macroalbuminuric as compared to normoalbuminuric diabetic patients (p<0.01). There was no significant difference between apo(a) in the four groups: 161 (10–1370), 191 (10–2080), 147 (10–942), 102 (10–1440) U/l (median (range)) in macro-, micro- and normoalbuminuric groups and healthy subjects. Serum total-cholesterol, HDL-cholesterol and LDL-cholesterol were not significantly different when comparing healthy subjects and each diabetic group. Apolipoprotein A-I was lower (p<0.05) in all diabetic groups as compared to healthy subjects (nephropathy vs healthy subjects): 1.50±0.25 vs 1.69±0.32 g/l (mean ± SD). Triglyceride was higher (p<0.05) in patients with nephropathy and microalbuminuria as compared to healthy subjects (nephropathy vs healthy subjects): 2.01 (0.66–14.7) vs 1.09 (0.41–2.75) mmol/l (median (range)). Apolipoprotein B was higher (p<0.02) in patients with nephropathy as compared to the other three groups (nephropathy vs healthy subjects): 1.54±0.47 vs 1.33±0.30 g/l. In conclusion, our case-control study has confirmed that Type 2 diabetic patients with increased urinary albumin excretion frequently suffer from dyslipidaemia and cardiovascular disease. However, our study revealed no significant elevation in serum concentration of apo(a) in patients with diabetic nephropathy, but numbers were small.     

Presenilin-1突变对全反式维甲酸诱导的PC12细胞生长和凋亡的影响

目的　观察Presenilin 1(PS 1)突变型L2 86V对全反式维甲酸 (RA)诱导的PC12细胞生长和凋亡的影响。　方法　运用脂质体介导的基因转染技术建立稳定表达PS 1WT和突变型L2 86V基因的细胞克隆 ,采用MTT法、流式细胞仪检测细胞生长曲线、生长周期及细胞凋亡情况。　结果　 (1)PS 1突变型L2 86V对RA诱导的PC12细胞生长具有抑制作用 ;主要表现为G1期细胞增多、S期细胞减少 ,细胞增殖指数降低 ;(2 )在正常培养条件下 ,对照组细胞凋亡率 (% )为 0 31±0 0 9、野生型组为 0 4 3± 0 11、L2 86V组为 0 6 2± 0 2 7;无血清培养 1、2、3d ,对照组细胞凋亡率 (% )分别为 0 5 5± 0 0 7、1 2 3± 0 4 7、5 5 9± 2 91;野生型组为 1 0 9± 0 73、3 2 1± 1 4 1、7 79± 2 78;L2 86V组为 4 6 5± 1 0 4、10 5 4± 3 18、14 4 7± 3 5 3;PS 1突变型L2 86V对RA诱导的PC12细胞均具有促进细胞凋亡的作用 ,以无血清培养时表现得更为明显。　结论　PS 1突变型L2 86V对RA诱导的PC12细胞生长具有抑制作用 ;在有和无血清培养条件下均具有促进细胞凋亡的作用 ,以无血清培养时表现得更为明显。     

Association of AGTR1 gene methylation and its genetic variant in Chinese farmer with hypertension: A case-control study

The objective was to determine the potential associations of the angiotensin II receptor type 1 (AGTR1) gene polymorphism, methylation, and lipid metabolism in Chinese farmers with hypertension.A case-control study was conducted in Wuzhi county of Henan province in China in 2013 to 2014. A total of 1034 local residents (35–74 years, 386 hypertensive cases, and 648 normotensive subjects) were enrolled in this study. Triglyceride (TG), total cholesterol (TC), high-density lipoprotein, and low-density lipoprotein were measured using automatic chemistry analyzer. The AGTR1 gene promoter methylation level was measured using quantitative methylation-specific polymerase chain reaction method. The single nucleotide polymorphism rs275653 was genotyped with TaqMan probe assay at an applied biosystems platform.The gender, body mass index (BMI), TG, TC, and family history of hypertension in the hypertension group were significantly higher than those in control group (P < .05). No significant difference was observed in the distribution of AGTR1 rs275653 polymorphism in the hypertension and controls (P > .05). The AGTR1 gene methylation in subjects carrying different genotypes was not significantly observed (P > .05). The logistic regression analysis found the AGTR1 gene methylation level was negative correlation with hypertension in the present study (odds ratio, 0.946, 95% confidence interval, 0.896–0.999) through adjusting for age, gender, BMI, education, smoking, alcohol drinking, fruit and vegetable intake, pickles intake, and family history of hypertension.The association of AGTR1 gene hypomethylation and essential hypertension was observed in Chinese farmers; no significant difference was observed in the distribution of AGTR1 rs275653 polymorphism.     

阿尔茨海默病患者分拣蛋白相关受体L1基因突变与mRNA水平的相关性分析

目的探讨散发型阿尔茨海默病患者(SAD)分拣蛋白相关受体L1(SORL1)基因突变与mRNA水平的相关性,为临床早期诊断阿尔茨海默病和预后提供理论依据。方法随机选择SAD患者55例(SAD组),另选健康体检者15例(对照组),取外周血进行SORL1基因mRNA水平检测以及全长cDNA测序分析,并将基因突变与未突变SORL1基因mRNA进行相关性分析。结果 SAD组有8例出现不同位点的SORL1基因突变,其发生概率为1 4.5%,较为常见的突变位点为2650以及2850碱基序列,而对照组未检测到SORL1基因突变。SAD组SORL1基因mRNA表达水平较对照组明显减低,差异有统计学意义(P<0.01);SAD组SORL1基因突变患者与未突变患者mRNA水平比较,差异无统计学意义(P>0.05)。结论 SORL1基因突变可能参与SAD的发生;SORL1基因突变与SORL1基因mRNA水平无明显相关性。     

不同认知水平的广泛性脑萎缩患者磁共振波谱分析

目的 观察不同认知水平的广泛性脑萎缩患者脑内生化物质含量的差异. 方法 按照美国精神障碍诊断与统计手册第4版(DSM-Ⅳ)和认知功能障碍的诊断标准将33名广泛性脑萎缩患者分为阿尔茨海默病(AD)组14例、遗忘型轻度认知功能损害(aMCI)组9例和认知功能正常组10例.所有研究对象接受神经心理量表检测,然后采用1.5-T MR系统对左侧额叶皮质和左侧海马进行氢质子磁共振波谱(1H-MRS)检测分析. 结果 广泛性脑萎缩AD组的左侧海马和左侧额叶皮层的N-乙酰天门冬氨酸(NAA)/肌酸(Cr)值较认知正常组分别降低10.2%和5.3%.胆碱复合物(Cho)/Cr值分别升高17.5%和16.7%,肌醇(MI)/Cr值分别升高39.5%和19.2%.与aMCI组比较,广泛性脑萎缩AD组的左侧海马NAA/Cr值降低6.4%,左侧海马和左侧额叶皮层的Cho/Cr值分别升高9.3%和12.3%,左侧海马和左侧额叶皮层的MI/Cr值分别升高30%和17%,而左侧额叶皮层的NAA/Cr值在两者间差异无统计学意义.广泛性脑萎缩aMCI组的左侧海马NAA/Cr值比正常组降低4.1%、Cho/Cr值比认知正常组升高7.5%,但是左侧额叶皮层的生化改变在两组间差异均无统计学意义. 结论 不同认知水平的广泛性脑萎缩患者存在脑内神经生化物质的变化.左侧海马NAA/Cr值的降低、左侧海马和左侧额叶皮质Cho/Cr和MI/Cr值的升高有助于预测aMCI进展为AD;左侧海马NAA/Cr降低和Cho/Cr升高有助于鉴别广泛性脑萎缩伴aMCI患者与广泛性脑萎缩认知功能正常的患者.     

探讨补体受体1基因与脑脊液中相关蛋白的关系

目的探讨补体成分(3b/4b)受体1基因[the complement component (3b/4b) receptor 1gene,CR1]多态性与脑脊液相关蛋白的关系。方法从美国国立老年研究所组织建立的阿尔茨海默病(AD)神经影像学研究数据库中选择812例受试者,其中48例AD患者为AD组,483例轻度认知障碍(MCI)患者为MCI组,281例正常对照(NC)者为NC组。用免疫分析试剂盒和xMAP Luminex试剂盒检测入选者脑脊液中β淀粉样蛋白(Aβ)、总tau(t-tau)蛋白和磷酸化tau(p-tau)蛋白水平。结果 AD组rs61522287位点基因突变后,GA基因型较GG基因型脑脊液Aβ42水平增加[(193.20±79.05)ng/L vs(137.90±37.28)ng/L,P=0.03762]。8个CR1的单核苷酸多态性(SNP)位点可以调节AD患者脑脊液中t-tau蛋白和p-tau蛋白水平。11个CR1的SNP突变位点参与调节MCI组患者tau蛋白水平。3个CR1的SNP位点基因突变增加MCI组脑脊液中Aβ42水平。rs41274776和rs12567945位点突变同时增加NC组脑脊液中p-tau和Aβ42水平。结论本研究率先明确了CR1的SNP位点与脑脊液蛋白(Aβ42,t-tau和p-tau)之间的关系,这对未来寻找AD的早期诊断,早期筛查提供了新思路。     

Prevalence of dyslipidaemia in statin-treated patients in South Africa: results of the DYSlipidaemia International Study (DYSIS)

Introduction and objectives

Cardiovascular disease (CVD) is the leading cause of mortality worldwide and increased levels of low-density lipoprotein cholesterol (LDL-C) are an important modifiable risk factor. Statins lower LDL-C levels and have been shown to reduce CVD risk. Despite the widespread availability of statins, many patients do not reach the lipid targets recommended by guidelines. We evaluated lipid goal attainment in statin-treated patients in South Africa and analysed variables contributing to poor goal attainment as part of the DYSlipidaemia International Study (DYSIS).

Methods

This cross-sectional, observational study enrolled 1 029 consecutive South African patients consulting officebased physicians. Patients were at least 45 years old, had to be treated with a stable dose of statins for at least three months and had been fasting for 12 hours. We evaluated lipid goal attainment and examined variables associated with residual dyslipidaemia [abnormal levels of LDL-C, highdensity lipoprotein cholesterol (HDL-C) and/or triglycerides (TG)].

Results

We found that 50.3% of the patients overall did not achieve target LDL-C levels and 73.5% of patients were at very high cardiovascular risk. In addition, 33.7% had low levels of HDL-C, while 45.3% had elevated TG levels despite statin therapy. Asian and mixed-ancestry patients but not black (vs Caucasian ethnicity), as well as obese individuals in South Africa were more likely to still have dyslipidaemia involving all three lipid fractions.

Conclusions

We observed that many patients in South Africa experienced persistent dyslipidaemia despite statin treatment, supporting the concept that there is a need for more intensive statin therapy or the development of novel treatment strategies. Measures aimed at combating obesity and other lifestyle-related risk factors are also vital for effectively controlling dyslipidaemia and reducing the burden of CVD.     

血清淀粉样蛋白A与冠心病合并糖尿病患者高密度脂蛋白胆固醇外流能力的相关性

目的 探讨冠心病合并糖尿病患者血浆高密度脂蛋白(HDL)胆固醇外流能力的变化及其影响因素.方法 选取北京大学第三医院心内科住院并经冠状动脉造影证实的冠心病患者140例为研究对象,依据是否合并糖尿病分为两组,即冠心病合并糖尿病组70例,单纯冠心病组70例.选取同期入院且经冠状动脉造影证实的非冠心病非糖尿病患者25例作为对...     

Increased cholesterylester transfer activity in complicated Type 1 (insulin-dependent) diabetes mellitus — its relationship with serum lipids

Summary In Type 1 (insulin-dependent) diabetes mellitus, macrovascular complications and the increased risk for cardiovascular disease in patients with microvascular complications may be related to alterations in plasma cholesterylester transfer. The activity of cholesterylester transfer protein, which mediates cholesterylester transfer between lipoproteins and lipoprotein lipid levels, was assessed in 7 normolipidaemic control subjects, 7 Type 1 diabetic control subjects without complications, 11 Type 1 diabetic patients with microvascular complications (retinopathy, incipient nephropathy) and in 7 Type 1 diabetic patients with macrovascular atherosclerotic lesions.The cholesterylester transfer activity was 30% higher in the diabetic groups with macrovascular and microvascular lesions than in the 2 control groups. Very low + low density lipoprotein cholesterol was higher in the 3 diabetic groups than in the non-diabetic control group. High density lipoprotein cholesterol was not different. The cholesterylester transfer activity was correlated positively with HbA1, urinary albumin excretion rate, serum cholesterol, very low + low density lipoprotein cholesterol and apolipoprotein B. The high density lipoprotein over very low + low density lipoprotein cholesterylester molar ratio was lower in the diabetic groups with micro- and macrovascular complications. A role for cholesterylester transfer activity in the lipoprotein abnormalities found in complicated Type 1 diabetes is suggested. A high cholesterylester transfer activity might be indicative of mechanisms which promote atherogenesis.     

Altered temporal patterns of anxiety in aged and amyloid precursor protein (APP) transgenic mice

Both normal aging and dementia are associated with dysregulation of the biological clock, which contributes to disrupted circadian organization of physiology and behavior. Diminished circadian organization in conjunction with the loss of cholinergic input to the cortex likely contributes to impaired cognition and behavior. One especially notable and relatively common circadian disturbance among the aged is "sundowning syndrome," which is characterized by exacerbated anxiety, agitation, locomotor activity, and delirium during the hours before bedtime. Sundowning has been reported in both dementia patients and cognitively intact elderly individuals living in institutions; however, little is known about temporal patterns in anxiety and agitation, and the neurobiological basis of these rhythms remains unspecified. In the present study, we explored the diurnal pattern of anxiety-like behavior in aged and amyloid precursor protein (APP) transgenic mice. We then attempted to treat the observed behavioral disturbances in the aged mice using chronic nightly melatonin treatment. Finally, we tested the hypothesis that time-of-day differences in acetylcholinesterase and choline acetyltransferase expression and general neuronal activation (i.e., c-Fos expression) coincide with the behavioral symptoms. Our results show a temporal pattern of anxiety-like behavior that emerges in elderly mice. This behavioral pattern coincides with elevated locomotor activity relative to adult mice near the end of the dark phase, and with time-dependent changes in basal forebrain acetylcholinesterase expression. Transgenic APP mice show a similar behavioral phenomenon that is not observed among age-matched wild-type mice. These results may have useful applications to the study and treatment of age- and dementia-related circadian behavioral disturbances, namely, sundowning syndrome.     

Sex-specific maternal calcium requirements for the prevention of nonalcoholic fatty liver disease by altering the intestinal microbiota and lipid metabolism in the high-fat-diet-fed offspring mice

ABSTRACT

The significance of maternal appropriate calcium intakes for energy metabolism in the offspring has been recognized. Nonalcoholic fatty liver disease (NAFLD) is considered as the hepatic manifestation of metabolic syndrome. So in this study, we proposed that there were long-term effects of maternal calcium status on the progress of NAFLD by altering the intestinal microbiota and lipid metabolism with attention to potential sex differences among the mouse offspring. Thirty-four-week female C57BL/6 J mice were subjected to obtain low, normal and high calcium reproductive diets throughout the gestation and lactation. After weaning, both the male and female mouse offspring were fed with the high-fat diet for 16 weeks, with the normal diet as control. Biochemical indicators in the plasma and hepatic tissue were measured using ELISA or enzymatic methods. The expression of lipid metabolism, inflammatory and fibrosis related genes was determined by RT-PCR. The intestinal microbiota was analyzed by 16S rRNA high-throughput sequencing. Maternal normal and low calcium intake could, respectively, inhibit the progress of high-fat diet induced NAFLD in the male and female mouse offspring, which was characterized by the least lipid droplets, inflammatory infiltration and fibrosis, the lowest concentrations of free fatty acids and triglyceridethe lowest expression of genes involving in de novo lipogenesis and the highest expression of genes related to lipid oxidation and hydrolysis, inflammatory, and fibrosis. Pyrosequencing of 16S rRNA genes revealed that the male mouse offspring with maternal normal calcium intake and the female mouse offspring with maternal low calcium intake, after the high-fat diet feeding, had distinct intestinal microbiota, which was closer to thosein mice with the normal diet feeding. Analysis of the functional features for the different microbiota was compatible with the expression of genes associated with lipogenesis, lipid oxidation and hydrolysis. Thus, there is a sex-specific manner for maternal calcium requirement to inhibit the progress of offspring NAFLD, that might be less for the female offspring and more for the male offspring.