Microfluidic-Based Cationic Cholesterol Lipid siRNA Delivery Nanosystem: Highly Efficient In Vitro Gene Silencing and the Intracellular Behavior

Safe and efficient delivery of small interfering RNA (siRNA) is essential to gene therapy towards intervention of genetic diseases. Herein, we developed a novel cationic cholesterol lipid derivative (CEL) in which cholesterol hydrophobic skeleton was connected to L-lysine cationic headgroup via a hexanediol linker as the non-viral siRNA delivery carrier. Well-organized CEL/siRNA nanocomplexes (100–200 nm) were prepared by microfluidic-assisted assembly of CEL and siRNA at various N/P ratios. The CEL and CEL/siRNA nanocomplexes have lower cytotoxicity compared with bPEI25k. Delightfully, we disclosed that, in Hela–Luc and H1299–Luc cell lines, the micro-fluidic-based CEL/siRNA nanocomplexes exhibited high siRNA transfection efficiency under both serum-free condition (74–98%) and low-serum circumstances (80–87%), higher than that of lipofectamine 2000. These nanocomplexes also showed high cellular uptake through the caveolae/lipid-raft mediated endocytosis pathway, which may greatly contribute to transfection efficiency. Moreover, the time-dependent (0–12 h) dynamic intracellular imaging demonstrated the efficient delivery to cytoplasm after lysosomal co-localization. The results indicated that the microfluidic-based CEL/siRNA nanosystems possessed good stability, low cytotoxicity, high siRNA delivery efficiency, rapid cellular uptake and caveolae/lipid raft-dependent internalization. Additionally, this study provides a simple approach for preparing and applying a “helper lipid-free” cationic lipid siRNA delivery system as potential nanotherapeutics towards gene silencing treatment of (tumor) diseases.     

核桃油对小鼠血脂及胆固醇的影响

采用不同剂量的核桃油对小鼠进行试验，研究其对血脂水平的影响。结果表明，核桃油试验组小鼠的血清甘油三脂（TG）、总胆固醇（TC）和动脉硬化指数（AI）均不同程度低于高脂模型组（P〈0.05或P〈0.01），而高密度脂蛋白（HDL-C）却显著（P〈0．05）高于高脂模型组，表明，核桃油确有显著降低血脂和降低动脉硬化危险性的作用。     

乳酸菌降胆固醇作用研究现状

乳酸菌及其相关制品具有降低介质以及血清中胆固醇的能力，已被体内及体外的大量实验所证实。然而，国内外对于乳酸菌降低胆固醇的作用机理尚存在不同的观点，主要体现在：（1）乳酸菌细胞直接吸收胆固醇；（2）乳酸菌的胆盐水解酶活性使胆盐由结合态转变为脱结合态，与胆固醇发生共沉淀；（3）其他理论，如抑制胆固醇合成途径中的酶等。对涉及上述作用机理的相关研究作了评述，提出和分析了研究中所存在的问题。     

胆固醇氧化酶降解鸭肠中胆固醇的工艺条件研究

目的:研究用胆固醇氧化酶降解鸭肠中胆固醇,为拓宽鸭肠的应用范围提供参考。方法:分光光度法。结果:通过单因素研究和响应曲面法分析,得到酶解胆固醇的最佳条件是,当料液比为1∶10时,酶解pH为7.08,酶解时间为3.04h,酶解温度为25℃,酶用量为0.12U/g,鸭肠中胆固醇的降解率达36.76%。结论:胆固醇氧化酶可有效降解鸭肠中胆固醇且感官品质基本不变。     

APOB CRISPR-Cas9 Engineering in Hypobetalipoproteinemia: A Promising Tool for Functional Studies of Novel Variants

Hypobetalipoproteinemia is characterized by LDL-cholesterol and apolipoprotein B (apoB) plasma levels below the fifth percentile for age and sex. Familial hypobetalipoproteinemia (FHBL) is mostly caused by premature termination codons in the APOB gene, a condition associated with fatty liver and steatohepatitis. Nevertheless, many families with a FHBL phenotype carry APOB missense variants of uncertain significance (VUS). We here aimed to develop a proof-of-principle experiment to assess the pathogenicity of VUS using the genome editing of human liver cells. We identified a novel heterozygous APOB-VUS (p.Leu351Arg), in a FHBL family. We generated APOB knock-out (KO) and APOB-p.Leu351Arg knock-in Huh7 cells using CRISPR-Cas9 technology and studied the APOB expression, synthesis and secretion by digital droplet PCR and ELISA quantification. The APOB expression was decreased by 70% in the heterozygous APOB-KO cells and almost abolished in the homozygous-KO cells, with a consistent decrease in apoB production and secretion. The APOB-p.Leu351Arg homozygous cells presented with a 40% decreased APOB expression and undetectable apoB levels in cellular extracts and supernatant. Thus, the p.Leu351Arg affected the apoB secretion, which led us to classify this new variant as likely pathogenic and to set up a hepatic follow-up in this family. Therefore, the functional assessment of APOB-missense variants, using gene-editing technologies, will lead to improvements in the molecular diagnosis of FHBL and the personalized follow-up of these patients.     

溶血磷脂制备含胆固醇微乳液的研究

溶血磷脂HLB较大，易形成O／W型乳化剂，可制备热力学稳定的微乳化体系。利用溶血磷脂作乳化剂，制备含胆固醇的微乳液，是以胆固醇为油相，溶血磷脂为乳化剂，正丙醇为共表面活性剂，制成微乳液。最终产物呈透明或半透明的均相混合物。     

胆固醇氧化酶高产菌株的筛选及鉴定

将从不同地点采集到的9 个样品进行分离筛选,从中分离到23 株以胆固醇为唯一碳源的菌株,确定其中1 株胞外胆固醇氧化酶高产菌株A3,其胆固醇氧化酶的活力为525U/L。经过形态学和生理生化实验初步鉴定,该菌株属于短杆菌属(Brevibacterium sp.)。     

Dietary effects on growth,plasma lipid and tissues of rats fed with non‐conventional oil of Telfairia occidentalis

The nutritional value of Telfairia occidentalis seed and seed oil available in local market was studied in rats. The average values of the proximate analysis on dry matter basis were 203 g kg^?1 crude protein, 536.3 g kg^?1 fat, 39 g kg^?1 ash, 95 g kg^?1 crude fibre and 43.7 g kg^?1 carbohydrate, while the moisture content was 75 g kg^?1. The mineral analyses showed that T occidentalis seeds contain potassium (3280 mg kg^?1), sodium (1230 mg kg^?1), magnesium (60.8 mg kg^?1), calcium (60.8 mg kg^?1) and iron (13.0 mg kg^?1). The oil extracted from the seed sample showed physicochemical properties that suggest its suitability for edible and industrial uses. The oil contained high levels of unsaturated fatty acids, oleic and linoleic, which accounted for 63.22% of total fatty acids recovered, while palmitic acid (27.4% of the oil by weight) was the most prevalent saturated fatty acid in the oil. No trace of eleostearic acid was detected in the oil. The effect on physical appearance, weight gain, organ weight, tissue and plasma cholesterol and triacylglycerol levels was determined in rats fed a diet containing 5% T occidentalis oil. These were compared with those of rats fed a control diet (0% T occidentalis oil) over an 8 week study period. Weekly monitoring of the rats showed good physical appearance and steady weight increase. The test rats were not anaemic and no mortality was recorded. While there was a significantly higher (p < 0.05) level of cholesterol in the heart of test rats compared with the control, there was a significantly lower (p < 0.05) plasma cholesterol level, suggesting that T occidentalis seed oil can be used to lower plasma levels of cholesterol. Histological examination of the sections of the heart, liver, kidney and spleen of the test rats revealed varying degrees of pathology. These include artherosclerotic changes in the walls of vasa vasori and lymphatics in the heart and hyaline degeneration of myofibrils, fatty infiltration of hepatocytes, and glomerular and tubular degeneration in the kidneys. These lesions may have been due to the effect of fatty acids, especially erucic acid, in the seed oil, which at 1.24% (less that the 5% conventional allowable limit for edible oils) may be intolerable for rats and may be unsuitable for human consumption. The seed oil may be used for industrial purposes such as soap making, lubrication and cosmetics for which it is well suited. Copyright © 2004 Society of Chemical Industry     

柿单宁的体外降胆固醇作用

采用分光光度法和气相色谱法研究了柿单宁对胰脂酶活性和胆固醇酯酶活性的抑制作用,以及柿单宁与胆汁酸、胆固醇的结合能力。结果表明：柿单宁对胰脂酶具有显著的抑制效果,其半数抑制浓度（half inhibitoryconcentration,IC50）为（0.445±0.021） mg/mL;抑制类型为非竞争性抑制,抑制常数Ki为0.406 mg/mL;柿单宁对胆固醇酯酶也具有显著的抑制作用,其半数抑制浓度IC50为（0.442±0.017）mg/mL,抑制类型为非竞争性抑制,抑制常数Ki为0.488 mg/mL。不同质量浓度的柿单宁对胆酸（cholic acid,CA）、脱氧胆酸（deoxycholic acid,DCA）都有一定的结合能力,10 mg/mL的单宁与CA、DCA的结合率分别为58%和94%;柿单宁对胆固醇胶束化溶液有较强的结合能力,0.20 mg/mL的单宁对其抑制率达到了67%。另外,在不同的pH值下,柿单宁对胆固醇都有一定的结合作用,其中pH 7.0时的结合能力明显强于pH 2.0时的结合能力。总之,柿单宁可抑制胰脂酶活性和胆固醇酯酶活性,并与胆固醇和胆汁酸结合,减少胃肠道对脂肪的吸收,这可能是柿单宁在体内发挥强降胆固醇作用的原因之一。     

固相萃取—GC/MS法测定油脂胆固醇含量的研究

建立了固相萃取- GC/MS法测定油脂中胆固醇含量的分析方法,用氟罗里硅土小柱提取和纯化油脂样品中的待测化合物后,GC/MS用HP -5毛细管柱分离,以选择离子模式(SIM)进行定性定 量分析.在0.1 ～ 25mg/L的浓度范围内浓度与峰面积具有良好的线性关系,相关系数(r)为0.9994,在1、5、20mg/...