老年帕金森病患者血清Aβ1-42、EGF和Hcy水平与认知功能障碍及疾病严重程度的关系及其预测价值分析

摘要 目的：探讨老年帕金森病（PD）患者血清中β淀粉样蛋白1-42（Aβ1-42）、表皮生长因子（EGF）和同型半胱氨酸（Hcy）的表达水平,分析其与认知功能障碍及疾病严重程度的关系,并探讨其对老年PD患者认知功能障碍的预测价值。方法：将2018年10月~2021年10月在河北北方学院附属第一医院进行治疗的老年PD患者120例研究对象纳入PD组,另选取同期在河北北方学院附属第一医院进行健康体检的120例老年健康体检者为对照组。检测PD组、对照组血清Aβ1-42、EGF、Hcy水平,比较各血清指标水平的差异。Spearman相关性分析法分析血清中Aβ1-42、EGF、Hcy与简易智力状态检查量表（MMSE）、蒙特利尔认知评估量表（MoCA）评分的相关性。接收者工作特征曲线（ROC）分析血清Aβ1-42、EGF、Hcy水平对老年PD患者认知功能障碍的预测价值。结果：PD组患者血清Aβ1-42、EGF水平低于对照组,Hcy水平高于对照组,差异有统计学意义（P<0.05）。随着PD分期的加重,老年PD患者的Aβ1-42、EGF水平随之降低,Hcy水平则随之升高（P<0.05）。PD伴认知功能障碍组的Aβ1-42、EGF水平和MMSE、MoCA评分低于PD认知功能正常组,Hcy则高于PD认知功能正常组,差异均有统计学意义（P<0.05）。Spearman相关性分析显示,Aβ1-42、EGF水平与MMSE、MoCA评分均呈正相关性（P<0.05）,Hcy水平与MMSE、MoCA评分均呈负相关性（P<0.05）。血清Aβ1-42、EGF、Hcy单独应用时对老年PD患者认知功能障碍均有一定的预测效能,而血清Aβ1-42、EGF、Hcy联合应用的曲线下面积（AUC）（0.95CI）为0.841（0.738～0.932）,预测效能高于单一指标检测。结论：老年PD患者血清Aβ1-42、EGF水平呈低表达,Hcy水平呈高表达,血清Aβ1-42、EGF、Hcy水平参与了老年PD患者病情的发生、进展过程,且与患者认知功能关联密切。血清Aβ1-42、EGF、Hcy水平联合检测在预测老年PD患者的认知功能障碍方面具有一定的临床价值。     

缺失plcg1基因引起成纤维细胞PI—3K信号上调

磷脂酶C－γ1（phospholipase C－γ1,PLC－γ1）与磷脂酰肌醇3－激酶（phosphatidylinositol-3 kinase,PI-3K）是生长因子调控细胞生长与增殖的两个重要信号中介。为探讨PLC－γ1在表皮生长因子（EGF）介导的细胞分裂信号中的代偿机制,用磷脂酶C（phospholipase C,PLC）特异性抑制剂U73122及PI－3K牧场划必抑制剂wortmannin处理剔除PLC－δ1基因plcg1(PLC-γ1^-/-）及野生型（PLC－γ1^ / )小鼠胚胎成纤维细胞,发现未经处理情况下两种细胞的克隆形成能力、细胞活力及EGF引起的DNA合成能力相似,且均可被U73122与wortmannin抑制,但与PLC－γ1^ / 细胞相比,PLC－γ1^－／－更依赖于PI－3K,而对PLC的依赖性却减小。Western印迹也表明EGF刺激后PI－3K的p85α亚单位酷氨酸磷酸化程度比野生型显著增高,PI－3K信号通路的激活出现上调,且PLC－γ1^-/-中无基近亲PLC－γ2的代偿表达。因此PLC－γ1^-/-中PLC－γ1的功能可能被PI－3K通路代偿,而PLC－γ2或其他PLC同工酶并不代偿其功能。结果表明EGF介导的信号能路的冗余性及PLC－γ1信号通路的可代偿性。     

甲状旁腺素、表皮生长因子及维生素A酸对UMR106细胞EGF受体的调节作用

本文研究了EGF、PTH和RA对UMR106细胞EGF受体的调节作用。结果显示PTH能上调EGF的受体,UMR106细胞经bPTH(1-34)处理3天,EGF受体的相对结合率与对照比较提高了40.3％,每个细胞的EGF受体数目从7.22×10~3增加到1.44×10~4,Kd从2.02×10~(-11)增加到3.68×10~(-11)mol/L。而RA则能下调EGF受体,以RA处理3天,EGF受体数目从7.22×10~3下降到4.28×10~3,Kd则从2.02×10~(-11)增加到4.17×10~(-11)mol/L。提示PTH和RA可能通过调变其EGF受体而分别起到正性和负性生长调节作用。     

抗肿瘤药物帕玛度胺类似物的合成

目的：设计并合成抗肿瘤药物帕玛度胺的3位N取代的新型类似物。方法：从3-硝基邻苯二甲酸（2）和N-（叔丁氧羰基）．L-谷氨酰胺（4）出发,经过六步反应得到目标化合物。3-硝基邻苯二甲酸（2）经脱水制得3-硝基邻苯二甲酸酐（3）。用N-（叔丁氧羰基）-L-谷氨酰胺（4）经闭环、脱保护制得3-氨基-2,6-哌啶二酮三氟乙酸盐（6）。4与6经缩合、钯碳催化氢化制得免疫调节剂Pomalidomide（8）,（8）经过酰化得到3-乙酰氨基-N-（2,6-二氧代-3-哌啶基）邻苯二甲酰亚胺（1）。以（4）计总收率约34．9％。结果：得到3-乙酰氨基-N-（2,6-二氧代-3-哌啶基）-邻苯二甲酰亚胺（1）,应用于细胞活性测试。结论：改进了帕玛度胺的合成工艺,得到了3位N乙酰化的新型帕玛度胺类似物（1）,初步研究显示（1）的生物活性与帕玛度胺接近。     

木瓜化学成分的研究

从木瓜[Chaenomeles speciosa（Sweet）Nakai]果实的乙酸乙酯提取物中分离并鉴定了4个化合物,利用现代波谱技术（MS、^1H NMR、^13C NMR）进行结构鉴定,它们分别为对苯二酚（1）、3,4-二羟基苯甲酸（2）、槲皮素（3）和3-羟基丁二酸甲酯（4）.2,2-二苯基-1-苦肼基（DPPH）自由基清除试验表明化合物（1～3）显示抗氧化活性.化合物1～4均为首次从该植物中获得.     

草珊瑚中一个新的苯基丙二醇类化合物

从金粟兰科植物草珊瑚（Sarcandra glabra）全株的水提取物中分离得到一个新的苯基丙二醇类化合物（2S）-3, 3-二-（4-羟基-3-甲氧基苯基）-丙-1, 2-二醇 （1）,以及一个已知化合物,银线草内酯E （2）,并利用NMR、MS、旋光值等波谱学技术鉴定其了结构。     

隔山消化学成分的研究

从隔山消的石油醚部分分离得到6个芳香类化合物,分别鉴定为2,4-二羟基苯乙酮（1-（2,4-dihy—droxph—enyl）ethanone,1）、奎乙酰苯（acetylquinol,2）、对羟基苯乙酮（1-（4-hydroxyphenyl）ethanone,3）、4-羟基-3-甲氧基苯乙酮（1-（4-hydroxy-3-methoxyphenyl）ethanone,4）、2,4-二羟基-5-甲氧基苯乙酮（1-（2,4-dihydroxy-5-methoxyphenyl）ethanone,5）、3-羟基4-甲氧基苯甲酸（3-hydroxy-4-methoxy—benzoicacid,6）。除化合物2外,其余5个化合物均首次从该植物中分离得到。     

柳叶白前化学成分研究

从柳叶白前根和茎中分离并鉴定了7个化合物,分别为β-谷甾醇（β-sitosterol,1）,2,4-二羟基苯乙酮（1-（2,4-dihydroxphenyl）ethanone,2）,间二苯酚（resorcinol,3）,4-羟基-3-甲氧基苯乙酮（1-（4-hydroxy-3-methoxyphenyl）e-hanone,4）,4-羟基苯乙酮（1-（4-hydroxyphenyl）ethanone,5）,齐墩果酸（oleanolic acid,6）,蔗糖（sucrose,7）。其中化合物2-7为首次从该植物中分得。     

肺泡Ⅱ型细胞表皮生长因子和转化生长因子α、β1及其受体表达的研究

分离培养成年大鼠的肺泡Ⅱ型细胞,通过斑点杂交、原位杂交和免疫组织化学染色,研究肺泡Ⅱ型细胞内表皮生长因子（EGF）、转化生长因子α和β1（TGFα、TGFβ1）及其受体基因的表达。结果显示肺泡Ⅱ型细胞可表达EGF、TGFα和TGFβ1,也可表达相应的EGF受体（EGFR）、TGFβ受体Ⅰ型和Ⅱ型（TβRⅠ、TβRⅡ）。表明肺泡Ⅱ型细胞是合成和分泌EGF、TGFα和TGFβ1的细胞之一;细胞凭借其EGFR、TβR的存在,其增殖与分化可能受EGF、TGFα和TGFβ1的旁分泌和自分泌两种途径调控。     

黄花石蒜中的黄酮类成分

通过对黄花石蒜（Lycoris aurea）乙醇提取物的乙酸乙酯溶解部分进行分离纯化,得到7个化合物.经物理方法和波谱分析,分别鉴定为（2S）-4＇-羟基-7-甲氧基黄烷（1）、（2S）-3＇,7-二羟基-4＇-甲氧基黄烷（2）、（2S）-4＇,7-二羟基黄烷（3）、（2S）-4＇,7-二羟基-8-甲基黄烷（4）、（2S）-4＇,7-二羟基-3＇-甲氧基-8-甲基黄烷（5）、（2S）-4＇,5,7-三羟基-8-甲基黄烷酮（6）、2,4＇-二羟基-4-甲氧基二氢查尔酮（7）.所有化合物均为首次从该属植物中分离得到,另外,本文首次归属了化合物5、6、7的13C NMR信号.     

Antibody-quantum dot conjugates exhibit enhanced antibacterial effect<Emphasis Type="Italic">vs.</Emphasis> unconjugated quantum dots

The effect of a 20-min exposure to antibody-quantum dot (Ab-QD) conjugates on colony counts of Escherichia coli was assessed by the spread-plate method and compared with exposure to unconjugated QDs having only amine or carboxyl groups on their surfaces. Under these conditions, Ab-QD conjugates generally exhibited >90% reduction in colony-forming units as compared to untreated E. coli and E. coli treated with unconjugated QDs after incubation for as long as 41 h. The antibacterial effect of Ab-QD conjugates vs. unconjugated QDs on Salmonella enterica subsp. enterica serovar Typhimurium was also assessed by means of a disk-diffusion technique which demonstrated greater growth inhibition (approximately 3 mm greater) by Ab-QD conjugate-impregnated disks than by unconjugated-QD-only-impregnated disks at a 10-microg disk load. At a 25-microg disk load, both treatment groups exhibited nearly equal growth inhibition.     

Biotin-4-fluorescein based fluorescence quenching assay for determination of biotin binding capacity of streptavidin conjugated quantum dots

The valency of quantum dot nanoparticles conjugated with biomolecules is closely related to their performance in cell tagging, tracking, and imaging experiments. Commercially available streptavidin conjugates (SAv QDs) are the most commonly used tool for preparing QD-biomolecule conjugates. The fluorescence quenching of biotin-4-fluorscein (B4F) provides a straightforward assay to quantify the number of biotin binding sites per SAv QD. The utility of this method was demonstrated by quantitatively characterizing the biotin binding capacity of commercially available amphiphilic poly(acrylic acid) Qdot ITK SAv conjugates and poly(ethylene glycol) modified Qdot PEG SAv conjugates with emission wavelengths of 525, 545, 565, 585, 605, 625, 655, 705, and 800 nm. Results showed that 5- to 30-fold more biotin binding sites are available on ITK SAv QDs compared to PEG SAv QDs of the same color with no systematic variation of biotin binding capacity with size.     

Effect of heteroatom-doped carbon quantum dots on the red emission of metal-conjugated phthalocyanines through hybridization

Quantum dots (QDs) are significant fluorescent materials for energy transfer studies with phthalocyanines (Pcs) and phthalocyanine (Pc)-like biomolecules (such as chlorophylls). Carbon-based QDs, especially, have been used in numerous studies concerning energy transfer with chlorophylls, but the numbers of studies concerning energy transfer between phthalocyanines and carbon-based QDs are limited. In this study, peripherally, hydroxythioethyl terminal group substituted metal-free phthalocyanine (H₂Pc) and zinc phthalocyanine (ZnPc) were noncovalently (electrostatic and/or π–π interaction) attached to carbon QDs containing boron and nitrogen to form QD-Pc nanoconjugates. The QD-Pc conjugates were characterized using different spectroscopic techniques (Fourier transform infrared spectroscopy and transmission electron microscopy). The absorption and fluorescence properties of QD-Pc structures in solution were studied. It was found that the quantum yields of the QDs slightly decreased from 30% to 25% upon doping the QDs with heteroatoms B and N. Förster resonance energy transfer efficiency was calculated as 33% for BCN-QD/ZnPc. For the other conjugates, almost no energy transfer from QDs to Pc cores was observed. It was shown that the energy transfer between QDs to Pc cores was completely different from the energy transfer between QDs and photosynthetic pigments, and therefore we concluded that heteroatom doping in the QD structure and the existence of zinc metal in the phthalocyanine structure is obligatory for an efficient energy transfer.     

Quantum Dots as alternatives to organic fluorophores for Cryptosporidium detection using conventional flow cytometry and specific monoclonal antibodies: lessons learned.

BACKGROUND: Significant developments in biological applications are occurring through the incorporation of Quantum Dots (QDs) as biological labels. The demonstration of QDs unique optical properties may have important implications for the study of environmental samples, where microorganisms of interest need to be isolated away from the background debris. METHODS: Flow cytometric analysis was used to determine the fluorescence intensity of oocysts after mAb staining by QDs or organic fluorophore conjugates. In addition, the level of non-specific binding to detrital particles within a control water concentrate was estimated using the optimal staining concentration determined for each mAb analyzed. RESULTS: Under 488 nm excitation, oocysts stained with QD-conjugates exhibited significantly lower fluorescence intensity than organic conjugates. Moreover, the level of non-specific binding by QD-conjugates to detrital particles present in the water concentrate was significantly higher that of the organic conjugates. CONCLUSIONS: While QDs are noted for their superior spectral characteristics, they have been shown here to be unsuitable for conventional flow cytometric detection of Cryptosporidium. Therefore, we conclude that in their current form, QD's are severely limited for fluorescent detection of pathogens in environmental applications.     

Synthesis and photophysical studies of CdTe quantum dot-monosubstituted zinc phthalocyanine conjugates

The linkage of unsymmetrically monosubstituted 4-aminophenoxy zinc phthalocyanine (ZnAPPc, 5) to CdTe quantum dots capped with mercaptopropionic acid (MPA), l-cysteine (l-cys) or thioglycolic acid (TGA) has been achieved using the coupling agents ethyl-N(3-dimethylaminopropyl) carbodiimide and N-hydroxy succinimide, which facilitate formation of an amide bond to form the QD-ZnAPPc-linked conjugate. The formation of the amide bond was confirmed using Raman and IR spectroscopies. Atomic force microscopy (AFM) and UV-Vis spectroscopy were used further to characterise the conjugate. Förster resonance energy transfer (FRET) resulted in stimulated emission of ZnAPPc in both the linked (QD-ZnAPPc-linked) and mixed (QD:ZnAPPc-mixed) conjugates. The linked l-cys and TGA QDs conjugates (QD-ZnAPPc-linked) gave the largest FRET efficiencies hence showing the advantages of covalent linking. Fluorescence quantum yields of QDs were decreased in QD:ZnAPPc-mixed and QD:ZnAPPc-linked.     

Effect of ligand density on the spectral, physical, and biological characteristics of CdSe/ZnS quantum dots

Chemical modification of the surface of CdSe/ZnS quantum dots (QDs) with small molecules or functional ligands often alters the characteristics of these particles. For instance, dopamine conjugation quenches the fluorescence of the QDs, which is a property that can be exploited for sensing applications if the conjugates are taken up into living cells. However, different sizes and/or preparations of mercaptocarboxylic acid solubilized QDs show very different properties when incubated with cells. It is unknown what physical parameters determine a QDs ability to interact with a cell surface, be endocytosed, escape from endosomes, and/or enter the nucleus. In this study, we examine the surface chemistry of QD-dopamine conjugates and present an optimized method for tracking the attachment of small biomolecules to the surface. It is found that the fluorescence intensity, surface charge, colloidal stability, and biological interactions of the QDs vary as a function of the density of dopamine on the surface. Successful targeting of QD-dopamine to dopamine receptor positive PC12 cells correlates with greater homogeneity of particle thiol layer, and a minimum number of ligands required for specific association can be estimated. These results will enable users to develop methods for screening QD conjugates for biological activity before proceeding to experiments with cell lines and animals.     

Capping of CdSe-ZnS quantum dots with DHLA and subsequent conjugation with proteins

We provide a detailed protocol for designing water-soluble CdSe-ZnS quantum dots (QDs) based on cap exchange of the native hydrophobic shell with dihydrolipoic acid (DHLA) ligands, and the preparation of functional QD bioconjugates for use in immunoassays. Our conjugation strategy is based on non-covalent self-assembly between DHLA-capped QDs and protein appended with either an electrostatic attachment domain (namely, the basic leucine zipper) or a polyhistidine tag. These bioconjugates combine the properties of the QD and attached biomolecule to create structures with desirable luminescent and biologically specific properties. This method also allows the preparation of mixed surface conjugates, which results in the conjugates gaining multiple biological activities. Conjugation of DHLA-capped QDs to maltose binding protein (MBP), the immunoglobulin-G-binding beta2 domain of streptococcal protein G (PG) and avidin will be described. MBP and PG were modified by genetic fusion with either a charged leucine zipper or a polyhistidine interaction domain.     

Synthesis of a quinazoline derivative: a new α₁-adrenoceptor ligand for conjugation to quantum dots to study α₁-adrenoceptors in living cells

Quantum dots (QDs) that are conjugated to small molecule derivatives of drugs and endogenous ligands may be useful tools to study the distribution and dynamic of membrane bound receptors, ion channels and transporters in live cells. In order to use these tools, it is necessary to functionalize QDs with bioactive ligands. In this paper, we successfully synthesized a ligand of α(1)-adrenoceptor that could be conjugated to QDs. In addition, the conjugation of the ligands to QDs and their biological activity were evaluated through binding assay with 30 nM QD conjugates in living human embryonic kidney 293 cells.     

In situ detection of salicylic acid binding sites in plant tissues

The determination of hormone‐binding sites in plants is essential in understanding the mechanisms behind hormone function. Salicylic acid (SA) is an important plant hormone that regulates responses to biotic and abiotic stresses. In order to label SA‐binding sites in plant tissues, a quantum dots (QDs) probe functionalized with a SA moiety was successfully synthesized by coupling CdSe QDs capped with 3‐mercaptopropionic acid (MPA) to 4‐amino‐2‐hydroxybenzoic acid (PAS), using 1‐ethyl‐3‐(3‐dimethyllaminopropyl) carbodiimide (EDC) as the coupling agent. The probe was then characterized by dynamic light scattering and transmission electron microscopy, as well as UV/vis and fluorescence spectrophotometry. The results confirmed the successful conjugation of PAS to CdSe QDs and revealed that the conjugates maintained the properties of the original QDs, with small core diameters and adequate dispersal in solution. The PAS–CdSe QDs were used to detect SA‐binding sites in mung bean and Arabidopsis thaliana seedlings in vitro and in vivo. The PAS–CdSe QDs were effectively transported into plant tissues and specifically bound to SA receptors in vivo. In addition, the effects of the PAS–CdSe QDs on cytosolic Ca²⁺ levels in the tips of A. thaliana seedlings were investigated. Both SA and PAS–CdSe QDs had similar effects on the trend in cytosolic‐free Ca²⁺ concentrations, suggesting that the PAS–CdSe QDs maintained the bioactivity of SA. To summarize, PAS–CdSe QDs have high potential as a fluorescent probe for the in vitro/in vivo labeling and imaging of SA receptors in plants. Copyright © 2014 John Wiley & Sons, Ltd.     

Colistin-functionalised CdSe/ZnS quantum dots as fluorescent probe for the rapid detection of Escherichia coli

Intensely fluorescent, colistin-functionalised CdSe/ZnS QDs (Colis-QDs) nanoparticles, are synthesized and used as sensitive probes for the detection of Escherichia coli, a Gram-negative bacteria. Colistin molecules are attached to the terminal carboxyl of the mercaptoacetic acid-capped QDs in the presence of 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) as amide bond promoters. The TEM analysis of bacteria treated with Colis-QDs conjugates showed the accumulation of Colis-QDs in the cell wall of E. coli. Under the recommended working conditions, the method provides a detection limit as few as 28 E. coli cells per mL, which is competitive which more elaborate detection systems. The simplicity of the method together with short analysis time (< 15 min, without including preparation and photoactivation of the Colis-QDs conjugate) make the proposed approach useful as quick bacteria screening system.