纳米二氧化硅对体外培养HaCaT细胞凋亡的影响

[目的]比较不同粒径(15、30、100 nm)纳米二氧化硅(nano-SiO2)和常规二氧化硅(Micro-SiO2)对人皮肤表皮细胞(HaCaT)生长的抑制作用及凋亡的影响。[方法]采用不同浓度(2.5、5、10μg/ml)不同粒径(15、30、100 nm)的nano-SiO2和10μg/ml的Micro-SiO2(1～5μm)染毒体外培养的HaCaT细胞24 h,同时设溶剂对照组(nano-SiO2和Micro-SiO2的分散液染毒组)。采用细胞计数试剂盒(CCK-8)法检测HaCaT细胞暴露于不同浓度、不同粒径的nano-SiO2后的细胞活力,用Annexinⅴ-PI双染法和Hoechest33342染色法测定细胞凋亡情况;使用2’,7’-二乙酰二氯荧光素荧光探针检测细胞内活性活性氧(ROS)水平。[结果]不同粒径的nano-SiO2纳米对HaCaT细胞的半数抑制浓度分别为(19.4±1.3)、(27.7±1.5)、(35.9±1.6)μg/ml。当染毒浓度固定时,随着nano-SiO2粒径的减小,凋亡率逐渐增加。当粒径固定时,胞内ROS的水平也随着nano-SiO2浓度的增大而增高。相关分析表明,细胞存活率和凋亡率与胞内活性氧的水平的相关性分别为-0.952和0.898(P<0.01)。[结论]nano-SiO2能抑制HaCaT细胞生长并可诱导其凋亡,这种现象的发生可能与胞内产生的活性氧水平有关。     

Polymeric nanoparticles as an oral delivery system for biocontrol agents for the brushtail possum (Trichosurus vulpecula)

Abstract

AIM: To investigate polymeric nanoparticles as an oral delivery system for protein biocontrol agents for control of the brushtail possum, Trichosurus vulpecula.

METHODS: Insulin-loaded poly(ethyl 2-cyanoacrylate) (PECA) nanoparticles were prepared using interfacial polymerisation, and characterised for size, zeta potential, and efficiency of encapsulation of insulin. In-vitro release of insulin-loaded PECA nanoparticles was quantified using reverse-phase highpressure liquid chromatography (HPLC). The in-vivo pharmacokinetics of insulin in PECA nanoparticles was investigated following I/V administration, and when injected directly into the caecum alone or in conjunction with the permeation enhancer EDTA. Blood samples were collected at intervals from ?5 to 180 minutes, and the concentration of insulin in plasma was quantified using a radioimmunoassay (RIA) validated for possum plasma.

RESULTS: Poly(ethyl 2-cyanoacrylate) nanoparticles were produced with a uniform particle size of 200–300 nm, and the mean entrapment of insulin was 78%. In-vitro release of insulin from the PECA nanoparticles was controlled, although incomplete, and approximately 30% of the insulin remained entrapped. The bioavailability of insulin when administered in a PECA nanoparticulate formulation injected directly into the caecum was <1%, and was not increased by addition of the permeation enhancer.

CONCLUSIONS: The nanoparticulate formulations investigated as part of this study resulted in low bioavailability of the peptide insulin in the brushtail possum.     

芒果皮提取物合成纳米银及抑菌性研究

以芒果皮提取物作为还原剂,采用生物法合成纳米银粒子。研究了合成过程中pH值、芒果皮提取物添加量和硝酸银浓度对合成粒径和形貌的影响,研究结果通过紫外可见分光光度法、X射线衍射和透射电镜进行了表征。通过表征,分析得到合成的纳米银的粒径介于7～27nm之间,其结晶为立体对称晶体,pH值为11,芒果皮提取物的添加量为0.1mL,硝酸银溶液浓度为0.5mM,反应条件为80℃,水浴15min时,合成的纳米银粒径最小。试验结果表明,生物合成的纳米银粒子的抗菌性良好。     

铁酸钴纳米粒子的化学共沉淀法制备及表征

采用弱碱 NH4 HCO3作为沉淀剂,CoCl·6H2 O和 FeCl3·6H2 O作为反应物,在一定摩尔比条件下,通过化学共沉淀法制备了铁酸钴纳米粒子,并采用 FT-IR、XRD、TEM和振动样品磁强计对其进行了表征。结果表明,制得的铁酸钴纳米粒子为尖晶石型结构,其粒径为20 nm左右,并且由于反应过程中弱碱 NH4 HCO3分解产生的CO2和 H2 O 气体对产物的阻隔作用,制得的铁酸钴纳米粒子的分散性较好,饱和磁化强度为55.5 Am2·kg-1。     

Caged Dexamethasone/Quercetin Nanoparticles,Formed of the Morphogenetic Active Inorganic Polyphosphate,are Strong Inducers of MUC5AC

Inorganic polyphosphate (polyP) is a widely distributed polymer found from bacteria to animals, including marine species. This polymer exhibits morphogenetic as well as antiviral activity and releases metabolic energy after enzymatic hydrolysis also in human cells. In the pathogenesis of the coronavirus disease 2019 (COVID-19), the platelets are at the frontline of this syndrome. Platelets release a set of molecules, among them polyP. In addition, the production of airway mucus, the first line of body defense, is impaired in those patients. Therefore, in this study, amorphous nanoparticles of the magnesium salt of polyP (Mg-polyP-NP), matching the size of the coronavirus SARS-CoV-2, were prepared and loaded with the secondary plant metabolite quercetin or with dexamethasone to study their effects on the respiratory epithelium using human alveolar basal epithelial A549 cells as a model. The results revealed that both compounds embedded into the polyP nanoparticles significantly increased the steady-state-expression of the MUC5AC gene. This mucin species is the major mucus glycoprotein present in the secreted gel-forming mucus. The level of gene expression caused by quercetin or with dexamethasone, if caged into polyP NP, is significantly higher compared to the individual drugs alone. Both quercetin and dexamethasone did not impair the growth-supporting effect of polyP on A549 cells even at concentrations of quercetin which are cytotoxic for the cells. A possible mechanism of the effects of the two drugs together with polyP on mucin expression is proposed based on the scavenging of free oxygen species and the generation of ADP/ATP from the polyP, which is needed for the organization of the protective mucin-based mucus layer.     

基于链替代扩增-纳米金比色直观检测单链核酸方法的研究

[目的]将链替代扩增技术与核酸修饰纳米金积聚变色的光学特性相结合,设计了一种新型的直观检测3′端暴露单链核酸的方法,实现对单链核酸的高灵敏度检测。[方法]设计一条含有硫代修饰酶切位点的单链核酸（ZDNA）,其酶切位点5′端是能将核酸修饰纳米金变色的Linker序列,3′端是能与Target单链核酸3′端完全互补的H序列。当无Target存在时,Linker会充分暴露,能使核酸修饰纳米金积聚呈现紫色;但是当Target存在时,会与H序列完全互补,作为ZDNA的引物进入链替代扩增循环,形成的新链不断与Linker序列互补,不能使核酸修饰纳米金积聚呈现红色,从而间接检测了Target单链核酸。[结果]通过一系列试验确定检测体系,具体为：40μl修饰纳米金溶液（0.52 nmol/L）加入10μl酶循环体系（ZDNA20 nmol/L,33 mmol/L Tris-HCl（pH值7.9）;10 mmol/L MgCl2;66 mmol/LNaCl;0.5 mmol/LdNTP;0.1 mg/ml BSA;0.05 U/μl klenow;1 U/μl Hinc II）,直观或紫外检测并绘制Target浓度与纳米金积聚程度的标准曲线,表明Target浓度在1~200 pmol/L的范围内呈现良好的线性关系,R2=0.946,最低检测限是1 pmol/L。[结论]链替代扩增-纳米金比色检测单链核酸方法简便、直观、成本低,与传统修饰纳米金比色检测DNA方法相比,灵敏度提高了104倍。     

Chitosan-Based Anti-Oxidation Delivery Nano-Platform: Applications in the Encapsulation of DHA-Enriched Fish Oil

Refined cobia liver oil is a nutritional supplement (CBLO) that is rich in polyunsaturated fatty acids (PUFAs), such as DHA and EPA; however, PUFAs are prone to oxidation. In this study, the fabrication of chitosan-TPP-encapsulated CBLO nanoparticles (CS@CBLO NPs) was achieved by a two-step method, including emulsification and the ionic gelation of chitosan with sodium tripolyphosphate (TPP). The obtained nanoparticles were inspected by dynamic light scattering (DLS) and showed a positively charged surface with a z-average diameter of between 174 and 456 nm. Thermogravimetric analysis (TGA) results showed the three-stage weight loss trends contributing to the water evaporation, chitosan decomposition, and CBLO decomposition. The loading capacity (LC) and encapsulation efficiency (EE) of the CBLO loading in CS@CBLO NPs were 17.77–33.43% and 25.93–50.27%, respectively. The successful encapsulation of CBLO in CS@CBLO NPs was also confirmed by the Fourier transform infrared (FTIR) spectroscopy and X-ray diffraction (XRD) techniques. The oxidative stability of CBLO and CS@CBLO NPs was monitored by FTIR. As compared to CBLO, CS@CBLO NPs showed less oxidation with a lower generation of hydroperoxides and secondary oxidation products after four weeks of storage. CS@CBLO NPs are composed of two ingredients that are beneficial for health, chitosan and fish oil in a nano powdered fish oil form, with an excellent oxidative stability that will enhance its usage in the functional food and pharmaceutical industries.     

一种适用于SELEX技术的单链DNA制备方法

为了建立一种简单、高效制备高纯度单链DNA方法,用于SELEX技术筛选过程中次级文库的筛选,对制备链霉亲和素磁珠的实验条件进行了优化,确定了对称PCR产物与链霉亲和素磁珠偶联的最佳时间和饱和度,并验证了不同浓度NaOH对解链效果的影响,建立了适于SELEX技术的单链DNA制备体系。利用荧光法和聚丙烯酰胺凝胶电泳对本实验所建立方法及传统不对称PCR法制备单链DNA的回收效率和纯度进行了比较,结果表明在最优实验条件下用磁性分离法制备的单链DNA纯度及回收效率均显著高于不对称PCR法。     

纳米磷酸钙作为猪瘟多肽疫苗佐剂的研究

为了探索纳米磷酸钙(NCaP)作为猪瘟多肽疫苗佐剂的可行性,本研究利用猪瘟多肽疫苗作为抗原,制备纳米磷酸钙吸附猪瘟多肽疫苗。将制备好的疫苗免疫兔后,分别检测兔抗体水平和攻毒后体温变化情况。检测结果显示,纳米磷酸钙对猪瘟多肽吸附率达70%;免疫后纳米磷酸钙佐剂组抗体水平低于弗氏佐剂,但与游离猪瘟多肽疫苗组相比,有较大提高;除弗氏佐剂组外,所有实验兔均有体温升高现象,纳米磷酸钙佐剂组体温升高的时间与空白组相比推迟16 h。结果表明,NCaP作为佐剂相对游离猪瘟多肽可提高其免疫效果,但其诱导的抗体水平低于弗氏佐剂组。     

外源SiNPs对盐胁迫下生姜幼苗生长和生理特性的影响

【目的】研究外源二氧化硅纳米颗粒（SiNPs）对盐胁迫下生姜幼苗生长和生理特性的影响,为SiNPs在生姜抗盐性中的应用提供理论依据与技术支撑。【方法】以竹根姜（Zingiber officinale Roscoe.cv.zhugen）盆栽苗为试材,设置用100 mg/L SiNPs溶液喷施全株5 d后再用蒸馏水浇灌植株根部的SiNP100处理、用蒸馏水喷施全株5 d后再用20 g/L NaCl溶液浇灌植株根部的NaCl处理以及用100 mg/L SiNPs溶液喷施全株5 d后再用20 g/L NaCl溶液浇灌植株根部的SiNP100+NaCl处理,以前后均用蒸馏水处理为对照（CK）。随机选取不同处理后的生姜幼苗,观测其生长形态、光合色素（叶绿素a（Chl a）、叶绿素b（Chl b）、叶绿素a+b（Chl(a+b)）和类胡萝卜素（Car））含量、叶绿素荧光参数（最大光化学效率（F_v/F_m）、实际光量子效率（ΦPSⅡ）、光化学淬灭系数（qP）和非光化学淬灭系数（NPQ））、光合作用参数（净光合速率（P_n）、蒸腾速率（T_r）和气孔导度（G_s））、丙二醛（MDA）和H₂O₂含量以及抗氧化酶（超氧化物歧化酶（SOD）、过氧化物酶（POD）和过氧化氢酶（CAT））活性。【结果】与CK相比,SiNP100处理生姜幼苗的根尖数显著增加,Chl a、Chl b和Chl(a+b)含量显著提高,NPQ显著下降,T_r显著增加,SOD和CAT活性显著升高,MDA和H₂O₂含量显著降低,其余指标均与CK差异不显著。在NaCl胁迫条件下,生姜幼苗根系生长受到抑制,叶片明显变黄,与CK相比,总根长、根系表面积、根尖数显著减少,4种光合色素含量均显著下降,F_v/F_m、ΦPSⅡ和qP显著降低,而NPQ显著升高,P_n、T_r和G_s显著降低,SOD、POD和CAT活性以及MDA和H₂O₂含量均显著升高。SiNP100预处理则可以缓解NaCl胁迫对生姜幼苗的损伤,总根长、根系表面积、根尖数较NaCl处理显著增加,叶片发黄症状也得到缓解;此外,与NaCl处理相比,SiNP100+NaCl处理生姜幼苗的Chl a、Chl b、Chl(a+b)含量和F_v/F_m、ΦPSⅡ、qP、P_n、T_r以及SOD、POD、CAT活性显著提高,而NPQ和MDA、H₂O₂含量显著下降,Car含量和G_s则与NaCl处理差异不显著。【结论】外源SiNPs预处理能通过促进植株根系生长、增加光合色素含量、提高光合作用效率、调节抗氧化酶活性、降低MDA和H₂O₂含量来缓解盐胁迫对生姜幼苗造成的生长抑制和氧化损伤,从而增强生姜植株的抗盐性。