评价荧光原位杂交技术用于临床快速产前诊断唐氏综合征的可行性

目的用羊水细胞培养技术比对荧光原位杂交技术(fluorescence in situ hybridization,FISH),评价FISH用于临床快速产前诊断唐氏综合征的可行性。方法采集124名孕妇18~24周的羊水标本,应用荧光标记21号染色体特殊位点的探针(locus-spe-cific probe,LSI)及X/Y染色体着丝粒探针(centromeric probe,CEP)对未培养的羊水间期细胞进行FISH;同步进行羊水细胞培养,通过染色体核型分析,并以核型分析为标准,对FISH技术进行评价。结果124份标本发生母血污染3例,培养失败1例,将其余120份羊水标本的FISH杂交结果与其染色体核型分析结果进行了比较。FISH分析羊水间期细胞性染色体数目正常者120例(XX 58例、XY 62例);LSI 21探针的FISH结果中21号染色体数目异常者2例,核型分析为典型的21三体,取脐血进行G显带染色体核型分析得以验证为47,XY, 21。产前诊断染色体正常者追踪至分娩,行新生儿外周血染色体检查结果皆为正常核型。结论荧光原位杂交技术可用于羊水间期细胞快速产前诊断唐氏综合征。     

乙型肝炎病毒前S1抗原、HBV-DNA含量与HBeAg的相关性分析

目的分析乙型肝炎病毒前S1抗原、HBV-DNA含量与HBeAg在反映乙肝病毒复制方面的相关性。方法收集HBsAg阳性的血清标本524例,使用ELISA法检测HBV-M(包括HBsAg、HBsAb、HBeAg、HBeAb、HBcAb及前S1抗原)。采用核酸扩增荧光定量(FQ-PCR)检测HBV-DNA。结果HBeAg、前S1抗原及HBV-DNA的阳性率依次是30.5%、54.6%、75.2%,HBeAg阳性率最低,HBV-DNA阳性率最高。结论在反应病毒复制水平上,乙型肝炎病毒前S1抗原、HBeAg与HBV-DNA有一定的相关性,但其检出率并不一致,临床应正确认识三者的检测价值及临床意义,合理选择检测项目。     

乙型肝炎病毒DNA与血清免疫标志物的关系分析

目的 探讨乙型肝炎病毒DNA(HBV DNA)与血清免疫标志物的关系.方法 荧光定量聚合酶链反应法 (FQ-PCR) 检测血清样本中HBV DNA含量;酶联免疫吸附法(ELISA)检测HBV血清标志物.结果 A组(HBsAg和HBeAg阳性)、B组(大三阳)、C组(HBsAg 、HBcAb阳性)和D组(小三阳)的HBV DNA阳性率分别为96.7%、87.6%、52.2%和38.2%,各组之间均有显著性差异(P＜0.05); A组中阳性样本的HBV DNA量显著高于其它组别(P＜0.05); 315份HBV DNA阳性标本全部携带有HBsAg;HBsAg( )HBeAg( )和HBsAg( )HBeAg(-)中HBV DNA阳性率分别为89.1%和44.0%,有显著性差异(P＜0.05).结论 HBsAg是HBV感染灵敏的血清标志物;FQ-PCR定量检测HBV DNA作为血清学方法的补充对乙肝的临床诊断具有重要意义.     

CFDA-SE和Hoechst两种荧光标记技术在嗅鞘细胞移植修复脊髓损伤中的应用

目的将CFDA—SE和Hoechst两种常用的细胞荧光标记技术在嗅鞘细胞移植修复脊髓损伤中的应用进行比较，选出一种更适合的方法进行推广．方法取20只脊髓完全横断的SD大鼠，其中10只移植用CFDA—SE标记的嗅鞘细胞，另10只移植用Hoechst标记的嗅鞘细胞．结果两种标记方法都能成功标记上，但Hoechst对移植后组织的污染率较高，CFDA—SE染色清楚，对受区组织干扰较少．结论CFDA—SE相比Hoechst在嗅鞘细胞移植修复脊髓损伤中进行荧光标记更清楚、更准确，值得推广．     

光谱法研究2-环己氨基噻吩并[2,3-d]嘧啶-4(3H)-酮与牛血清白蛋白的相互作用

目的为新型噻吩并嘧啶的开发与应用提供重要信息。方法应用荧光光谱、紫外可见光谱方法研究了2-环己氨基噻吩并[2,3-d]嘧啶-4(3H)-酮(HJA)与牛血清白蛋白(BSA)的相互作用。结果HJA能强烈猝灭BSA的荧光强度,其荧光猝灭机理为动态猝灭。在此基础上计算了二者相互作用的结合常数、结合位点数及热力学参数等。结论HJA分子与BSA分子以摩尔比1∶1结合,其结合反应主要是熵驱动,主要作用力是疏水力。     

荧光标记法比较4种表面处理方式种植体骨矿化沉积率的研究

目的荧光标记法观察比较4种表面处理方式种植体周围骨矿化沉积率的差异。方法微创拔除4只Beagle犬双侧下颌4颗前磨牙,待拔牙创愈合3个月后,植入奥齿泰种植体40颗,其中机械形态表面(ma-chined morphology,MM)组4颗,喷砂加酸蚀(sand blasted with alumina and acid etched,SA)组、可吸收性研磨介质(resorbable blasting media,RBM)组和生物化学喷砂酸蚀组(biochemistry sand blasted with alumina and acid etched,Bio-SA)组各12颗,愈合3个月,处死前13、14 d和3、4 d分别注射盐酸四环素和钙黄绿素,制取标本并置于70%酒精固定,采用塑料包埋技术制作骨磨片,荧光显微镜下观察,测量骨矿化沉积率。结果螺纹间及远离螺纹区域新骨矿化沉积率在4组间的差异均没有统计学意义(P>0.05);单独比较每组种植体螺纹间及远离区域骨矿化沉积率的差异性,RBM组及SA组组内差异没有统计学意义(P>0.05),而MM组和Bio-SA组组内差异具有统计学意义(P<0.05)。结论采用塑料包埋技术,通过荧光标记法能成功观察到新骨矿化沉积的情况;种植体植入3个月后,4种不同粗糙程度的种植体螺纹间及远离螺纹区域骨矿化沉积率没有差异性,而经机械形态表面处理的种植体和生物化学喷砂酸蚀处理的种植体,螺纹间骨矿化沉积比远离螺纹区域快。     

Comparative biodistribution in mice of cyanine dyes loaded in lipid nanoparticles

Two near infrared cyanine dyes, DiD (1,1′-dioctadecyl-3,3,3′,3′-tetramethylindotricarbocyanine perchlorate) and ICG (Indocyanine Green) were loaded in lipid nanoparticles (LNP). DiD-LNP and ICG-LNP presented similar physicochemical characteristics (hydrodynamic diameter, polydispersity, zeta potential), encapsulation efficiency, and colloidal stability when stored in PBS buffer. However, whereas DiD had similar biodistribution than cholesteryl-1-¹⁴C-oleate ([¹⁴C]CHO, a constituent of the nanoparticle used as a reference radiotracer), ICG displayed a different biodistribution pattern, similar to that of the free dye, indicative of its immediate leakage from the nanovector after blood injection. NMR spectroscopy using Proton NOE (Nuclear Overhauser Effect) measurements showed that the localization of the dye in the lipid nanoparticles was slightly different: ICG, more amphiphilic than DiD, was found both inside the lipid core and at particle interface, whereas DiD, more hydrophobic, appeared exclusively located inside the particle core. The ICG release rate from the particles was 7% per 1 month under storage conditions (4 °C, dark, 10% of lipids), whereas no leakage could be detected for DiD. ICG leakage increased considerably in the presence of BSA 40 g/L (45% leakage in 24 h at 100 mg/mL of lipids), because of the high affinity of the fluorophore for plasma proteins. On the contrary, no DiD leakage was observed, until high dilution of the nanoparticles which triggered their dissociation (45% leakage in 24 h at 1 mg/mL of lipids). Altogether, the subtle difference in dye localization into the nanoparticles, the partial dissociation of the LNP in diluted media, and more importantly the high ICG affinity for plasma proteins, accounted for the differences observed in the fluorescence biodistribution after tail vein injection of the dye-loaded nanoparticles.     

Analysis of corticosteroid and antiepileptic drug treatment effects on heme biosynthesis mRNA expression in lower-grade gliomas: Potential implications for 5-ALA metabolization

ObjectivesIntraoperative visualization of gliomas with 5-aminolevulinic acid (5-ALA) induced fluorescence constitutes a powerful technique. While visible fluorescence is typically observed in high-grade gliomas, fluorescence is considerably less common in lower-grade gliomas (LGGs) WHO grade II&III. Whereas the exact mechanisms determining fluorescence in LGGs are not fully understood, metabolization of non-fluorescent 5-ALA to fluorescent Protoporphyrin IX by specific heme biosynthesis enzymes/transporters has been identified as relevant mechanism influencing fluorescence behavior. Furthermore, recent in-vitro studies have suggested preoperative treatment with corticosteroids and anti-epileptic drugs (AED) as potential factors influencing 5-ALA induced fluorescence.MethodsThe goal of this study was thus to investigate the effect of preoperative corticosteroid/AED treatment on heme biosynthesis mRNA expression in a clinically relevant patient population. For this purpose, we analyzed the mRNA expression levels of specific heme biosynthesis factors including ALAD, HMBS, UROS, UROD, CPOX, PPOX, FECH, ABCB6, ACG2, SLC15A1 and SLC15A2, ABCB1, ABCB10 in a cohort of LGGs from “The Cancer Genome Atlas”.ResultsAltogether, 403 patients with available data on preoperative corticosteroid/AED treatment and heme biosynthesis mRNA expression were identified. Regarding corticosteroid treatment, no significant differences in expression of any of the 11 investigated heme biosynthesis factors were found. In contrast, a marginal yet statistically significant increase in SLC15A1 levels and decrease in ABCB6 levels were observed in patients with preoperative AED treatment.ConclusionWhile no significant differences in heme biosynthesis mRNA expression were observed according to preoperative corticosteroid treatment, changes in SLC15A1 as well as ABCB6 expression were detected in patients treated with AED. However, since these alterations were minor and have opposing effects on 5-ALA metabolization, our findings do not support a distinct effect of AED and corticosteroid treatment on heme biosynthesis regulation in LGGs.