Advances in osmotic opening of the blood-brain barrier to enhance CNS chemotherapy

The blood-brain barrier (BBB) to water-soluble drugs and macromolecules can be opened in vivo by infusing a hypertonic solution of arabinose or mannitol into the carotid artery for 30 sec. Opening involves widening of tight junctions between endothelial cells of the cerebrovasculature and is mediated by endothelial cell shrinkage, vascular dilatation associated with removal of water from brain, and modulation of the contractile state of the endothelial cytoskeleton and junctional proteins by increased intracellular calcium. A 10-fold increase in BBB permeability to intravascular substances, lasting about 10 min following osmotic exposure, reflects both increased diffusion and bulk fluid flow from blood into brain. Furthermore, recent evidence indicates that the duration of peak BBB opening can be extended beyond 30 min, by pre-treatment with a Na⁺/Ca²⁺ channel blocker. In experimental animals, the osmotic method has been used to grant wide access to brain of water-soluble drugs, peptides, antibodies, boron compounds for neutron capture therapy, viral vectors for gene therapy and enzymes. Ongoing multi-centre clinical studies suggest that the method, when used with intra-arterially administered anticancer drugs, can prolong survival in patients with malignant brain tumours, with minimal morbidity. However, controlled clinical trials are critical to see if the osmotic procedure with intra-arterial drugs enhances survival in brain tumour patients compared with intra-arterial drug alone.     

A monoclonal antibody to the galactan from Helix pomatia snails recognizes beta-(1,6)-linked D-galactose residues

Using the hybridoma technique, a series of four monoclonal antibodies to the galactan isolated from albumin glands of wine yard snails (Helix pomatia) could be generated. Characterization of the binding properties of one of these antibodies revealed a specificity for beta-(1,6)-glycosidically linked D-galactose residues. This could be demonstrated by reaction of the antibody with beta-(1,6)-D-galactans and by the D-galactose-inhibitable binding to group B streptococci of type II, which possess beta-(1,6)-linked D-galactose as immuno-determinant group.     

UPLC-MS/MS测定参芎葡萄糖注射液中阿拉伯糖、果糖、甘露糖和葡萄糖

目的:建立超高效液相色谱-三重四级杆质谱联用仪(UPLC-MS/MS)法测定参芎葡萄糖注射液中阿拉伯糖、果糖、甘露糖、葡萄糖的含量测定方法,为科学评价参芎葡萄糖注射液的质量提供依据,为保障临床安全用药奠定基础。方法:采用国产GDX-403固相萃取柱净化,Waters ACQUITY UPLC BEH Xbridge Amide色谱柱(2.1 mm×100 mm,1.7μm),柱温35℃进行分析;[(0.1%氨水-乙腈)-(0.1%氨水-水)](85∶15)为流动相进行分离,流速0.3 mL·min-1,以超高效液相色谱-三重四级杆质谱联用仪(UPLC-MS/MS)测定参芎葡萄糖注射液中阿拉伯糖、甘露糖、果糖和葡萄糖含量。结果:建立了参芎葡萄糖注射液中阿拉伯糖、果糖、甘露糖、葡萄糖的含量测定方法,所测定的样品中和混合对照品中阿拉伯糖、果糖、甘露糖、葡萄糖相应的浓度范围与峰面积呈良好的线性关系,重复性、精密度良好,加样回收率分别为98.43%,102.13%,100.72%,101.75%,其RSD分别为2.4%和1.3%,3.1%,2.7%。5个批次参芎葡萄糖注射液中阿拉伯糖和甘露糖含量稳定。结论:该操作方法简便,专属性强,与总糖的含量测定相比更加科学稳定,可用于参芎葡萄糖注射液的质量控制。     

人参囊泡调控肿瘤免疫微环境的活性成分研究

目的 探讨人参Panx ginseng来源的细胞外囊泡（ginseng-derived nanoparticles,GDNPs）在调控肿瘤相关巨噬细胞表型及抑制黑色素瘤生长方面的潜在物质基础。方法 采用纳米颗粒跟踪分析仪、透射电镜测试及紫外-可见分光光度法全面表征GDNPs及其主要成分（蛋白质、多糖和皂苷）的含量;通过qRT-PCR和流式细胞术检测GDNPs与其含有的多糖、皂苷成分对骨髓来源巨噬细胞（bone marrow-derived macrophage,BMDM）表型的调控影响。收集不同极化程度的BMDM条件培养基（conditional medium,CM）孵育小鼠皮肤黑色素瘤B16F10细胞,CCK-8法测定不同CM处理后B16F10细胞活力,验证活性成分对肿瘤免疫微环境的调控作用;通过PMP柱前衍生法定量分析GDNPs活性成分组成。结果 透射电镜下观察GDNPs形态结构良好,含量测定结果为2.46×10¹¹颗粒的GDNPs含有4.31 mg蛋白质、4.46 mg多糖、1.22 mg皂苷。qRT-PCR和流式细胞术实验结果显示GDNPs多糖可以逆转M2型巨噬细胞表型,向M1方向极化。GDNPs多糖诱导巨噬细胞极化后的CM显著抑制了B16F10细胞活力。PMP柱前衍生法分析GDNPs多糖成分由葡萄糖、半乳糖、阿拉伯糖组成,其物质的量比为4.72∶1.07∶2.15。结论 揭示了GDNPs中多糖成分在调控肿瘤免疫微环境的关键作用,为进一步的机制研究和临床应用提供了实验依据。     

急支糖浆多糖的成分分析及质量评价

目的 研究急支糖浆多糖组分的化学成分组成、重均相对分子质量以及单糖组成,并比较不同批次急支糖浆多糖组分,为其质量控制提供依据。方法 采用比色法、高效凝胶色谱法(high performance gel permeation chromatography,HPGPC)和高效阴离子交换色谱法(high performance anion exchange chromatography,HPAEC)测定急支糖浆多糖的基本化学组成、重均相对分子质量及分散系数、单糖组成。结果 15批急支糖浆多糖的中性多糖、糖醛酸和蛋白质的平均质量分数分别为47.60%、33.38%、9.30%;重均相对分子质量及分散系数分别为11 757~26 367和1.89~2.65。15批急支糖浆多糖中均含有岩藻糖、鼠李糖、阿拉伯糖、半乳糖、葡萄糖、甘露糖、木糖、半乳糖醛酸及葡萄糖醛酸,平均质量分数分别为1.94、20.13、115.54、99.25、117.45、13.62、6.03、215.23、9.61 µg/mg,物质的量比结果为1.5:14.4:100.0:71.6:84.7:9.8:5.2:144.1:6.4。将急支糖浆与川贝枇杷糖浆、杏苏止咳糖浆、小儿热速清糖浆和小儿肺热咳喘口服液进行比较,结果发现单糖组成物质的量比结合HPAEC指纹图谱可用于急支糖浆多糖的质量控制。结论 15批急支糖浆多糖的化学组成、重均相对分子质量及单糖组成具有较高的一致性,HPAEC指纹图谱具有特异性,可用作急支糖浆多糖的质控指标。     

党参多糖单糖组成与其对HepG2细胞毒活性的相关分析

目的探究党参多糖的单糖组成与其对肝癌HepG2细胞毒活性之间的相关性。方法 26批不同产地的党参样品,采用水提醇沉法提取多糖,苯酚硫酸法测多糖的量、气相色谱法和间羟基联苯法同时测定半乳糖醛酸的量、糖腈乙酸酯衍生化法分析单糖种类及量、三甲基硅醚衍生化法测果糖的量,MTT法研究党参多糖对HepG2细胞的细胞毒活性,采用聚类分析法对26批党参样品进行聚类分析,以偏最小二乘法(PLS)探究样品中各单糖种类和量与其对HepG2细胞毒活性的相关性。结果 26批党参多糖样品均显示一定的HepG2细胞毒活性,且以甘肃文县的8号党参多糖的细胞毒活性最强(其抑制率为36.36%)。26批党参多糖中各类单糖的量存在差异。聚类分析结果表明,党参多糖的单糖种类及量不能作为党参分类的指标。党参多糖的单糖组成与HepG2细胞毒活性的相关性研究表明,半乳糖醛酸、鼠李糖、阿拉伯糖、半乳糖、果糖与HepG2细胞毒活性呈正相关,而甘露糖、木糖和葡萄糖与HepG2细胞毒活性呈负相关。结论党参多糖对HepG2的细胞毒活性与其单糖的种类存在相关性,含有半乳糖醛酸相对较高的党参多糖的细胞毒活性较强。     

An acidic glycoconjugate from Lythrum salicaria L. with controversial effects on haemostasis

Aim of the study

Lythrum salicaria L. belongs to the small Lythraceae family of 22 genera, which range in habit from herbs to shrubs and trees found with worldwide distribution (Heywood, 1993). The generic name of Lythrum derived from Greek “luthron”—blood, possibly referring to the color of the flowers or to the one of its herbal use as an astringent to stop bleeding ( [26], [17] and [18]). The flowering parts and the flowering branch tips are used in traditional medicine and pharmaceuticals internally in a form of decoctions or as extracts for treatment of diarrhea, chronic intestinal catarrhs, hemorrhoids and eczema, or externally to treat varicose veins, venous insufficiency and gums ( [Mantle et al., 2000] and [Rauha et al., 2000]). The aim of this study was to isolate the plant glycoconjugate from flowering parts of Lythrum salicaria, and to verify its influence on blood coagulation process.

Materials and methods

From the air-dried flowering parts of this plant a water-soluble glycoconjugate has been isolated by hot alkaline extraction followed by neutralization and purification by multi-steps extraction with organic solvents, dialysis and concentration. The plant isolate was tested in vitro on anticoagulant activity on human plasma, and on Wistar rats blood system in vivo as well as ex vivo.

Results

A dark brown isolate was obtained in the yield of 8% of starting material (w/w) as a macromolecular compound with M_w ∼ 12,500. Chemical analysis revealed the presence of carbohydrates (30%), phenolics (1 g contained 1.2 mM of gallic acid equivalent) and proteins (0.8%). The result of compositional analyses of carbohydrate part revealed the predominance of uronic acids (∼66%), galactose (∼12%), rhamnose (∼10%) and arabinose (∼9%) residues indicating thus the presence of pectic type of polymers, i.e. galacturonan and/or rhamnogalacturonan associated with arabinogalactan in Lythrum glycoconjugate. In vitro and ex vivo experiments showed complete inhibition of plasma clot formation, however, the application of Lythrum glycoconjugate in vivo showed controversial effect on animal blood system in comparison with in vitro ones, i.e. pro-coagulant activity.

Conclusion

The in vivo results give a scientific explanation for the traditional use of Lythrum salicaria as a styptic agent. It seems that pro-coagulant activity of this complex could be probably connected with the other factors in blood circulation system, like platelets.     

防风中1个新多糖的结构及免疫调节活性研究

目的 对防风Saposhnikovia divaricata中分离得到均一多糖的结构及其免疫调节活性进行研究。方法 采用DEAE-纤维素、Sephadex G-75等柱色谱法,对防风多糖进行系统分离纯化,采用ESI-MSⁿ、GC-MS、NMR等谱学技术,对分离得到的防风多糖SP800203的相对分子质量分布、单糖组成、寡糖片段、糖残基类型和糖苷键连接方式等进行分析,确定其结构。采用细胞实验和斑马鱼实验,研究防风多糖SP800203的免疫调节活性。结果 从防风中分离得到防风多糖SP800203,糖醛酸质量分数为75.73%,相对分子质量约为7.14×10⁴,由鼠李糖、半乳糖、阿拉伯糖和半乳糖醛酸组成,单糖物质的量比为2.8∶6.7∶6∶84.5。主链由半乳糖醛酸聚合而成,2条支链分别由阿拉伯糖、半乳糖醛酸、半乳糖、鼠李糖,以及阿拉伯糖、半乳糖醛酸、半乳糖聚合而成,二者分别通过β、α糖苷键与主链相连。防风多糖SP800203可以促进巨噬细胞一氧化氮、肿瘤坏死因子-α、白介素-1β和白介素-6的释放,增加斑马鱼的免疫细胞密度和巨噬细胞数量。结论 防风多糖SP800203为从防风中分离得到的新的均一多糖,具有良好的免疫调节活性;研究结果为阐明防风免疫调节作用机制奠定了基础,为防风临床应用及进一步研究与开发提供科学依据。     

鼠伤寒沙门氏菌阿拉伯糖抗性正向突变试验的改进

改进了鼠伤寒沙门氏菌(SV50)阿拉伯糖抗性正向突变试验的细菌培养和实验条件,并对影响实验的因素进行了探索。用LB培养基代替DM培养基,振荡培养(90rpm)3-4h,控制因细菌浓度在1×10~7/平板左右,能够获得较稳定的自发突变值(75～149/平板)和较高的诱变比值(MR)。软琼脂培养基中加入一定量葡萄糖或色氨酸,能够提高实验的敏感性,增加组氨酸或苏氨酸或尿嘧啶的量对实验无干扰作用。本试验适合于初筛化学物的诱变性,并可试用于体液及食品的诱变性检测。