Development of vaccine for the trypanosomiasis by using glycosphingolipids antigen.

利用鞘糖脂抗原开发锥虫病疫苗。

• 批准号: 13660325
• 负责人: WATARAI Shinobu
• 金额: $ 2.24万
• 依托单位: Osaka Prefecture University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13660325/
• 关键词: Protozoan disease; Trypanosomiasls; Vaccine; Glycosphingolipids; Gangliosides; Carbohydrate antigen; Liposomes; Monoclonal antibody; Trypanosoma brucei

To identify the novel antigens that can effectively induce the immune response in order to develop an effective vaccine for the prevention of trypanosomiasis, neutral glycosphingolipids (GSLs) and gangliosdes were isolated from Trypanosoma brucei and analyzed by thin-layer chromatography (TLC) and TLC/secondary ion mass spectrometry (TLC/SIMS) or TLC immunostaining test. Three species of neutral GSLs, N-1, N-2 and N-3, were separated on TLC. The TLC/SIMS analysis of N-1 of the parasites revealed a series of (M-H)^- ions from m/z 698 to m/z 825 representing the molecular mass range of ceramide monohexoside (CMH) (GlcCer or galactosylceramide). On the other hand, the TLC/SIMS spectra of N-2 GSL revealed a series of (M-H)^- ions from m/z 944 to m/z 987 indicating the molecular mass range of LacCer. In the TLC/SIMS analysis of N-3 GSL, however, the characteristic molecular ions that can elucidate the structure of N-3 GSL were not obtained. Both GlcCer and LacCer were detected on the cell s … More urface of T. brucei by antibodies. In ganglioside fraction, four species of gangliosides, G-1, G-2, G-3 and G-4, were separated on TLC. G-1 ganglioside showed the reactivity to the anti-GM3 monoclonal antibody. G-2 was recognized by the anti-GM1 monoclonal antibody. G-3 gave reaction with the monoclonal antibody to GD1a G-4 had the reactivity to anti-GD1b monoclonal antibody. GM3, GM1, GD1a and GD1b were detected on the cell surface of T. brucei by using 4 kinds of monodonal antibodies. Furthermore, to investigate whether immune response against GSLs on T. brucei is effective against the inhibition of T. brucei infection, BALB/c mice were immunized with liposome containing ganglioside mixture (GM1, GD1a and GD1b) and challenged with T. brucei. All of control (non-immunized) mice died 6 days after challenge. However, none of mice immunized with ganglioside mixture-containing liposome died 40 days after challenge. In addition, no parasites were detectable in survived mice. These results suggest that induction of immune response against gangliosides on T. brucei would be effective for the prevention of T. brucei infection. Less

为了鉴定能有效诱导免疫应答的新型抗原，以开发预防锥虫病的有效疫苗，从布鲁氏锥虫中分离中性鞘糖脂（GSLs）和神经节苷脂，并采用薄层色谱（TLC）、TLC/次级离子质谱（TLC/SIMS）或TLC免疫染色试验进行分析。在TLC上分离出中性GSLs N-1、N-2和N-3。对N-1的TLC/SIMS分析显示，m/z 698 ~ m/z 825的一系列（m - h）^-离子代表了神经酰胺单己糖（CMH） （glcer或半乳糖神经酰胺）的分子质量范围。另一方面，N-2 GSL的TLC/SIMS光谱显示了一系列（m - h）^-离子，从m/z 944到m/z 987，表明LacCer的分子质量范围。然而，在N-3 GSL的TLC/SIMS分析中，并没有获得能够阐明N-3 GSL结构的特征分子离子。glcer和LacCer均在布鲁氏t细胞表面被抗体检测到。在神经节苷脂组分中，通过薄层色谱分离出G-1、G-2、G-3和G-4 4种神经节苷脂。G-1神经节苷脂对抗gm3单克隆抗体具有反应性。G-2被抗gm1单克隆抗体识别。G-3与抗gd1b单克隆抗体反应，G-4对抗gd1b单克隆抗体有反应性。采用4种单抗检测布鲁氏体细胞表面GM3、GM1、GD1a和GD1b。此外，为了研究GSLs对布鲁氏t的免疫反应是否有效抑制布鲁氏t感染，我们用含有神经节苷脂混合物（GM1， GD1a和GD1b）的脂质体免疫BALB/c小鼠，并用布鲁氏t攻击。对照组（未免疫）小鼠在攻击后6天全部死亡。然而，用含有神经节脂苷脂质体的混合物免疫的小鼠在攻击后40天没有死亡。此外，在存活的小鼠中没有检测到寄生虫。提示诱导机体对神经节苷类的免疫应答可有效预防布氏体感染。少

项目成果

Tana: "Induction of intestinal IgA and IgG antibodies preventing adhesion of verotoxin-producing Escherichia coli to Caco-2 cells by oral immunization with liposomes."Lett. Appl. Microbiol.. 36. 135-139 (2003)

Tana：“通过脂质体口服免疫诱导肠道 IgA 和 IgG 抗体，防止产生维罗毒素的大肠杆菌粘附到 Caco-2 细胞。”

Watarai, S., et al.: "Isolation and characterization of gangliosides from Trypanosoma brucei"Vet. Parasitol.. (発表予定).

Watarai, S. 等人：“布氏锥虫神经节苷脂的分离和表征”Vet Parasitol..（待出版）。

Watarai, S.: "Inhibitiry effect of anti-ganglioside antibodies on in vitro and in vivo growth of Trypanosoma brucei."Vet. Parasitol.. manuscript in preparation..

Watarai, S.：“抗神经节苷脂抗体对布氏锥虫体外和体内生长的抑制作用。”兽医。

Watarai, S., et al.: "Protective effect of anti-ganglioside antibodies against experimental Trypanosomes brucei infection in mice"Vet. Parasitol.. (発表予定).

Watarai, S. 等人：“抗神经节苷脂抗体对小鼠实验性布氏锥虫感染的保护作用”Vet Parasitol..（待提交）。

Watarai, S.: "Protective effect of anti-ganglioside antibodies against experimental Trypanosoma brucei infection in mice."Vet. Parasitol.. manuscript in preparation..

Watarai, S.：“抗神经节苷脂抗体对小鼠实验性布氏锥虫感染的保护作用。”兽医。