CpG寡核苷酸和霍乱毒素联合STAg滴鼻免疫小鼠诱导的弓形虫特异性免疫应答

发布时间：2018-01-14 07:07

  本文关键词：CpG寡核苷酸和霍乱毒素联合STAg滴鼻免疫小鼠诱导的弓形虫特异性免疫应答　出处：《山西医科大学》2006年硕士论文　论文类型：学位论文

  更多相关文章： STAg CpG ODN CT 刚地弓形虫 黏膜免疫 黏膜佐剂 IgG抗体 IgA抗体

【摘要】： 目的确定CpG寡核苷酸(CpG oligonucleotide,CpG ODN)作为可溶性速殖子抗原(soluble tachyzoite antigen,STAg)的最佳剂量,观察最佳剂量的CpG ODN和STAg鼻内免疫小鼠所诱导的免疫应答的持续时间,以及CpG ODN和霍乱毒素(cholera toxin,CT)联合佐剂的协同作用,弓形虫复合黏膜疫苗研制提供理论和实验依据。 方法第一部分:5～6周龄BALB/c小鼠50只随机分为5组,每组10只,实验组以0、1、5、10μg的CpG ODN联合20μg STAg滴鼻免疫小鼠,对照组以PBS滴鼻,间隔2周免疫2次。末次免疫后30d处死全部小鼠,观察脾、Peyer’s patch(PP)、肠系膜淋巴结(MLN)和IEL淋巴细胞增殖情况。检测血清IgG和粪便IgA含量。 第二部分:雌性BALB/c小鼠72只随机分为实验组和对照组,每组36只。实验组以10μg CpG ODN联合20μg STAg滴鼻免疫,对照组以PBS滴鼻。间隔2周免疫2次,于末次免疫后第1、2、3、4、5、6周分别随机处死小鼠(6只/组)。测定小肠冲洗液、鼻咽冲洗液、肺冲洗液、阴道冲洗液IgA和血清IgG。分离脾和IEL进行淋巴细胞计数。 第三部分:5～6周龄BALB/c小鼠30只,雌雄各半,随机分为4个实验组和PBS对照组。实验组分别用20μg STAg、20μg STAg+10μg CpG ODN、20μg STAg+ 1μgCT、20μg STAg+ 1μgCT +10μg CpG ODN滴鼻免疫,间隔2周免疫2次。对照组用PBS以相同程序免疫。末次免疫后第20d处死小鼠。收集血清和肠冲洗液,用ELISA法检测血清弓形虫特异性IgG和肠液IgA含量。分离脾淋巴细胞,用MTT法检测淋巴细胞增殖活性。 结果不同剂量的CpG ODN联合STAg滴鼻免疫小鼠后,10μg CpG ODN组小鼠肠相关淋巴组织PP、MLN和IEL以及脾淋巴细胞增殖明显(P0.05)。10μg组血清IgG远高于对照组(P0.05),粪便IgA高于对照组(P0.05)。 10μg CpG ODN联合STAg滴鼻免疫小鼠后脾淋巴细胞和IEL增生明显。脾淋巴细胞在第2、3、4、5周显著高于对照组(P 0.001)。IEL数量分别在第2、4周出现两个峰值,第2、3、4周显著高于对照组(P 0.001)。实验组小鼠各冲洗液中IgA平均水平高于对照组。小肠冲洗液中弓形虫特异性IgA抗体水平在各个时间点均高于对照组。与对照组相比,第2、3周(P 0.001),4周(P 0.05)有显著差异。鼻咽冲洗液中IgA水平在第1周时最高,各时间点虽高于对照组,
[Abstract]:Objective to determine the CpG oligonucleotide CpG oligonucleotide. CpG ODN was the best dosage of soluble tachyzoite antigen STAG. The duration of immune response induced by intranasal immunization with CpG ODN and STAg, and the duration of CpG ODN and cholera toxin were observed. CT) combined with adjuvant can provide theoretical and experimental basis for the development of Toxoplasma gondii complex mucosal vaccine. Methods in the first part, 50 BALB/c mice aged 6 weeks were randomly divided into 5 groups with 10 mice in each group. Mice were immunized with 10 渭 g CpG ODN combined with 20 渭 g STAg. The control group was immunized with PBS twice every 2 weeks. All the mice were killed 30 days after the last immunization and the spleen was observed. The proliferation of Peyer's, mesenteric lymph node (MLN) and IEL lymphocytes were detected. The levels of serum IgG and stool IgA were measured. The second part: 72 female BALB/c mice were randomly divided into experimental group (n = 36) and control group (n = 36). The experimental group was immunized with 10 渭 g CpG ODN and 20 渭 g STAg. The control group was immunized with PBS nasal drip twice every 2 weeks. The mice were killed at random for 6 weeks at the first week after the last immunization. The small intestine flushing fluid and nasopharyngeal lavage fluid were measured. Lung lavage fluid, vaginal lavage fluid IgA and serum IgG. spleen and IEL were separated for lymphocyte count. Part three: 30 BALB/c mice, half male and half female, were randomly divided into 4 experimental groups and PBS control groups. The experimental groups were given 20 渭 g STAg respectively. 20 渭 g STAg, 10 渭 g CpG, 20 渭 g STAg, 1 渭 gCT. 20 渭 g STAg 1 渭 gCT 10 渭 g CpG ODN were intranasal immunized. The control group was immunized with PBS in the same procedure. The mice were killed on the 20th day after the last immunization. Serum and intestinal irrigation fluid were collected. Serum Toxoplasma gondii specific IgG and intestinal fluid IgA were detected by ELISA method, spleen lymphocytes were isolated and lymphocyte proliferative activity was detected by MTT method. Results 10 渭 g CpG ODN mice were immunized with different doses of CpG ODN combined with STAg nasal drip. The levels of serum IgG in MLN and IEL and splenic lymphocyte proliferation were significantly higher in P0.05 渭 g group than in control group (P0.05 渭 g), and in fecal IgA was higher than that in control group (P0.05 渭 g). The splenic lymphocytes and IEL proliferation of mice immunized with 10 渭 g CpG ODN combined with STAg nasal drip were obvious. At 5 weeks, the number of IEL was significantly higher than that of the control group (P 0.001). The number of IEL had two peaks and two peaks at the 4th week and the third peak value at the 2nd week, respectively. At 4 weeks, it was significantly higher than that in control group (P 0.001). The average level of IgA in each flushing fluid of experimental group was higher than that of control group, and the level of Toxoplasma gondii specific IgA antibody in small intestinal lavage fluid was higher than that in control group at all time points. The level of IgA in nasopharyngeal lavage fluid was the highest at the first week, although it was higher than that in the control group at each time point.
【学位授予单位】：山西医科大学
【学位级别】：硕士
【学位授予年份】：2006
【分类号】：R392

【参考文献】

相关期刊论文 前10条

1 韩剑峰,殷国荣;弓形虫诱导的黏膜细胞免疫应答研究进展[J];免疫学杂志;2003年05期

2 黄炳成,陈少卿,付婷霞,魏庆宽,李瑾,胡迎新;不同佐剂的弓形虫亚单位疫苗免疫效果观察[J];中国热带医学;2004年01期

3 韩剑峰,殷国荣,刘红丽,申金雁,元海军,沈[?琼;经口感染弓形虫BALB/c小鼠血清及肠道分泌物中IgA抗体含量的检测[J];热带医学杂志;2004年01期

4 殷国荣,元海军,杨亚波,周永安,沈[?琼,孟晓丽;TSo联合IFN-γ鼻内免疫小鼠诱导的弓形虫IgA抗体特异性免疫应答[J];热带医学杂志;2004年06期

5 刘红丽,殷国荣,沈，

本文编号：1422527