胃黏膜局部免疫反应在以壳聚糖为佐剂的幽门螺杆菌疫苗免疫保护中的作用  

The role of local immune response in gastric mucosa in the protection induced by Helicobacter. pylori vaccine with chitosan as adjuvant

作　　者：谢勇[1] 龚燕锋[1] 周南进[2] 陈江[1] 周小江[1] 吕农华[1] 王崇文[1] 

机构地区：[1]江西医学院第一附属医院消化研究所,南昌330006 [2]江西省医学科学研究所

出　　处：《中华医学杂志》2005年第37期2629-2635,共7页National Medical Journal of China

基　　金：国家自然科学基金资助项目(30460052);江西省学科带头人培养项目基金资助(K010501);江西省卫生厅重点招标项目基金资助(031070);江西省教育厅基金资助项目

摘　　要：目的探讨以壳聚糖为佐剂的幽门螺杆菌(Hp)疫苗的免疫保护作用及其机理。方法BALB/C小鼠随机分为9组:①空白对照组;②壳聚糖酸溶液组;③壳聚糖颗粒组;④Hp抗原组;⑤Hp抗原+壳聚糖酸溶液组;⑥Hp抗原+壳聚糖颗粒组;⑦Hp抗原+霍乱毒素(CT)组;⑧Hp抗原+壳聚糖酸溶液+霍乱毒素组;⑨Hp抗原+壳聚糖颗粒+霍乱毒素组,各组于第0、7、14、21天灌胃各免疫1次,免疫后4周给予1×109/ml的SS1Hp菌液0.5ml/只进行攻击,隔日1次,共2次。4周后,采用定量Hp培养和病理改良Giemsa染色法检测胃黏膜内Hp感染。用酶联免疫吸附实验(ELISA)检测唾液和胃黏膜内抗HpIgA,用定量ELISA法检测胃黏膜内Th1和Th2细胞因子,用SP免疫组织化学法检测胃黏膜内分泌型IgA(sIgA)。结果(1)以壳聚糖为佐剂的Hp疫苗的免疫保护率达60%,与以霍乱毒素为佐剂的Hp疫苗的免疫保护率(58.33%)相似,显著高于单纯Hp抗原组及其他不含Hp抗原组(P<0.01或P<0.05),同时以霍乱毒素+壳聚糖为佐剂的Hp疫苗的保护率为84.62%、85.71%,其Hp的定植评分显著低于无佐剂组及以霍乱毒素为佐剂组(P<0.01或P<0.05)。(2)胃黏膜内sIgA及特异性抗HpIgA水平在壳聚糖为佐剂组与以霍乱毒素为佐剂组无差别(P>0.05),显著高于无佐剂组,而壳聚糖与霍乱毒素联合应用组显著高于单以霍乱毒素为佐剂组(P<0.01或P<0.05)。(3)胃黏膜内白细胞介素(IL)2含量在攻击前各组间差异无统计学意义(P>0.05),攻击后含佐剂组显著高于对照组(P<0.01或P<0.05),且攻击后含Hp抗原各组均显著高于攻击前(P<0.05)。(4)胃黏膜内IL10含量攻击前以壳聚糖为佐剂组显著高于无佐剂组(P<0.01或P<0.05),攻击后各组之间差异无统计学意义(P>0.05),且攻击后含佐剂组均显著低于攻击前(P<0.01)。(5)胃黏膜内IL4含量攻击前以壳聚糖为佐剂组显著高于无佐剂组(P<0.05),攻击后以壳聚糖颗粒为佐剂组显著高于以霍乱毒素为佐剂组(Objective To study the immunological protection of Helicobacter pylori （H. pylori ） vaccine with ehitosan as adjuvant and it＇s mechanism. Methods One-grade female BALB/c mice were randomly divided into nine groups and immunized by ①PBS alone,②ehitosan solution alone, ③ehitosan particles alone, ④H. pylori antigen alone, ⑤H. pylori antigen plus ehitosan solution, ⑥H. pylori antigen plus chitosan particles, ⑦H. pylori antigen plus CT, ⑧H. pylori antigen plus chitosan solution and cholera toxin （CT）, ⑨H. pylori antigen plus chitosan particles and CT orally respectively once a week for four weeks. At 4 weeks after the last immunization, these mice were challenged by alive H. pylori（ 1 × 10^9/ml） twice at two days intervals. At 4 weeks after the last challenge, these mice were all killed and Gastric mucosa were embedded in paraffin, sectioned and assayed with Giemsa stain. The other gastric mucosa were used to quantitatively culture H. pylori. An ELISA was used to detect anti-H, pylori IgA in saliva and gastric mucosa and a quantitative ELISA was used to detect IL-2, IL-4, IL-10 content in gastric mucosa, and SP immunohistochemical method was used to detect secretory immunoglobulin A （slgA） in gastric mucosa. Results （1） In the groups with chitosan as adjuvant, 60% mice could achieve immunological protection, which was according to that with CT as adjuvant （58.33%） , and was significantly higher than H. pylori antigen alone and other groups without H. pylori antigen （P 〈 0. 01 or P 〈 0. 05 ） . While the rates of protection in the groups with chitosan plus CT as adjuvant were 84. 62%, 85.71% and the H. pylori colonization score in it was significantly lower than that in the groups with CT as adjuvant and without adjuvants（P 〈0. 01 or P 〈0.05）. （2）the labeling index for slgA-positive lumen of glands and special antiH. pylori IgA levels in gastric mucosa in the groups with chitosan as an adjuvant had no difference with those in the group with CT as an adjuv

关 键 词：胃黏膜 免疫反应 壳聚糖 佐剂 幽门螺杆菌 疫苗 免疫保护 

分 类 号：R392[医药卫生—免疫学]