论文部分内容阅读
目的探讨制备脂质体包裹重组幽门螺杆菌(Hp)热休克蛋白60(Hsp60)口服疫苗的方法,并用Hp感染的小鼠模型评价其在预防Hp感染中的作用。方法将PET-22(+)/Hsp60在BL21(DE3)大肠杆菌表达,Ni-NTA琼脂糖树脂纯化Hsp60重组蛋白,用薄膜分散法制备以卵磷脂和胆固醇为膜组分包裹的Hsp60重组蛋白口服疫苗,并用透射电镜测定其粒径。75只BALB/C小鼠分为5组,分别通过灌胃方法给予PBS、空白脂质体、Hsp60重组蛋白+霍乱霉素(CT)、脂质体包裹Hsp60重组蛋白、脂质体包裹Hsp60重组蛋白+CT,每周1次共4次,末次攻击2周再用活Hp攻击3次,3周后处死小鼠,行胃组织快速尿素酶试验、Hp的定植半定量、炎症程度及其炎症活动度的评分。结果可溶性表达产物占全菌总蛋白的27%,经纯化获得纯度为95%的重组蛋白,制备的脂质体粒径为(0.7±0.4)μm。PBS组和空白脂质体组保护率均为0,而Hsp60重组蛋白+CT组、脂质体包裹Hsp60重组蛋白组、脂质体包裹Hsp60重组蛋白+CT组的保护率分别为73.3%、66.7%和86.7%,且均能使免疫小鼠胃粘膜Hp感染数目明显减少,炎症反应减轻。结论口服脂质体能分地代替免疫佐剂,作为Hp疫苗的免疫佐剂,将具有广泛的应用前景。
OBJECTIVE: To study the preparation of lipopolysaccharide (Hp) heat shock protein 60 (Hsp60) oral vaccine and evaluate its effect in preventing Hp infection by using mouse model of Hp infection. Methods PET-22 (+) / Hsp60 was expressed in E. coli BL21 (DE3) and Hsp60 recombinant protein was purified by Ni-NTA agarose resin. The recombinant protein Hsp60 was encapsulated with lecithin and cholesterol as membrane components Vaccines and their size were measured by transmission electron microscopy. Seventy five BALB / C mice were divided into five groups. PBS, blank liposome, Hsp60 recombinant protein + cholera toxin (CT), liposome-encapsulated Hsp60 recombinant protein, liposome-encapsulated Hsp60 recombinant Protein + CT once a week for 4 times, the last attack for 2 weeks and then live Hp challenge 3 times, 3 weeks after the mice were sacrificed, gastric tissue rapid urease test, Hp colonization semi-quantitative, degree of inflammation and inflammation Activity score. Results The soluble expression product accounted for 27% of the total bacterial protein. The recombinant protein was purified with purity of 95%. The size of the liposome was (0.7 ± 0.4) μm. The protection rates of PBS group and blank liposome group were all 0, while those of Hsp60 recombinant protein + CT group, liposome-encapsulated Hsp60 recombinant protein group and liposome-encapsulated Hsp60 recombinant protein + CT group were 73.3%, 66.7 % And 86.7%, respectively, and can significantly reduce the number of gastric mucosal Hp infection in immunized mice and reduce the inflammatory response. Conclusion Oral liposome can replace the immune adjuvant, as a vaccine adjuvant Hp vaccine, will have a wide range of applications.