小鼠疟原虫感染经氯喹治疗痊愈后再感染免疫特征分析

Immune characteristics of Plasmodium reinfections in mice following chloro⁃quine cure of primary Plasmodium infections

目的探讨疟原虫感染小鼠在原虫血症高峰期经氯喹治愈后,再感染同/异种疟原虫的疾病进程和免疫保护特征。方法用非致死型约氏疟原虫17XNL株(P.y 17XNL株)感染C57BL/6小鼠,感染率达高峰时(第9天)半数小鼠以氯喹治疗,其余小鼠自然痊愈。痊愈后小鼠于初次感染90 d后分别采用等量致死型约氏疟原虫17XL株(P.y 17XL株)或伯氏疟原虫ANKA株(P.b ANKA株)再次感染,采用吉姆萨薄血膜染色法观察原虫血症水平变化,并应用酶联免疫吸附试验和流式细胞术分别检测再感染前后小鼠血清中IgG抗体水平和脾细胞中记忆性T细胞亚群比例。结果初次感染P.y 17XNL株后,自愈与氯喹治愈组小鼠血清IgG抗体水平分别为(5.047±0.924)、(4.429±0.624)pg/mL,差异无统计学意义(t=0.437,P>0.05);但均显著高于无感染正常组小鼠的(1.624±0.280)pg/mL(F=22.522,P<0.01)。自愈和氯喹治愈组小鼠再感染P.y 17XL株或P.b ANKA株并痊愈后,各组小鼠血清IgG抗体水平分别为(15.487±1.173)、(14.644±1.523)、(15.965±1.150)pg/mL和(15.185±1.333)pg/mL,差异无统计学意义(F=0.542,P>0.05);但均高于初次感染组,差异有统计学意义(F=67.383,P<0.01)。感染P.y 17XNL株后自愈及氯喹治愈组小鼠再感染P.y 17XL株或P.b ANKA株并痊愈后,各组小鼠CD4+记忆性T细胞比例分别为(34.023±2.289)%、(35.608±1.779)%、(34.208±2.106)%和(32.820±1.930)%,CD8+记忆性T细胞比例分别为(17.103±1.627)%、(17.873±1.425)%、(17.935±2.092)%和(18.918±2.823)%,组间差异均无统计学意义(F=0.944、0.390,P均>0.05);但均高于初次感染后小鼠,且差异有统计学意义(CD4+记忆性T细胞,F=50.532,P<0.01;CD8+记忆性T细胞,F=21.751,P<0.01)。结论小鼠疟疾经氯喹治疗痊愈不影响宿主在再感染时产生有效免疫保护力。初次感染后,小鼠对同种和异种疟原虫再感染均具有一定保护性且对同种疟原虫再感染的抵抗力强于异种疟原虫。

Objective To investigate the disease progression and immunoprotective characteristics in mice re⁃infected with homogeneous/heterogeneous Plasmodium strains following cure of Plasmodium infections with chloroquine at the peak of parasit⁃emia.Methods C57BL/6 mice were infected with the non⁃lethal P.yoelii 17XNL strain,and half of mice were given treatment with chloroquine at the peak of parasitemia(9 days post⁃infection),while the other mice were self⁃cured naturally.Then,all cured mice were re⁃infected with the equivalent lethal P.yoelii 17XL or P.berghei ANKA strain 90 days following primary Plasmodium infections.The parasitemia levels during primary infections and reinfections were measured by microscopic examinations of Gi⁃emsa⁃stained thin blood films,and the levels of the IgG antibody in sera and the percentages of memory T cell subsets in spleen cells were detected in mice using ELISA and flow cytometry before and after parasite reinfections,respectively.Results Follow⁃ing primary infections with the P.yoelii 17XNL strain,the serum IgG antibody levels were(5.047±0.924)pg/mL in the self⁃cured mice and(4.429±0.624)pg/mL in the chloroquine⁃treated mice,respectively(t=0.437,P>0.05),which were both signif⁃icantly higher than that in the uninfected mice(1.624 pg/mL±0.280 pg/mL)(F=22.522,P<0.01).There was no significant dif⁃ference in the serum IgG antibody level among self⁃cured and chloroquine⁃treated mice re⁃infected with the P.yoelii 17XL strain or the P.berghei ANKA strain(F=0.542,P>0.05);however,the serum IgG antibody levels were all significantly higher in self⁃cured and chloroquine⁃treated mice re⁃infected with the P.yoelii 17XLstrain[(15.487±1.173)pg/mLand(15.965±1.150)pg/mL]or the P.berghei ANKA strain[(14.644±1.523)pg/mL and(15.185±1.333)pg/mL]relative to primary infections(F=67.383,P<0.01).There was no significant difference in the proportion of CD4+[(34.208±2.106),(32.820±1.930),(34.023±2.289),(35.608±1.779)pg/mL]or CD8+T memory cells[(17.935±2.092),(18.918±2.823),(17.103±1.627),(17.873±1.425)pg/mL]in self⁃cured and chloroquine⁃treated mice with primary infections with the P.yoelii 17XNL strain followed by re⁃infections with the P.yoelii 17XL strain or the P.berghei ANKA strain(F=0.944 and 0.390,both P>0.05);however,the proportions of the CD4+or CD8+T memory cells were significantly greater in self⁃cured and chloroquine⁃treated mice with primary infections with the P.yoelii 17XNL strain followed by re⁃infections with the P.yoelii 17XL strain or the P.berghei ANKA strain than in mice with primary infections(F=50.532 and 21.751,both P<0.01).Conclusions The cure of murine Plasmodium infections with chloroquine does not affect the production of effective immune protections in mice during parasite re⁃infections.Following a pri⁃mary infection,mice show a protection against re⁃infections with either homogeneous or heterogeneous Plasmodium strains,and a higher⁃level resistance to re⁃infections with homogeneous parasite strains is found than with heterogeneous strains.