表达柔嫩艾美耳球虫HMGB1基因质粒DNA的免疫效果分析

doi:10.11843/j.issn.0366-6964.2016.02.019

摘要/Abstract

摘要：

旨在评价表达柔嫩艾美耳球虫（E.tenella）HMGB1基因质粒DNA免疫对同源株攻虫的免疫保护效果。将EtHMGB1基因插入pcDNA3.1(-)/myc-His A真核表达载体中，并在Hela细胞中进行体外瞬时表达。设计了pcDNA3.1(-)/myc-EtHMGB1免疫组、pcDNA3.1(-)/myc-His A免疫组、重组蛋白质+FCA佐剂免疫组、pcDNA3.1(-)/myc-EtHMGB1+重组蛋白质联合免疫组、FCA佐剂免疫组、未免疫攻虫组和未免疫未攻虫组7个试验组。分别于14和21日龄对鸡进行两次免疫，28日龄时除未免疫未攻虫组外各组用1×10⁴个E.tenella孢子化卵囊攻虫，以平均增重、卵囊产量和病变记分作为免疫保护效果的评价指标。结果显示，成功构建了pcDNA3.1(-)/myc-EtHMGB1重组质粒，转染后该质粒可在Hela细胞中进行瞬时表达。该重组质粒单独免疫、重组蛋白质单独免疫或重组质粒与重组蛋白质联合免疫均对鸡肠道病变改善效果不明显，但可显著提高增重，降低卵囊产量，尤以重组质粒与重组蛋白质联合免疫效果最佳，显示Prime-boost免疫策略可提高该重组质粒的免疫保护效果。上述结果显示HMGB1可作为未来球虫疫苗研制的良好候选抗原之一。

Abstract:

This experiment was conducted to assess the immuno-efficacy of the eukaryotic plasmid.The high mobility group box1 gene from Eimeria tenella was inserted into expression vector pcDNA3.1(-)/myc-His A，and then constructed the eukaryotic expression recombinant plasmid pcDNA3.1(-)/myc-EtHMGB1 and expressed transiently the objective protein in Hela cells.Seven experimental groups including pcDNA3.1(-)/myc-EtHMGB1 immunization group，pcDNA3.1(-)/myc-His A immunization group，recombinant antigen with adjuvant immunization group，pcDNA3.1(-)/myc-EtHMGB1 with recombinant antigen prime-boost immunization group，FCA adjuvant immunization group，unimmunized and challenged group and unimmunized and unchallenged group were designed.Chickens were vaccinated at the age of 14 and 21 days.All birds except the unchallenged control group were challenged orally with 1×10⁴E.tenella sporulated oocysts at 28 days.Gain of body weight，fecal oocyst output and lesion scores were assessed as measures of protective immunity.The results showed that the plasmid pcDNA3.1(-)/myc-EtHMGB1 was successfully constructed，and the transient expression product of EtHMGB1 could be detected by IFA and Western blot.Challenge experiments demonstrated that pcDNA3.1(-)/myc-EtHMGB1 immunization，recombinant antigen with adjuvant immunization and pcDNA3.1(-)/myc-EtHMGB1 with recombinant antigen prime-boost immunization could all increase body weight gains and reduce oocyst excretion，but all three methods did not have an effect on cecal lesion.The prime-boost immunization strategy were superior to the other method based on the immune effects.These results suggest that EtHMGB1 could be proposed as an anti-apicomplexan vaccine candidate.

中图分类号:

S852.723

顾有方，李文超，汪凯，靳二辉，胡倩倩，李升和，陈会良. 表达柔嫩艾美耳球虫HMGB1基因质粒DNA的免疫效果分析[J]. 畜牧兽医学报, 2016, 47(2): 354-360.

GU You-fang，LI Wen-chao，WANG Kai，JIN Er-hui，HU Qian-qian，LI Sheng-he，CHEN Hui-liang . Protective Immunity Induced by a Plasmid Encoding Eimeria tenella High Mobility Group box1 Gene against Homologous Challenge[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(2): 354-360.

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参考文献

［1］BLAKE D P，BILLINGTON K J，COPESTAKE S L，et al.Genetic mapping identifies novel highly protective antigens for an apicomplexan parasite［J］.PLoS Pathog，2011，7(2)：e1001279.
［2］LI W C，ZHANG X K，DU L，et al.Eimeria maxima：efficacy of recombinant Mycobacterium bovis BCG expressing apical membrane antigen1 against homologous infection［J］.Parasitol Res，2013，112(11)：3825-3833.
［3］SONG X，HUANG X，YAN R，et al.Efficacy of chimeric DNA vaccines encoding Eimeria tenella 5401 and chicken IFN-γ or IL-2 against coccidiosis in chickens［J］.Exp Parasitol，2015，156：19-25.
［4］SONG X，REN Z，YAN R，et al.Induction of protective immunity against Eimeria tenella，Eimeria necatrix，Eimeria maxima and Eimeria acervulina infections using multivalent epitope DNA vaccines［J］.Vaccine，2015，33(24)：2764-2770.
［5］LANGE S S，MITCHELL D L，VASQUEZ K M.High mobility group protein B1 enhances DNA repair and chromatin modification after DNA damage［J］.Proc Natl Acad Sci U S A，2008，105(30)：10320-10325.
［6］EL GAZZAR M，YOZA B K，CHEN X，et al.Chromatin-specific remodeling by HMGB1 and linker histone H1 silences proinflammatory genes during endotoxin tolerance［J］.Mol Cell Biol，2009，29(7)：1959-1971.
［7］曹利利.犬新孢子虫“警报素”介导的宿主Th1型免疫应答的研究［D］.长春：吉林大学，2010.
CAO L L.Regulation of host Th1 immune response by the alarmins of Neospora caninum［D］.Changchun：Jilin University，2010.(in Chinese)
［8］JOHNSON J，REID W M.Anticoccidial drugs：lesion scoring techniques in battery and floor-pen experiments with chickens［J］.Exp Parasitol，1970，28(1)：30-36.
［9］GNANASEKAR M，VELUSAMY R，HE Y X，et al.Cloning and characterization of a high mobility group box 1 (HMGB1) homologue protein from Schistosoma mansoni［J］.Mol Biochem Parasitol，2006，145(2)：137-146.
［10］KUMAR K，SINGAL A，RIZVI M M，et al.High mobility group box (HMGB) proteins of Plasmodium falciparum：DNA binding proteins with pro-inflammatory activity［J］.Parasitol Int，2008，57(2)：150-157.
［11］ABHYANKAR M M，HOCHREITER A E，HERSHEY J，et al.Characterization of an Entamoeba histolytica high-mobility-group box protein induced during intestinal infection［J］.Eukaryot Cell，2008，7(9)：1565-1572.
［12］ZHANG L，LIU R，SONG M，et al.Eimeria tenella：interleukin 17 contributes to host immunopathology in the gut during experimental infection［J］.Exp Parasitol，2013，133(2)：121-130.
［13］陈海宁，戚南山，廖申权，等.Prime-boost 免疫策略在抗顶复门原虫感染中的应用［J］.畜牧与兽医，2015，47(1)：130-134.
CHEN H N，QIN N S，LIAO S Q，et al.The application of Prime-boost immunization strategy in anti-infection of apicomplexan protozoa［J］.Animal Husbandry＆Veterinary Medicine，2015，47(1)：130-134.(in Chinese)
［14］YU L，YAMAGISHI J，ZHANG S，et al.Protective effect of a prime-boost strategy with plasmid DNA followed by recombinant adenovirus expressing TgAMA1 as vaccines against Toxoplasma gondii infection in mice［J］.Parasitol Int，2012，61（3)：481-486.
［15］GLYNN A，FREYTAG L C，CLEMENTS J D.Effect of homologous and heterologous prime-boost on the immune response to recombinant plague antigens［J］.Vaccine，2005，23(16)：1957-1965.
［16］LEE S H，LILLEHOJ H S，JANG S I，et al.Embryo vaccination of chickens using a novel adjuvant formulation stimulates protective immunity against Eimeria maxima infection［J］.Vaccine，2010，28(49)：7774-7778.
［17］吴绍强，蒋金书，刘群，等.柔嫩艾美耳球虫BJ 株核酸疫苗的构建及其免疫保护效果研究［J］.中国兽医杂志，2004，40(9)：3-6.
WU S Q，JIANG J S，LIU Q，et al.Construction of the DNA vaccine pCT and pCTE and their efficacy against experimental coccidiosis in chickens［J］.Chinese Journal of Veterinary Medicine，2004，40(9)：3-6.(in Chinese)
［18］刘秉春，崔新洁，罗新松，等.核酸疫苗初免-蛋白疫苗加强的免疫策略提高日本血吸虫核酸疫苗免疫效果［J］.生物工程学报，2013，29(6)：814-822.
LIU B C，CUI X J，LUO X S，et al.DNA prime followed by protein boost enhances the protective efficacy against Schistosoma japonicum infection in mice［J］.Chinese Journal of Biotechnology，2013，29(6)：814-822.(in Chinese)
［19］MIN J，QU D F，LI C Z，et al.Enhancement of protective immune responses induced by Toxoplasma gondii dense granule antigen 7(GRA7) against toxoplasmosis in mice using a prime-boost vaccination strategy［J］.Vaccine，2012，30(38)：5631-5636.

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