细粒棘球绦虫囊化原头蚴AQP基因转录动态及异源表达实验研究

谭青青; 吴宏烨; 高剑; 李锴; 范俊杰; 李想; 廖鹏; 曹纹侨; 叶彬

doi:10.16016/j.1000-5404.202006204

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细粒棘球绦虫囊化原头蚴AQP基因转录动态及异源表达实验研究

基础医学 | 更新时间：2026-01-09

- 细粒棘球绦虫囊化原头蚴AQP基因转录动态及异源表达实验研究
- Transcription profile and heterologous expression of Echinococcus granulosus aquaporins genes in the encysted protoscolex
- 第三军医大学学报 2020年42卷第21期页码：2081-2092
- 作者机构：
  
  1.400016 重庆，重庆医科大学基础医学院：病原生物学教研室
  3.400016 重庆，重庆医科大学基础医学院：分子医学与肿瘤研究中心
  2.830011 乌鲁木齐，新疆医科大学寄生虫学教研室
- 作者简介：
  
  [叶彬，E-mail: yebin@cqmu.edu.cn YE Bin, E-mail: yebin@cqmu.edu.cn
- 基金信息：
  
  国家自然科学基金面上项目(81672045)
- DOI：10.16016/j.1000-5404.202006204
  中图分类号： R341;R383.33;R394.3
- 收稿：2020-06-22，
  
  修回：2020-08-11，
  
  纸质出版：2020-11-15
- 稿件说明：
移动端阅览
谭青青, 吴宏烨, 高剑, 等. 细粒棘球绦虫囊化原头蚴AQP基因转录动态及异源表达实验研究[J]. 第三军医大学学报, 2020,42(21):2081-2092.

Qingqing TAN, Hongye WU, Jian GAO, et al. Transcription profile and heterologous expression of Echinococcus granulosus aquaporins genes in the encysted protoscolex[J]. Journal of Third Military Medical University, 2020, 42(21): 2081-2092.
谭青青, 吴宏烨, 高剑, 等. 细粒棘球绦虫囊化原头蚴AQP基因转录动态及异源表达实验研究[J]. 第三军医大学学报, 2020,42(21):2081-2092. DOI： 10.16016/j.1000-5404.202006204.

Qingqing TAN, Hongye WU, Jian GAO, et al. Transcription profile and heterologous expression of Echinococcus granulosus aquaporins genes in the encysted protoscolex[J]. Journal of Third Military Medical University, 2020, 42(21): 2081-2092. DOI： 10.16016/j.1000-5404.202006204.

摘要

目的

探索细粒棘球蚴水通道蛋白(

Echinococcus granulosus

aquaporins

AQPs)在棘球蚴液形成过程中的作用。

方法

收集体外培养不同时期的棘球蚴标本和BALB/c小鼠体内接种的继发棘球蚴标本，RT-qPCR方法检测12个

AQPs基因在体内、体外培养不同发育时期的转录表达。以细粒棘球蚴cDNA为模板，扩增

AQP、

AQP1、

AQP3、

AQP4-1、

AQP4-4和

AQP9-1 6个基因编码区片段，同时，通过化学合成方法扩增另外6个

AQPs(

AQPAn.G、

AQPFA-CHIP、

AQP4-2、

AQP4-3、

AQP4-5、

AQP9-2)基因编码区片段，并在非洲爪蟾卵母细胞中检测这12个

AQPs基因的透水功能，Western blot检测

AQPs在卵母细胞膜上的表达。

结果

体外培养的原头蚴及感染小鼠的继发棘球蚴体积均随培养时间的延长而逐渐增大，棘球蚴囊内透明液体增多。RT-qPCR结果显示12个

AQPs基因在体内、体外培养不同发育时间的转录表达水平均较低。注射

AQPs基因的非洲爪蟾卵母细胞的体积变化率和透水系数与阴性对照组差异

无统计学意义，这12个

AQPs基因在卵母细胞中未表现出水通道功能。Western blot检测12个

AQPs基因在卵母细胞总蛋白、胞质蛋白与胞膜蛋白上的表达，结果均未观察到蛋白在预期的水通道蛋白理论分子量处表达。

结论

12个

AQPs基因在原头蚴囊化形成棘球蚴过程中转录表达水平均较低，这些基因在爪蟾卵母细胞的异源表达均未表现出通透水的功能。

Abstract

Objective

To explore the role of

Echinococcus granulosus

aquaporins (

AQPs) in the accumulation of hydatid fluid.

Methods

The secondary hydatids cultured

in vitro

and inoculated in BALB/c mice were collected respectively

and the relative transcription expression levels of 12

AQPs genes from the hydatid cysts cultured

in vitro

and in mice were detected by RT-qPCR. Six gene coding region fragments of

AQP

AQP1

AQP3

AQP4-1

AQP4-4 and

AQP9-1 were amplified using cDNA templates. The other 6 genes coding region fragments of

AQPs (

AQPAn.G

AQPFA-CHIP

AQP4-2

AQP4-3

AQP4-5 and

AQP9-2) were amplified with chemical synthesis. The water permeability of these 12

AQPs genes were detected in

Xenopus laevis

(

X. laevis

)

and the expression of

AQPs on the oocytes membranes was detected by Western blotting.

Results

The volume of hydatid cultured

in vitro

and

in vivo

was increased gradually with the extension of culture time

and there was more and more transparent hydatid fluid in hydatid cyst. The results of RT-qPCR showed that the relative transcription levels of

AQPs genes in the hydatid cysts were very low or even undetectable except for

AQP3 and

AQP9-1 at the beginning of culture

in vitro

. There were no significant differences in relative volume change rates and water permeability coefficients of

X. laevis

oocytes injected with

AQPs genes when compared with the negative control

and these

AQPs genes did not show water channel functions in the oocytes. Western blot assay displayed that there was no expression of

AQPs genes in total

cytoplasmic and membrane protein on the oocytes.

Conclusion

The transcription levels of

AQPs genes are quite low in the encystation process of protoscolex

and their heterologous expression do not show water channel functions in

X. laevis

oocytes.

关键词

Keywords

references

ALAM-ELDIN Y H, BADAWY A F. Destructive effect of gamma irradiation on Echinococcus granulosus metacestodes[J ] . Parasitol Res, 2015, 114(8): 3145-3150. DOI: 10.1007/s00436-015-4533-9.

XIN Q, YUAN M M, LI H P, et al . In vitro and in vivo effects of 3-bromopyruvate against Echinococcus metacestodes[J ] . Vet Res, 2019, 50(1): 96. DOI: 10.1186/s13567-019-0710-7..

ZHANG W B, ZHANG Z Z, WU W P, et al . Epidemiology and control of echinococcosis in central Asia, with particular reference to the People's Republic of China[J ] . Acta Trop, 2015, 141: 235-243. DOI: 10.1016/j.actatropica.2014.03.014.

伍卫平, 王虎, 王谦, 等.2012-2016年中国棘球蚴病抽样调查分析[J].中国寄生虫学与寄生虫病杂志, 2018, 36(1): 1-14.

WU W P, WANG H, WANG Q, et al . A nationwide sampling survey on echinococcosis in China during 2012-2016[J ] . Chin J Parasitol Parasit Dis, 2018, 36(1): 1-14.

王慧.棘球蚴囊液蛋白质组学及主要分泌蛋白对宿主免疫调节的影响[D].乌鲁木齐: 新疆医科大学, 2015.

WANG H. Proteomic analysis of hydatid cyst fiuid and effect of mainly secreted protein on host immune regulation[D]. Urumchi: Xinjiang Medical University, 2015.

ABASCAL F, IRISARRI I, ZARDOYA R. Diversity andevolution of membrane intrinsic proteins[J]. Biochim Biophys Acta, 2014, 1840(5): 1468-1481. DOI: 10.1016/j.bbagen.2013.12.001.

FARLORA R, VALENZUELA-MUNOZ V, CHAVEZ-MARDONES J, et al . Aquaporin family genes exhibit developmentally-regulated and host-dependent transcription patterns in the sea louse Caligus rogercresseyi[J ] . Gene, 2016, 585(1): 119-127. DOI: 10.1016/j.gene.2016.03.035.

MA G X, JIANG A Y, LUO Y F, et al . Aquaporin 1 is located on the intestinal basolateral membrane in Toxocara canis and might play a role in drug uptake[J ] . Parasit Vectors, 2019, 12(1): 243. DOI: 10.1186/s13071-019-3500-1..

NI Z X, CUI J M, ZHANG N Z, et al . Structural and evolutionary divergence of aquaporins in parasites[J ] . Mol Med Rep, 2017, 15(6): 3943-3948. DOI: 10.3892/mmr.2017.6505.

WANG F, YE B. Bioinformatics analysis and construction of phylogenetic tree of aquaporins from Echinococcus granulosus [J ] . Parasitol Res, 2016, 115(9): 3499-3511. DOI: 10.1007/s00436-016-5114-2.

王芬.细粒棘球绦虫棘球蚴的水通道蛋白基因研究[D].重庆: 重庆医科大学, 2018.

WANG F. Studies on aquaporins genes of hydatid of Echinococcus granulosus [D ] . Chongqing: Chongqing Medical University, 2018.

吴宏烨, 李锴, 王芬, 等.原头蚴体外形成棘球蚴的优化培养体系研究[J].第三军医大学学报, 2019, 41(5):415-423. DOI: 10.16016/j.1000-5404.201808123.

WU H Y, LI K, WANG F, et al . Study on the culture systems for transformations of hydatid cysts from protoscoleces in vitro [J ] . J Third Mil Med Univ, 2019, 41(5):415-423. DOI: 10.16016/j.1000-5404. 201808123..

王芬, 刘许诺, 吴宏烨, 等.原头蚴成囊过程中内参基因的确定及水通道候选基因的转录表达[J].第三军医大学学报, 2018, 40(17): 1533-1541. DOI: 10.16016/j.1000-5404.201803049.

WANG F, LIU X N, WU H Y, et al . Identification of optimal r eference genes during development of hydatid cysts from protoscoleces and transcriptional expression of candidate aquaporins[J ] . J Third Mil Med Univ, 2018, 40(17): 1533-1541. DOI: 10.16016/j.1000-5404.201803049.

吴宏烨.细粒棘球绦虫的6种水通道蛋白基因研究[D].重庆: 重庆医科大学, 2019.

WU H Y. Study on six aquaporins genes of Echinococcus granulosus [D ] .Chongqing: Chongqing Medical University, 2019.

ZHOU S Y, HU W, DENG X M, et al . Overexpression of the wheat aquaporin gene, Ta AQP7, enhances drought tolerance in transgenic tobacco[J ] . PLoS ONE, 2012, 7(12): e52439. DOI: 10.1371/journal.pone.0052439..

MONTALVETTI A, ROHLOFF P, DOCAMPO R. A functional aquaporin co-localizes withthe vacuolar proton pyrophosphatase to acidocalcisomes and the contractile vacuole complex of Trypanosoma cruzi [J ] . J Biol Chem, 2004, 279(37):38673-38682.DOI:10.1074/jbc.M406304200.

郭刚, 张文宝.胆汁酸盐对细粒棘球绦虫分化和发育的调控[J].中国高原医学与生物学杂志, 2018, 39(2): 129-131. DOI: 10.13452/j.cnki.jqmc.2018.02.012.

GUO G, ZHANG W B. Regulation of bile salts on differentiation and development of Echinococcus granulosus [J ] . Chin High Altitude Med Biol, 2018, 39(2): 129-131. DOI: 10 .13452/j.cnki.jqmc.2018.02.012.

WANG H, LI J, GUO B P, et al . In vitro culture of Echinococcus multilocularis producing protoscoleces and mouse infection with the cultured vesicles[J ] . Filaria J, 2016, 9(1): 411. DOI: 10.1186/s13071-016-1687-y..

MORTEZAEI S, AFGAR A, MOHAMMADI M A, et al . The effect of albendazole sulfoxide on the expression of miR-61 and let-7 in different in vitro developmental stages of Echinococcus granulosus [J ] . Acta Trop, 2019, 195: 97-102. DOI: 10.1016/j.actatropica.2019.04.031..

PEREZ DI GIORGIO J, SOTO G, ALLEVA K, et al . Prediction of aquaporin function by integrating evolutionary and functional analyses[J ] . J Membr Biol, 2014, 247(2): 107-125. DOI:10.1007/s00232-013-9618-8.

SONG J, MAK E, WU B H, et al . Parasite aquaporins: current developments in drug facilitation and resistance[J ] . Biochim Biophys Acta, 2014, 1840(5): 1566-1573. DOI:10.1016/j.bbagen.2013.10.014.

MURATA K, MITSUOKA K, HIRAI T, et al . Structural determinants of water permeation through aquapor in-1[J ] . Nature, 2000, 407(6804): 599-605. DOI:10.1038/35036519.

GOMES D, AGASSE A, THIÉBAUD P, et al . Aquaporins are multifunctional water and solute transporters highly divergent in living organisms[J ] . Biochim Biophys Acta, 2009, 1788(6):1213-1228. DOI:10.1016/j.bbamem.2009.03.009.

VERKMAN A S, MITRA A K. Structure and function of aquaporin water channels[J]. Am J Physiol Ren Physiol, 2000, 278(1):F13-F28. DOI:10.1152/ajprenal.2000.278.1.F13.

范俊杰, 吴宏烨, 李锴, 等.细粒棘球绦虫原头蚴水通道蛋白编码基因的转录组分析[J].中国病原生物学杂志, 2018, 13(11):1187-1192, 1197.DOI:10.13350/j.cjpb.181103.

FAN J J, WU H Y, LI K, et al . Characterization of the transcriptome of aquaporin-encoding genes in the protoscoleces of Echinococcus granulosus [J ] . J Pathogen Biol, 2018, 13(11):1187-1192, 1197. DOI:10.13350/j.cjpb.181103..

PIKE S, MATTHES M S, MCSTEEN P, et al . Using Xenopus laevis oocytes to functionally characterize plant transporters[J ] . Curr Protoc Plant Biol, 2019, 4(1): e20087. DOI:10.1002/cppb.20087..

HUANG Y Z, LI W, LU W G, et al . Cloning and in vitro characterization of a Schistosoma japonicum aquaglyceroporin that functions in osmoregulation[J ] . Sci Rep, 2016, 6: 35030. DOI:10.1038/srep35030..

PAVLOVIC-DJURANOVIC S, SCHULTZ J E, BEITZ E. A single aquaporin gene encodes a water/glycerol/urea facilitator in Toxoplasma gondii with similarity to plant tonoplast intrinsic proteins[J ] . FEBS Lett, 2003, 555(3): 500-504. DOI:10.1016/s0014-5793(03)01313-9.

邹珊珊, 王萍, 魏威, 等.携带GFP和FLAG标签的真核表达载体介导细胞珠蛋白在细胞内表达的定位研究[J].热带医学杂志, 2014, 14(4): 456-460, 548.

ZOU S S, WANG P, WEI W, et al . The difference of localization of GFP-tagged/FLAG-tagged eukaryotic expression vector of cytogiobin in the cells[J ] . J Trop Med, 2014, 14(4): 456-460, 548.

孔佑宾, 王冰, 张华, 等.植物蛋白融合HA标签通用载体构建与应用[J].中国农业科技导报, 2017, 19(6): 131-137.

KONG Y B, WANG B, ZHANG H, et al . Construction and amplification of protein fusion HA tag universal expression vector in plant[J ] . J Agric Sci Technol, 2017, 19(6): 131-137.

HEDFALK K, PETTERSSON N, OBERG F, et al . Production, characterization and crystallization of the Plasmodium falciparum aquaporin[J ] . Protein Expr Purif, 2008, 59(1): 69-78. DOI: 10.1016/j.pep.2008.01.004..

FINN R N, CERDÀ J. Evolution and functional diversity of aquaporins[J]. Biol Bull, 2015, 229(1): 6-23. DOI:10.1086/BBLv229n1p6.

AHMADPOUR D, GEIJER C, TAMÁS M J, et al . Yeast reveals unexpected roles and regulatory features of aquaporins and aquaglyceroporins[J ] . Biochim Biophys Acta, 2014, 1840(5):1482-1491. DOI:10.1016/j.bbagen.2013.09.027.

ZHENG H J, ZHANG W B, ZHANG L, et al . The genome of the hydatid tapeworm Echinococcus granulosus [J ] . Nat Genet, 2013, 45(10): 1168-1175. DOI:10.1038/ng.2757..

WANG N, ZHU H, ZHAN J F, et al . Cloning, expression, characterization, and immunological properties of citrate synthase from Echinococcus granulosus [J ] . Parasitol Res, 2019, 118(6): 1811-1820. DOI:10.1007/s00436-019-06334-6..

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