6个wnt基因家族基因在细粒棘球绦虫原头节和成虫的差异表达分析

Differential Expression of Six wnt Gene Family Members in Echinococcus granulosus Protoscoleces and Adult Worms

目的探讨无翅型小鼠乳房肿瘤病毒整合位点家族[wingless-type mouse mammary tumor virus(MMTV)integration site family,wnt]成员wnt1、wnt2、wnt4、wnt5、wnt11A和wnt11B基因在细粒棘球绦虫(Echinococcus granulosus)原头节和成虫的m RNA差异转录及组织分布。方法用SYBR GreenⅠq RT-PCR方法检测wnt1、wnt2、wnt4、wnt5、wnt11A和wnt11B基因在细粒棘球绦虫原头节和成虫的m RNA相对转录情况。用RNA荧光探针以全量组织荧光原位杂交方法检测6个基因在原头节的组织分布情况。结果 q RT-PCR检测结果显示,wnt1和wnt2基因m RNA在原头节和成虫的相对转录水平分别为1.00、1.49和1.00、2.53,在成虫的相对转录水平分别为原头节的1.49倍(P>0.05)、2.53倍(P<0.05)。wnt4、wnt5、wnt11A和wnt11B m RNA在原头节和成虫的相对转录水平分别为1.00、0.04,1.00、0.03,1.00、0.07和1.00、0.97,在原头节的相对转录水平分别为成虫的25.00倍(P<0.01)、33.33倍(P<0.01)、14.29倍(P<0.01)和1.03倍(P>0.05),原头节和成虫间wnt4、wnt5、wnt11A表达量的差异有统计学意义,而wnt11B的差异则无统计学意义。全量组织荧光原位杂交结果显示,在原头节中,wnt1主要分布于表皮组织,wnt2主要分布于吸盘部位,wnt4主要分布于吸盘和顶突部位,wnt5、wnt11B主要分布于吸盘和表皮,wnt11A主要分布于顶突和小刺。结论 wnt2基因在细粒棘球绦虫成虫的m RNA相对转录水平高于原头节,wnt4、wnt5、wnt11A在原头节的m RNA相对转录水平高于成虫,6个wnt基因均分布于细粒棘球绦虫原头节的前端区域。

Objective To investigate the differential mRNA expression and tissue distribution of wnt [wingless-type mouse mammary tumor virus（MMTV）integration site family,wnt] gene members wnt1, wnt2, wnt4, wnt5, wnt11A and wnt11B in protoscoleces and adult worms of Echinococcus granulosus. Methods The mRNA expression of wnt1, wnt2, wnt4, wnt5, wnt11A and wnt11B was determined by qRT-PCR. Tissue distribution of wnt1, wnt2, wnt4, wnt5, wnt11A and wnt11B in Echinococcus granulosus protoscoleces was determined by the whole-mount in situ hybridization. Results The qRT-PCR results showed that the mRNA expression levels of wnt1 and wnt2 in the adult worms were 1.49 （P〉0.05） and 2.53 folds（P〈0.05） of those in the protoscoleces, respectively. The mRNA expression levels of wnt4, wnt5, wnt11A and wnt11B in the protoscoleces were 25.00（P〈0.01）, 33.33（P〈0.01）, 14.29（P〈0.01） and 1.03 folds（P〉0.05） of those in the adult worms, respectively. In brief, there was no significant difference of mRNA expression in wnt2 and wnt11B between protoscoleces and adult, but there was a significant difference of mRNA expression in wnt1, wnt4, wnt5 and wnt11A between protoscoleces and adults. Results of the whole-mount in situ hybridization showed that in protoscoleces wnt1 was mainly localized in the epidermal tissue, wnt2 in suckers, wnt4 in suckers and rostellum, wnt5 and wnt11B in suckers and epidermal tissue, and wnt11A in rostellum and hooks. Conclusion The mRNA expression of wnt2 in adult E. granulosus was higher than that in protoscoleces, and the mRNA expression ofwnt4, wnt5, wnt11A and wnt11B in protoscoleces was higher than that in the adult worms. The six wnt gene family members were all distributed in the forward region of protoscoleces.