| Objective: To study the role of microRNA(mi RNA)-mediatted Hippo pathway in reproductive organ damage / malformation in mice by vinclozolin.Methods:(1)Use bioinformatics to predict whether Yap1 was the target gene of miR132 and miR195 a.(2)Vinclozolin was orally administered to the pregnant mice in the treatment group with 400 mg/ kg·bw during gestation days 12-18,while those of the control group were treated with equal volume of soybean oil.The shape and body mass of the mice were observed after 1,3 and 7-week-old of birth.Then the penis,testis of the male mice,the uterus and ovaries of the female mice were taken and their length,size,and volume were measured and calculated,and observe pathological changes.(3)Real-time PCR was used to detect the mRNA abundance of Hippo pathway,cell cycle,apoptosis and hormone receptorrelated genes,and expression abundance of miR132 and miR195 a in 1,3,and 7-week-old of birth uterus,ovaries,penis,and testis.(4)The expression of Yap and pYap in uterus,ovaries,penis and testis was detected by Western blot.Results:(1)Yap1 is the target gene of miR132 and mi R195 a.(2)At 1,3,and 7 weeks of age,the body mass of the mice in the experimental group was lower than that in the control group(except 7-week-old female offspring).At 3 and 7 weeks of age,the uterus and ovarian tissues development malformation,the male mice showed penis malformation and "small testicles".The uterus and ovarian tissues development of female offsprings in the experimental group at the age of 3 and 7 weeks were retarded.the urethra of the experimental group did not form a closed structure,and the layers of the layers were disordered.The testicular spermatogenesis were impaired.(3)miR132 expression was up-regulated in the experimental group including 1,7-week-old uterus,7-week-old ovaries and 1,3,7-week-old penis,testis(P<0.05).miR195 a expression was up-regulated in the experimental group including 1-week-old testis,1,3,7-week-old uterus,penis and 7-week-old ovaries(P<0.05).(4)The upregulated genes in the experimental group including 1-week-old uterus Mst1,Sav1,Lats1,Lats2,Ccnc,Jnk2,Jnk3,Caspase 9,Caspase 12,1-week-old ovaries Mst1,Sav1,Lats2,Mob1,Ccnc,1-week-old penis Mst1,Sav1,Ccna1,Jnk1,1-week-old testis Mst1,Mob1,Ccna1,Ccnc,Ccnd1,Ccne1,3-week-old uterus Mst1,Lats1,Mob1,Ccna1,Ccnc,Ccnd1,Ccne1,Jnk2,Jnk3,Caspase 3,Caspase 12,3-week-old ovaries Mst1,Lats2,Mob1,Ccna1,Ccnc,Ccne1,Jnk1,3-week-old penis Mst1,Ccna1,Ccnb1,Ccnc,Ccnd1,Ccne1,Jnk3,Caspase 3,3-week-old testis Mst1,Sav1,Lats1,Mob1,Ccnb1,Ccnc,Ccnd1,Ccne1,Jnk3,Caspase 3,7-week-old uterus Mst1,Mob1,Ccne1,Caspase 3,7-week-old ovaries Mst1,Lats1,Caspase 3,Caspase 9,7-week-old penis Mst1,Lats1 and 7-week-old testis Mst1,Sav1,Ccnb1,Ccnd1,Ccne1,Jnk1,Jnk2,Jnk3 were observed(P<0.05).The down-regulated genes in the experimental group including 1-week-old uterus Yap,Gper1,1-week-old ovaries,penis and 3-week-old penis Yap,Ar,1-week-old testis Yap,Esr1,Esr2,Gper1,Ar,Jnk1,Jnk3,Caspase 3,Caspase 9,Caspase 12,3-week-old uterus,testis and 7-week-old ovaries Yap,Taz,3-week-old ovaries Yap,Gper1,Esr1,Esr2,Ar,7-week-old uterus Yap,Taz,Ccnd1,7-week-old penis Taz,Jnk1,Jnk3,Caspase 3,Caspase 12 and 7-week-old testis Yap were observed(P<0.05).(5)At 1,3,and 7 weeks of age,the protein expression of Yap in the ovaries and penis tissue groups were lower than those in the control group(P<0.05),and the protein expression of p Yap were higher than those in the control group(P<0.05).No Yap and pYap in the uterus and testis were detected at 1,3,and 7 weeks of age.Conclusions:(1)Pregnant ICR mice were given vinclozolin can induce uterus and ovaries malformation in female offsprings,penis malformation," small testicles " in male mice.(2)The ovaries injury and penis malformation of offspring mice may be related to the Hippo pathway activated by miR132 and miR195 a. |
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