| Objective: To study Effect of different concentration of zinc on palatal cell proliferation during palatine process fusion;Feeding pregnant mouse with different concentrations of zinc,to observe the expression level of Sp family gene(Sp8,Sp5)and verify relativity between Sp family gene and cleft palate.Lay the foundation for the follow-up study of the role and mechanism of zinc in the process of cleft palate.Methods:The pregnant females were randomly divided into four groups according to the concentration of zinc in the feed.The normal zinc group(zinc containing 30mg/kg),zinc deficiency group(zinc deficiency feed),low zinc group(zinc containing 5mg/kg),zinc rich group(zinc containing 100mg/kg).Experiment 1:Death of the pregnant mice at ED14.5,and the development and morphology of the palate of fetal mice were observed under HE staining microscope.Select good slice specimens for PCNA staining,The proliferation of cells in the fusion stage of the palatine embryo was observed,and then the marker index(LI)was used to analyze the cell proliferation.Experiment 2:Death of the pregnant mice at ED14.5,Scissor the palate of embryo in the stereomicroscope,and study the proembryonic process of the palatine palatine,the expression level of Sp8 and Sp5 gene was observed by Real-Time PCR technique.Results:Experiment 1:the number of embryos in the normal zinc group was 39,of which there were 1 cleft palate,0 stillbirths,and 2.5% cleft palate.The number of embryos fed zinc deficient group: 38,the occurrence of cleft palate have 15,stillbirth 9,39.4%incidence of cleft palate;low zinc diet group embryo number: 41,which occurs in cleft palate 12,stillbirth 5,29.2% incidence of cleft palate;zinc;feeding group: 48,the number of embryos the occurrence of cleft palate have 3,stillbirth 0,6.2% incidence of cleft palate.HE staining display at ED14.5,The left and right palate processes in both the normal zinc and the zinc rich groups were raised and started to contact,however,the bilateral palatine process in the zinc deficiency group and the low zinc group did not lift up or the unilateral palatine process was not raised on both sides of the tongue.art of the section found that the palatine process was raised but because of its shortness,the palatine could not be fused in the middle of the palate.Observation of cell proliferation by PCNA staining:The positive rate of proliferating cells in the normal zinc and zinc rich groups of ED14.5 was significantly higher than that in the zinc deficiency group and the low zinc group,differences are statistically significant(P<0.05).Experiment 2:Real-Time PCR result prompt:Compared with the normal zinc group,the expression of Sp5 m RNA in the palatine process in the zinc deficiency group increased significantly at ED14.5(P<0.05),The expression level of Sp8 m RNA decreased slightly,but the difference was not statistically significant(P>0.05).Compared with the normal zinc group,the expression of Sp5 m RNA in the palatine process in the zinc rich group decreased significantly(P<0.05),The expression level of Sp8 m RNA was slightly higher,and the difference was not statistically significant(P>0.05).Conclusion:(1)The content of trace element zinc is closely related to the occurrence of cleft palate,and the deficiency of zinc is very easy to cause cleft palate.(2)The maternal zinc deficiency during pregnancy can inhibit embryonic palate,mesenchymal cell proliferation,resulting in the palatal morphology variation,lifting power weakened,and ultimately the formation of cleft palate(3).Zinc deficiency can increase the expression of Sp5 in the fusion stage of offspring,and the expression of Sp5 can be reduced in the fusion stage of offspring embryos,suggesting that Sp5 plays a negative role in regulating the development of palate... |
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