The Influence Of Zinc On Apoptosis Of Palatal Process Fusion And The Study Of Sp Family Candidate Gene Based On Cleft Palate

Objective: Through this experimental study,it was recognized that the effect and influence of trace element zinc on the development of palatal tissue during the fusion stage of palatal tissue and further screening of the candidate gene of zinc finger protein Sp family between cleft palate and normal development of palatal tissue,with a view to providing theoretical basis for the prevention of cleft palate in humans.Methods:1.Four groups of pregnant mice were fed with zinc-rich feed(zinc 100 mg/kg),normal zinc feed(zinc 30 mg/kg),low zinc feed(zinc 5 mg/kg)and zinc deficient feed(zinc 0mg/kg).Each group of 3 pregnant mice took off the neck to be put to death at ED14.5and the embryo head was made of paraffin embedded and paraffin sections.The morphological development of palatal tissue during the fusion of palatal embryogenesis was observed by hematoxylin and eosin staining and TUNEL staining to observe cell apoptosis in palatine fusion stage of palatal tissue respectively.2.For the rich zinc group,the normal zinc group and the zinc deficiency group,each group of 3 pregnant mice took off the neck to be put to death at ED14.5 and the fetal mice were taken out.Under the stereoscopic microscope,surgical instruments and vessels treated with DEPC were used to separate the palatal tissue and then the total RNA was extracted from the palatal tissue by Trizol method.After the quality check is qualified,we send the qualified samples to the gene company for quality inspection again.Through the agilent expression spectrum chip hybridization,there are three groups of agilent chips' expression information were collected and the expression differences between the groups were compared,screening differential gene using 2 times foldchange and analysising whether there are differences in gene expression of zinc finger protein Sp family.Results:1.The palatal tissue of the rich zinc group was well developed to form the complete palatal plate.The bilateral palatal tissues of the normal zinc group were in contact with the fusion.ow zinc groups' bilateral palatal tissue completed rising to the near and far distance,can't form cleft palate fusion.The bilateral palatal tissue in the zinc deficiency group was vertical to growth and was short in development and it cannot be fuse to form cleft palate.2.Rich zinc group of palatal plate can be seen scattered apoptotic cells in mesenchymal cells.The apoptotic cells of the normal zinc group were slightly higher and focus on the palatine raphe and surrounding stroma cells of palatal plate in the middle area.Low zinc groups' bilateral palatal tissue mesenchymal cells have more apoptosis cells.Zinc deficiency group of bilateral palatal tissue mesenchymal cells have a large number of apoptotic cells.3.Rich zinc group compared with zinc deficiency cleft palate group 2 times differentially expressed genes' number is 663.Rich zinc group compared with normal zinc group 2 times differentially expressed genes' number is 97.Zinc deficiency cleft palate group compared with normal zinc group 2 times differentially expressed genes' number is 97.4.Rich zinc group compared with zinc deficiency cleft palate group 2 times differentially expressed genes' number is 663,there were 513 up-regulated genes and 150down-regulated genes.Sp5 was found to be involved in the screening of these significantly down-regulated differential genes and the average expression level of Sp5 in the zinc rich group was 6.787452825 times lower than zinc deficiency cleft palate group and P< 0.05 was statistically significant.Conclusion:1.In the absence of zinc,the apoptotic cells of the palatal tissue were more and the palatal tissue was poorly developed,which obstructed the development of the palatal tissue.In the condition of rich zinc,the epithelium of the palatal suture disappeared early,the apoptotic cells of the palatal interstitial cells were the least and the palatal tissue developed well to form the complete palatal plate,which promoted the development of the palatal tissue.Trace element zinc plays a key role in the development of palatal process.2.Zinc finger protein Sp5 may be a candidate gene for the cleft palate and Sp5 may play a negative role in the development of palatal tissue.