Effects Of Citral On Morphologic Changes And Expression Of Rarβ、tgfβ3during Palate Development In Chicken Embryo

Objective:The objective of our experiment is to use low concentrations of citral, which is the inhibitor of retinaldehyde dehydrogenase, and observe the effect of citral during the development of chick embryonic palate and the morphology changes on frontonasal Process. We investigated the expression of retinoic acid receptor beta and transforming growth factor beta3base on citral. These are having an effect on the chick embryonic frontonasal process. Explore the effect of retinoic acid and transforming growth factor beta3base on the development of chick embryonic palate.Methods:(1)Citral-soaked beads were implanted into the nasal pit of stage20chicken embryos, which established the citral-effected stabilized development in chicken embryo. The chicken embryo has a stable and a continuing developmental environment. The chicken embryos were divided into8groups, where each group contains20chick embryos. There were citral (CT) experimental groups, control group with dimethyl sulfoxide (DMSO) and the last group was normal group. In experimental groups SM2beads were soaked in several concentrations of CT (0.001g/ml、0.01g/ml、0.05g/ml、0.1g/ml、0.5g/ml、0.88g/ml) and the control beads were soaked in DMSO.(2)A11the experimental eggs were incubated at38℃until stage38.The chicken embryonic specimens were fixed in4%paraformaldehyde.Next he chicken embryonic specimens were stained in alizarin red or alcian blue solution. Analyzed and evaluated the morphological of the craniofacial skeletonand cartilage defects based on citral. We chose one suitable solution as the perfect experimental solution.(3) Another part of the experimental eggs were incubated at38℃until stage24、25、26、27. In the0.5g/ml CT groups and DMSO groups,we took20specimens at each stages. Making3μm sections by normal method, we investigate the expression patterns of the retinoic acid receptor beta and transforming growth factor beta3through Immunohistochemistry.Results:Different concentrations of CT have different effects on the chick craniofacial skeleton during the development of chick embryonic palate. There were no differences in the CT and DMSO groups. There were two specimens with lose nasal bone in the0.1g/ml CT group. All of the specimens in the0.5g/ml and0.88g/ml CT groups had detectable effect.0.5g/ml CT can decrease the Rara and TGFa3, and when compared with the control group it had a significant difference (P<0.05).Conclusions:Citral locally inhibits the retinaldehyde dehydrogenase blocking RA synthesis in the facial development of embryo. The result was a specific loss of derivatives from the lateral nasal prominences. When we implanted the0.5g/ml CT-soaked beads into the chick nasal pit, citral down-regulated the expression of retinoic acid receptor a and transforming growth factor beta3, which obstructed nasofrontal process growth. The final result lead to the palatine process being formed incompletely introducing the unilateral palate defect. We modified the embryonic facial process at the early stage of embryonic development. Exogenous factor be restricted at the maxillofacial region decreasing the maternal interference factors. It is easy to control the required conditions for building the cleft palate animal models in chicken embryos. We could directly observe the craniofacial development in early embryonic stage through the eggshell windows. Using chick embryo as a kind of cleft palate animal model is a good supplement of model in the mammal research.