Spatial And Temporal Differences And Significance Of 3M Syndrome-associated Proteins And LC3 Proteins In Normal Mouse Growth Plates

Objective:In this study,the thickness of each growth band of normal mice was observed at 0-4 weeks of age.The expression positions and levels of CUL7,CCDC8 and LC3 proteins in growth plates of mice aged 0-4 weeks were changed.To elucidate the role of CUL7,CCDC8 proteins and autophagy in the proliferation and differentiation of normal mouse ti-bial growth plate.Methods:1.The changes of resting zone,proliferation zone,hypertrophic zone and overall thickness of tibia growth plate in normal mice of 0-4 weeks were observed by HE staining.2.The expression positions and levels of Cul7,CCDC8 and LC3 roteins in the tibial growth plate of normal mice aged 0-4 weeks were observed by immunohistochemical staining.3.The total protein expression of CCDC8 and LC3 in the tibial gowth plate of normal mice aged 0-4 weeks was detected by Western blot.Results:1.The thickness of each band and total thickness of tibial growth plate of mice aged 0-4 weeks gradually thinned with the increase of week of age(the difference was statistically significant,F=477.291,P < 0.001).The thickness of the stationary zone decreased most significantly,followed by the proliferative zone and the hypertrophic zone.2.CUL7 protein was expressed in the cytoplasm and membrane of tibial growth plate in the resting zone,proliferative zone and hypertrophic zone of 0to 4 weeks of normal mice.The expression level and total expression level of CUL7 protein in each zone of growth plate cartilage gradually decreased with the increase of week of age(the difference was statistically significant,F=369.61,P < 0.001).The expression of CUL7 protein decreased most significantly in the proliferative zone,followed by the stationary zone and the hypertrophic zone.3.CCDC8 protein was mainly expressed in the proliferation zone and hypertrophic zone cell membrane and nuclear membrane of the growth plate at0-2 weeks,and mainly expressed in the hypertrophic zone cell membrane and nuclear membrane at 3-4 weeks.The expression of CCDC8 protein in the proliferating zone decreased with the increase of week of age,and the expression of CCDC8 protein in the hypertrophic zone increased with the increase of week of age(the difference was statistically significant,F=453.67,P < 0.001).The total expression of CCDC8 protein in growth plates decreased with the increase of week of a-ge.4.The LC3 protein is mainly expressed in the nuclear membrane in the resting zone and proliferation zone in the early stage,and in the cytoplasm and nuclear membrane in the proliferation zone and the hypertrophic zone in the later stage.As the age increases,the expression of LC3 protein in the resting zone and the proliferation zone gradually decreases.The expression of LC3 protein in the hypertrophic zone increases first,and then gradually decreases.The total LC3 protein expression in the growth plate decreased with increasing age(the difference was statistically significant,F=617.43,P<0.001).Conclusions:1.The thickness and total thickness of the growth plate of tibia decreased with the increase of weeks of age,especially the thickness of the stationary zone and the proliferative zone,which indicated that the chondrocytes of the stationary zone and proliferative zone played an important role in promoting the linear growth of long bone.2.CUL7 protein was expressed in the cytoplasm and membrane of chondrocytes in the growth plate resting zone,proliferating zone and hypertrophic zone.CCDC8 protein was expressed in the cell membrane and nuclear membrane of growth plate proliferation zone and hypertrophic zone LC3 was expressed in the cytoplasm and nuclear membrane of the quiescent zone and the proliferative zone at the early stage,and mainly in the proliferative zone and the hypertrophic zone at the late stage.3.CUL7 and CCDC8 proteins expressed in growth plate chondrocytes can promote the proliferation and differentiation of chondrocytes.4.Autophagy plays a role in the process of growth plate chondrocyte proliferation and differentiation.