| Background:All-trans retinoic acid is an intermediate metabolite of vitamin A in vivo,which plays an important role in regulating the morphological formation of vertebrate embryogenesis.In target tissues,all-trans retinoic acid activates specific nuclear receptors and regulates the transcription of multiple genes to achieve the main physiological functions of vitamin A,including regulating cell proliferation and differentiation and maintaining normal immune function.The epiphyseal plate,also known as the growth plate,is a unique structure of children and is an important part of the growth and development of the growing bone.A series of events such as proliferation,hypertrophy and apoptosis of epiphyseal chondrocytes determine the longitudinal growth potential of bones.When the epiphyseal plate is damaged by various internal and external factors,the height of children in adulthood will be affected and malformation may occur.In addition,children’s body functions are not yet perfect,so they are susceptible to the effects of drugs in the process of organ developmentAs living standards have improved,vitamin A deficiency has improved markedly in rapidly developing countries,while vitamin A overdose has crept in.From the perspective of clinical use,preventive supplementation with vitamin A supplements or cod liver oil(mainly vitamin A and vitamin D)without professional guidance is on the riseThis experiment to explore in vitro high concentrations of all trans retinoic acid on SD rat epiphyseal cartilage cell biology behavior and function,and explore in excess all trans retinoic acid gastric SD rats after tibial growth plate in the body and brain,heart,lungs,liver,kidney,spleen and other organs,in the hope to improve on vitamin A and all trans retinoic acid treatment of relevant knowledge,to provide the clinical application of the new train of thought,also for the future of the study provide valuable information.Part I Effects of all-trans retinoic acid on the biological behavior and function of epiphyseal chondrocytesObjective:To investigate the effect of high-concentration all-trans retinoic acid(atRA)on the proliferation,morphology,apoptosis,migration and function of chondrocytes in growth plate of SD rats and the effect of excessive atRA on the growth plateMethods:SD rat epiphyseal cartilage cells as the research object,atRA for interference factors,adopt CCK-8,cytometry,HE staining,the Annexin V-FITC cells apoptosis flow detection technique,Hoechst staining,cell scratches,Transwell experiment evaluation respectively after atRA treatment cell proliferation,cycle,morphology,apoptosis and migration,Western-blot detection proteoglycan,collagen-Ⅱ,collagen-X protein changes;In three-week-male SD rats as experimental object,were randomly divided into control group,40 mg/kg·d-1 atRA group,60 mg/kg·d-1 atRA group,80 mg/kg·d-1 atRA group,for ten consecutive days atRA lavage treatment,measuring each SD rats lavage on the first day,the end of the tenth day,after ten days HE staining was carried out on the tibial growth plate processingResults:After the effect of high concentration atRA on the epiphyseal chondrocytes of SD rats,the proliferation ability was decreased and the cycle of epiphyseal chondrocytes of SD rats was blocked in S phase(P<0.01).The cell morphology changed from triangle and polygon to long strip.The occurrence of apoptosis increased(P<0.01).The migration ability was inhibited(P<0.05).Western-blot results show proteoglycan,collagen-Ⅱ,collagen-X related protein expression than the control group were significantly decreased(P<0.01).After atRA intragastric administration in SD rats,compared with the control group,both the 60 mg/kg·d-1 atRA group and the 80 mg/kg·d-1 atRA group had shorter length of head and tail(P<0.01),no statistical significance was found in 40 mg/kg·d-1 atRA group.HE staining showed that the total thickness of growth plate in the atRA gavage group was narrowed or even closed,and the thickness of proliferative and hypertrophic zones was decreased.Conclusion:This study confirms the high concentrations in vitro all-trans retinoic acid can epiphyseal cartilage cells of SD rats inhibition of proliferation,migration,at the same time can induce apoptosis,reduce the related functional protein expression,also excessive in SD rats that all-trans retinoic acid can affect the normal growth and development of growth plate,eventually make the growth plate part or all of the early closure,in turn,affect the growth of SD rat body length.Part Ⅱ Effects of all-trans retinoic acid on parenchymal organ injury in developmental SD ratsObjective:To investigate the effects of overdose all-trans retinoic acid(atRA)on brain,heart,lung,liver,kidney and spleen in Developmental SD rats.Methods:Forty-eight male SD rats,3-week-old,were divided into control group and three treatment groups at the doses of 40,60,and 80 mg/kg·d-1 of atRA,respectively,12 rats each group,the rats were administrated by gavage every day for 10 days.The rats body weight were weighed every day.After treatment,the rats were sacrificed and then rat heart,lung,liver,kidney,and spleen tissues were weighed,respectively.The organ index was calculated.The organs were then stained with HE.Results:After atRA gavage,compared with control group,the 40 mg/kg·d-1 atRA group had higher kidney index and body weight change was not statistically significant,the 60 mg/kg·d-1 atRA group had lower body weight,higher heart and kidney index,and lower spleen weight,and the 80 mg/kg·d-1 atRA group had significantly lower body weight,higher brain,heart and kidney index,and lower brain and spleen weight.HE staining showed that compared with the control group,SD rats in the 40,60 and 80 mg/kg·d-1 atRA groups had thickened alveolar wall,vacuol-like changes in renal tubular epithelial cells,and more macrophages in the spleen,but no significant histological changes in the brain,liver and heart.Conclusion:This study preliminarily confirmed that excessive all-trans retinoic acid in vivo could damage the lungs,kidneys and spleen in Developmental SD rats. |
