Effects of All-Trans Retinoic Acid on the Biological Behavior and Function of Epiphyseal Chondrocytes

WANG Xia^1,2, BAI Haodi^1,2, ZHOU Yin², LIU Xing^1*

(¹Department of orthopedics; Ministry of Education Key Laboratory of Child Development and Disorders, National Clinical Research Center for Children Health and Diseases (Chongqing); China International Science and Technology Cooperation Base of Child Development and Critical Disorders; Children’s Hospital of Chongqing Medical University, Chongqing 400016, China; ²Chongqing Key Laboratory of Pediatrics, Chongqing Engineering Research Center of Stem Cell Therapy, Chongqing 400016, China)

Abstract:

This study investigated the effect of high concentration of ATRA（ all-trans retinoic acid) in vitro on the biological characteristics and function of epiphyseal cartilage cells in SD rats and the effect of ATRA in vivo on tibia growth plate in SD rats. The epiphyseal chondrocytes of SD rats were studied and ATRA was used as the intervention factor. CCK-8, cytometry, HE staining, Annexin V-FITC apoptosis flow cytometry, Hoechst staining, cell scratches and Transwell experiment were used to evaluate the cell proliferation, cycle, morphology, apoptosis and migration after ATRA treatment. The changes of proteoglycan, collagen-II and collagen-X related functional proteins were detected by Western blot. Male SD rats at 3-weeks were divided into control group, 60 mg/kg·d ATRA group and 80 mg/kg·d ATRA group for continuous gavage for 10 days. The length of the head and tail of each SD rat was measured on the first day and the tenth day of gavage, and the tibia growth plate was stained with HE after 10 days of treatment. The results showed that, compared with the control group, the proliferation ability was weakened and the cell cycle was blocked in S phase (P<0.01). Compared with the control group, the occurrence of apoptosis increased (P<0.01) and the migration ability was inhibited (P<0.05). Western blot results showed that the expressions of proteoglycan, collagen-II and collagen-X were significantly lower than those of the control group (P<0.01). Compared with the control group, the head and tail length of the ATRA treatment group were all shorter (P<0.01). HE staining of tibia growth plate showed that the growth plate in the ATRA gavage group was narrowed or even closed. This study confirmed that high concentration of ATRA could inhibit cell proliferation, inhibit cell migration, induce cell apoptosis, and reduce the expression of related functional proteins. Excessive ATRA has been shown to affect the endochondral osteogenesis of growth plates in SD rats. Finally, ATRA caused the epiphyseal plate to be partially or completely closed in advance, thus affecting the growth of body length of SD rats.

CSTR: 32200.14.cjcb.2020.02.0008