The Role Of Autophagy In The Pathogenesis Of Glucocorticoid-Induced Osteoporosis

Objective To study the effect of different dose of Dex impact on bone density,osteogenesis and autophagy in rats at different times,and to investigate the role and possible mechanism in glucocorticoid-induced osteoporosis(GIOP).Methods 120 SD rats in 3-month were randomly divided into 4 groups of 30 each:control group(Cont,saline),low dose Dex group(1 mg/kg,Dex),medium dose Dex group(2.5 mg/kg,Dex)and high dose Dex group(5 mg/kg,Dex).Dex was injected intramuscularly twice a week for 4-,9-,and 12-week.Bone mineral density(BMD)was measured using dual energy X ray absorptiometry.The expression of ALP,Osteocalcin,Collagen I,LC3,Beclin1,ATG5,AKT,m TOR m RNA in bone tissue was measured with In-situ hybridyzation.The expression of ALP,Osteocalcin,Collagen I,LC3,Beclin1,ATG5,AKT,P-AKT,m TOR,P-m TOR protein in bone tissue was measured with immunohistochemistry.Results1.Before the Dex treatment,the total BMD were no difference among the four groups(P(29)0.05);After 4-week intervention,the total BMD in different dose of Dex were no difference compare with control group(P(29)0.05);After 9-week treatment,the total BMD in different dose of Dex groups were lower than control group(P<0.05),the BMD was significantly lower in low dose Dex group than those of high dose Dex group(P<0.05);After 12-week intervention,the total BMD was significant differences in four groups,the BMD were significantly lower in high dose Dex group than low dose Dex group and medium dose Dex group(P<0.05).2.The expression of OCN,Collagen I m RNA and OCN protein in the low dose Dex group were higher than the control group(P < 0.05)for 4 weeks,the expression of ALP,OCN and I collagen m RNA and protein in the other groups was no difference(P > 0.05).The expression of ALP,OCN,Collagen I m RNA and protein were lower in the medium dose Dex group / high dose Dex group than the control group(P < 0.05),and then decreased with the increase of dosage,the expression of Collagen I,OCN protein and Collagen I m RNA in the low dose Dex group were not different from the control group(P > 0.05);After treated with Dex for 12-week,the expression of ALP,OCN,Collagen I m RNA and protein in Dex group were lower than the control group(P < 0.05),the expression of ALP,OCN,Collagen I m RNA and protein were decreased gradually with the dose and duration increasing(P < 0.05),bone formation was decreased in time-and dose-dependent in the osteoblasts.3.After treated Dex for 4-weeks,the expression of LC3,Beclin1,ATG5 m RNA and protein in the low dose Dex group and medium dose Dex group were higher than the control group(P < 0.05),the LC3,Beclin1 m RNA and proteins in high dose Dex group were higher than the control group(P < 0.05).For 9-week,the expression of m RNA and protein in LC3,Beclin1,ATG5 in the medium dose Dex group and high dose Dex group were decreased(P < 0.05),the expression of LC3,ATG5 m RNA and protein in the low dose Dex group were not different from the control group(P > 0.05)but the Beclin1 m RNA and protein expression were lower than the control group(P < 0.05).After 12-weeks treatment,the expression of LC3,ATG5,Beclin1 m RNA and protein in different dose of Dex groups were lower than the control group(P < 0.05),The expression of LC3,ATG5,Beclin1 m RNA and protein wrere decreased gradually with the increasing of dosage and duration.These results indicated that Dex induced autophagy in osteoblast at the early treatment,lower Dex promoted the autophagy.Whereas the prolonged and excessive Dex may inhibited the autophagy in osteoblast by time-and dose-dependent.4.Except the expression of m TOR m RNA in medium dose Dex group after 9-week treatment and AKT m RNA in high dose Dex group after 12-week treatment were lower than the control group(P < 0.05),others groups’ AKT/ m TOR m RNA and protein were no difference after treated for 4-,9-,12-weeks.The expression of P-AKT,P-m TOR in different dose of Dex groups were lower than the control group(P<0.05),Dex had dose-dependent inhibited the phosphorylation of AKT/ m TOR.Conclusion Dex may induce autophagy by inhibiting the AKT/m TOR pathway in treatment of 4 weeks,autophagy can protect the bone metabolism of rats.However,the autophagic activity and bone formation in osteoblast,and bone mass were decreased with the increasing of duration and dosage.Autophagy may has a protective effect in GIOP.