| Objective : To investigate whether long non-coding RNA myocyte enhancer factors 2C-antisense RNA 1(lnc RNA MEF2C-AS1)can affect the bone mass and bone mineral density in normal and osteoporosis rats by down-regulating the expression levels of myocyte enhancer factors 2C(MEF2C)and sclerostin protein(SOST)in vivo.Methods:(1)Forty-six five-month-old SD female rats were randomly divided into sham operations(SHAM)and ovariectomized group(OVX),with 23 rats in each group.Bilateral ovaries were removed by dorsal approach in the OVX group,and adipose tissue of the same size around ovaries was removed by the same method in the SHAM group.At 12 weeks after the operation,rats in each group were randomly dissected to verify the success of the osteoporosis model.(2)The remaining rats were randomly divided into sham control(SHAM)group 、 SHAM+MEF2C-AS1 group 、 ovarian control(OVX)group 、OVX+MEF2C-AS1 group,with 8 rats in each group.Then the corresponding lentivirus vector of MEF2C-AS1 and an equivalent empty lentivirus vector were injected.(3)After 8 weeks,when the rats were executed,the bilateral femurs and blood were collected for Western blot,ELISA analysis and micro-CT imaging.Results:(1)At 12 weeks after the operation,in order to verify the success of the osteoporosis rat model,Micro-CT was used to detect the changes in bone mass of each group.It was found that the Bone volume(BV)、Percent bone volume(BV/TV)、Trabecular number(Tb.N)、Trabecular thickness(Tb.Th)and bone mineral density(BMD)in OVX group were decreased compared with SHAM group(all p<0.05),while the Trabecular separation(Tb.Sp)was significantly higher than that of the SHAM group(p<0.05),which confirming that the rat osteoporosis model was successfully established.(2)After 8 weeks of lentivirus vector injection,in order to test the effect of MEF2C-AS1 on MEF2 C in rats,the Western blot analysis was used to find that the MEF2 C protein expression levels in the SHAM group,OVX group,SHAM+MEF2C-AS1 group,OVX+MEF2C-AS1 group were different(p<0.05).The MEF2 C protein in OVX group was significantly higher than in SHAM group(p<0.05);the protein level of MEF2 C in SHAM+MEF2C-AS1 group was slightly lower than that in SHAM group,the difference was not statistically significant(p>0.05).However,the MEF2 C protein in OVX+MEF2C-AS1 group decreased compared with OVX group(p<0.05).(3)The ELISA analysis showed that the MEF2 C protein expression levels in the SHAM group,OVX group,SHAM+MEF2C-AS1 group,OVX+MEF2C-AS1 group were different(p<0.05).The SOST protein expression in OVX group was higher than that in SHAM group(p<0.01);the protein level of SOST in SHAM+MEF2C-AS1 group was lower than that in SHAM group,the difference was not statistically significant(p>0.05);the SOST protein in OVX+MEF2C-AS1 group decreased compared with OVX group(p<0.01).(4)The effect of MEF2C-AS1 on the bone mass of rats was further detected by Micro-CT imaging,the Bone volume(BV)、Percent bone volume(BV/TV)、bone mineral density(BMD)、Tissue mineral density(TMD)、Trabecular thickness(Tb.Th)、Trabecular number(Tb.N)、Cortical bone thickness(Ct.Th)in SHAM+MEF2C-AS1 group were higher than those in SHAM group,but the differences were not statistically significant(all p>0.05).The Bone volume(BV)、Percent bone volume(BV/TV)、Trabecular number(Tb.N)、Trabecular thickness(Tb.Th)、bone mineral density(BMD)in OVX+MEF2C-AS1 group were higher than OVX control group(p<0.05).Conclusion:(1)5-month-old SD female rats can successfully replicate the osteoporosis model after 12 weeks of ovariectomy.(2)lnc RNA MEF2C-AS1 can inhibit the expression of sclerostin by down-regulating MEF2 C protein expression,thereby promoting bone formation,increasing bone mineral density,improving bone microstructure of the trabecular in rats with osteoporosis after ovariectomization,which has potential therapeutic significance for osteoporosis.However,the effect of increasing bone mass in normal rats is not significant. |
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