| Objective:In this study,a rat model of type 2 diabetes mellitus was established by high-fat diet and stteptozootocin injection.The role of exercise intervention in improving the damage of bone microstructure and bone mass loss was studied,and the mechanism of exercise promoting bone synthesis and preventing osteoporosis in diabetic rats was explored from the perspective of Irisin regulating the changes of ERK/MAPK pathway.Methods:Thirty-two 8-week-old Wistar rats were purchased from the Experimental Animal Center of Yangzhou University and randomly divided into control group(C)and diabetic group(DM).After 1 week of adaptive feeding,DM group was given pure high-fat diet for 4 weeks.After 4 weeks,DM group was intraperitoneally injected with 30mg/kg STZ,and C group was intraperitoneally injected with the same dose of sodium citrate buffer.After the modeling,the blood samples were collected from the caudal vein to measure the blood glucose on the 3rd and 7th day,and the glycemic index≥16.7mmol/L was considered as the successful modeling.After modeling,rats had a static rest for a week and were randomly divided into diabetes control group(D),diabetes+exercise group(DE),diabetes+exercise+RGDyk(Irisin receptor pathway blocker)group(DER).After 1 week of treadmill adaptation training,rats in exercise group began formal training at a treadmill running speed of 18m/min for 45 minutes each time.Five times a week for eight weeks.During exercise,the rats in DER group were given caudal intravenous injection of RGDyk twice a week,with a dose of 2.5 mg/kg and a concentration of 1mg/ml,and the rats in DE group were injected with the same dose of DMSO solution,blood glucose and body weight were measured once a week,and all the rats were anesthetised to death after the last week of training,and blood and femur were collected.The morphometric indexes of rat femur were detected by microCT,the contents of serum Irisin and BAP were detected by ELISA kit,the number of OB in femur tissue was detected by ALP staining,and the mRNA expressions of RUNX2,ATF4,FNDC5 and ERK in femur were detected by RT-PCR.Western Blot was used to detect the expression of RUNX2,ATF4,Irisin and ERK proteins in rat femur.SPSS25.0 was used for statistical analysis of the data.The data were reported by mean±standard deviation.P<0.05 was statistically significant.Results:(1)Changes in body weight.There was no significant difference in body weight between the four groups before the experiment.After exercise intervention,compared with group C,the body weight of rats in group D was significantly decreased(P<0.05),the body weight of rats in group DE after exercise intervention was slightly increased(P<0.05),and the blocking of Irisin receptor pathway by RGDyk caused a significant decrease in body weight of rats in group DER(P<0.05).(2)Changes of blood glucose,serum Irisin and BAP.After the 8-week exercise intervention experiment,blood glucose level in group D was significantly increased compared with group C(P<0.05);Blood glucose level in DE group after exercise intervention was significantly decreased compared with group D(P<0.05);Compared with DE group,the blood glucose level in DER group increased,but there was no significant difference.Compared with group C,serum Irisin concentration of rats in group D was significantly decreased(P<0.05);After 8 weeks of exercise intervention,serum Irisin in DE group was significantly higher than that in D group(P<0.05);Compared with DE group,serum Irisin in DER group was significantly increased by blocking Irisin receptor pathway with RGDyk(P<0.05).Compared with group C,the serum BAP concentration in group D was significantly decreased(P<0.05);Compared with group D,exercise intervention resulted in a significant increase in BAP concentration in DE group(P<0.05).Compared with DE group,BAP in DER group was significantly decreased(P<0.05).(3)Changes in bone microstructure.Compared with group C,the cancellous bone BMD(P<0.05),BV/TV(P<0.05),Tb.Th(P<0.05),Tb.N(P<0.05)were significantly decreased in group D,and Tb.Sp was significantly increased(P<0.05).Compared with group D,BMD(P<0.05),BV/TV(P<0.05),Tb.Th(P<0.05),Tb.N(P<0.05)were significantly increased in DE group,and Tb.Sp was significantly decreased(P<0.05).Compared with DE group,BMD(P<0.05),BV/TV(P<0.05)and Tb.Th(P<0.05)in DER group were significantly decreased,while Tb.N showed a downward trend,but the difference was not obvious,and Tb.Sp increased,but the difference was not statistically significant.Compared with group C,cortical bone BMD(P<0.05),BV/TV(P<0.05),Tb.Th(P<0.05)and Tb.N(P<0.05)in group D were significantly decreased,while Tb.Sp was significantly increased(P<0.05);Compared with group D,BMD(P<0.05),BV/TV(P<0.05)and Tb.Th(P<0.05)of rats in DE group were significantly increased,Tb.N was increased but not significantly changed,and Tb.Sp was significantly decreased(P<0.05);Compared with DE group,BV/TV in DER group was significantly decreased(P<0.05),Tb.Th and Tb.N were decreased without significant difference,and Tb.Sp was increased without statistical significance.(4)Changes in ALP staining of femur and ALP+OB quantity.Compared with group C,the number of ALP+OB in femur of group D was significantly decreased(P<0.05).Compared with group D,the number of ALP+OB in DE group after 8 weeks of training was significantly increased(P<0.05);However,blocking Irisin receptor pathway with RGDyk in DER group resulted in the disappearance of the stimulative effect of exercise to increase the amount of ALP+OB(P<0.05).(5)mRNA expression changes.Compared with group C,RUNX2(P<0.05),ATF4(P<0.05),FNDC5(P<0.05)and ERK(P<0.05)in femur of group D were significantly decreased.Compared with group D,RUNX2(P<0.05),ATF4(P<0.05)and FNDC5(P<0.05)in DE group were significantly increased after 8-week exercise intervention.Compared with DE group,mRNA expression levels of RUNX2(P<0.05),ATF4(P<0.05)and ERK(P<0.05)in femur of DER group were significantly decreased by blocking Irisin receptor pathway with RGDyk,while mRNA expression of FNDC5 was not significantly changed(P>0.05).(6)Protein expression changes.Compared with group C,the protein expressions of RUNX2(P<0.05),ATF4(P<0.05),Irisin(P<0.05)and ERK(P<0.05)in femur of group D were significantly decreased.Compared with group D,RUNX2(P<0.05),ATF4(P<0.05),Irisin(P<0.05)and ERK(P<0.05)in DE group were significantly increased after 8-week exercise intervention.Compared with DE group,RUNX2(P<0.05),ATF4(P<0.05)and ERK(P<0.05)in DER group were significantly decreased,but Irisin changes were not statistically significant(P>0.05).Conclusion:(1)Exercise can improve the loss of bone mineral density,bone microtissue destruction and bone mass loss caused by diabetes.(2)Exercise significantly promoted the synthesis of osteoblasts and the expression of their transcription factors,and this promoting effect could be blocked by RGDyk,suggesting that Irisin signaling system may be involved in the effect of exercise in increasing the synthesis of osteoblasts.(3)Exercise can increase OB synthesis,promote bone formation,improve bone microstructure and reduce bone loss by promoting Irisin expression and up-regulating ERK/MAPK signal. |
PDF Full Text Request