Protective Effects Of Sitagliptin On Bone In Type 2 Diabetic Rats And Its Relationship With Oxidative Stress

Objective 1. To explore the effects of sitagliptin on BMD, bone histomorphology and bone turnover markers in rats with type 2 diabetes induced by streptozotocin(STZ) plus high fat and high sugar diet. 2. To observe the relationship between the protective effects of sitagliptin on bone in type 2 diabetic rats and oxidative stress; 3. To explore the relationship between the protective effects of sitagliptin on bone in type 2 diabetic rats and the expression of p-ERK1/2/PPARγ.Methods 1. Rat model of type 2 diabetes: SD rats were fed with high fat and high sugar diet for 8 weeks, and injected intraperitoneally with STZ(30mg/kg). Subjects with the fast blood glucose(FBG) ≥16.7 mmol/L in three consecutive days after 72 hours were successfully modeled. 2. Sitagliptin intervention: After modeled, SD rats were randomly divided into three groups: NC group, T2 DM group and SIT group(mean n = 10). Subjects in SIT group were given a gavage of sitagliptin, while the NC group and T2 DM group were given a gavage of distilled water of the same volume with the drug. 3. The detection of relevant indicators: blood glucose and weight were detected conventional method; Intraperitoneal glucose tolerance test(IPGTT) was conducted and calculate the area under the curve(AUC); DEXA was used to measure BMD; HE staining was used to observe the femur histomorphology; Radioimmunoassay was used to detect the serum insulin; ELISA was used to detect serum OCN and TRACP-5b activity; AMP-buffer was used to detect the serum ALP level. 4. The detection of gene and protein: RT-PCR and western blot were used to detect the m RNA and protein expression of femur osteoblast p-ERK1/2, PPARγ, p47 phox, Bcl-2 and Bax respectively. Results 1. After modeled, FBG of DM group was significantly higher than NC group(P<0.05), mean FPG was(18.46±1.67) mmol/L, and serum INS was lower than NC group(P<0.05). 2. Sitagliptin intervention 20 weeks, FBG of T2 DM and SIT group were higher than NC group(P<0.05), while FBG of SIT group was lower than T2 DM group; Weight of T2 DM group was lower than NC group(P<0.05). Serum INS of T2 DM group and SIT were lower than NC group(P<0.05), and serum INS of SIT group was higher than T2 DM group(P<0.05). Each point of blood sugar and AUC of IPGTT in T2 DM and SIT group were higher than NC group(P<0.05); Each point of blood sugar and AUC of SIT group were beween NC and T2 DM group(P<0.05). 3. The expression of bone formation marker ALP and OCN in T2 DM group were lower than NC group(p<0.05), bone sorption marker TRACP-5b in T2 DM group was higher than NC group(p<0.05); ALP of SIT group was lower than NC group and higher than T2 DM group(p<0.05); OCN of SIT group was higher than T2 DM group(p<0.05), TRACP-5b of SIT group was lower than T2 DM group. BMD of lower limbs and spine in T2 DM group were lower than NC group(p<0.05); BMD of lower limbs in SIT group was higher than T2 DM group(p<0.05), and the BMD of spine in SIT group were between NC and T2 DM group, had no statistical significance(p>0.05). 4. Histomorphology analysis of bone tissue: T2 DM group: thin bone cortex, disorder bone trabecular, wide spacing, marrow cavity significant expansion, the number of osteoblasts reduced, the bone lacunar cells significantly increased. SIT group: bone plate level was clear, cortex structure was dense, bone trabecular was order, spacing slightly narrow, marrow cavity narrow, bone cells were clear and complete, few bone lacunar cells. 5. The expression of femur osteoblast p-ERK1/2, PPARγ, p47 phox and Bax/Bcl-2 m RNA and protein were higher than NC group(p<0.05); The expression of p-ERK1/2, p47 phox and Bax/Bcl-2 in SIT group were lower than group T2DM(P<0.05), whereas compared with T2 DM group, and the expression of PPARγ m RNA and protein in SIT group had no significant difference(P>0.05). Conclusion 1. Sitagliptin can reduce the blood sugar effectively and improve the function of islet in rats with type 2 diabetes induced by STZ plus high fat and high sugar diet. 2. Sitagliptin can increase bone mineral density, improve bone microstructure, promote bone formation and inhibit bone sorption in type 2 diabetic rats. 3. The protective effects of sitagliptin on bone in type 2 diabetic rats probably be related with the antagonism of the expression of osteoblast oxidative stress-related genes p-ERK1/2, p47 phox and Bax / Bcl-2. 4. The protective effects of sitagliptin on bone has no relationship with the expression of osteoblast PPARγ in type 2 diabetic rats.