Effects of insulin glargine on fracture healing and osteocalcin expression in type 2 diabetic rats

doi:10.3969/j.issn.2095-4344.2017.36.003

Abstract

Abstract:

BACKGROUND: Insulin analogues have been extensively applied in the treatment of diabetes mellitus. Insulin glargine has a higher affinity for insulin like growth factor 1 receptor compared with human insulin. Further research is needed to ensure whether insulin and its analogues exert same effects on fracture healing in type 2 diabetes mellitus.
OBJECTIVE: To observe the osteocalcin expression and callus formation in the healing of fracture in type 2 diabetic rats induced by human insulin and insulin glargine, to observe the difference between two treatment methods, and to explore the related mechanisms.
METHODS: Sixty-four Sprague-Dawley rats were randomly assigned into human insulin group (group A), insulin glargine group (group B), diabetes mellitus group (group C) and control group (group D). Rats in the groups A, B and C were fed with high-sugar and high-fat diet for 4 weeks followed by intraperitoneal injection of small-dosage streptozotocin twice, to establish the rat models of type 2 diabetes mellitus. After the right tibia of each rat was broken, insulin glargine and Novolin 30R were used in the groups A and B, respectively. Fracture healing was observed on X-ray, callus formation and number of osteoblasts were observed by microscope, and serum level of osteocalcin was measured by ELISA method at 1, 2, 4, and 6 weeks after modeling.
RESULTS AND CONCLUSION: X-ray results revealed better fracture healing in the groups A, B and D than the group C. Osteoblast proliferation in callus was significantly better in the groups A, B and D than in the group C. Serum level of osteocalcin in each group was on the rise, which was significantly higher in the groups A, B and D than the group C (P < 0.05), but had no significant difference among groups A, B and D (P > 0.05). In summary, insulin glargine can increase the serum level of osteocalcin, accelerate the callus formation, and improve the healing of fracture in type 2 diabetic rats. Furthermore, there is no significant difference in the therapeutic efficacy between insulin glargine and human insulin.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Insulin, Diabetes Mellitus, Type 2, Tibial Fractures, Osteocalcin, Tissue Engineering

CLC Number:

R318

Zheng Bai1, Xin Bing2, Huang Dong2, Liu Yong-tao2, Liu Guo-dong1, Sun Bai-han1, Yuan Feng2, He Yu-ze2. Effects of insulin glargine on fracture healing and osteocalcin expression in type 2 diabetic rats[J]. Chinese Journal of Tissue Engineering Research, 2017, 21(36): 5752-5756.

Figures/Tables 1

2.1 实验动物数量分析 64只大鼠随机分为4组，其中48只建立2型糖尿病大鼠模型，造模过程中2只大鼠血糖不达标，1只大鼠甘精胰岛素干预治疗过程中血糖自行恢复正常，予以排除，实验过程中及时补足。4组大鼠均制作右胫骨骨折模型，无感染死亡，均纳入实验分析，每组16只。 2.2 X射线观察结果第1周：各组骨折线明显，治疗组骨膜增厚较糖尿病组明显。第2周：人胰岛素组、甘精胰岛素组、骨折对照组可见少量稀疏连续骨痂影，糖尿病组末见骨痂影。第4周：人胰岛素组、甘精胰岛素组、骨折对照组可见稀松连续骨痂影，骨密度增强，糖尿病组少许稀松骨痂影。第6周：人胰岛素组、甘精胰岛素组、骨折对照组可见连续骨痂影，骨密度增强，骨折线模糊。糖尿病组稀松连续骨痂影，骨折线可见(图1)。 2.3 苏木精-伊红染色观察结果第1周：人胰岛素组、甘精胰岛素组、骨折对照组可见成骨细胞数目增多，纤维性骨痂形成，骨折对照组成骨细胞数量低于人胰岛素组、甘精胰岛素组、骨折对照组，未见纤维骨痂形成。第2周：人胰岛素组、甘精胰岛素组、骨折对照组骨折断端可见纤维骨痂及软骨痂，新生血管形成，糖尿病组纤维骨痂稀松，软骨痂形成明显少于其余各组。第4周：人胰岛素组、甘精胰岛素组、骨折对照组骨折断端可见大量软骨骨痂形成，纤维母细胞数量减少。糖尿病组纤维骨痂，纤维母细胞无明显减少。第6周：人胰岛素组、甘精胰岛素组、骨折对照组骨折断端可见大量软骨骨痂，骨性骨痂形成，纤维骨痂消失。糖尿病组可见稀松软骨骨痂，骨性骨痂形成(图2)。 2.4 血清骨钙素及血糖水平各组大鼠骨折后血清骨钙素水平呈上升趋势，在第2周达高峰，第4周出现回落，第6周继续回落。通过方差齐性检验，各组骨钙素质量浓度方差齐。人胰岛素组、甘精胰岛素组予胰岛素干预治疗后随机血糖水平维持在8-16 mmol/L；糖尿病组未予干预治疗，血糖水平较高，波动在22-33 mmol/L；骨折对照组随机血糖水平保持在3.5-6.5 mmol/L。因为血糖对骨钙素的表达有影响[3]，以大鼠平均血糖水平作为协变量，采用协方差分析结果显示，各时间点，不同组别骨钙素质量浓度差异有显著性意义(P < 0.05)；对各组骨钙素质量浓度进行两两比较，发现糖尿病组低于其余3组(P < 0.05)，但人胰岛素组、甘精胰岛素组、骨折对照组3组间差异无显著意义(P > 0.05)，见表1，2。"

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