利拉鲁肽对糖尿病前期大鼠胰岛海马胆碱能神经刺激肽前体蛋白的影响

Effects of Liraglutide on HCNP Precursor Protein Expression in Islet of Prediabetes Rats

目的明确利拉鲁肽对糖尿病前期OLETF大鼠的阻抑作用,观察对胰腺海马胆碱能神经刺激肽前体蛋白(hippocampal cholinergicneurostimulating peptide precursor protein,HCNP-pp)的影响。方法检测空腹及餐后2小时血糖,将处于糖耐量减低阶段12～14周龄OLETF大鼠随机分为3组,安慰剂组(PBO组)、100μg/kg利拉鲁肽处理组(L-100组)、200μg/kg利拉鲁肽处理组(L-200组),LETO大鼠为阴性对照组(LETO组)。12周后检测大鼠体重、空腹血糖、餐后2 h血糖和胰岛素,以免疫组织化学染色分析胰岛细胞形态学变化,应用蛋白印迹技术检测HCNP-pp及胰高血糖素样多肽-1受体(GLP-1R)蛋白含量;应用实时定量PCR检测HCNP-pp和LGP-1R、胆碱乙酰转移酶(ChAT)、3型毒蕈碱样受体(M3R)基因表达水平。结果PBO组OLETF大鼠空腹、餐后2 h血糖符合糖尿病诊断标准,而L-100组、L-200组及LETO组大鼠均未进展为糖尿病状态。PBO组OLETF大鼠体重及空腹胰岛素均明显高于L-100组、L-200组和LETO组(P<0.01)。与PBO组相比,L-200组、LETO组大鼠胰腺胰岛素阳性细胞表达密度明显增多(P<0.01),而与L-100组差别无统计学意义(P>0.05);L-100组、L-200组及LEITO组大鼠胰腺HCNP-pp蛋白相对含量明显减少(P<0.05或P<0.01);L-100组、L-200组及LEITO组大鼠胰腺GLP-1R蛋白相对含量明显增多(P<0.01);L-100组、L-200组和LETO组大鼠胰腺HCNP-pp mRNA水平明显下降(P<0.01);L-200组和LETO组大鼠胰GLP-1R,ChAT、M3R mRNA水平均明显增多(P<0.01);L-100组大鼠GLP-1R和ChAT mRNA水平均明显增多(P<0.01),而M3R mRNA水平则无变化(P>0.05)。结论利拉鲁肽可阻抑糖尿病前期进展,可能与胰腺HCNP-pp表达下降、GLP-1R表达增高及胰岛β细胞密度增加相关。利拉鲁肽改善胰岛素分泌状态,可能与胰腺HCNP增多、ChAT和M3R表达增高相关。

Objective To define the prohibitive effect of liraglutide in IGT-OLETF and to analyse changes of HCNP-pp. Methods IGT- OLETF were divided into 3 groups at 12- 14 weeks of age. Rats received a single intraperitoneal injection of either saline vehicle or liraglutide （100 ug/kg,200 ug/kg） twice daily for 12 weeks,LETO as normal control group. All rats were sacrificed before determining weight, insulin and glucose. The measurement of islet cell morphology by immunohistochemistry, real time quantity- PCR and Western Blot were applied to determine the expression of gene and protein. Results PBO group rats blood glucose meet the criteria for diabetes,however L-100 group,L-200 group and LETO group rats were not to develop for diabetes. Body weight and fasting serum insulin （FINS） of PBO group rats were higher in PBO rats than L-100,L-200 and LETO group（P 〈 0. 01）. Compared with the PBO group,islet positive cell density（PCD） improved significantly in L-200 group and LETO group（P〈 0. 01）,however, the difference is not statistically significant in L- 100 group（P 〉 0.05）. The relative content of HCNP- pp were higher in PBO group than L- 100（ P 〈 0.05） ,L-200（P〈0.01） and LETO group（P〈 0.01）. The relative content of GLP-1R were lower in PBO group than L-100,L- 200 and LETO group（P 〈0. 01）. The expression of HCNP-pp gene were higher in PBO group than L-200 and LETO group; however, the conversly results in ChAT,M3R and GLP-1R（P〈0.01）. The expression of M3R was not different statistically significant between PBO group and L- 100 group（ P 〉 0.05）. Conclusion Liraglutide prohibit the progression of prediabetes maybe correlation with the decreased HCNP- pp and the increased GLP- I R, Insulin- positive cells density in pancreas. Liraglutide improves the model of insulin secretion maybe correlate with the increased HCNP,ChAT and M3R.