川芎嗪与氨基胍延长胰岛素抵抗大鼠存活期的机制研究
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摘要
目的:通过链脲佐菌素(STZ)诱导当日出生的Wistar大鼠建立胰岛素抵抗(IR)模型,观察该模型32周龄时的生存率。应用川芎嗪(TMP)与氨基胍(AG)治疗,探讨TMP+AG对该模型存活期的影响及其机理。
方法:
1.实验动物及分组:出生当日Wistar大鼠,腹腔注射STZ(90mg/kg)复制IR模型。正常伺养至8周龄,空腹血糖≥7.0mmol/L或糖耐量试验餐后2小时血糖≥11.1mmol几者选入本研究。设立正常对照组(CN组)、胰岛素抵抗对照组(IR组)、二甲双胍(MET组)及川芎嗪+氨基胍(TMP+AG组)治疗组。
2.空腹血糖(FPG)、空腹胰岛素(FINS)和糖化血清蛋白(GSP)检测:分别于造模第8周、24周和32周断尾取空腹血,检测FPG,留取血清检测FINS和GSP,根据公式(FPG·FINS)/22.5计算胰岛素抵抗指数(IRI)。采用果糖胺法检测各组GSP含量。
3.生化指标检测:于实验32周,摘眼球取血,离心留取血清,全自动生化分析仪测定血清中总胆固醇(TC)、甘油三酯(TG)、尿素氮(BUN)和肌酐(Cr)。
4.外周血白细胞原位杂交:于实验结束时取肝素抗凝血约1mL,检测外周血白细胞klotho和IGF-1R mRNA,比较各组之间差异。
5.病理学检测:肾组织学切片分别行苏木素—伊红(HE)染色、糖原(PAS)染色,观察肾组织形态学变化。
6.免疫组织化学染色:观察肾组织klotho蛋白、磷酸化的胰岛素生长因子1受体(IGF-1R)蛋白表达。
7.实时定量RT-PCR:分别检测肾组织晚期糖基化终末产物受体(RAGE)、IGF-1R和klotho mRNA,脑组织IGF-1R和klotho mRNA,肝组织IGF-1R mRNA和外周血淋巴细胞的IGF-1R和klotho mRNA的表达水平。
结果:
1.各组实验动物生存率:IR组为35%(7/20),MET组为70%(14/20),TMP+AG组为85%(17/20),CN组为100%(20/20)。IR组、MET组和TMP+AG组生存率均明显低于CN组(P＜0.05),MET组和TMP+AG组生存率均明显高于IR组(P＜0.05),MET组和TMP+AG组间差异无统计学意义。
2.FPG、FINS和GSP检测结果:8周龄时,模型组的FPG、FINS、IRI和GSP含量均高于CN组(P＜0.05)。用药8周(16周龄)时,MET组和TMP+AG组FPG、FINS、IRI和GSP均低于IR组(P＜0.05),高于CN组(P＜0.05);TMP+AG组GSP含量低于MET组(P＜0.05)。用药24周(32周龄),MET组和TMP+AG组FPG和FINS较IR组进一步降低(P＜0.05);MET组和TMP+AG组FPG与CN组差异无统计学意义,FINS和IRI仍高于CN组;TMP+AG组与CN组间GSP差异无统计学意义;MET组GSP低于IR组,但仍高于CN组(P＜0.05)。
3.生化指标检测结果:各组血清TG、TC和Cr差异无统计学意义,MET组和TMP+AG组的BUN较IR组明显下降(P＜0.05)。
4.外周血白细胞原位杂交结果:IR组IGF-1R mRNA阳性细胞率明显低于CN组(P＜0.05),TMP+AG组明显低于IR组(P＜0.05),而MET组高于IR组(P＜0.05);IR组Klotho mRNA阳性细胞率明显低于CN组(P＜0.05),TMP+AG组、MET组均高于IR组(P＜0.05),TMP+AG组又高于MET组(P＜0.05)。
5.病理学检测结果:HE染色结果显示,IR组两例局部轻度系膜细胞增生,其余未见异常。PAS染色结果显示,IR组两例可见少量肾小管上皮细胞胞浆内出现糖原沉积,其余未见异常。
6.免疫组化结果:IR组肾脏远曲小管和集合小管IGF-1R蛋白表达较CN组减少(P＜0.05),MET组和TMP+AG组较IR组明显升高(P＜0.05),TMP+AG组与CN组间差异无统计学意义,MET组高于CN组(P＜0.05)。IR组肾组织Klotho蛋白含量较CN组明显下降(P＜0.05),MET组和TMP+AG组明显高于IR组(P＜0.05),MET组与TMP+AG组间差异无统计学意义。
7.实时定量RT-PCR结果:
(1)肾组织RAGE mRNA表达
IR组显著高于CN组(P＜0.05)。TMP+AG组明显低于IR组(P＜0.05),与CN组差异无统计学意义。MET组低于IR组(P＜0.05),高于CN组(P＜0.05)。
(2)IGF-1R mRNA表达:
IR组脑、肝、外周血淋巴细胞较CN组明显降低(P＜0.05)。TMP+AG组脑、肝组织较IR组明显升高(P＜0.05),外周血淋巴细胞较IR组降低(P＜0.05);CN组、IR组和TMP+AG组间肾组织表达均无明显差异。MET组肾、脑和外周血淋巴细胞表达均较IR组明显升高(P＜0.05),肝组织的表达较IR组升高,但差异无统计学意义。
(3)Klotho mRNA表达:
IR组肾、脑、外周血淋巴细胞较CN组明显降低(P＜0.05)。TMP+AG组肾、脑组织较IR组明显升高(P＜0.05),与CN组比较无统计学差异。TMP+AG组外周血淋巴细胞与IR组差异无统计学意义。MET组肾、脑、外周血淋巴细胞较IR组明显升高(P＜0.05),其中MET组肾组织的表达高于TMP+AG组(P＜0.05),而在脑组织则低于TMP+AG组(P＜0.05)。
结论:
1.STZ可以诱导出生当日Wistar大鼠产生IR。该模型FPG、FINS、IRI和GSP都随时间延长而升高,其存活期明显缩短。该模型多种组织中Klotho mRNA均表达下降,提示其高死亡率与Klotho基因表达降低密切相关。
2.TMP+AG可提高IR大鼠多种器官组织Klotho mRNA表达,抑制IR大鼠肾组织RAGE mRNA表达,并明显降低大鼠死亡率。提示TMP+AG延长IR大鼠存活时间的作用主要与调控Klotho及RAGE基因表达有关。其确切机制尚待进一步探讨。
3.该实验IR大鼠多种器官组织IGF-1R mRNA表达降低,应用TMP+AG后不同器官的IGF-1R mRNA表达改变不一致,提示IR大鼠存活期的缩短,与IGF-1R信号的转录抑制有关,尚未发现TMP+AG延长IR大鼠存活期的作用与调控IGF-1R信号转录存在直接关系。
Objective:To establish an animal modal of insulin resistant in newborn Wistar rats induced by streptozotocin(STZ)and to observe its survival rate in 32 weeks,to explore the mechanism on the prolongation of survival period effect of the combined therapy with tetramethylpyrazine and aminoguanidine.
Methods:
1.Animal and group division
The insulin resistant model of newborn Wistar rats was duplicated with streptozocin(90mg/kg).The rats were raised for 8 weeks,and those whose fasting plasma glucose higher than or equal to 7.0mmol/L or whose 2 hours postprandial blood glucose higher than or equal to 11.1 mmol/L in glucose tolerance test were selected.Rats were divided into 4 groups randomly: normal control(CN)group,insulin resistant(IR)group,the dimethybiguanide(MET)group,and the combination of tetramethylpyrazine and aminoguanidine(TMP+AG)group.
2.The measurement of fasting blood glucose(FPG)、fasting insulin(FINS) and glucosylated serum protein(GSP)Measurement
FPG、FINS and GSP were detected in 8~(th),24~(th),32~(th)weeks after treatment from the tail vein,insulin resistance index(IRI)was count by calculation formula of FPG×FINS/22.5.
3.Measurement of blood biochemical index
At the end of the experiment,the serum total cholesterol(TC), triglyceride(TG),blood urea nitrogen(BUN),creatinine(Cr)were detected.
4.In situ hybridization Test
Detection of Klotho and insulin like growth factol-1 receptor (IGF-1R)mRNA in the peripheral blood leukocytes by in situ hybridization.
5.Histological and Morphological Analyses
The hematoxylin-eosin(HE)staining and periodic acid Schiff(PAS) staining were applied in the pathological observation.
6.Immunohistochemistry staining
Klotho and phosphated IGF-1R protein were detected by immunohistochemistry staining.
7.Real-Time Fluorescent Quantitative Polymerase Chain Reaction (RT-PCR)
The mRNA levels of advanced glycation end products receptors (RAGE),IGF-1R and Klotho were quantified by RT-PCR using SYBR Green technology.
Results:
1.Survival rate of each group
At the end of the experiment,the Survival rate in IR group,MET group, TMP+ AG group and CN group were 35%,70%,85%and 100%respectively.The survival rate in CN group was significantly higher than that in the other three groups(P＜0.05),The survival rate in MET group and TMP+ AG group were higher than IR group(P＜0.05),but there was no statistically difference between the two treated groups.
2.FPG、FINS and GSP Result
The FPG、FINS、IRI and GSP in modal groups were higher than CN group at the age of 8 weeks(P＜0.05);The index above in MET group and TMP+ AG group were lower than IR group at the age of 16 weeks(P＜0.05),higher than CN group(P＜0.05);At the age of 32 weeks,The FPG and FINS in MET group and TMP+ AG group were more decreased compared with IR group at the age of 24 weeks(P＜0.05),the FPG had no significance with CN group,FINS and IRI in the two groups were higher than CN group(P＜0.05),there was no significant difference in GSP levels between TMP+ AG group and CN group.
3.Blood biochemical index Result
The differences of serum TG,TC and Cr among groups were insignificant, the BUN in MET group and TMP+AG group was decreased significantly compared to IR group.
4.In situ hybridization Result
The positive rate of IGF-1R mRNA of IR group was significantly lower than CN group(P＜0.05),which was higher in IR group than that in TMP+AG group(P＜0.05),and lower than that in MET group(P＜0.05).The positive rate of Klotho mRNA of IR group was significantly lower than CN group (P＜0.05).The expression of Klotho mRNA in both MET group and TMP+AG group were higher than IR group(P＜0.05),which is higher in TMP+AG group compared to MET group(P＜0.05).
5.Histological and Morphological Analyses
HE staining showed that two cases appeared slight mesangial cells hyperplasia and the others showing no abnormal.The epithelial cells of renal tubule occurred a little of glycogen deposition in PAS staining in IR group.
6.Immunohistochemistry Result
The expression of IGF-1R in the distal convoluted renal tubule and collecting duct from IR group was significantly lower than CN group (P＜0.05),IGF-1R expression in both MET group and TMP+AG group was higher than IR group(P＜0.05).There was no significant difference between TMP+ AG group and CN group.The IGF-IR of MET group is higher than that of CN group.The Klotho expression from IR group was deceased compared with CN group(P＜0.05),which was increased in both MET group and TMP+AG group compared to IR group(P＜0.05),and no differences in the two treated groups were found.
7.RT-PCR Analyses
(1)The expression of RAGE mRNA
The expression of RAGE mRNA of IR group in renal tissues was increased significantly compared to CN group(P＜0.05),which was higher than that of TMP+ AG group(P＜0.05),and there was no statistical difference between TMP+ AG group and CN group.The RAGE mRNA of MET group was lower than that of IR group(P＜0.05),higher than that of CN group(P＜0.05).
(2)The expression of IGF-1R mRNA
The expression of IGF-IR mRNA in brain,hepatic tissues and peripheral blood lymphocytes from IR group was significantly lower than CN group (P＜0.05),which was increased in brain and hepatic tissues of TMP+ AG group than that of IR group(P＜0.05),but decreased in blood lymphocytes than IR group(P＜0.05).No statistical differences in the expression of IGF-1R mRNA in renal tissues of CN group,MET group and TMP+ AG group were found.IGF-1R mRNA expression of MET group in renal,brain tissues and blood lymphocytes were significantly increased than IR group expression (P＜0.05).
(3)The expression of Klotho mRNA
The Klotho mRNA in renal,brain tissues and peripheral blood lymphocytes from IR group was significantly lower than that of CN group (P＜0.05),which was increased in renal,brain tissues from TMP+AG group compared with IR group(P＜0.05).No differences of Klotho was detected in blood lymphocytes between TMP+ AG group and IR group.The Klotho mRNA in renal,brain tissues and peripheral blood lymphocytes from MT group was significantly increased than IR group(P＜0.05),which was higher in renal tissues than that in TMP+ AG group(P＜0.05),but lower in brain tissues than that in TMP+ AG group(P＜0.05).
Conclusions:
1.The insulin resistant modal of newborn was established with streptozotocin intraperitoneally.The level of FPG、FINS、IRI and GSP were increased with the time going on.Meanwhile the survival period was decreased.The expression of Klotho mRNA in several tissues were decreased in this model,which suggested that the high morality of IR rats was closely related with Klotho expression.
2.Combination of tetramethylpyrazine and aminoguanidine can upregulate the expression of Klotho mRNA in several tissues of IR rats, inhabite the expression of RAGE mRNA of renal tissues,and significantly prolonge survival period.The results indicated that the mechanism of TMP+AG for the prolongation of survival period was related to regulating the mRNA level of Klotho and RAGE.The exact mechanism needs to be further explored.
3.The expression of IGF-1R mRNA was decreased in several tissues,which indicated that the mechanism of shortening survival period of IR rats was associated with down-regulated of IGF-1R mRNA.The expression of IGF-1R mRNA changed differently in different tissues.It was not found that the mechanism of TMP+AG for the prolongation of survival period was directly related with regulating the mRNA level of IGF-1R.
方法:
1.实验动物及分组:出生当日Wistar大鼠,腹腔注射STZ(90mg/kg)复制IR模型。正常伺养至8周龄,空腹血糖≥7.0mmol/L或糖耐量试验餐后2小时血糖≥11.1mmol几者选入本研究。设立正常对照组(CN组)、胰岛素抵抗对照组(IR组)、二甲双胍(MET组)及川芎嗪+氨基胍(TMP+AG组)治疗组。
2.空腹血糖(FPG)、空腹胰岛素(FINS)和糖化血清蛋白(GSP)检测:分别于造模第8周、24周和32周断尾取空腹血,检测FPG,留取血清检测FINS和GSP,根据公式(FPG·FINS)/22.5计算胰岛素抵抗指数(IRI)。采用果糖胺法检测各组GSP含量。
3.生化指标检测:于实验32周,摘眼球取血,离心留取血清,全自动生化分析仪测定血清中总胆固醇(TC)、甘油三酯(TG)、尿素氮(BUN)和肌酐(Cr)。
4.外周血白细胞原位杂交:于实验结束时取肝素抗凝血约1mL,检测外周血白细胞klotho和IGF-1R mRNA,比较各组之间差异。
5.病理学检测:肾组织学切片分别行苏木素—伊红(HE)染色、糖原(PAS)染色,观察肾组织形态学变化。
6.免疫组织化学染色:观察肾组织klotho蛋白、磷酸化的胰岛素生长因子1受体(IGF-1R)蛋白表达。
7.实时定量RT-PCR:分别检测肾组织晚期糖基化终末产物受体(RAGE)、IGF-1R和klotho mRNA,脑组织IGF-1R和klotho mRNA,肝组织IGF-1R mRNA和外周血淋巴细胞的IGF-1R和klotho mRNA的表达水平。
结果:
1.各组实验动物生存率:IR组为35%(7/20),MET组为70%(14/20),TMP+AG组为85%(17/20),CN组为100%(20/20)。IR组、MET组和TMP+AG组生存率均明显低于CN组(P＜0.05),MET组和TMP+AG组生存率均明显高于IR组(P＜0.05),MET组和TMP+AG组间差异无统计学意义。
2.FPG、FINS和GSP检测结果:8周龄时,模型组的FPG、FINS、IRI和GSP含量均高于CN组(P＜0.05)。用药8周(16周龄)时,MET组和TMP+AG组FPG、FINS、IRI和GSP均低于IR组(P＜0.05),高于CN组(P＜0.05);TMP+AG组GSP含量低于MET组(P＜0.05)。用药24周(32周龄),MET组和TMP+AG组FPG和FINS较IR组进一步降低(P＜0.05);MET组和TMP+AG组FPG与CN组差异无统计学意义,FINS和IRI仍高于CN组;TMP+AG组与CN组间GSP差异无统计学意义;MET组GSP低于IR组,但仍高于CN组(P＜0.05)。
3.生化指标检测结果:各组血清TG、TC和Cr差异无统计学意义,MET组和TMP+AG组的BUN较IR组明显下降(P＜0.05)。
4.外周血白细胞原位杂交结果:IR组IGF-1R mRNA阳性细胞率明显低于CN组(P＜0.05),TMP+AG组明显低于IR组(P＜0.05),而MET组高于IR组(P＜0.05);IR组Klotho mRNA阳性细胞率明显低于CN组(P＜0.05),TMP+AG组、MET组均高于IR组(P＜0.05),TMP+AG组又高于MET组(P＜0.05)。
5.病理学检测结果:HE染色结果显示,IR组两例局部轻度系膜细胞增生,其余未见异常。PAS染色结果显示,IR组两例可见少量肾小管上皮细胞胞浆内出现糖原沉积,其余未见异常。
6.免疫组化结果:IR组肾脏远曲小管和集合小管IGF-1R蛋白表达较CN组减少(P＜0.05),MET组和TMP+AG组较IR组明显升高(P＜0.05),TMP+AG组与CN组间差异无统计学意义,MET组高于CN组(P＜0.05)。IR组肾组织Klotho蛋白含量较CN组明显下降(P＜0.05),MET组和TMP+AG组明显高于IR组(P＜0.05),MET组与TMP+AG组间差异无统计学意义。
7.实时定量RT-PCR结果:
(1)肾组织RAGE mRNA表达
IR组显著高于CN组(P＜0.05)。TMP+AG组明显低于IR组(P＜0.05),与CN组差异无统计学意义。MET组低于IR组(P＜0.05),高于CN组(P＜0.05)。
(2)IGF-1R mRNA表达:
IR组脑、肝、外周血淋巴细胞较CN组明显降低(P＜0.05)。TMP+AG组脑、肝组织较IR组明显升高(P＜0.05),外周血淋巴细胞较IR组降低(P＜0.05);CN组、IR组和TMP+AG组间肾组织表达均无明显差异。MET组肾、脑和外周血淋巴细胞表达均较IR组明显升高(P＜0.05),肝组织的表达较IR组升高,但差异无统计学意义。
(3)Klotho mRNA表达:
IR组肾、脑、外周血淋巴细胞较CN组明显降低(P＜0.05)。TMP+AG组肾、脑组织较IR组明显升高(P＜0.05),与CN组比较无统计学差异。TMP+AG组外周血淋巴细胞与IR组差异无统计学意义。MET组肾、脑、外周血淋巴细胞较IR组明显升高(P＜0.05),其中MET组肾组织的表达高于TMP+AG组(P＜0.05),而在脑组织则低于TMP+AG组(P＜0.05)。
结论:
1.STZ可以诱导出生当日Wistar大鼠产生IR。该模型FPG、FINS、IRI和GSP都随时间延长而升高,其存活期明显缩短。该模型多种组织中Klotho mRNA均表达下降,提示其高死亡率与Klotho基因表达降低密切相关。
2.TMP+AG可提高IR大鼠多种器官组织Klotho mRNA表达,抑制IR大鼠肾组织RAGE mRNA表达,并明显降低大鼠死亡率。提示TMP+AG延长IR大鼠存活时间的作用主要与调控Klotho及RAGE基因表达有关。其确切机制尚待进一步探讨。
3.该实验IR大鼠多种器官组织IGF-1R mRNA表达降低,应用TMP+AG后不同器官的IGF-1R mRNA表达改变不一致,提示IR大鼠存活期的缩短,与IGF-1R信号的转录抑制有关,尚未发现TMP+AG延长IR大鼠存活期的作用与调控IGF-1R信号转录存在直接关系。
Objective:To establish an animal modal of insulin resistant in newborn Wistar rats induced by streptozotocin(STZ)and to observe its survival rate in 32 weeks,to explore the mechanism on the prolongation of survival period effect of the combined therapy with tetramethylpyrazine and aminoguanidine.
Methods:
1.Animal and group division
The insulin resistant model of newborn Wistar rats was duplicated with streptozocin(90mg/kg).The rats were raised for 8 weeks,and those whose fasting plasma glucose higher than or equal to 7.0mmol/L or whose 2 hours postprandial blood glucose higher than or equal to 11.1 mmol/L in glucose tolerance test were selected.Rats were divided into 4 groups randomly: normal control(CN)group,insulin resistant(IR)group,the dimethybiguanide(MET)group,and the combination of tetramethylpyrazine and aminoguanidine(TMP+AG)group.
2.The measurement of fasting blood glucose(FPG)、fasting insulin(FINS) and glucosylated serum protein(GSP)Measurement
FPG、FINS and GSP were detected in 8~(th),24~(th),32~(th)weeks after treatment from the tail vein,insulin resistance index(IRI)was count by calculation formula of FPG×FINS/22.5.
3.Measurement of blood biochemical index
At the end of the experiment,the serum total cholesterol(TC), triglyceride(TG),blood urea nitrogen(BUN),creatinine(Cr)were detected.
4.In situ hybridization Test
Detection of Klotho and insulin like growth factol-1 receptor (IGF-1R)mRNA in the peripheral blood leukocytes by in situ hybridization.
5.Histological and Morphological Analyses
The hematoxylin-eosin(HE)staining and periodic acid Schiff(PAS) staining were applied in the pathological observation.
6.Immunohistochemistry staining
Klotho and phosphated IGF-1R protein were detected by immunohistochemistry staining.
7.Real-Time Fluorescent Quantitative Polymerase Chain Reaction (RT-PCR)
The mRNA levels of advanced glycation end products receptors (RAGE),IGF-1R and Klotho were quantified by RT-PCR using SYBR Green technology.
Results:
1.Survival rate of each group
At the end of the experiment,the Survival rate in IR group,MET group, TMP+ AG group and CN group were 35%,70%,85%and 100%respectively.The survival rate in CN group was significantly higher than that in the other three groups(P＜0.05),The survival rate in MET group and TMP+ AG group were higher than IR group(P＜0.05),but there was no statistically difference between the two treated groups.
2.FPG、FINS and GSP Result
The FPG、FINS、IRI and GSP in modal groups were higher than CN group at the age of 8 weeks(P＜0.05);The index above in MET group and TMP+ AG group were lower than IR group at the age of 16 weeks(P＜0.05),higher than CN group(P＜0.05);At the age of 32 weeks,The FPG and FINS in MET group and TMP+ AG group were more decreased compared with IR group at the age of 24 weeks(P＜0.05),the FPG had no significance with CN group,FINS and IRI in the two groups were higher than CN group(P＜0.05),there was no significant difference in GSP levels between TMP+ AG group and CN group.
3.Blood biochemical index Result
The differences of serum TG,TC and Cr among groups were insignificant, the BUN in MET group and TMP+AG group was decreased significantly compared to IR group.
4.In situ hybridization Result
The positive rate of IGF-1R mRNA of IR group was significantly lower than CN group(P＜0.05),which was higher in IR group than that in TMP+AG group(P＜0.05),and lower than that in MET group(P＜0.05).The positive rate of Klotho mRNA of IR group was significantly lower than CN group (P＜0.05).The expression of Klotho mRNA in both MET group and TMP+AG group were higher than IR group(P＜0.05),which is higher in TMP+AG group compared to MET group(P＜0.05).
5.Histological and Morphological Analyses
HE staining showed that two cases appeared slight mesangial cells hyperplasia and the others showing no abnormal.The epithelial cells of renal tubule occurred a little of glycogen deposition in PAS staining in IR group.
6.Immunohistochemistry Result
The expression of IGF-1R in the distal convoluted renal tubule and collecting duct from IR group was significantly lower than CN group (P＜0.05),IGF-1R expression in both MET group and TMP+AG group was higher than IR group(P＜0.05).There was no significant difference between TMP+ AG group and CN group.The IGF-IR of MET group is higher than that of CN group.The Klotho expression from IR group was deceased compared with CN group(P＜0.05),which was increased in both MET group and TMP+AG group compared to IR group(P＜0.05),and no differences in the two treated groups were found.
7.RT-PCR Analyses
(1)The expression of RAGE mRNA
The expression of RAGE mRNA of IR group in renal tissues was increased significantly compared to CN group(P＜0.05),which was higher than that of TMP+ AG group(P＜0.05),and there was no statistical difference between TMP+ AG group and CN group.The RAGE mRNA of MET group was lower than that of IR group(P＜0.05),higher than that of CN group(P＜0.05).
(2)The expression of IGF-1R mRNA
The expression of IGF-IR mRNA in brain,hepatic tissues and peripheral blood lymphocytes from IR group was significantly lower than CN group (P＜0.05),which was increased in brain and hepatic tissues of TMP+ AG group than that of IR group(P＜0.05),but decreased in blood lymphocytes than IR group(P＜0.05).No statistical differences in the expression of IGF-1R mRNA in renal tissues of CN group,MET group and TMP+ AG group were found.IGF-1R mRNA expression of MET group in renal,brain tissues and blood lymphocytes were significantly increased than IR group expression (P＜0.05).
(3)The expression of Klotho mRNA
The Klotho mRNA in renal,brain tissues and peripheral blood lymphocytes from IR group was significantly lower than that of CN group (P＜0.05),which was increased in renal,brain tissues from TMP+AG group compared with IR group(P＜0.05).No differences of Klotho was detected in blood lymphocytes between TMP+ AG group and IR group.The Klotho mRNA in renal,brain tissues and peripheral blood lymphocytes from MT group was significantly increased than IR group(P＜0.05),which was higher in renal tissues than that in TMP+ AG group(P＜0.05),but lower in brain tissues than that in TMP+ AG group(P＜0.05).
Conclusions:
1.The insulin resistant modal of newborn was established with streptozotocin intraperitoneally.The level of FPG、FINS、IRI and GSP were increased with the time going on.Meanwhile the survival period was decreased.The expression of Klotho mRNA in several tissues were decreased in this model,which suggested that the high morality of IR rats was closely related with Klotho expression.
2.Combination of tetramethylpyrazine and aminoguanidine can upregulate the expression of Klotho mRNA in several tissues of IR rats, inhabite the expression of RAGE mRNA of renal tissues,and significantly prolonge survival period.The results indicated that the mechanism of TMP+AG for the prolongation of survival period was related to regulating the mRNA level of Klotho and RAGE.The exact mechanism needs to be further explored.
3.The expression of IGF-1R mRNA was decreased in several tissues,which indicated that the mechanism of shortening survival period of IR rats was associated with down-regulated of IGF-1R mRNA.The expression of IGF-1R mRNA changed differently in different tissues.It was not found that the mechanism of TMP+AG for the prolongation of survival period was directly related with regulating the mRNA level of IGF-1R.
引文
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