STZ处理树鼩胰岛β细胞再生及EGF、IGF-1表达的研究
摘要
目的:建立树鼩胰岛β细胞损伤模型,证实树鼩胰岛β细胞的再生及其与EGF、IGF-1表达的关系。
方法:将适应饲养环境的32只成年树鼩随机分为两组,包括STZ处理组28只和正常对照组4只。2周内以STZ(每次120mg/kg)总计三次腹腔注射后再观察10天,根据血糖情况分为处理组A(血糖正常)5只,处理组B(血糖先升高后恢复正常) 8只,处理组C(血糖轻度升高但未达糖尿病标准)6只,处理组D(糖尿病组)5只。对照组4只,予以腹腔注射酸化生理盐水5ml/kg。3次注射STZ后3d、1w、10d检测血糖、称体重及实验前后空腹血胰岛素(FINS)、并计算胰岛素敏感指数;留取胰腺标本,光镜下观察胰岛病理形态学改变,应用HE染色观察胰岛炎症程度、以免疫组织化学方法检测胰岛素、胰岛组织中细胞再生相关因子Pdx-1、Ngn3、EGF、IGF-1的表达。
结果:(1)A组、B组胰岛基本保持正常组织形态,细胞体积较正常,岛形较饱满,C组少量胰岛细胞胞浆内有空泡变性,D组胰岛β细胞数量明显减少,胰岛空虚。(2)与D组相比,B组树鼩胰岛表达INS显著升高(P<0.05),血清胰岛素水平上升,血糖水平下降。(3)STZ处理组的ISI均显著低于正常对照组(P<0.05);与D组比较,A、B、C组树鼩的ISI均显著升高(P<0.05),A、B组ISI显著高于C组,(P<0.05),但A、B组之间没有统计学差异(P>0.05)。(4)STZ处理组的PDX-1和Ngn3表达低于正常对照组(P<0.05);B组与C组、D组相比显著升高(P<0.05),但与A组没有统计学差异(P>0.05)。(5)STZ处理组的EGF和IGF-1表达低于正常对照组(P<0.05);B组与C组、D组相比显著升高(P<0.05),但与A组没有统计学差异(P>0.05)。
结论:STZ处理后B组树鼩可能通过增加胰岛表达Pdx-1、Ngn3、EGF、IGF-1等相关因子,对实验性糖尿病树鼩残存β细胞的增殖和再生起到一定的促进作用,进而改善血糖代谢和胰岛素抵抗,并在一定程度上促进胰岛功能的恢复,降低血糖。
Objective:Establishment of tree shrew model of pancreaticβcell injury, confirmed that the tree shrew isletβ-cells regeneration and the relationship with EGF、IGF-1 expression.
Methods:Feeding the environment will adapt the 32 adult tree shrews were divided into two groups , including the 28 STZ treated group and control group 4.2 weeks to STZ(each 120mg/kg) injected a total of three times and then observed for 10 days,Were divided into treatment group A according to blood glucose( normal blood glucose)5,treatment group B(increased blood sugar returned to normal after the first)8,treatment group C(blood glucose increased slightly but less than diabetes standard)6,treatment group D(diabetic group) 5.E group,control group 4 .Control group received intraperitoneal injection of acidified saline 5ml/kg.STZ3 times after injection 3d,1w, 10d detection Blood glucose,weighed,Before the injection and death of insulin, and calculate Insulin sensitivity index. Pancreatic specimens collected,Observed under light microscope morphological changes of islet,HE staining of isletinflammation,Insulin was detected by immunohistochemistry, Measurement of islet Tissue cell regeneration of factor Pdx-1, Ngn3, EGF, IGF-1 expression.
Results:(1)A group, B group remained normal islet morphology, cell volume compared to normal, relatively full island-shaped, C group had a small amount of the cytoplasm of islet degeneration, D group significantly reduced the number of pancreaticβcells, islet empty.(2)Compared with the D group, B group tree shrew islet Expression of INS was significantly higher (P <0.05),B group tree shrew increase in serum insulin levels, glucose levels decrease.(3)STZ treated group ISI was significantly lower than the control group (P <0.05); with the D group, A, B, C group tree shrew ISI were significantly higher (P <0.05), where A, B group and C group significant difference, ISI increased more significantly (P <0.05), A, B did not differ between groups (P> 0.05).(4)STZ treatment of PDX-1 and Ngn3 protein antigen lower than the control group (P <0.05); compared with C group, D group B group was significantly higher (P <0.05),There was no difference with the A group (P> 0.05).(5)STZ treatment of EGF and IGF-1 protein antigen lower than the control group (P <0.05); compared with C group, D group B group was significantly higher (P <0.05),There was no difference with the A group (P> 0.05).
Conclution: STZ treated tree shrew islets may be expressed through the promotion of Pdx-1, Ngn3, EGF, IGF-1 and other related factors For the Tree shrew experimental Diabetes residualβ-cell proliferation and renewable play a role in promoting. Improve glucose metabolism and insulin resistance, and to some extent, promote the recovery of islet function and lower blood sugar.
方法:将适应饲养环境的32只成年树鼩随机分为两组,包括STZ处理组28只和正常对照组4只。2周内以STZ(每次120mg/kg)总计三次腹腔注射后再观察10天,根据血糖情况分为处理组A(血糖正常)5只,处理组B(血糖先升高后恢复正常) 8只,处理组C(血糖轻度升高但未达糖尿病标准)6只,处理组D(糖尿病组)5只。对照组4只,予以腹腔注射酸化生理盐水5ml/kg。3次注射STZ后3d、1w、10d检测血糖、称体重及实验前后空腹血胰岛素(FINS)、并计算胰岛素敏感指数;留取胰腺标本,光镜下观察胰岛病理形态学改变,应用HE染色观察胰岛炎症程度、以免疫组织化学方法检测胰岛素、胰岛组织中细胞再生相关因子Pdx-1、Ngn3、EGF、IGF-1的表达。
结果:(1)A组、B组胰岛基本保持正常组织形态,细胞体积较正常,岛形较饱满,C组少量胰岛细胞胞浆内有空泡变性,D组胰岛β细胞数量明显减少,胰岛空虚。(2)与D组相比,B组树鼩胰岛表达INS显著升高(P<0.05),血清胰岛素水平上升,血糖水平下降。(3)STZ处理组的ISI均显著低于正常对照组(P<0.05);与D组比较,A、B、C组树鼩的ISI均显著升高(P<0.05),A、B组ISI显著高于C组,(P<0.05),但A、B组之间没有统计学差异(P>0.05)。(4)STZ处理组的PDX-1和Ngn3表达低于正常对照组(P<0.05);B组与C组、D组相比显著升高(P<0.05),但与A组没有统计学差异(P>0.05)。(5)STZ处理组的EGF和IGF-1表达低于正常对照组(P<0.05);B组与C组、D组相比显著升高(P<0.05),但与A组没有统计学差异(P>0.05)。
结论:STZ处理后B组树鼩可能通过增加胰岛表达Pdx-1、Ngn3、EGF、IGF-1等相关因子,对实验性糖尿病树鼩残存β细胞的增殖和再生起到一定的促进作用,进而改善血糖代谢和胰岛素抵抗,并在一定程度上促进胰岛功能的恢复,降低血糖。
Objective:Establishment of tree shrew model of pancreaticβcell injury, confirmed that the tree shrew isletβ-cells regeneration and the relationship with EGF、IGF-1 expression.
Methods:Feeding the environment will adapt the 32 adult tree shrews were divided into two groups , including the 28 STZ treated group and control group 4.2 weeks to STZ(each 120mg/kg) injected a total of three times and then observed for 10 days,Were divided into treatment group A according to blood glucose( normal blood glucose)5,treatment group B(increased blood sugar returned to normal after the first)8,treatment group C(blood glucose increased slightly but less than diabetes standard)6,treatment group D(diabetic group) 5.E group,control group 4 .Control group received intraperitoneal injection of acidified saline 5ml/kg.STZ3 times after injection 3d,1w, 10d detection Blood glucose,weighed,Before the injection and death of insulin, and calculate Insulin sensitivity index. Pancreatic specimens collected,Observed under light microscope morphological changes of islet,HE staining of isletinflammation,Insulin was detected by immunohistochemistry, Measurement of islet Tissue cell regeneration of factor Pdx-1, Ngn3, EGF, IGF-1 expression.
Results:(1)A group, B group remained normal islet morphology, cell volume compared to normal, relatively full island-shaped, C group had a small amount of the cytoplasm of islet degeneration, D group significantly reduced the number of pancreaticβcells, islet empty.(2)Compared with the D group, B group tree shrew islet Expression of INS was significantly higher (P <0.05),B group tree shrew increase in serum insulin levels, glucose levels decrease.(3)STZ treated group ISI was significantly lower than the control group (P <0.05); with the D group, A, B, C group tree shrew ISI were significantly higher (P <0.05), where A, B group and C group significant difference, ISI increased more significantly (P <0.05), A, B did not differ between groups (P> 0.05).(4)STZ treatment of PDX-1 and Ngn3 protein antigen lower than the control group (P <0.05); compared with C group, D group B group was significantly higher (P <0.05),There was no difference with the A group (P> 0.05).(5)STZ treatment of EGF and IGF-1 protein antigen lower than the control group (P <0.05); compared with C group, D group B group was significantly higher (P <0.05),There was no difference with the A group (P> 0.05).
Conclution: STZ treated tree shrew islets may be expressed through the promotion of Pdx-1, Ngn3, EGF, IGF-1 and other related factors For the Tree shrew experimental Diabetes residualβ-cell proliferation and renewable play a role in promoting. Improve glucose metabolism and insulin resistance, and to some extent, promote the recovery of islet function and lower blood sugar.
引文
1.叶任高,陆再英.内科学.6版北京:人民卫生出版社,2006:759-763.
2.李鸿,罗敏.Betacellulin与胰岛β细胞再生.国际内分泌代谢杂志,2006,26:25l一253.
3. Yang L,An HX,Deng XL,et al.Roliglitazone reverses insulin secretion altered by chronic exposure to free fatty acid via IRS-2-associated phosphatidylinosito 3-kinase pathway.Acta Pharmaclo Sin , 2003 , 24(5):429-434.
4. Bobb GB,Getty RE,Williamson WM,et al.Spontaneo-us diabetes mellitus in tree threw.Urogate evertti.Diabetes,1966,(15):327- 330.
5.冼苏,黄松,苏建家等.链脲佐菌素诱导树鼩糖尿病动物模型的研究广西医科大学学报,2000,17(6):945-948.
6. Junod A,Lamber AE Orci L,et al.Studies of the diabetogenic action of streptozotocin[J].PTOC Soc Exp Biol Med,1967,125:201-203.
7. Rackietan N,Rackietan ML,Nadkarni MR.Rtudies on diabetogenic actionofstreptozotocin(NSC37917)CancerChemotherapyReports,1993,29:91-95.
8. SimaA,Garcia-Salinas R,Basu P.The BB Wistar rat:an experimental model for thestudy of diabetic retinopathy Metabolism,1983,32(l l):136-138.
9. Gardiner TA,Stitt AW,Anderson HR,et al.Selective loss ovascular smooth muscle cells in the retinal microcirculation of diabetic dogs[J].Br J Ophthalmol,1994,78:54-57.
10. Like AA,Rossini AA.Streptozotocin-induced pancreatic insulitis:new modelof diabetes mellitus.Science,1976,30;19(4251):415-417.
11. Kim Y T,et al.Immunologic studies on the induction of diabetes in experimental animals.Diabetes,1984;33:771.
12. M Govendir PJ Canfield,DB Church,et al.Cellular proliferation in the canine pancreas after d , l-etionine dosage as detected by double immunohistochemical labeling.Diabetes,2003,40(5):129-135.
13. Antella L,Kyomka K,De Riso L,ct a1.Calcium,protease action,and the regulation of the cell cycle.Cel Calcium,1998;23(2-3):123-130.
14. GERRISH K , VAN VELKINBURGH J C.Conservedtranscriptional regulatory domains of the pd][1 gene[J].Mol Endocrinol,2004,18(3):533—548.
15. George M,Ayuso E,Casellas A.βcell expression of IGF-1 leads to recovery from type 1 diabetes.J clin Invest,2002,109(9):1153-1163.
16.徐芳,信艳红,段艳冰等表皮生长因子的临床应用,药学实践杂志,2002,06-0324-03.
17. Maake C,Reinecke M.Immunohistochemical localization of insulin-like growth factor 1and 2 in the endocrine pancreas of rat,dog,and man,and their coexistence with classical islet hormones.Maake C,Reinecke M. Cell Tissue Res,1993,273(2);249-59.
18. Fannie E.Smith,Kenneth M.Rosen,Lydia Villa-Komaroff,et al.Enhanced insulin-like growth factor 1 gene expression in generating rat pancreas. Proc.Natl.Acad.Sci.USA,1999,88:6152-6156.
19. Hayakava H.Induction and involvement of endogenous IGF-1 inPancrreas regeneration after partial pancreatectomy in the dog.J Endocrinol ,1996,149:259-267.
20. Hill DJ,Petrik I.Insulin Like growth factors prevent cytkine-mediated celldeath in isolated islets of Langerhans from Prediabetic nonobese diabetic mice.J Endocrinol,1999,161:153-161.
1. Hardikar AA,Bhonde RR.Modulating experimental diabetes by treatment with cytosolic extract from the regenerating pancreas[J].Diabetes Res Clin Pract,1999,46(3):203.
2. Lombardo YB,Drago S,Chicco A et al.Long term administration of asucrose rich diet to normal rats:relationship between metabolic and hormonal profiles and morphological changes in the endocrinepancreas[J].Metabolism, 1996,45:1527.
3. Kritzik MR,Krahl T,Good A,et al.Transcription factor expression during pancreatic islet regenration[J].Mol Cell Endocrinol,2000,164:99.
4. Rosenberg L,Vink A,Pittenger GL,et al.Islet-cell regeneration in the diabetic hamster pancreas with restoration of normoglycemia can be induced by local growth factor[J].Diabetologia,1996,39:256.
5. Wang RN,Kloppel G,Bouwens L.Duct to islet-cell differentiation and islet growth in the pancreas of duct-ligated adult rats[J].Diabetologia,1995, 38:1405.
6. Bonner-Weir S,BAXTER L,Schuppin G,et al.A second pathway for regeneration of adult exocrine and endocrine pancreas.A possible Recapitulation of embryonic development[J].Diabetes,1993,42(12):1715.
7. Docherty K.Current Opinion in Pharmacology[J].2001:197:638-646.
8. Zhao M,Amiel SA,Christie MR.Insulin—producing ceils derived from human pancreatic non-endoerine cel cultures streptozotecin Induced hyperglycaemia in mice[J].Diabetologia.2005,48(1 0):2051-2 061.
9. Sharma A,Zangen DH,Reitz P,et a1.rI1he homeodomain pmtdn PDX-1 increases after an early burst of proliferation during pancreatic regeneration[J].Diabetes,1999,48(3):507- 513.
10.白日星,田井琦,等.新生鼠胰岛干细胞分离、培养和分化的实验研究[J].中国普通外科杂志,2004,13(10).
11.李艳华,白慈贤,等.成人骨髓间充质干细胞体外定向诱导分化为胰岛样细胞团的研究[J].自然科学进展,2006,13(6).
12. Stephen J.Brand,Sven Tagerud,Philip Lambert,Sheila G,et al.Pharmacologi- cal Tretment of Chronic Diabetes by Stimulating Pancreaticβ-Cell Regeneration with Systemic Co-administration of EGF and Gastrin[J]. Pharmacology&Toxicology,2002,91,414-420.
13. Antella L,Kyomka K,De Riso L,ct a1.Calcium,protease action,and the regulation of the cell cycle.Cel Calcium,1998;23(2—3):123-130.
14. GERRISH K,VAN VELKINBURGH J C.Conservedtranscriptional regulatory domains of the pd][1 gene[J].Mol Endocrinol,2004,18(3):533—548.
2.李鸿,罗敏.Betacellulin与胰岛β细胞再生.国际内分泌代谢杂志,2006,26:25l一253.
3. Yang L,An HX,Deng XL,et al.Roliglitazone reverses insulin secretion altered by chronic exposure to free fatty acid via IRS-2-associated phosphatidylinosito 3-kinase pathway.Acta Pharmaclo Sin , 2003 , 24(5):429-434.
4. Bobb GB,Getty RE,Williamson WM,et al.Spontaneo-us diabetes mellitus in tree threw.Urogate evertti.Diabetes,1966,(15):327- 330.
5.冼苏,黄松,苏建家等.链脲佐菌素诱导树鼩糖尿病动物模型的研究广西医科大学学报,2000,17(6):945-948.
6. Junod A,Lamber AE Orci L,et al.Studies of the diabetogenic action of streptozotocin[J].PTOC Soc Exp Biol Med,1967,125:201-203.
7. Rackietan N,Rackietan ML,Nadkarni MR.Rtudies on diabetogenic actionofstreptozotocin(NSC37917)CancerChemotherapyReports,1993,29:91-95.
8. SimaA,Garcia-Salinas R,Basu P.The BB Wistar rat:an experimental model for thestudy of diabetic retinopathy Metabolism,1983,32(l l):136-138.
9. Gardiner TA,Stitt AW,Anderson HR,et al.Selective loss ovascular smooth muscle cells in the retinal microcirculation of diabetic dogs[J].Br J Ophthalmol,1994,78:54-57.
10. Like AA,Rossini AA.Streptozotocin-induced pancreatic insulitis:new modelof diabetes mellitus.Science,1976,30;19(4251):415-417.
11. Kim Y T,et al.Immunologic studies on the induction of diabetes in experimental animals.Diabetes,1984;33:771.
12. M Govendir PJ Canfield,DB Church,et al.Cellular proliferation in the canine pancreas after d , l-etionine dosage as detected by double immunohistochemical labeling.Diabetes,2003,40(5):129-135.
13. Antella L,Kyomka K,De Riso L,ct a1.Calcium,protease action,and the regulation of the cell cycle.Cel Calcium,1998;23(2-3):123-130.
14. GERRISH K , VAN VELKINBURGH J C.Conservedtranscriptional regulatory domains of the pd][1 gene[J].Mol Endocrinol,2004,18(3):533—548.
15. George M,Ayuso E,Casellas A.βcell expression of IGF-1 leads to recovery from type 1 diabetes.J clin Invest,2002,109(9):1153-1163.
16.徐芳,信艳红,段艳冰等表皮生长因子的临床应用,药学实践杂志,2002,06-0324-03.
17. Maake C,Reinecke M.Immunohistochemical localization of insulin-like growth factor 1and 2 in the endocrine pancreas of rat,dog,and man,and their coexistence with classical islet hormones.Maake C,Reinecke M. Cell Tissue Res,1993,273(2);249-59.
18. Fannie E.Smith,Kenneth M.Rosen,Lydia Villa-Komaroff,et al.Enhanced insulin-like growth factor 1 gene expression in generating rat pancreas. Proc.Natl.Acad.Sci.USA,1999,88:6152-6156.
19. Hayakava H.Induction and involvement of endogenous IGF-1 inPancrreas regeneration after partial pancreatectomy in the dog.J Endocrinol ,1996,149:259-267.
20. Hill DJ,Petrik I.Insulin Like growth factors prevent cytkine-mediated celldeath in isolated islets of Langerhans from Prediabetic nonobese diabetic mice.J Endocrinol,1999,161:153-161.
1. Hardikar AA,Bhonde RR.Modulating experimental diabetes by treatment with cytosolic extract from the regenerating pancreas[J].Diabetes Res Clin Pract,1999,46(3):203.
2. Lombardo YB,Drago S,Chicco A et al.Long term administration of asucrose rich diet to normal rats:relationship between metabolic and hormonal profiles and morphological changes in the endocrinepancreas[J].Metabolism, 1996,45:1527.
3. Kritzik MR,Krahl T,Good A,et al.Transcription factor expression during pancreatic islet regenration[J].Mol Cell Endocrinol,2000,164:99.
4. Rosenberg L,Vink A,Pittenger GL,et al.Islet-cell regeneration in the diabetic hamster pancreas with restoration of normoglycemia can be induced by local growth factor[J].Diabetologia,1996,39:256.
5. Wang RN,Kloppel G,Bouwens L.Duct to islet-cell differentiation and islet growth in the pancreas of duct-ligated adult rats[J].Diabetologia,1995, 38:1405.
6. Bonner-Weir S,BAXTER L,Schuppin G,et al.A second pathway for regeneration of adult exocrine and endocrine pancreas.A possible Recapitulation of embryonic development[J].Diabetes,1993,42(12):1715.
7. Docherty K.Current Opinion in Pharmacology[J].2001:197:638-646.
8. Zhao M,Amiel SA,Christie MR.Insulin—producing ceils derived from human pancreatic non-endoerine cel cultures streptozotecin Induced hyperglycaemia in mice[J].Diabetologia.2005,48(1 0):2051-2 061.
9. Sharma A,Zangen DH,Reitz P,et a1.rI1he homeodomain pmtdn PDX-1 increases after an early burst of proliferation during pancreatic regeneration[J].Diabetes,1999,48(3):507- 513.
10.白日星,田井琦,等.新生鼠胰岛干细胞分离、培养和分化的实验研究[J].中国普通外科杂志,2004,13(10).
11.李艳华,白慈贤,等.成人骨髓间充质干细胞体外定向诱导分化为胰岛样细胞团的研究[J].自然科学进展,2006,13(6).
12. Stephen J.Brand,Sven Tagerud,Philip Lambert,Sheila G,et al.Pharmacologi- cal Tretment of Chronic Diabetes by Stimulating Pancreaticβ-Cell Regeneration with Systemic Co-administration of EGF and Gastrin[J]. Pharmacology&Toxicology,2002,91,414-420.
13. Antella L,Kyomka K,De Riso L,ct a1.Calcium,protease action,and the regulation of the cell cycle.Cel Calcium,1998;23(2—3):123-130.
14. GERRISH K,VAN VELKINBURGH J C.Conservedtranscriptional regulatory domains of the pd][1 gene[J].Mol Endocrinol,2004,18(3):533—548.