Stimulation of pancreatic β-cell replication by incretins involves transcriptional induction of cyclin D1 via multiple signalling pathways
Stimulation of pancreatic β-cell replication by incretins involves transcriptional induction of cyclin D1 via multiple signalling pathways
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DOI：
10.1677/joe.1.06160
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发表时间：
2006-03-01
影响因子：
4
通讯作者：
Moldrup, A
中科院分区：
文献类型：
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作者：
Friedrichsen, BN;Neubauer, N;Moldrup, A
胰升糖素激素，即胰升糖素样肽-1(GLP-1)和葡萄糖依赖的促胰岛素肽(GIP)，被认为是P细胞生长因子，因此对维持适当的P细胞质量可能是至关重要的。我们研究了胰岛素诱导新生大鼠胰岛细胞原代单层培养中β细胞复制的分子机制。GLP-1、GIP和长效GLP-1衍生物利拉鲁肽在10-100 nM作用24 h可使P-细胞复制增加50-80%，而类似浓度的GLP-1对P细胞复制无明显影响。GLP-1和GIP的刺激作用可被腺苷环化酶激活剂Forskolin在10 nM时有效模拟(类似于增加90%)，并与对人生长激素(HGH)的生长反应相加(类似于增加170-250%)，表明分别使用不同的细胞内信号通路通过胰岛素和细胞因子导致有丝分裂。对GLP-1和GIP的反应都被蛋白激酶A(PKA)抑制剂H89完全阻断。此外，磷酸肌醇3-激酶(PI3K)抑制剂Wortmannin和丝裂原活化蛋白激酶(MEK)抑制剂PD98059均抑制GLP-1和GIP刺激的增殖。P38丝裂原活化蛋白激酶(MAPK)抑制剂SB203580对GLP-1和GIP刺激的细胞增殖均无抑制作用。细胞周期蛋白D在许多细胞类型中作为G0/G1-S相变的分子开关，我们先前已经在胰岛素瘤细胞系INS-1中证明了hGH诱导细胞周期蛋白D2的表达。GLP-1呈时间依赖性地诱导INS-1E细胞周期蛋白D1mRNA和蛋白水平升高，而对细胞周期蛋白D2水平无明显影响。而GLP-1刺激对细胞周期蛋白D3的影响较小。瞬时将Cyclin D1启动子-荧光素酶报告结构导入胰岛单层细胞或INS-I细胞，GLP-1和GIP的转录活性增加约2-3倍，Forsklin的转录活性增加4-7倍。然而，用hGH处理两种细胞类型均不影响细胞周期蛋白DI启动子的活性。H89、Wortmannin和PD98059可抑制GLP-1对细胞周期蛋白D-1前体的刺激作用。我们的结论是，胰岛素诱导的β细胞复制依赖于石油cAMP/PKA、p42 MAPK和PI3K活性，这可能涉及细胞周期蛋白M的转录诱导。GLP-1、GIP和利拉鲁肽可能具有增加人类P细胞复制的潜力，这将对石油长期治疗糖尿病产生重大影响。
The incretin hormones, glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic peptide (GIP), have been suggested to act as P-cell growth factors and may therefore be of critical importance for the maintenance of a proper P-cell mass. We have investigated the molecular mechanism of incretin-induced beta-cell replication in primary monolayer cultures of newborn rat islet cells. GLP-1, GIP and the long-acting GLP-1 derivative, liraglutide, increased P-cell replication 50-80% at 10-100 nM upon a 24 h stimulus, whereas glucagon at a similar concentration had no significant effect. The stimulatory effect of GLP-1 and GIP was efficiently mimicked by the adenylate cyclase activator, forskolin, at 10 nM (similar to 90% increase) and was additive (similar to 170-250% increase) with the growth response to human growth hormone (hGH), indicating the use of distinct intracellular signalling pathways leading to mitosis by incretins and cytokines, respectively. The response to both GLP-1 and GIP was completely blocked by the protein kinase A (PKA) inhibitor, H89. In addition, the phosphoinositol 3-kinase (PI3K) inhibitor wortmannin and the mitogen-activated protein kinase kinase (MEK) inhibitor PD98059, both inhibited GLP-1- and GIP-stimulated proliferation. The p38 mitogen-activated protein kinase (MAPK) inhibitor, SB203580, had no inhibitory effect oil either GLP-1 or GIP stimulated proliferation. Cyclin Ds act as molecular switches for the G0/G1-S phase transition in many cell types and we have previously demonstrated hGH-induced cyclin D2 expression in the insulinoma cell line, INS-1. GLP-1 time-dependently induced the cyclin D1 mRNA and protein levels in INS-1E, whereas the cyclin D2 levels were unaffected. However, minor effect of GLP-1 stimulation was observed on the cyclin D3 mRNA levels. Transient transfection of a cyclin D1 promoter-luciferase reporter construct into islet monolayer cells or INS-I cells revealed approximately a 2-3 fold increase of transcriptional activity in response to GLP-1 and GIP, and a 4-7 fold increase in response to forskolin. However, treatment of either cell type with hGH had no effect on cyclin D I promoter activity. The stimulation of the cyclin D-1 prornoter by GLP-1 was inhibited by H89, wortmannin, and PD98059. We conclude that incretin-induced beta-cell replication is dependent oil cAMP/PKA, p42 MAPK and PI3K activities, which may involve transcriptional induction of cyclin M. GLP-1, GIP and liraglutide may have the potential to increase P-cell replication in humans which would have significant impact oil long-term diabetes treatment.