Colonic PDGFRα Overexpression Accompanied Forkhead Transcription Factor FOXO3 Up-Regulation in STZ-Induced Diabetic Mice
Colonic PDGFRα Overexpression Accompanied Forkhead Transcription Factor FOXO3 Up-Regulation in STZ-Induced Diabetic Mice
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STZ 诱导的糖尿病小鼠结肠 PDGFR α 过表达伴随叉头转录因子 FOXO3 上调
DOI：
10.1159/000480335
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发表时间：
2017-01-01
影响因子：
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通讯作者：
Xu, Wenxie
中科院分区：
文献类型：
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作者：
Lu, Hongli;Zhang, Chunmei;Xu, Wenxie
背景：结肠运输紊乱引起的便秘是糖尿病患者的主要并发症。PDGFR α(+)（血小板源性生长因子受体α阳性）细胞在结肠运动的抑制调节中起关键作用，FOXO3（叉头转录因子3）具有广泛的生物学功能。本研究旨在探讨FOXO3与链脲佐菌素（STZ）诱导的糖尿病小鼠PDGFR α(+)细胞增殖的关系。方法：采用免疫组化、定量RT-RCR、Western blotting等技术评价特异性蛋白表达；ChIP法鉴定FOXO3蛋白与PDGFR α启动子之间的相互作用；慢病毒转染短发夹rna （shRNAs）过表达下调FOXO3。结果：stz诱导的糖尿病小鼠结肠近端平滑肌组织中PDGFR α和Ki67免疫反应性显著升高。PDGFR α mRNA和蛋白在结肠平滑肌组织中的表达水平均显著升高，而PDGFR β的表达水平不变。同时，PDGF配体PDGF α和PDGF β在糖尿病结肠平滑肌组织中的表达均显著升高。在全细胞和细胞核提取物中，FOXO3蛋白的表达也显著增加；P-FOXO3（磷酸化FOXO3）蛋白的表达显著降低。与50 mmol/L葡萄糖孵育NIH细胞12 h、24 h和48 h时，PDGFR α的表达均显著升高，全细胞和细胞核提取物中FOXO3蛋白的表达均显著升高。P-FOXO3蛋白的表达明显降低。FOXO3可以结合PDGFR α启动子上的一个位点，在NIH细胞中，用FOXO3短发夹RNA （shRNA）敲低内源性FOXO3表达可显著降低PDGFR α的基础表达。磷酸化Akt在糖尿病结肠肌肉组织中的表达显著下调。结论：上述结果提示糖尿病诱导的结肠PDGFR α(+)细胞增殖受FOXO3上调介导。在stz诱导的糖尿病小鼠中，FOXO3的上调可能是通过抑制PI3K/Akt信号通路来诱导的。PDGFRa（+）细胞增殖可能成为糖尿病性结肠转运障碍临床治疗的新靶点。(C) 2017作者：s . Karger AG，巴塞尔出版
Background: Colonic transit disorder-induced constipation is a major complication in diabetic patients. PDGFR alpha(+) (platelet-derived growth factor receptor alpha-positive) cells play critical roles in the inhibitory regulation of colonic motility, and FOXO3 (forkhead transcription factor 3) has a broad range of biological functions. The present study was designed to investigate the relationship between FOXO3 and PDGFR alpha(+) cell proliferation in streptozotocin (STZ)induced diabetic mice. Methods: The major experimental techniques used in this paper are immunohistochemistry, quantitative RT-RCR and Western blotting for the evaluation of specific protein expression; ChIP assay for identifying the interaction between FOXO3 protein and the PDGFR alpha promotor; and lentiviral transfection for the overexpression of short hairpin RNAs (shRNAs) to down-regulate FOXO3. Results: In proximal colonic smooth muscle tissue of STZ-induced diabetic mice, there was a significant increase in PDGFR alpha and Ki67 immunoreactivity. PDGFR alpha mRNA and protein expression levels were both significantly increased in colonic smooth muscle tissue, but PDGFR beta expression was unchanged. Meanwhile, the expression of PDGF ligands, including both PDGF alpha and PDGF beta, was significantly increased in diabetic colonic smooth muscle tissue. In whole cell and nuclear extracts, the expression of FOXO3 protein was also significantly increased; however, the expression of P-FOXO3 (phosphorylated FOXO3) protein was significantly decreased. When NIH cells were incubated with 50 mmol/L glucose for 12 h, 24 h and 48 h, the expression of PDGFR alpha significantly increased, and in whole cell and nuclear extracts, the expression of FOXO3 protein was significantly increased. However, the expression of P-FOXO3 protein was significantly decreased. FOXO3 could bind to a site on the PDGFR alpha promoter, and the basal expression of PDGFR alpha was significantly reduced when endogenous FOXO3 expression was knocked down with FOXO3 short hairpin RNA (shRNA) in NIH cells. The expression of phosphorylated Akt was significantly down-regulated in diabetic colonic muscle tissue. Conclusions: These results suggest that diabetes-induced colonic PDGFR alpha(+) cell proliferation is mediated by FOXO3 up-regulation. FOXO3 up-regulation may be induced by inhibiting the PI3K/Akt signaling pathway in STZ-induced diabetic mice. PDGFRa(+) cell proliferation could be a new target for clinical therapy of diabetes-induced colonic transit disorder. (C) 2017 The Author(s) Published by S. Karger AG, Basel