SUMO化修饰C/EBPα在高氧暴露所致支气管肺发育不良早产大鼠中的动态表达及作用
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摘要
目的探讨小泛素相关修饰物(SUMO)化修饰CCAAT增强子结合蛋白α(C/EBPα)在支气管肺发育不良(BPD)早产大鼠中的表达及其作用。方法将18只早产大鼠随机分为空气组和高氧组(n=9),建立高氧暴露早产大鼠BPD模型。分别在4 d、7 d及14 d,每组各取3只大鼠采集肺组织,糖原染色观察肺组织分化情况;免疫组化检测Ki67表达;Western blot检测SUMO1及C/EBPα蛋白表达;免疫共沉淀检测SUMO化C/EBPα表达。结果与同时间点空气组相比,高氧组大鼠肺组织糖原染色强度4 d时减弱,7 d及14 d时明显增强;随高氧暴露时间延长,高氧组大鼠肺组织中Ki67表达逐渐升高,且均高于同时间点空气组;与空气组相比,高氧组C/EBPα蛋白4 d时表达增加,7 d及14 d时表达降低(P<0.05);高氧组SUMO1及SUMO化C/EBPα蛋白表达呈逐渐上升趋势,均明显高于同时间点空气组(P<0.05)。高氧组SUMO化C/EBPα表达与糖原染色强度及核增殖抗原Ki67均呈正相关(r=0.529、0.671,P<0.05)。结论高氧暴露所致早产大鼠BPD模型中,肺泡上皮细胞过度增殖与分化障碍可能与SUMO化C/EBPα表达增加相关。
Objective To study the expression of SUMO-modified CCAAT enhancer binding protein α(C/EBPα) in preterm rat model of bronchopulmonary dysplasisa(BPD) induced by hyperoxia exposure and its role. Methods Eighteen preterm rats were randomly divided into an air group and a hyperoxia group(n=9 each). The model of BPD was prepared in preterm rats exposed to hyperoxia. The rats from the two groups were sacrificed on postnatal days 4, 7 and 14 respectively(3 rats at each time) and lung tissues were harvested. Periodic acid-Schiff(PAS) staining was used to observe the differentiation of rat lung tissues. Ki67 expression was detected by immunohistochemistry. Western blot was used to measure the protein expression of small ubiquitin-related modifier-1(SUMO1) and C/EBPα. A co-immunoprecipitation assay was performed to measure the protein expression of SUMO-modified C/EBPα. Results Compared with the air group, the hyperoxia group showed a decreased glycogen content in the lung tissue on postnatal day 4, and an increased content on postnatal days 7 and 14. Over the time of hyperoxia exposure, the hyperoxia group showed an increased expression of Ki67 in the lung tissue compared with the air group at all time points. Compared with the air group, the protein expression of C/EBPα increased on postnatal day 4 and decreased on postnatal days 7 and 14 in the hyperoxia group(P<0.05). The hyperoxia group had significantly upregulated expression of SUMO1 and SUMO-modified C/EBPα compared with the air group at all time points(P<0.05). In the hyperoxia group, the protein expression of SUMO-modified C/EBPα was positively correlated with the glycogen content(r=0.529, P<0.05) and the expression of Ki67(r=0.671, P<0.05). Conclusions Hyperoxia may induce over-proliferation and differentiation disorders of alveolar epithelial cells in preterm rat model of BPD, possibly through an increased expression of SUMO-modified C/EBPα.
Objective To study the expression of SUMO-modified CCAAT enhancer binding protein α(C/EBPα) in preterm rat model of bronchopulmonary dysplasisa(BPD) induced by hyperoxia exposure and its role. Methods Eighteen preterm rats were randomly divided into an air group and a hyperoxia group(n=9 each). The model of BPD was prepared in preterm rats exposed to hyperoxia. The rats from the two groups were sacrificed on postnatal days 4, 7 and 14 respectively(3 rats at each time) and lung tissues were harvested. Periodic acid-Schiff(PAS) staining was used to observe the differentiation of rat lung tissues. Ki67 expression was detected by immunohistochemistry. Western blot was used to measure the protein expression of small ubiquitin-related modifier-1(SUMO1) and C/EBPα. A co-immunoprecipitation assay was performed to measure the protein expression of SUMO-modified C/EBPα. Results Compared with the air group, the hyperoxia group showed a decreased glycogen content in the lung tissue on postnatal day 4, and an increased content on postnatal days 7 and 14. Over the time of hyperoxia exposure, the hyperoxia group showed an increased expression of Ki67 in the lung tissue compared with the air group at all time points. Compared with the air group, the protein expression of C/EBPα increased on postnatal day 4 and decreased on postnatal days 7 and 14 in the hyperoxia group(P<0.05). The hyperoxia group had significantly upregulated expression of SUMO1 and SUMO-modified C/EBPα compared with the air group at all time points(P<0.05). In the hyperoxia group, the protein expression of SUMO-modified C/EBPα was positively correlated with the glycogen content(r=0.529, P<0.05) and the expression of Ki67(r=0.671, P<0.05). Conclusions Hyperoxia may induce over-proliferation and differentiation disorders of alveolar epithelial cells in preterm rat model of BPD, possibly through an increased expression of SUMO-modified C/EBPα.
引文
[1]Kalikkot Thekkeveedu R,Guaman MC,Shivanna B.Bronchopulmonary dysplasia:A review of pathogenesis and pathophysiology[J].Respir Med,2017,132:170-177.
[2]Xue M,Li X,Chen W.Hypoxia regulates the expression and localization of CCAAT/enhancer binding proteinαby hypoxia inducible factor-1αin bladder transitional carcinoma cells[J].Mol Med Rep,2015,21(3):345-351.
[3]Yang G,Hinson MD,Bordner JE,et al.Silencing hyperoxiainduced C/EBPɑin neonatal mice improves lung architecture via enhanced proliferation of alveolar epithelial cells[J].Am J Physiol Lung Cell Mol Physiol,2011,301(2):L187-L196.
[4]马禕文,吕翔.抑制SUMO特异性蛋白酶1在小鼠急性肺损伤中的保护作用[J].中国临床医学,2015,22(6):738-740.
[5]Sato Y,Miyake K,Kaneoka H,et al.Sumoylation of CCAAT/enhancer-binding protein alpha and its functional roles in hepatocyte differentiation[J].J Biol Chem,2006,281(31):21629-21639.
[6]Yuan H,Zhang T,Liu X,et al.Sumoylation of CCAAT/enhancer-binding proteinαis implicated in hematopoietic stem/progenitor cell development through regulating runx1 in zebrafish[J].Sci Rep,2015,5:9011.
[7]Chen YD,Liu JY,Lu YM,et al.Functional roles of C/EBPɑand SUMO-modification in lung development[J].Int J Mol Med,2017,40(4):1037-1046.
[8]Warner BB,Stuart LA,Papes RA,et al.Functional and pathological effects of prolonged hyperoxia in neonatal mice[J].Am J Physiol,1998,275(1 Pt 1):L110-L117.
[9]Pan B,Xue X,Zhang D,et al.SOX4 arrests lung development in rats with hyperoxia-induced bronchopulmonary dysplasia by controlling EZH2 expression[J].Int J Mol Med,2017,40(6):1691-1698.
[10]Roos AB,Berg T,Barton JL,et al.Airway epithelial cell differentiation during lung organogenesis requires C/EBPαand C/EBPβ[J].Dev Dyn,2012,241(5):911-923.
[11]卢衍敏,卢红艳,刘姜艳,等.高氧调节早产大鼠肺泡Ⅱ型上皮细胞CCAAT增强子结合蛋白α和肺泡表面活性蛋白的表达[J].细胞与分子免疫学杂志,2017,33(6):767-771.
[12]Torres F,González-Candia A,Montt C,et al.Melatonin reduces oxidative stress and improves vascular function in pulmonary hypertensive newborn sheep[J].J Pineal Res,2015,58(3):362-373.
[13]Jiang Y,Wang J,Tian H,et al.Increased SUMO-1 expression in response to hypoxia:Interaction with HIF-1αin hypoxic pulmonary hypertension[J].Int J Mol Med,2015,36(1):271-281.
[14]Geletu M,Balkhi MY,Peer Zada AA,et al.Target proteins of C/EBPalphap30 in AML:C/EBPalphap30 enhances sumoylation of C/EBPalphap42 via up-regulation of Ubc9[J].Blood,2007,110(9):3301-3309.
[15]Mackert JR,Qu P,Min Y,et al.Dual negative roles of C/EBPαin the expansion and pro-tumor fuctions of MDSCs[J].Sci Rep,2017,7(1):14048.
[16]Hankey W,Silver M,Sun BS,et al.Differential effects of sumoylation on the activities of CCAAT enhancer binding protein alpha(C/EBPα)p42 versus p30 may contribute in part,to aberrant C/EBPαactivity in acute leukemias[J].Hematol Rep,2011,3(1):e5.
[17]Gerlach JC,Over P,Foka HG,et al.Role of transcription factor CCAAT/enhancer-binding protein alpha in human fetal liver cell types in vitro[J].Hepatol Res,2015,45(8):919-932.
[18]Xu Y,Saegusa C,Schehr A,et al.C/EBPalpha is required for pulmonary cytoprotection during hyperoxia[J].Am J Physiol Lung Cell Mol Physiol,2009,297(2):L286-L298.
[19]Lu GD,Leung CH,Yan B,et al.C/EBPalpha is up-regulated in a subset of hepatocellular carcinomas and plays a role in cell growth and proliferation[J].Gastroenterology,2010,139(2):632-643.
[2]Xue M,Li X,Chen W.Hypoxia regulates the expression and localization of CCAAT/enhancer binding proteinαby hypoxia inducible factor-1αin bladder transitional carcinoma cells[J].Mol Med Rep,2015,21(3):345-351.
[3]Yang G,Hinson MD,Bordner JE,et al.Silencing hyperoxiainduced C/EBPɑin neonatal mice improves lung architecture via enhanced proliferation of alveolar epithelial cells[J].Am J Physiol Lung Cell Mol Physiol,2011,301(2):L187-L196.
[4]马禕文,吕翔.抑制SUMO特异性蛋白酶1在小鼠急性肺损伤中的保护作用[J].中国临床医学,2015,22(6):738-740.
[5]Sato Y,Miyake K,Kaneoka H,et al.Sumoylation of CCAAT/enhancer-binding protein alpha and its functional roles in hepatocyte differentiation[J].J Biol Chem,2006,281(31):21629-21639.
[6]Yuan H,Zhang T,Liu X,et al.Sumoylation of CCAAT/enhancer-binding proteinαis implicated in hematopoietic stem/progenitor cell development through regulating runx1 in zebrafish[J].Sci Rep,2015,5:9011.
[7]Chen YD,Liu JY,Lu YM,et al.Functional roles of C/EBPɑand SUMO-modification in lung development[J].Int J Mol Med,2017,40(4):1037-1046.
[8]Warner BB,Stuart LA,Papes RA,et al.Functional and pathological effects of prolonged hyperoxia in neonatal mice[J].Am J Physiol,1998,275(1 Pt 1):L110-L117.
[9]Pan B,Xue X,Zhang D,et al.SOX4 arrests lung development in rats with hyperoxia-induced bronchopulmonary dysplasia by controlling EZH2 expression[J].Int J Mol Med,2017,40(6):1691-1698.
[10]Roos AB,Berg T,Barton JL,et al.Airway epithelial cell differentiation during lung organogenesis requires C/EBPαand C/EBPβ[J].Dev Dyn,2012,241(5):911-923.
[11]卢衍敏,卢红艳,刘姜艳,等.高氧调节早产大鼠肺泡Ⅱ型上皮细胞CCAAT增强子结合蛋白α和肺泡表面活性蛋白的表达[J].细胞与分子免疫学杂志,2017,33(6):767-771.
[12]Torres F,González-Candia A,Montt C,et al.Melatonin reduces oxidative stress and improves vascular function in pulmonary hypertensive newborn sheep[J].J Pineal Res,2015,58(3):362-373.
[13]Jiang Y,Wang J,Tian H,et al.Increased SUMO-1 expression in response to hypoxia:Interaction with HIF-1αin hypoxic pulmonary hypertension[J].Int J Mol Med,2015,36(1):271-281.
[14]Geletu M,Balkhi MY,Peer Zada AA,et al.Target proteins of C/EBPalphap30 in AML:C/EBPalphap30 enhances sumoylation of C/EBPalphap42 via up-regulation of Ubc9[J].Blood,2007,110(9):3301-3309.
[15]Mackert JR,Qu P,Min Y,et al.Dual negative roles of C/EBPαin the expansion and pro-tumor fuctions of MDSCs[J].Sci Rep,2017,7(1):14048.
[16]Hankey W,Silver M,Sun BS,et al.Differential effects of sumoylation on the activities of CCAAT enhancer binding protein alpha(C/EBPα)p42 versus p30 may contribute in part,to aberrant C/EBPαactivity in acute leukemias[J].Hematol Rep,2011,3(1):e5.
[17]Gerlach JC,Over P,Foka HG,et al.Role of transcription factor CCAAT/enhancer-binding protein alpha in human fetal liver cell types in vitro[J].Hepatol Res,2015,45(8):919-932.
[18]Xu Y,Saegusa C,Schehr A,et al.C/EBPalpha is required for pulmonary cytoprotection during hyperoxia[J].Am J Physiol Lung Cell Mol Physiol,2009,297(2):L286-L298.
[19]Lu GD,Leung CH,Yan B,et al.C/EBPalpha is up-regulated in a subset of hepatocellular carcinomas and plays a role in cell growth and proliferation[J].Gastroenterology,2010,139(2):632-643.