AKT/FOXO1信号通路在心力衰竭小鼠骨骼肌萎缩中的作用
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摘要
目的:探讨AKT/FOXO1信号通路在心力衰竭(HF)小鼠骨骼肌萎缩中的作用。方法:采用主动脉弓横向结扎8周,复制小鼠HF动物模型。采用实时定量-聚合酶链式反应和Western blot技术检测HF小鼠胫骨前肌内,E3连接酶的2个肌肉萎缩特异性指标Atrogin-1和MuRF1,对胫骨前肌中转录因子FOXO1和激酶AKT的磷酸化水平和总蛋白水平进行测定,并比较磷酸化蛋白和总蛋白的比率。结果:与对照组小鼠相比,TAC 8周诱导HF小鼠的胫骨前肌肌纤维变小,质量减轻。RT-PCR分析结果显示,HF小鼠胫骨前肌内的Atrogin-1和MuRF1的mRNA表达明显上调(P<0.01)。Western blot分析结果显示,HF小鼠胫骨前肌组织中Atrogin-1和MuRF1的蛋白相对表达量HF组较对照组明显增加(P<0.01)。HF组小鼠胫骨前肌中p-FOXO1的表达水平较对照组增加,FOXO1的总蛋白水平显著下降;p-AKT的蛋白表达较对照组增加(P<0.05),AKT的总蛋白水平差异无统计学意义。p-FOXO1/FOXO1和p-AKT/AKT比率HF组明显高于对照组(P<0.01)。结论:AKT/FOXOs信号通路参与HF后骨骼肌萎缩的过程并发挥重要作用。
Objective: To investigate the role of AKT/FOXO1 signaling pathway in skeletal muscle atrophy in mouse with heart failure. Methods: Transverse aortic constriction for 8 weeks was used to establish heart failure( HF) model. The mRNA expression of Atrogin-1 and MuRF1 were measured by realtime-polymerase chain reaction( realtime-PCR). The protein expression of Atrogin-1,MuRF1,phosohorylated-FOXO1,FOXO-1,phosphorylated-AKT and total AKT were measured by Western blot. Results: Compared with the control group,the mRNA expressions of Atrogin-1 and MuRF1 significantly increased in tibialis anterior( TA)muscle of the HF group( P < 0. 05). Meanwhile the protein expression of Atrogin-1 and MuRF1 significantly increased in the HF group( P < 0. 01). In addition,the protein expression levels of FOXO1 decreased in HF group and the protein expression levels AKT were not changed in control and HF group. The ratio of phosohorylated-FOXO1 to total FOXO1 and the ratio of phosphorylated-AKT to total AKT significantly increased in TA muscle of the HF group( P < 0. 05). Conclusion: The phosphorylated level of AKT/FOXO1 skeletal muscle are significantly increased in HF mouse,suggesting that AKT/FOXO1 signaling pathway plays a crucial role in skeletal muscle atrophy of HF.
Objective: To investigate the role of AKT/FOXO1 signaling pathway in skeletal muscle atrophy in mouse with heart failure. Methods: Transverse aortic constriction for 8 weeks was used to establish heart failure( HF) model. The mRNA expression of Atrogin-1 and MuRF1 were measured by realtime-polymerase chain reaction( realtime-PCR). The protein expression of Atrogin-1,MuRF1,phosohorylated-FOXO1,FOXO-1,phosphorylated-AKT and total AKT were measured by Western blot. Results: Compared with the control group,the mRNA expressions of Atrogin-1 and MuRF1 significantly increased in tibialis anterior( TA)muscle of the HF group( P < 0. 05). Meanwhile the protein expression of Atrogin-1 and MuRF1 significantly increased in the HF group( P < 0. 01). In addition,the protein expression levels of FOXO1 decreased in HF group and the protein expression levels AKT were not changed in control and HF group. The ratio of phosohorylated-FOXO1 to total FOXO1 and the ratio of phosphorylated-AKT to total AKT significantly increased in TA muscle of the HF group( P < 0. 05). Conclusion: The phosphorylated level of AKT/FOXO1 skeletal muscle are significantly increased in HF mouse,suggesting that AKT/FOXO1 signaling pathway plays a crucial role in skeletal muscle atrophy of HF.
引文
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[17]Zhao J,Brault JJ,Schild A,et al.Fox O3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells.Cell Metab,2007,6:472-483.
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[19]Milan G,Romanello V,Pescatore F,et al.Regulation of autophagy and the ubiquitin-proteasome system by the Fox O transcriptional network during muscle atrophy.Nat Commun 2015,6:66-70.
[20]Brocca L,Toniolo L,Reggiani C,et al.Fox O-dependent atrogenes vary among catabolic conditions and play a key role in muscle atrophy induced by hindlimb suspension.J Physiol,2017,595:1143-1158.
[21]Brunet A,Bonni A,Zigmond MJ,et al.Akt promotes cell survival by phosphorylating and inhibiting a Forkhead transcription factor.Cell,1999,96:857-868.
[22]Egerman MA,Glass DJ.Signaling pathways controlling skeletal muscle mass.Crit Rev Biochem Mol Biol,2014,49:59-68.
[23]Rommel C,Bodine SC,Clarke BA,et al.Mediation of IGF-1-induced skeletal myotube hypertrophy by PI(3)K/Akt/m TOR and PI(3)K/Akt/GSK3 pathways.Nat Cell Biol,2001,3:1009-1013.
[2]Rossignol P,Masson S,Barlera S,et al.Loss in body weight is an independent prognostic factor for mortality in chronic heart failure:insights from the GISSI-HF and Val-He FT trials.Eur J Heart Fail,2015,17:424-433.
[3]刘婷婷,王月丽,李平,等.基于小鼠横向主动脉缩窄模型建立及超声影像的动态观察.心肺血管病杂志,2015,34:575-579.
[4]阿希,李玉琳,王绿娅,等.炎症小体Nlrp3在高血压小鼠心肌纤维化中的作用.心肺血管病杂志,2015,34:496-500.
[5]张健,张宇辉.多中心、前瞻性中国心力衰竭注册登记研究-病因、临床特点和治疗情况初步分析.中国循环杂志,2015,30:413-416.
[6]Strassburg S,Springer J,Anker SD.Muscle wasting in cardiac cachexia.Int J Biochem Cell Biol,2005,37:1938-1947.
[7]von Haehling S,Ebner N,Dos Santos MR,et al.Muscle wasting and cachexia in heart failure:mechanisms and therapies.Nat Rev Cardiol,2017,14:323-341.
[8]Glass DJ.Signaling pathways perturbing muscle mass.Curr Opin Clin Nutr Metab Care,2010,13:225-229.
[9]Bodine SC,Baehr LM.Skeletal muscle atrophy and the E3 ubiquitin ligases MuRF1 and MAFbx/atrogin-1.Am J Physiol Endocrinol Metab,2014,307:E469-484.
[10]Frost RA,Nystrom GJ,Jefferson LS,et al.Hormone,cytokine,and nutritional regulation of sepsis-induced increases in atrogin-1and MuRF1 in skeletal muscle.Am J Physiol Endocrinol Metab,2007,292:E501-512.
[11]Testelmans D,Crul T,Maes K,et al.Atrophy and hypertrophy signalling in the diaphragm of patients with COPD.Eur Respir J,2010,35:549-556.
[12]Li YP,Chen Y,John J,et al.TNF-alpha acts via p38MAPK to stimulate expression of the ubiquitin ligase atrogin1/MAFbx in skeletal muscle.FASEB J,2005,19:362-370.
[13]Schiaffino S,Mammucari C.Regulation of skeletal muscle growth by the IGF1-Akt/PKB pathway:insights from genetic models.Skelet Muscle,2011,1:4.
[14]韩锋锋,徐卫国,崔志磊,等.AKT/FOXOs/Atrogin-1/MuRF1信号通路在慢性阻塞性肺疾病大鼠骨骼肌萎缩中的作用.中国呼吸与危重监护杂志,2013,12:217-222.
[15]Kaestner KH,Knochel W,Martinez DE.Unified nomenclature for the winged helix/forkhead transcription factors.Genes Dev,2000,14:142-146.
[16]Hu P,Geles KG,Paik JH,et al.Codependent activators direct myoblast-specific Myo D transcription.Dev Cell,2008,15:534-546.
[17]Zhao J,Brault JJ,Schild A,et al.Fox O3 coordinately activates protein degradation by the autophagic/lysosomal and proteasomal pathways in atrophying muscle cells.Cell Metab,2007,6:472-483.
[18]Handayaningsih AE,Iguchi G,Fukuoka H,et al.Reactive oxygen species play an essential role in IGF-I signaling and IGF-I-induced myocyte hypertrophy in C2C12 myocytes.Endocrinology,2011,152:912-921.
[19]Milan G,Romanello V,Pescatore F,et al.Regulation of autophagy and the ubiquitin-proteasome system by the Fox O transcriptional network during muscle atrophy.Nat Commun 2015,6:66-70.
[20]Brocca L,Toniolo L,Reggiani C,et al.Fox O-dependent atrogenes vary among catabolic conditions and play a key role in muscle atrophy induced by hindlimb suspension.J Physiol,2017,595:1143-1158.
[21]Brunet A,Bonni A,Zigmond MJ,et al.Akt promotes cell survival by phosphorylating and inhibiting a Forkhead transcription factor.Cell,1999,96:857-868.
[22]Egerman MA,Glass DJ.Signaling pathways controlling skeletal muscle mass.Crit Rev Biochem Mol Biol,2014,49:59-68.
[23]Rommel C,Bodine SC,Clarke BA,et al.Mediation of IGF-1-induced skeletal myotube hypertrophy by PI(3)K/Akt/m TOR and PI(3)K/Akt/GSK3 pathways.Nat Cell Biol,2001,3:1009-1013.