小檗碱抑制NF-κB p65改善肝脏糖异生的作用研究
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摘要
目的:观察小檗碱对核转录因子NF-κB p65蛋白活性的影响,探究小檗碱改善cAMP/PKA肝脏糖异生信号的作用及机制。方法:雄性ICR小鼠随机分为正常对照组、胰高血糖素模型组和小檗碱组,每组8只。采用小鼠腹腔注射胰高血糖素模型来观察小檗碱对糖异生信号的影响。RT-PCR法定量检测肝组织中糖异生关键酶PGC-1α、G6Pase和PEPCK基因表达。Western Blot法检测肝组织中p-P65(Ser468)、PDE4B和p-PKA substrate蛋白表达,检测肝组织细胞核中Ac-P65(Lys310)、p65及细胞胞浆中Ac-P65(Lys310)的蛋白表达。结果:小檗碱可明显抑制肝组织中p65蛋白活性及其细胞核转位,增加PDE4B蛋白表达,减少胰高血糖素介导的PKA底物水平磷酸化和肝糖异生关键酶的基因表达(P<0.05,P<0.01)。结论:小檗碱能够通过抑制NF-κB p65的蛋白活性阻断cAMP/PKA信号,从而改善胰高血糖素介导的肝脏糖异生。
Objective: To observe the role of berberine on the activity of nuclear transcription factor NF-κB p65, and to explore the effect and mechanism of berberine on the hepatic cAMP/PKA gluconeogenesis signaling. Methods: Male ICR mice were randomly divided into control group, glucagon model group and berberine group. Mice intraperitoneally injection of glucagon were used to investigate the role of berberine on the gluconeogenesis signaling. Gene expressions of key gluconeogenic enzymes PGC-1α, G6 Pase and PEPCK in the liver tissue were detected by RT-PCR. The protein expressions of p-P65(Ser468), PDE4 B and p-PKA substrate in the liver tissue were assayed with Western blot. Besides, nuclear Ac-P65(Lys310), p65 expressions and cytosol Ac-P65(Lys310) expression in the liver tissue were examined. Results: Berberine significantly inhibited p65 protein activity and its nuclear translocation in the liver tissue, increased PDE4 B protein expression and thus reduced glucagon-mediate PAK substrates phosphorylation and gene expressions of key gluconeogenic enzymes. Conclusion: Berberine blocked cAMP/PKA signaling via inhibiting NF-κB p65 protein activity, and thus alleviated glucagon-driven hepatic gluconeogenesis.
Objective: To observe the role of berberine on the activity of nuclear transcription factor NF-κB p65, and to explore the effect and mechanism of berberine on the hepatic cAMP/PKA gluconeogenesis signaling. Methods: Male ICR mice were randomly divided into control group, glucagon model group and berberine group. Mice intraperitoneally injection of glucagon were used to investigate the role of berberine on the gluconeogenesis signaling. Gene expressions of key gluconeogenic enzymes PGC-1α, G6 Pase and PEPCK in the liver tissue were detected by RT-PCR. The protein expressions of p-P65(Ser468), PDE4 B and p-PKA substrate in the liver tissue were assayed with Western blot. Besides, nuclear Ac-P65(Lys310), p65 expressions and cytosol Ac-P65(Lys310) expression in the liver tissue were examined. Results: Berberine significantly inhibited p65 protein activity and its nuclear translocation in the liver tissue, increased PDE4 B protein expression and thus reduced glucagon-mediate PAK substrates phosphorylation and gene expressions of key gluconeogenic enzymes. Conclusion: Berberine blocked cAMP/PKA signaling via inhibiting NF-κB p65 protein activity, and thus alleviated glucagon-driven hepatic gluconeogenesis.
引文
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[16]Jiang Q,Liu P,Wu X,et al.Berberine attenuates lipopolysaccharideinduced extracelluar matrix accumulation and inflammation in rat mesangial cells:Involvement of NF-κB signaling pathway.Mol Cell Endocrinol,2011,331(1):34-40
[17]Jiang G,Zhang B B.Glucagon and regulation of glucose metabolism.Am J Physiol-Endoc M,2003,284(4):E671-E678
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[19]Abdollahi M,Chan T S,Subrahmanyam V,et al.Effects of phosphodiesterase 3,4,5 inhibitors on hepatocyte cAMP levels,glycogenolysis,gluconeogenesis and susceptibility to a mitochondrial toxin.Mol Cell Biochem,2003,252(1-2):205-211
[20]Ke B,Zhao Z,Ye X,et al.Inactivation of NF-κB p65(RelA)in liver improves insulin sensitivity and inhibits cAMP/PKA pathway.Diabetes,2015,64(10):3355-3362
[21]Johanns M,Lai Y C,Hsu M F,et al.AMPK antagonizes hepatic glucagon-stimulated cyclic AMP signalling via phosphorylation-induced activation of cyclic nucleotide phosphodiesterase 4B.Nat Commun,2016,7:10856
[2]Chung S T,Hsia D S,Chacko S K,et al.Increased gluconeogenesis in youth with newly diagnosed type 2 diabetes.Diabetologia,2015,58(3):596-603
[3]Cahill G F Jr.Starvation in man.New Engl J Med,1970,282:668-675
[4]Miller R A,Chu Q,Xie J,et al.Biguanides suppress hepatic glucagon signalling by decreasing production of cyclic AMP.Nature,2013,494:256-260
[5]Altarejos J Y,Montminy M.CREB and the CRTC co-activators:Sensors for hormonal and metabolic signals.Nat Rev Mol Cell Bio,2011,12:141-151
[6]Matulewicz N,Karczewska-Kupczewska M.Insulin resistance and chronic inflammation.Postep Hig Med Dosw,2016,70:1245-1258
[7]Zand H,Morshedzadeh N,Naghashian F.Signaling pathways linking inflammation to insulin resistance.Diabetes Metab Syndr,2017,11(Suppl 1):S307-S309
[8]金美英,崔镇海,朴春丽,等.基于IRE1α/JNK通路探析解毒通络调肝方对2型糖尿病调控机制.中华中医药杂志,2018,33(12):5353-5357
[9]Xia X,Yan J,Shen Y,et al.Berberine improves glucose metabolism in diabetic rats by inhibition of hepatic gluconeogenesis.PLoS One,2011,6(2):e16556
[10]Zhang M,Lv X,Li J,et al.Sodium caprate augments the hypoglycemic effect of berberine via AMPK in inhibiting hepatic gluconeogenesis.Mol Cell Endocrinol,2012,363(1-2):122-130
[11]Jiang S J,Dong H,Li J B,et al.Berberine inhibits hepatic gluconeogenesis via the LKB1-AMPK-TORC2 signaling pathway in streptozotocin-induced diabetic rats.World J Gastroentero,2015,21(25):7777-7785
[12]Donath M Y.Targeting inflammation in the treatment of type 2diabetes.Diabetes Obes Meta,2013,15(Suppl 3):193-196
[13]毛竹君,翁思颖,王磊,等.小檗碱调控IR-Hep G2细胞mi R-29a-3p及胰岛素受体信号通路的作用机制研究.中华中医药杂志,2018,33(12):5559-5562
[14]Gratas-Delamarche A,DerbréF,Vincent S,et al.Physical inactivity,insulin resistance,and the oxidative-inflammatory loop.Free Radic Res,2014,48(1):93-108
[15]Yuan M,Konstantopoulos N,Lee J,et al.Reversal of obesity-and dietinduced insulin resistance with salicylates or targeted disruption of Ikkβ.Science,2001,293(5535):1673-1677
[16]Jiang Q,Liu P,Wu X,et al.Berberine attenuates lipopolysaccharideinduced extracelluar matrix accumulation and inflammation in rat mesangial cells:Involvement of NF-κB signaling pathway.Mol Cell Endocrinol,2011,331(1):34-40
[17]Jiang G,Zhang B B.Glucagon and regulation of glucose metabolism.Am J Physiol-Endoc M,2003,284(4):E671-E678
[18]Soderling S H,Beavo J A.Regulation of cAMP and cGMP signaling:New phosphodiesterases and new functions.Curr Opin Cell Biol,2000,12(2):174-179
[19]Abdollahi M,Chan T S,Subrahmanyam V,et al.Effects of phosphodiesterase 3,4,5 inhibitors on hepatocyte cAMP levels,glycogenolysis,gluconeogenesis and susceptibility to a mitochondrial toxin.Mol Cell Biochem,2003,252(1-2):205-211
[20]Ke B,Zhao Z,Ye X,et al.Inactivation of NF-κB p65(RelA)in liver improves insulin sensitivity and inhibits cAMP/PKA pathway.Diabetes,2015,64(10):3355-3362
[21]Johanns M,Lai Y C,Hsu M F,et al.AMPK antagonizes hepatic glucagon-stimulated cyclic AMP signalling via phosphorylation-induced activation of cyclic nucleotide phosphodiesterase 4B.Nat Commun,2016,7:10856
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