Watt Matthew J, Holmes Anna G, Pinnamaneni Srijan K, Garnham Andrew P, Steinberg Gregory R, Kemp Bruce E, Febbraio Mark A
Cellular & Molecular Metabolism Laboratory, School of Medical Sciences, RMIT University, PO Box 71, Bundoora 3083, Victoria, Australia.
Am J Physiol Endocrinol Metab. 2006 Mar;290(3):E500-8. doi: 10.1152/ajpendo.00361.2005. Epub 2005 Sep 27.
Hormone-sensitive lipase (HSL) is important for the degradation of triacylglycerol in adipose and muscle tissue, but the tissue-specific regulation of this enzyme is not fully understood. We investigated the effects of adrenergic stimulation and AMPK activation in vitro and in circumstances where AMPK activity and catecholamines are physiologically elevated in humans in vivo (during physical exercise) on HSL activity and phosphorylation at Ser(563) and Ser(660), the PKA regulatory sites, and Ser(565), the AMPK regulatory site. In human experiments, skeletal muscle, subcutaneous adipose and venous blood samples were obtained before, at 15 and 90 min during, and 120 min after exercise. Skeletal muscle HSL activity was increased by approximately 80% at 15 min compared with rest and returned to resting rates at the cessation of and 120 min after exercise. Consistent with changes in plasma epinephrine, skeletal muscle HSL Ser(563) and Ser(660) phosphorylation were increased by 27% at 15 min (P < 0.05), remained elevated at 90 min, and returned to preexercise values postexercise. Skeletal muscle HSL Ser(565) phosphorylation and AMPK signaling were increased at 90 min during, and after, exercise. Phosphorylation of adipose tissue HSL paralleled changes in skeletal muscle in vivo, except HSL Ser(660) was elevated 80% in adipose compared with 35% in skeletal muscle during exercise. Studies in L6 myotubes and 3T3-L1 adipocytes revealed important tissue differences in the regulation of HSL. AMPK inhibited epinephrine-induced HSL activity in L6 myotubes and was associated with reduced HSL Ser(660) but not Ser(563) phosphorylation. HSL activity was reduced in L6 myotubes expressing constitutively active AMPK, confirming the inhibitory effects of AMPK on HSL activity. Conversely, in 3T3-L1 adipocytes, AMPK activation after epinephrine stimulation did not prevent HSL activity or glycerol release, which coincided with maintenance of HSL Ser(660) phosphorylation. Taken together, these data indicate that HSL activity is maintained in the face of AMPK activation as a result of elevated HSL Ser(660) phosphorylation in adipose tissue but not skeletal muscle.
激素敏感性脂肪酶（HSL）对于脂肪组织和肌肉组织中三酰甘油的降解很重要，但这种酶的组织特异性调节尚未完全明确。我们在体外以及在人体体内生理条件下（运动期间）AMPK活性和儿茶酚胺升高的情况下，研究了肾上腺素能刺激和AMPK激活对HSL活性以及PKA调节位点Ser(563)和Ser(660)以及AMPK调节位点Ser(565)磷酸化的影响。在人体实验中，在运动前、运动期间15分钟和90分钟以及运动后120分钟采集骨骼肌、皮下脂肪和静脉血样本。与静息状态相比，运动15分钟时骨骼肌HSL活性增加了约80%，在运动停止时和运动后120分钟恢复到静息水平。与血浆肾上腺素的变化一致，运动15分钟时骨骼肌HSL的Ser(563)和Ser(660)磷酸化增加了27%（P<0.05），在90分钟时仍保持升高，运动后恢复到运动前水平。运动期间90分钟以及运动后，骨骼肌HSL的Ser(565)磷酸化和AMPK信号传导增加。体内脂肪组织HSL的磷酸化与骨骼肌的变化相似，只是运动期间脂肪组织中HSL的Ser(660)升高了80%，而骨骼肌中升高了35%。在L6肌管和3T3-L1脂肪细胞中的研究揭示了HSL调节中重要的组织差异。AMPK抑制L6肌管中肾上腺素诱导的HSL活性，并与HSL的Ser(660)而非Ser(563)磷酸化减少有关。在组成型激活AMPK的L6肌管中，HSL活性降低，证实了AMPK对HSL活性的抑制作用。相反，在3T3-L1脂肪细胞中，肾上腺素刺激后AMPK激活并未阻止HSL活性或甘油释放，这与HSL的Ser(660)磷酸化的维持一致。综上所述，这些数据表明，由于脂肪组织而非骨骼肌中HSL的Ser(660)磷酸化升高，HSL活性在AMPK激活的情况下得以维持。
