钙蛋白酶活化的mTORC2 / Akt通路介导哮喘气道平滑肌重塑
2016/12/27
背景：过敏性哮喘以炎症和气道重塑为特征。气道重塑可表现为细胞外基质（ECM）的过度沉积以及平滑肌增厚，它与气道反应性增高和哮喘严重程度相关。钙蛋白酶是钙依赖性内肽酶家族，其在ECM重塑中起重要作用。然而，钙蛋白酶在气道平滑肌重塑中的作用尚不清楚。
目的：探索钙蛋白酶在哮喘气道重塑中的作用以及潜在机制。
方法：通过卵蛋白致敏和激发制备大鼠哮喘模型。在模型中使用钙蛋白酶条件敲除小鼠。从大鼠气道的平滑肌束中分离气道平滑肌细胞（ASMCs）。选择哮喘患者的细胞因子IL-4，IL-5，TNF-α和TGF-β1，以及血清来处理ASMCs。分析ASMCs中胶原蛋白-I合成，细胞增殖和Akt的磷酸化。
结果：使用钙蛋白酶敲除小鼠抑制了钙蛋白酶，从而减弱大鼠哮喘模型中的气道平滑肌重塑。来自哮喘患者的细胞因子IL-4，IL-5，TNF-α和TGF-β1以及血清增加ASMCs中的胶原蛋白-I合成，细胞增殖和Akt的磷酸化，它能被钙蛋白酶抑制剂MDL28170阻断。此外，MDL28170能降低细胞因子诱导Rictor蛋白产生过多，它是哺乳动物雷帕霉素靶复合物2（mammalian target of rapamycin complex 2，mTORC2）最重要的组成部分。在小鼠哮喘模型中，阻断mTORC2信号通路可防止细胞因子诱导的Akt磷酸化，胶原蛋白-I合成和ASMCs的增殖，从而减弱气道平滑肌重塑。
结论和临床相关性：我们的研究结果表明，钙蛋白酶通过mTORC2 / Akt信号通路介导细胞因子诱导的胶原蛋白-I合成和ASMCs的增殖，从而调节哮喘的气道平滑肌重塑。
Background: Allergic asthma is characterized by inflammation and airway remodelling. Airway remodelling with excessive deposition of extracellular matrix (ECM) and larger smooth muscle mass are correlated with increased airway responsiveness and asthma severity. Calpain is a family of calcium-dependent endopeptidases, which plays an important role in ECM remodelling. However, the role of calpain in airway smooth muscle remodelling remains unknown.
Objective: To investigate the role of calpain in asthmatic airway remodelling as well as the underlying mechanism.
Methods: The mouse asthma model was made by ovalbumin sensitization and challenge. Calpain conditional knockout mice were studied in the model. Airway smooth muscle cells (ASMCs) were isolated from smooth muscle bundles in airway of rats. Cytokines IL-4, IL-5, TNF-α, and TGF-β1, and serum from patients with asthma were selected to treated ASMCs. Collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs were analysed.
Results: Inhibition of calpain using calpain knockout mice attenuated airway smooth muscle remodelling in mouse asthma models. Cytokines IL-4, IL-5, TNF-α, and TGF-β1, and serum from patients with asthma increased collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs, which were blocked by the calpain inhibitor MDL28170. Moreover, MDL28170 reduced cytokine-induced increases in Rictor protein, which is the most important component of mammalian target of rapamycin complex 2 (mTORC2). Blockage of the mTORC2 signal pathway prevented cytokine-induced phosphorylation of Akt, collagen-I synthesis, and cell proliferation of ASMCs and attenuated airway smooth muscle remodelling in mouse asthma models.
Conclusions and Clinical Relevance: Our results indicate that calpain mediates cytokine-induced collagen-I synthesis and proliferation of ASMCs via the mTORC2/Akt signalling pathway, thereby regulating airway smooth muscle remodelling in asthma.
目的：探索钙蛋白酶在哮喘气道重塑中的作用以及潜在机制。
方法：通过卵蛋白致敏和激发制备大鼠哮喘模型。在模型中使用钙蛋白酶条件敲除小鼠。从大鼠气道的平滑肌束中分离气道平滑肌细胞（ASMCs）。选择哮喘患者的细胞因子IL-4，IL-5，TNF-α和TGF-β1，以及血清来处理ASMCs。分析ASMCs中胶原蛋白-I合成，细胞增殖和Akt的磷酸化。
结果：使用钙蛋白酶敲除小鼠抑制了钙蛋白酶，从而减弱大鼠哮喘模型中的气道平滑肌重塑。来自哮喘患者的细胞因子IL-4，IL-5，TNF-α和TGF-β1以及血清增加ASMCs中的胶原蛋白-I合成，细胞增殖和Akt的磷酸化，它能被钙蛋白酶抑制剂MDL28170阻断。此外，MDL28170能降低细胞因子诱导Rictor蛋白产生过多，它是哺乳动物雷帕霉素靶复合物2（mammalian target of rapamycin complex 2，mTORC2）最重要的组成部分。在小鼠哮喘模型中，阻断mTORC2信号通路可防止细胞因子诱导的Akt磷酸化，胶原蛋白-I合成和ASMCs的增殖，从而减弱气道平滑肌重塑。
结论和临床相关性：我们的研究结果表明，钙蛋白酶通过mTORC2 / Akt信号通路介导细胞因子诱导的胶原蛋白-I合成和ASMCs的增殖，从而调节哮喘的气道平滑肌重塑。
(黄丹 张红萍 王刚 四川大学华西医院中西医结合科呼吸病组 610041 摘译)
（Clinical & Experimental Allergy; 2016;(00)1-14）
（Clinical & Experimental Allergy; 2016;(00)1-14）
Calpain-activated mTORC2/Akt pathway mediates airway smooth muscle remodelling in asthma
S.-S. Rao;Q. Mu;Y. Zeng;P.-C. Cai;F. Liu;J. Yang;Y. Xia;Q. Zhang;L.-J. Song;
L.-L. Zhou; F.-Z. Li; Y.-X. Lin; J. Fang; P. A. Greer; H.-Z. Shi; W.-L. Ma; Y. Su;
H. Ye
Clinical & Experimental Allergy; 2016;(00)1-14Background: Allergic asthma is characterized by inflammation and airway remodelling. Airway remodelling with excessive deposition of extracellular matrix (ECM) and larger smooth muscle mass are correlated with increased airway responsiveness and asthma severity. Calpain is a family of calcium-dependent endopeptidases, which plays an important role in ECM remodelling. However, the role of calpain in airway smooth muscle remodelling remains unknown.
Objective: To investigate the role of calpain in asthmatic airway remodelling as well as the underlying mechanism.
Methods: The mouse asthma model was made by ovalbumin sensitization and challenge. Calpain conditional knockout mice were studied in the model. Airway smooth muscle cells (ASMCs) were isolated from smooth muscle bundles in airway of rats. Cytokines IL-4, IL-5, TNF-α, and TGF-β1, and serum from patients with asthma were selected to treated ASMCs. Collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs were analysed.
Results: Inhibition of calpain using calpain knockout mice attenuated airway smooth muscle remodelling in mouse asthma models. Cytokines IL-4, IL-5, TNF-α, and TGF-β1, and serum from patients with asthma increased collagen-I synthesis, cell proliferation, and phosphorylation of Akt in ASMCs, which were blocked by the calpain inhibitor MDL28170. Moreover, MDL28170 reduced cytokine-induced increases in Rictor protein, which is the most important component of mammalian target of rapamycin complex 2 (mTORC2). Blockage of the mTORC2 signal pathway prevented cytokine-induced phosphorylation of Akt, collagen-I synthesis, and cell proliferation of ASMCs and attenuated airway smooth muscle remodelling in mouse asthma models.
Conclusions and Clinical Relevance: Our results indicate that calpain mediates cytokine-induced collagen-I synthesis and proliferation of ASMCs via the mTORC2/Akt signalling pathway, thereby regulating airway smooth muscle remodelling in asthma.
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基质金属蛋白酶-1活化促进气道平滑肌生长并加重哮喘严重程度
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