Liang Minrui, Lv Jiaoyan, Chu Haiyan, Wang Jiucun, Chen Xiangjun, Zhu Xiaoxia, Xue Yu, Guan Ming, Zou Hejian
Division of Rheumatology, Huashan Hospital, Shanghai 200040, China; Institute of Rheumatology, Immunology and Allergy, Fudan University, Shanghai 200040, China.
Department of Immunology, School of Basic Medical Sciences, Fudan University, Shanghai 200032, China.
J Dermatol Sci. 2014 Nov;76(2):104-11. doi: 10.1016/j.jdermsci.2014.08.002. Epub 2014 Sep 7.
The mammalian target of rapamycin (mTOR) regulates cellular activity in many diseases, but the complex interplay with PI3K/Akt pathway may hampers its function.
This study was undertaken to determine the activity of PI3K/Akt/mTOR signaling in the fibroblasts from systemic sclerosis (SSc) patients, and compare the effects of vertical inhibiting PI3K/Akt/mTOR by BEZ235 and inhibiting mTOR alone by rapamycin in fibroblast activation and in two complementary established mouse model of SSc.
Pharmaceutical specific inhibitors BEZ235 and rapamycin were used to vertical inhibit PI3K/Akt/mTOR signaling and mTOR signaling alone in cultured fibroblasts and in mice. SSc mouse model was established by daily injecting bleomycin subcutaneously or by overexpression of constitutively active type I TGF-β receptor (TβRI(ca)). To delineate the mechanisms underlying the antifibrotic effects of BEZ235 and rapamycin, activity of PI3K/Akt/mTOR signaling was analyzed by determining the expressions of phosphorylated Akt, GSK-3β, mTOR and S6 ribosomal protein (S6).
Primary dermal fibroblasts demonstrated hyperactivity of PI3K/Akt and mTOR signaling. mTOR inhibitor rapamycin failed to inhibit dermal fibrosis in an established SSc mouse model. However, administration of a dual inhibitor for PI3K/Akt and mTOR signaling BEZ235 attenuated dermal fibrosis by reversing increased dermal thickness and collagen deposition in two SSc mouse models. Furthermore, BEZ235 showed superior inhibitory effect on fibroblast activation relative to rapamycin in vitro. Also both BEZ235 and rapamycin could prevent the phosphorylation of mTOR and S6 completely. BEZ235 also blocked the activation of Akt and GSK-3β dramatically, whereas rapamycin has been shown to increase further upregulation of phosphorylated Akt on Ser473 both in vitro and in vivo.
These data show that blocking PI3K/Akt/mTOR with BEZ235 leads to superior inhibitory effect for dermal fibrosis, suggesting that vertical inhibition of PI3K/Akt/mTOR signaling may have therapeutic potential for SSc.
雷帕霉素的哺乳动物靶点（mTOR）在多种疾病中调节细胞活性，但与PI3K/Akt信号通路复杂的相互作用可能会妨碍其功能。
本研究旨在确定系统性硬化症（SSc）患者成纤维细胞中PI3K/Akt/mTOR信号的活性，并比较BEZ235垂直抑制PI3K/Akt/mTOR和雷帕霉素单独抑制mTOR对成纤维细胞活化以及两种互补的已建立的SSc小鼠模型的影响。
使用药物特异性抑制剂BEZ235和雷帕霉素分别在培养的成纤维细胞和小鼠中垂直抑制PI3K/Akt/mTOR信号和单独抑制mTOR信号。通过每天皮下注射博来霉素或过表达组成型活性I型转化生长因子-β受体（TβRI(ca)）建立SSc小鼠模型。为了阐明BEZ235和雷帕霉素抗纤维化作用的潜在机制，通过测定磷酸化Akt、糖原合成酶激酶-3β（GSK-3β）、mTOR和S6核糖体蛋白（S6）的表达来分析PI3K/Akt/mTOR信号的活性。
原发性皮肤成纤维细胞表现出PI3K/Akt和mTOR信号的过度活化。mTOR抑制剂雷帕霉素未能在已建立的SSc小鼠模型中抑制皮肤纤维化。然而，给予PI3K/Akt和mTOR信号的双重抑制剂BEZ235可通过逆转两种SSc小鼠模型中增加的皮肤厚度和胶原沉积来减轻皮肤纤维化。此外，在体外，BEZ235对成纤维细胞活化的抑制作用优于雷帕霉素。而且BEZ235和雷帕霉素均可完全阻止mTOR和S6的磷酸化。BEZ235还显著阻断Akt和GSK-3β的活化，而雷帕霉素在体外和体内均已显示会进一步增加Ser473位点磷酸化Akt的上调。
这些数据表明，用BEZ235阻断PI3K/Akt/mTOR对皮肤纤维化具有更好的抑制作用，提示垂直抑制PI3K/Akt/mTOR信号可能对SSc具有治疗潜力。
