Duvelisib attenuates bleomycin-induced pulmonary fibrosis via inhibiting the PI3K/Akt/mTOR signalling pathway

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive interstitial lung disease that seriously threatens the health of patients. The pathogenesis of IPF is still unclear, and there is a lack of effective therapeutic drugs. Myofibroblasts are the main effector cells of IPF, leading to excessive deposition of extracellular matrix (ECM) and promoting the progression of fibrosis. Inhibiting the excessive activation and relieving autophagy blockage of myofibroblasts is the key to treat IPF. PI3K/Akt/mTOR pathway plays a key regulatory role in promoting fibroblast activation and autophagy inhibition in lung fibrosis. Duvelisib is a PI3K inhibitor that can simultaneously inhibit the activities of PI3K-δ and PI3K-γ, and is mainly used for the treatment of relapsed/refractory chronic lymphocytic leukaemia (CLL) and small lymphocytic lymphoma tumour (SLL). In this study, we aimed to examine the effects of Duvelisib on pulmonary fibrosis. We used a mouse model of bleomycin-induced pulmonary fibrosis to evaluate the effects of Duvelisib on pulmonary fibrosis in vivo and further explored the potential pharmacological mechanisms of Duvelisib in lung fibroblasts in vitro. The in vivo experiments showed that Duvelisib significantly alleviated bleomycin-induced collagen deposition and improved pulmonary function. In vitro and in vivo pharmacological experiments showed that Duvelisib dose-dependently suppressed lung fibroblast activation and improved autophagy inhibition by inhibiting the phosphorylation of PI3K, Akt and mTOR. Our results indicate that Duvelisib can alleviate the severity of pulmonary fibrosis and provide potential drugs for the treatment of pulmonary fibrosis.     

IPF Fibroblasts Are Desensitized to Type I Collagen Matrix-Induced Cell Death by Suppressing Low Autophagy via Aberrant Akt/mTOR Kinases

Idiopathic pulmonary fibrosis (IPF) is a chronic, lethal interstitial lung disease in which the aberrant PTEN/Akt axis plays a major role in conferring a survival phenotype in response to the cell death inducing properties of type I collagen matrix. The underlying mechanism by which IPF fibroblasts become desensitized to polymerized collagen, thereby eluding collagen matrix-induced cell death has not been fully elucidated. We hypothesized that the pathologically altered PTEN/Akt axis suppresses autophagy via high mTOR kinase activity, which subsequently desensitizes IPF fibroblasts to collagen matrix induced cell death. We found that the autophagosome marker LC3-2 expression is suppressed, while mTOR activity remains high when IPF fibroblasts are cultured on collagen. However, LC3-2 expression increased in response to IPF fibroblast attachment to collagen in the presence of rapamycin. In addition, PTEN over-expression or Akt inhibition suppressed mTOR activity, thereby increasing LC3-2 expression in IPF fibroblasts. Furthermore, the treatment of IPF fibroblasts over-expressing PTEN or dominant negative Akt with autophagy inhibitors increased IPF fibroblast cell death. Enhanced p-mTOR expression along with low LC3-2 expression was also found in myofibroblasts within the fibroblastic foci from IPF patients. Our data show that the aberrant PTEN/Akt/mTOR axis desensitizes IPF fibroblasts from polymerized collagen driven stress by suppressing autophagic activity, which produces a viable IPF fibroblast phenotype on collagen. This suggests that the aberrantly regulated autophagic pathway may play an important role in maintaining a pathological IPF fibroblast phenotype in response to collagen rich environment.     

Acetylshikonin induces autophagy‐dependent apoptosis through the key LKB1‐AMPK and PI3K/Akt‐regulated mTOR signalling pathways in HL‐60 cells

Acetylshikonin (ASK) is a natural naphthoquinone derivative of traditional Chinese medicine Lithospermum erythrorhyzon. It has been reported that ASK has bactericidal, anti‐inflammatory and antitumour effects. However, whether ASK induces apoptosis and autophagy in acute myeloid leukaemia (AML) cells and the underlying mechanism are still unclear. Here, we explored the roles of apoptosis and autophagy in ASK‐induced cell death and the potential molecular mechanisms in human AML HL‐60 cells. The results demonstrated that ASK remarkably inhibited the cell proliferation, viability and induced apoptosis in HL‐60 cells through the mitochondrial pathway, and ASK promoted cell cycle arrest in the S‐phase. In addition, the increased formation of autophagosomes, the turnover from light chain 3B (LC3B) I to LC3B II and decrease of P62 suggested the induction of autophagy by ASK. Furthermore, ASK significantly decreased PI3K, phospho‐Akt and p‐p70S6K expression, while enhanced phospho‐AMP‐activated protein kinase (AMPK) and phospho‐liver kinase B1(LKB1) expression. The suppression of ASK‐induced the conversion from LC3B I to LC3B II caused by the application of inhibitors of AMPK (compound C) demonstrated that ASK‐induced autophagy depends on the LKB1/AMPK pathway. These data suggested that the autophagy induced by ASK were dependent on the activation of LKB1/AMPK signalling and suppression of PI3K/Akt/mTOR pathways. The cleavage of the apoptosis‐related markers caspase‐3 and caspase‐9 and the activity of caspase‐3 induced by ASK were markedly reduced by inhibitor of AMPK (compound C), an autophagy inhibitor 3‐methyladenine (3‐MA) and another autophagy inhibitor chloroquine (CQ). Taken together, our data reveal that ASK‐induced HL‐60 cell apoptosis is dependent on the activation of autophagy via the LKB1/AMPK and PI3K/Akt‐regulated mTOR signalling pathways.     

mTORC1 activation decreases autophagy in aging and idiopathic pulmonary fibrosis and contributes to apoptosis resistance in IPF fibroblasts

Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, and usually lethal disease associated with aging. However, the molecular mechanisms of the aging process that contribute to the pathogenesis of IPF have not been elucidated. IPF is characterized by abundant foci of highly active fibroblasts and myofibroblasts resistant to apoptosis. Remarkably, the role of aging in the autophagy activity of lung fibroblasts and its relationship with apoptosis, as adaptive responses, has not been evaluated previously in this disease. In the present study, we analyzed the dynamics of autophagy in primary lung fibroblasts from IPF compared to young and age‐matched normal lung fibroblasts. Our results showed that aging contributes for a lower induction of autophagy on basal conditions and under starvation which is mediated by mTOR pathway activation. Treatment with rapamycin and PP242, that target the PI3K/AKT/mTOR signaling pathway, modified starvation‐induced autophagy and apoptosis in IPF fibroblasts. Interestingly, we found a persistent activation of this pathway under starvation that contributes to the apoptosis resistance in IPF fibroblasts. These findings indicate that aging affects adaptive responses to stress decreasing autophagy through activation of mTORC1 in lung fibroblasts. The activation of this pathway also contributes to the resistance to cell death in IPF lung fibroblasts.     

IL‐17A‐producing T cells exacerbate fine particulate matter‐induced lung inflammation and fibrosis by inhibiting PI3K/Akt/mTOR‐mediated autophagy

Fine particulate matter (PM2.5) is the primary air pollutant that is able to induce airway injury. Compelling evidence has shown the involvement of IL‐17A in lung injury, while its contribution to PM2.5‐induced lung injury remains largely unknown. Here, we probed into the possible role of IL‐17A in mouse models of PM2.5‐induced lung injury. Mice were instilled with PM2.5 to construct a lung injury model. Flow cytometry was carried out to isolate γδT and Th17 cells. ELISA was adopted to detect the expression of inflammatory factors in the supernatant of lavage fluid. Primary bronchial epithelial cells (mBECs) were extracted, and the expression of TGF signalling pathway‐, autophagy‐ and PI3K/Akt/mTOR signalling pathway‐related proteins in mBECs was detected by immunofluorescence assay and Western blot analysis. The mitochondrial function was also evaluated. PM2.5 aggravated the inflammatory response through enhancing the secretion of IL‐17A by γδT/Th17 cells. Meanwhile, PM2.5 activated the TGF signalling pathway and induced EMT progression in bronchial epithelial cells, thereby contributing to pulmonary fibrosis. Besides, PM2.5 suppressed autophagy of bronchial epithelial cells by up‐regulating IL‐17A, which in turn activated the PI3K/Akt/mTOR signalling pathway. Furthermore, IL‐17A impaired the energy metabolism of airway epithelial cells in the PM2.5‐induced models. This study suggested that PM2.5 could inhibit autophagy of bronchial epithelial cells and promote pulmonary inflammation and fibrosis by inducing the secretion of IL‐17A in γδT and Th17 cells and regulating the PI3K/Akt/mTOR signalling pathway.     

A Critical Role for the mTORC2 Pathway in Lung Fibrosis

A characteristic of dysregulated wound healing in IPF is fibroblastic-mediated damage to lung epithelial cells within fibroblastic foci. In these foci, TGF-β and other growth factors activate fibroblasts that secrete growth factors and matrix regulatory proteins, which activate a fibrotic cascade. Our studies and those of others have revealed that Akt is activated in IPF fibroblasts and it mediates the activation by TGF-β of pro-fibrotic pathways. Recent studies show that mTORC2, a component of the mTOR pathway, mediates the activation of Akt. In this study we set out to determine if blocking mTORC2 with MLN0128, an active site dual mTOR inhibitor, which blocks both mTORC1 and mTORC2, inhibits lung fibrosis. We examined the effect of MLN0128 on TGF-β-mediated induction of stromal proteins in IPF lung fibroblasts; also, we looked at its effect on TGF-β-mediated epithelial injury using a Transwell co-culture system. Additionally, we assessed MLN0128 in the murine bleomycin lung model. We found that TGF-β induces the Rictor component of mTORC2 in IPF lung fibroblasts, which led to Akt activation, and that MLN0128 exhibited potent anti-fibrotic activity in vitro and in vivo. Also, we observed that Rictor induction is Akt-mediated. MLN0128 displays multiple anti-fibrotic and lung epithelial-protective activities; it (1) inhibited the expression of pro-fibrotic matrix-regulatory proteins in TGF-β-stimulated IPF fibroblasts; (2) inhibited fibrosis in a murine bleomycin lung model; and (3) protected lung epithelial cells from injury caused by TGF-β-stimulated IPF fibroblasts. Our findings support a role for mTORC2 in the pathogenesis of lung fibrosis and for the potential of active site mTOR inhibitors in the treatment of IPF and other fibrotic lung diseases.     

Inhibition of PI3K prevents the proliferation and differentiation of human lung fibroblasts into myofibroblasts: the role of class I P110 isoforms

Idiopathic pulmonary fibrosis (IPF) is a progressive fibroproliferative disease characterized by an accumulation of fibroblasts and myofibroblasts in the alveolar wall. Even though the pathogenesis of this fatal disorder remains unclear, transforming growth factor-β (TGF-β)-induced differentiation and proliferation of myofibroblasts is recognized as a primary event. The molecular pathways involved in TGF-β signalling are generally Smad-dependent yet Smad-independent pathways, including phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt), have been recently proposed. In this research we established ex-vivo cultures of human lung fibroblasts and we investigated the role of the PI3K/Akt pathway in two critical stages of the fibrotic process induced by TGF-β: fibroblast proliferation and differentiation into myofibroblasts. Here we show that the pan-inhibitor of PI3Ks LY294002 is able to abrogate the TGF-β-induced increase in cell proliferation, in α- smooth muscle actin expression and in collagen production besides inhibiting Akt phosphorylation, thus demonstrating the centrality of the PI3K/Akt pathway in lung fibroblast proliferation and differentiation. Moreover, for the first time we show that PI3K p110δ and p110γ are functionally expressed in human lung fibroblasts, in addition to the ubiquitously expressed p110α and β. Finally, results obtained with both selective inhibitors and gene knocking-down experiments demonstrate a major role of p110γ and p110α in both TGF-β-induced fibroblast proliferation and differentiation. This finding suggests that specific PI3K isoforms can be pharmacological targets in IPF.     

RhoGDI2与PI3K/Akt/mTOR信号通路在肺癌细胞中的相关性研究

目的探讨肿瘤转移相关因子RhoGDI2与PI3K/Akt/mTOR信号通路在肺癌侵袭转移过程中的作用及相关机制。方法利用PI3K/Akt/mTOR信号通路上特异性的抑制剂,采用MTT法,伤口愈合实验及侵袭实验观察不同浓度药物对肺癌95D细胞生长侵袭转移能力的影响,通过Western Blot方法观察RhoGDI2蛋白水平的变化。结果PI3K抑制剂LY294002及mTOR抑制剂Rapamycin都能抑制肺癌细胞95D的侵袭转移能力,联合应用抑制作用更强。PI3K抑制剂LY294002处理组RhoGDI2蛋白的表达量增加,且随浓度增加RhoGDI2蛋白表达也增加。mTOR抑制剂Rapamycin组,在低浓度时增加RhoGDI2蛋白的表达,但增大Rapamycin的浓度,RhoGDI2蛋白的表达反而降低。低浓度LY294002组和Rapa-mycin组联合应用可以明显增加RhoGDI2蛋白的表达。结论PI3K/Akt/mTOR信号通路中Akt的活化与RhoGDI2密切相关,RhoGDI2可能直接或间接通过与Akt的相互作用参与调节肺癌的侵袭转移的过程。     

PI3K/Akt/mTOR信号通路与自噬及肿瘤的关系

自噬是一种以胞质内出现双层膜结构包裹长寿命蛋白和细胞器的自噬体为特征的细胞“自我消化”过程,在维持细胞内稳态、发育、肿瘤发生和感染中发挥重要作用。近来,诸多研究表明,自噬作为一把“双刃剑”,对肿瘤的发生发展既有促进作用,也有抑制作用。PI3K/Akt/mTOR通路由PI3激酶(PI3K)、蛋白激酶B（PKB/Akt）和哺乳动物类雷帕霉素靶蛋白（mTOR）3个作用分子组成,是一个中心的调节机构,对肿瘤细胞的生长与增殖有促进作用,同时对自噬进行抑制。本文就PI3K/Akt/mTOR通路与自噬及肿瘤发生发展的关系作一综述。     

Regulation of Autophagy and Ubiquitinated Protein Accumulation by bFGF Promotes Functional Recovery and Neural Protection in a Rat Model of Spinal Cord Injury

The role of autophagy in the recovery of spinal cord injury remains controversial; in particular, the mechanism of autophagy regulated degradation of ubiquitinated proteins has not been discussed to date. In this study, we investigated the protective role of basic fibroblast growth factor (bFGF) both in vivo and in vitro and demonstrated that excessive autophagy and ubiquitinated protein accumulation is involved in the rat model of trauma. bFGF administration improved recovery and increased the survival of neurons in spinal cord lesions in the rat model. The protective effect of bFGF is related to the inhibition of autophagic protein LC3II levels; bFGF treatment also enhances clearance of ubiquitinated proteins by p62, which also increases the survival of neuronal PC-12 cells. The activation of the downstream signals of the PI3K/Akt/mTOR pathway by bFGF treatment was detected both in vivo and in vitro. Combination therapy including the autophagy activator rapamycin partially abolished the protective effect of bFGF. The present study illustrates that the role of bFGF in SCI recovery is related to the inhibition of excessive autophagy and enhancement of ubiquitinated protein clearance via the activation of PI3K/Akt/mTOR signaling. Overall, our study suggests a new trend for bFGF drug development for central nervous system injuries and sheds light on protein signaling involved in bFGF action.     

PI3K/Akt/mTOR signaling orchestrates the phenotypic transition and chemo-resistance of small cell lung cancer

Small cell lung cancer(SCLC) is a phenotypically heterogeneous disease with an extremely poor prognosis,which is mainly attributed to the rapid development of resistance to chemotherapy. However, the relation between the growth phenotypes and chemo-resistance of SCLC remains largely unclear. Through comprehensive bioinformatic analyses, we found that the heterogeneity of SCLC phenotype was significantly associated with different sensitivity to chemotherapy. Adherent or semiadherent SCLC cells were enriched with activation of the PI3K/Akt/mTOR pathway and were highly chemoresistant. Mechanistically,activation of the PI3K/Akt/mTOR pathway promotes the phenotypic transition from suspension to adhesion growth pattern and confers SCLC cells with chemo-resistance. Such chemo-resistance could be largely overcome by combining chemotherapy with PI3K/Akt/mTOR pathway inhibitors. Our findings support that the PI3K/Akt/mTOR pathway plays an important role in SCLC phenotype transition and chemo-resistance,which holds important clinical implications for improving SCLC treatment.     

Vertical inhibition of the PI3K/Akt/mTOR pathway for the treatment of osteoarthritis

Osteoarthritis is characterized by degenerative alterations of articular cartilage including both the degradation of extracellular matrix and the death of chondrocytes. The PI3K/Akt pathway has been demonstrated to involve in both processes. Inhibition of its downstream target NF‐kB reduces the degradation of extracellular matrix via decreased production of matrix metalloproteinases while inhibition of mTOR increased autophagy to reduce chondrocyte death. However, mTOR feedback inhibits the activity of the PI3K/Akt pathway and inhibition of mTOR could result in increased activity of the PI3K/Akt/NF‐kB pathway. We proposed that the use of dual inhibitors of PI3K and mTOR could be a promising approach to more efficiently inhibit the PI3K/Akt pathway than rapamycin or PI3K inhibitor alone and produce better treatment outcome. J. Cell. Biochem. 114: 245–249, 2013. © 2012 Wiley Periodicals, Inc.     

Mitochondrial uncoupling protein‐2 reprograms metabolism to induce oxidative stress and myofibroblast senescence in age‐associated lung fibrosis

Mitochondrial dysfunction has been associated with age‐related diseases, including idiopathic pulmonary fibrosis (IPF). We provide evidence that implicates chronic elevation of the mitochondrial anion carrier protein, uncoupling protein‐2 (UCP2), in increased generation of reactive oxygen species, altered redox state and cellular bioenergetics, impaired fatty acid oxidation, and induction of myofibroblast senescence. This pro‐oxidant senescence reprogramming occurs in concert with conventional actions of UCP2 as an uncoupler of oxidative phosphorylation with dissipation of the mitochondrial membrane potential. UCP2 is highly expressed in human IPF lung myofibroblasts and in aged fibroblasts. In an aging murine model of lung fibrosis, the in vivo silencing of UCP2 induces fibrosis regression. These studies indicate a pro‐fibrotic function of UCP2 in chronic lung disease and support its therapeutic targeting in age‐related diseases associated with impaired tissue regeneration and organ fibrosis.     

A proteomics approach to identifying key protein targets involved in VEGF inhibitor mediated attenuation of bleomycin‐induced pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a progressive lung disease with a life expectancy of less than 5 years post diagnosis for most patients. Poor molecular characterization of IPF has led to insufficient understanding of the pathogenesis of the disease, resulting in lack of effective therapies. In this study, we have integrated a label‐free LC‐MS based approach with systems biology to identify signaling pathways and regulatory nodes within protein interaction networks that govern phenotypic changes that may lead to IPF. Ingenuity Pathway Analysis of proteins modulated in response to bleomycin treatment identified PI3K/Akt and Wnt signaling as the most significant profibrotic pathways. Similar analysis of proteins modulated in response to vascular endothelial growth factor (VEGF) inhibitor (CBO‐P11) treatment identified natural killer cell signaling and PTEN signaling as the most significant antifibrotic pathways. Mechanistic/mammalian target of rapamycin (mTOR) and extracellular signal‐regulated kinase (ERK) were identified to be key mediators of pro‐ and antifibrotic response, where bleomycin (BLM) treatment resulted in increased expression and VEGF inhibitor treatment attenuated expression of mTOR and ERK. Using a BLM mouse model of pulmonary fibrosis and VEGF inhibitor CBO‐P11 as a therapeutic measure, we identified a comprehensive set of signaling pathways and proteins that contribute to the pathogenesis of pulmonary fibrosis that can be targeted for therapy against this fatal disease.     

PI3K/Akt/mTOR信号通路及临床相关肿瘤抑制剂

哺乳动物雷帕霉素靶（mTOR）和蛋白激酶B（Akt/PKB）与肿瘤发生的密切关系已被广泛地认可.mTOR是一种丝/苏氨酸激酶,可以通过影响mRNA转录、代谢、自噬等方式调控细胞的生长.它既是PI3K的效应分子,也可以是PI3K的反馈调控因子.mTORC1 和mTORC2是mTOR的两种不同复合物. 对雷帕霉素敏感的mTORC1受到营养、生长因子、能量和应激4种因素的影响.生长因子通过PI3K/Akt信号通路调控mTORC1是最具特征性调节路径.而mTORC2最为人熟知的是作为Akt473磷酸化位点的上游激酶. 同样,Akt/PKB在细胞增殖分化、迁移生长过程中发挥着重要作用. 随着Thr308和Ser473两个位点激活,Akt/PKB也得以全面活化.因此,mTORC2-Akt-mTORC1的信号通路在肿瘤形成和生长中是可以存在的.目前临床肿瘤治疗中,PI3K/Akt/mTOR是重要的靶向治疗信号通路.然而,仅抑制mTORC1活性,不是所有的肿瘤都能得到预期控制.雷帕霉素虽然能抑制mTORC1,但也能反馈性地增加PI3K信号活跃度,从而影响治疗预后.近来发现的第二代抑制剂可以同时抑制mTORC1/2和PI3K活性,这种抑制剂被认为在肿瘤治疗上颇具前景.本综述着重阐述了PI3K/Akt/mTOR信号通路的传导、各因子之间的相互调控以及相关抑制剂的发展.     

KAI1/CD82 decreases Rac1 expression and cell proliferation through PI3K/Akt/mTOR pathway in H1299 lung carcinoma cells

Although the KAI1/CD82 protein has been reported to inhibit cell metastasis in many studies, its mechanism of action has not yet been fully elucidated. In the present study, we investigated the possible effects of KAI1/CD82 on the metastatic phenotype in H1299 lung carcinoma cells. These studies were based on the pivotal role that the acquisition of motile phenotype plays on the initial steps of metastasis. KAI1/CD82‐mediated morphological changes were observed using phase contrast microscopy. We report here, that a KAI1/CD82‐induced phenotypic change was involved in the decrease of Rac1 expression and GTPase activity. However, we found that KAI1/CD82 did not regulate Rac1 mRNA levels. This suggests the existence of another regulatory mechanism of Rac1 protein maturation or activation. To identify the signaling pathway of Rac1 regulation, we investigated the PI3K/Akt/mTOR pathway, since the PI3K/Akt pathway regulates Rac1 activation and mTOR is known to play a regulatory role in protein translation. H1299/CD82‐transfectants showed lower mTOR expression and cell growth than the control group. The data obtained from this study suggested that KAI1/CD82 decreased the metastatic phenotype of H1299 lung carcinoma cells by down‐regulating Rac1 expression through the PI3K/Akt/mTOR pathway. Copyright © 2008 John Wiley & Sons, Ltd.     

Ribosomal protein L22‐like1 promotes prostate cancer progression by activating PI3K/Akt/mTOR signalling pathway

Prostate cancer (PCa) is one of the most common malignancies in men. Ribosomal protein L22‐like1 (RPL22L1), a component of the ribosomal 60 S subunit, is associated with cancer progression, but the role and potential mechanism of RPL22L1 in PCa remain unclear. The aim of this study was to investigate the role of RPL22L1 in PCa progression and the mechanisms involved. Bioinformatics and immunohistochemistry analysis showed that the expression of RPL22L1 was significantly higher in PCa tissues than in normal prostate tissues. The cell function analysis revealed that RPL22L1 significantly promoted the proliferation, migration and invasion of PCa cells. The data of xenograft tumour assay suggested that the low expression of RPL22L1 inhibited the growth and invasion of PCa cells in vivo. Mechanistically, the results of Western blot proved that RPL22L1 activated PI3K/Akt/mTOR pathway in PCa cells. Additionally, LY294002, an inhibitor of PI3K/Akt pathway, was used to block this pathway. The results showed that LY294002 remarkably abrogated the oncogenic effect of RPL22L1 on PCa cell proliferation and invasion. Taken together, our study demonstrated that RPL22L1 is a key gene in PCa progression and promotes PCa cell proliferation and invasion via PI3K/Akt/mTOR pathway, thus potentially providing a new target for PCa therapy.     

Astragaloside IV modulates TGF‐β1‐dependent epithelial‐mesenchymal transition in bleomycin‐induced pulmonary fibrosis

Epithelial‐mesenchymal transition (EMT) plays an important role in idiopathic pulmonary fibrosis (IPF). Astragaloside IV (ASV), a natural saponin from astragalus membranaceus, has shown anti‐fibrotic property in bleomycin (BLM)‐induced pulmonary fibrosis. The current study was undertaken to determine whether EMT was involved in the beneficial of ASV against BLM‐induced pulmonary fibrosis and to elucidate its potential mechanism. As expected, in BLM‐induced IPF, ASV exerted protective effects on pulmonary fibrosis and ASV significantly reversed BLM‐induced EMT. Intriguing, transforming growth factor‐β1 (TGF‐β1) was found to be up‐regulated, whereas Forkhead box O3a (FOXO3a) was hyperphosphorylated and less expressed. However, ASV treatment inhibited increased TGF‐β1 and activated FOXO3a in lung tissues. TGF‐β1 was administered to alveolar epithelial cells A549 to induce EMT in vitro. Meanwhile, stimulation with TGF‐β1‐activated phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) pathway and induced FOXO3a hyperphosphorylated and down‐regulated. It was found that overexpression of FOXO3a leading to the suppression of TGF‐β1‐induced EMT. Moreover, ASV treatment, similar with the TGF‐β1 or PI3K/Akt inhibitor, reverted these cellular changes and inhibited EMT in A549 cells. Collectively, the results suggested that ASV significantly inhibited TGF‐β1/PI3K/Akt‐induced FOXO3a hyperphosphorylation and down‐regulation to reverse EMT during the progression of fibrosis.     

Bone marrow‐derived mesenchymal stem cells enhance autophagy via PI3K/AKT signalling to reduce the severity of ischaemia/reperfusion‐induced lung injury

Autophagy, a type II programmed cell death, is essential for cell survival under stress, e.g. lung injury, and bone marrow‐derived mesenchymal stem cells (BM‐MSCs) have great potential for cell therapy. However, the mechanisms underlying the BM‐MSC activation of autophagy to provide a therapeutic effect in ischaemia/reperfusion‐induced lung injury (IRI) remain unclear. Thus, we investigate the activation of autophagy in IRI following transplantation with BM‐MSCs. Seventy mice were pre‐treated with BM‐MSCs before they underwent lung IRI surgery in vivo. Human pulmonary micro‐vascular endothelial cells (HPMVECs) were pre‐conditioned with BM‐MSCs by oxygen‐glucose deprivation/reoxygenation (OGD) in vitro. Expression markers for autophagy and the phosphoinositide 3‐kinase/protein kinase B (PI3K/Akt) signalling pathway were analysed. In IRI‐treated mice, administration of BM‐MSCs significantly attenuated lung injury and inflammation, and increased the level of autophagy. In OGD‐treated HPMVECs, co‐culture with BM‐MSCs attenuated endothelial permeability by decreasing the level of cell death and enhanced autophagic activation. Moreover, administration of BM‐MSCs decreased the level of PI3K class I and p‐Akt while the expression of PI3K class III was increased. Finally, BM‐MSCs‐induced autophagic activity was prevented using the inhibitor LY294002. Administration of BM‐MSCs attenuated lung injury by improving the autophagy level via the PI3K/Akt signalling pathway. These findings provide further understanding of the mechanisms related to BM‐MSCs and will help to develop new cell‐based therapeutic strategies in lung injury.     

Berberine regulates mesangial cell proliferation and cell cycle to attenuate diabetic nephropathy through the PI3K/Akt/AS160/GLUT1 signalling pathway

High glucose (HG) is one of the basic factors of diabetic nephropathy (DN), which leads to high morbidity and disability. During DN, the expression of glomerular glucose transporter 1 (GLUT1) increases, but the relationship between HG and GLUT1 is unclear. Glomerular mesangial cells (GMCs) have multiple roles in HG‐induced DN. Here, we report prominent glomerular dysfunction, especially GMC abnormalities, in DN mice, which is closely related to GLUT1 alteration. In vivo studies have shown that BBR can alleviate pathological changes and abnormal renal function indicators of DN mice. In vitro, BBR (30, 60 and 90 μmol/L) not only increased the proportion of G1 phase cells but also reduced the proportion of S phase cells under HG conditions at different times. BBR (60 μmol/L) significantly reduced the expression of PI3K‐p85, p‐Akt, p‐AS160, membrane‐bound GLUT1 and cyclin D1, but had almost no effect on total protein. Furthermore, BBR significantly declined the glucose uptake and retarded cyclin D1‐mediated GMC cell cycle arrest in the G1 phase. This study demonstrated that BBR can inhibit the development of DN, which may be due to BBR inhibiting the PI3K/Akt/AS160/GLUT1 signalling pathway to regulate HG‐induced abnormal GMC proliferation and the cell cycle, supporting BBR as a potential therapeutic drug for DN.