scholarly journals Activated mTORC1-SIRT6 Mediates Myofibroblast Differentiation and Idiopathic Pulmonary Fibrosis

2021 ◽  
Author(s):  
Ji Zhang ◽  
Yi Hu ◽  
Huiping Huang ◽  
Qun Liu ◽  
Yang Du ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Signaling Pathway ◽  
Mtor Signaling ◽  
Signalling Pathway ◽  
Lung Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Dependent Manner ◽  
Mtor Signaling Pathway ◽  
Mtorc1 Pathway ◽  
Tgf Β1

Abstract BackgroundIdiopathic pulmonary fibrosis (IPF) is characterised by accumulation of myofibroblasts and deposition of extracellular matrix proteins. Fibroblast-to-myofibroblast transdifferentiation and myofibroblast hyperproliferation plays a major role in pulmonary fibrosis. Moreover, mTOR signaling pathway and SIRT6 play a critical role in pulmonary fibrosis. However, the mechanisms whether SIRT6 affect the myofibroblasts differentiation during IPF remain unclear.MethodWe investigated myofibroblast differentiation using a bleomycin-induced mouse pulmonary fibrosis model and TGF-b1 induced human fetal lung fibroblasts (MRC5) in vitro. We used both SIRT6 siRNA and rapamycin to study the role of SIRT6 and mTOR signaling pathway in the normal human lung fibroblasts and the myofibroblasts from human IPF lungs.ResultsOur data show that high level of SIRT6 was detected in IPF samples, and SIRT6 was significantly upregulated by TGF-β1 in a time and concentration-dependent manner. SIRT6 expression and activation of mTORC1 signalling pathway were upregulated in fibrotic lung tissues and primary lung fibroblasts isolated from patients with IPF and bleomycin-challenged mice. Furthermore, rapamycin treatment inhibited mTORC1 pathway activity and SIRT6 protein expression. SIRT6 SiRNA failed to mediate the activity of mTORC1 pathway and autophagy induction. However, SIRT6 knockdown could promote TGF-b1 induced pro-fibrotic cytokines.ConclusionActivated mTORC1 signalling pathway regulated SIRT6 overexpression. Deficiency of SIRT6 mediated myofibroblasts differentiation through induced pro-fibrotic cytokines production in the present of TGF-β1. The study indicated that manipulations of SIRT6 expression may provide a new therapeutic strategy to prevent and reverse the progression of pulmonary fibrosis.

scholarly journals mTORC1 Signaling Pathway Mediated SIRT6 Overexpression in TGF-β1-Induced Pulmonary Fibrosis

2021 ◽  
Author(s):  
Ji Zhang ◽  
Yi Hu ◽  
Huiping Huang ◽  
Qun Liu ◽  
Yang Du ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Signaling Pathway ◽  
Critical Role ◽  
Mtor Signaling ◽  
Lung Fibroblasts ◽  
Dependent Manner ◽  
Mtor Signaling Pathway ◽  
Mtorc1 Signaling ◽  
Mtorc1 Pathway ◽  
Tgf Β1

Abstract Fibroblast-to-myofibroblast transdifferentiation and myofibroblast hyperproliferation play a major role in Idiopathic pulmonary fibrosis (IPF). It was also reported that mTOR signaling pathway and SIRT6 have a critical role in pulmonary fibrosis. However, the mechanisms whether mTOR signaling pathway and SIRT6 affect the myofibroblasts differentiation in IPF remain unclear. The results show that SIRT6 is significantly upregulated by TGF-β1 with a time and concentration-dependent manner in MRC5 line and primary lung fibroblasts isolated from IPF patients. SIRT6 protein is also increased in IPF fibrotic lung tissues and bleomycin-challenged mice lung tissues. Also, the activity of mTOR signaling is activated in MRC5 and primary lung fibroblasts. Furthermore, the inhibitor of mTOR, rapamycin treatment significantly suppress mTORC1 pathway activity and SIRT6 protein expression. SIRT6 siRNA failed to mediate the activity of mTORC1 pathway and autophagy induction. Finally, deficiency of SIRT6 could promote TGF-β1 induced pro-fibrotic cytokines. In summary, the study have suggested that SIRT6 is a downstream of mTORC1 signaling pathway in the pulmonary fibrosis caused by TGF-β1-induced. Deficiency of SIRT6 mediated myofibroblasts differentiation through induced pro-fibrotic cytokines production but not induced-autophagy. It was indicated that manipulations of SIRT6 expression may provide a new therapeutic strategy to reverse the progression of pulmonary fibrosis.

scholarly journals Transforming growth factor (TGF)-β1-induced miR-133a inhibits myofibroblast differentiation and pulmonary fibrosis

2019 ◽  
Vol 10 (9) ◽  
Author(s):  
Peng Wei ◽  
Yan Xie ◽  
Peter W. Abel ◽  
Yapei Huang ◽  
Qin Ma ◽  
...  
Keyword(s):  
Growth Factor ◽  
Pulmonary Fibrosis ◽  
Transforming Growth Factor ◽  
Tissue Growth ◽  
Negative Regulator ◽  
Lung Fibroblasts ◽  
Luciferase Reporter ◽  
Myofibroblast Differentiation ◽  
Dependent Manner ◽  
Tgf Β1

Abstract Transforming growth factor (TGF)-β1, a main profibrogenic cytokine in the progression of idiopathic pulmonary fibrosis (IPF), induces differentiation of pulmonary fibroblasts to myofibroblasts that produce high levels of collagen, leading to concomitantly loss of lung elasticity and function. Recent studies implicate the importance of microRNAs (miRNAs) in IPF but their regulation and individual pathological roles remain largely unknown. We used both RNA sequencing and quantitative RT-PCR strategies to systematically study TGF-β1-induced alternations of miRNAs in human lung fibroblasts (HFL). Our data show that miR-133a was significantly upregulated by TGF-β1 in a time- and concentration-dependent manner. Surprisingly, miR-133a inhibits TGF-β1-induced myofibroblast differentiation whereas miR-133a inhibitor enhances TGF-β1-induced myofibroblast differentiation. Interestingly, quantitative proteomics analysis indicates that miR-133a attenuates myofibroblast differentiation via targeting multiple components of TGF-β1 profibrogenic pathways. Western blot analysis confirmed that miR-133a down-regulates TGF-β1-induced expression of classic myofibroblast differentiation markers such as ɑ-smooth muscle actin (ɑ-SMA), connective tissue growth factor (CTGF) and collagens. miRNA Target Searcher analysis and luciferase reporter assays indicate that TGF-β receptor 1, CTGF and collagen type 1-alpha1 (Col1a1) are direct targets of miR-133a. More importantly, miR-133a gene transferred into lung tissues ameliorated bleomycin-induced pulmonary fibrosis in mice. Together, our study identified TGF-β1-induced miR-133a as an anti-fibrotic factor. It functions as a feed-back negative regulator of TGF-β1 profibrogenic pathways. Thus, manipulations of miR-133a expression may provide a new therapeutic strategy to halt and perhaps even partially reverse the progression of IPF.

scholarly journals Butyrate Prevents TGF-β1-Induced Alveolar Myofibroblast Differentiation and Modulates Energy Metabolism

Metabolites ◽  
2021 ◽  
Vol 11 (5) ◽  
pp. 258
Author(s):  
Hyo Yeong Lee ◽  
Somi Nam ◽  
Mi Jeong Kim ◽  
Su Jung Kim ◽  
Sung Hoon Back ◽  
...  
Keyword(s):  
Energy Metabolism ◽  
Pulmonary Fibrosis ◽  
Transforming Growth Factor ◽  
Mitochondrial Dynamics ◽  
Tca Cycle ◽  
Lung Fibroblasts ◽  
Myofibroblast Differentiation ◽  
Pulmonary Fibroblasts ◽  
Matrix Deposition ◽  
Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a serious lung disease characterized by excessive collagen matrix deposition and extracellular remodeling. Signaling pathways mediated by fibrotic cytokine transforming growth factor β1 (TGF-β1) make important contributions to pulmonary fibrosis, but it remains unclear how TGF-β1 alters metabolism and modulates the activation and differentiation of pulmonary fibroblasts. We found that TGF-β1 lowers NADH and NADH/NAD levels, possibly due to changes in the TCA cycle, resulting in reductions in the ATP level and oxidative phosphorylation in pulmonary fibroblasts. In addition, we showed that butyrate (C4), a short chain fatty acid (SCFA), exhibits potent antifibrotic activity by inhibiting expression of fibrosis markers. Butyrate treatment inhibited mitochondrial elongation in TGF-β1-treated lung fibroblasts and increased the mitochondrial membrane potential (MMP). Consistent with the mitochondrial observations, butyrate significantly increased ADP, ATP, NADH, and NADH/NAD levels in TGF-β1-treated pulmonary fibroblasts. Collectively, our findings indicate that TGF-β1 induces changes in mitochondrial dynamics and energy metabolism during myofibroblast differentiation, and that these changes can be modulated by butyrate, which enhances mitochondrial function.

scholarly journals Ginsenoside Rg3 Suppresses Proliferation and Induces Apoptosis in Human Osteosarcoma

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Yi Li ◽  
Jinying Lu ◽  
Furong Bai ◽  
Yanan Xiao ◽  
Yiran Guo ◽  
...  
Keyword(s):  
Western Blot ◽  
Signaling Pathway ◽  
Mtor Signaling ◽  
Assay Method ◽  
Osteosarcoma Cell ◽  
Dependent Manner ◽  
Ginsenoside Rg3 ◽  
Mtor Signaling Pathway ◽  
Human Osteosarcoma ◽  
Human Osteosarcoma Cells

Osteosarcoma is the most common primary malignancy of bone in children and the elderly. Recently, more and more researches have demonstrated that Ginsenoside Rg3 (Rg3) is involved in chemotherapy resistance in many cancer, making it a promising Chinese herbal monomer for oncotherapy. In this study, we investigated the efficacy of Rg3 in human osteosarcoma cell lines (MG-63, U-2OS, and SaOS-2). Cell proliferation was measured by CCK8 assay. The migration of cells was examined using the scratch assay method. Quantification of apoptosis was assessed further by flow cytometry. In addition, the expression of apoptosis-related genes (caspase9, caspase3, Bcl2, and Bax) were investigated using RT-PCR. We further investigated the protein level expression of Bcl 2, cleaved-caspase3, and PI3K/AKT/mTOR signaling pathway factors by Western blot assay. Our results revealed that Rg3 inhibited the proliferation and migration of human osteosarcoma cells and induced apoptosis in a concentration- and time-dependent manner. Western blot results showed that Rg3 reduced the protein expression of Bcl2 and PI3K/AKT/mTORbut increased the levels of cleaved-caspase3. Therefore, we hypothesized Rg3 inhibits the proliferation of osteosarcoma cell line and induces their apoptosis by affecting apoptosis-related genes (Bcl2, caspase3) as well as the PI3K/AKT/mTOR signaling pathway. To conclude, Rg3 is a new therapeutic agent against osteosarcoma.

scholarly journals Osthole Attenuates Bleomycin-Induced Pulmonary Fibrosis by Modulating NADPH Oxidase 4-Derived Oxidative Stress in Mice

2021 ◽  
Vol 2021 ◽  
pp. 1-12
Author(s):  
Lijun Fang ◽  
Wei Wang ◽  
Jiazheng Chen ◽  
Anju Zuo ◽  
Hongmei Gao ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Pulmonary Fibrosis ◽  
Lung Fibroblasts ◽  
Ros Generation ◽  
Dependent Manner ◽  
Active Constituent ◽  
Concentration Dependent Manner ◽  
Nadph Oxidase 4 ◽  
Tgf Β1

Idiopathic pulmonary fibrosis (IPF) is a chronic progressive lung disease characterized by the extensive accumulation of myofibroblasts and collagens. However, the exact mechanism that underlies this condition is unclear. Growing evidence suggests that NADPH oxidases (NOXs), especially NOX4-derived oxidative stress, play an important role in the development of lung fibrosis. Bleomycin (BLM) is a tumor chemotherapeutic agent, which has been widely employed to establish IPF animal models. Osthole (OST) is an active constituent of the fruit of Cnidium ninidium. Here, we used an in vivo mouse model and found that OST suppressed BLM-induced body weight loss, lung injury, pulmonary index increase, fibroblast differentiation, and pulmonary fibrosis. OST also significantly downregulated BLM-induced NOX4 expression and oxidative stress in the lungs. In vitro, OST could inhibit TGF-β1-induced Smad3 phosphorylation, differentiation, proliferation, collagen synthesis, NOX4 expression, and ROS generation in human lung fibroblasts in a concentration-dependent manner. Moreover, NOX4 overexpression could prevent the above effects of OST. We came to the conclusion that OST could significantly attenuate BLM-induced pulmonary fibrosis in mice, via the mechanism that involved downregulating TGF-β1/NOX4-mediated oxidative stress in lung fibroblasts.

scholarly journals Melatonin-induced Cell Rejuvenation from Long-term Ex-vivo passaging Functioned by its Restoration of Damaged Cell Autophagic Processes

2020 ◽  
Author(s):  
Yi-Zhou Tan ◽  
Xin-Yue Xu ◽  
Ji-Min Dai ◽  
Yuan Yin ◽  
Xiao-Tao He ◽  
...  
Keyword(s):  
Stem Cells ◽  
Signaling Pathway ◽  
Cellular Senescence ◽  
Ex Vivo ◽  
Mtor Signaling ◽  
Beclin 1 ◽  
Dependent Manner ◽  
Mtor Signaling Pathway ◽  
Associated Proteins

Abstract Background: Stem cells undergone long-term ex-vivo expansion are most likely functionally compromised (namely cellular senescence) in terms of their stem cell properties and therapeutic potentials. Due to the ability to attenuate cellular senescence, melatonin (MLT) has been proposed as an adjuvant across long-term cell expansion protocols, but the underlying mechanism remains largely unknown. Methods: Human periodontal ligament stem cells (PDLSCs) were isolated and cultured ex-vivo for 15 passages, and passage 2, 7 and 15 cells were used to interrogate the cellular senescence and alteration in cell autophagy during long-term expansion. The cellular senescence features were evidenced by senescence-associated β-galacotosidase (SA-β-gal) activity and the expression of senescence-related proteins including p53, p21, p16 and γ-H2AX. Electronic microscope was used to observe the autophagic vesicles. Adenovirus mRFP-GFP-LC3 was transfected to indicate the alteration of autophagic flux during long-term expansion, and the autophagy-associated proteins Atg7, Beclin-1, LC3-II and p62 were evaluated by Western blot. Results: It was found that long-term in-vitro passaging led to an accumulated SA-β-gal, elevated expressions of p53, p21, p16 and γ-H2AX, along with downregulated autophagy-associated proteins Atg7, Beclin-1 and LC3 as well as a mounting autophagy substrate p62. In accordance with expectation, supplemented with MLT not only ameliorated cells to a younger state but also restored the impaired autophagy level in senescent cells. Additionally, we demonstrated that autophagy inhibitor could block such MLT-induced cell rejuvenation. When the underlying signaling pathways involved was interrogated, we found that MLT receptor (MT) participated in mediating MLT-related autophagy restoration by regulating PI3K/AKT/mTOR signaling pathway.Conclusions: The present study suggests that MLT may rejuvenate long-term expansion-caused cellular senescence by restoring autophagy, more likely via PI3K/AKT/mTOR signaling pathway in an MT-dependent manner. This is the first report identifying the MT-dependent PI3K/AKT/mTOR signaling involved in MLT-induced autophagy alteration, pointing to a potential target for using autophagy-restoring agents such as MLT to develop optimized clinical-scale cell production protocols.

scholarly journals 5-Aminosalicylic acid Attenuates Paraquat-induced Lung Fibroblasts Activation and Pulmonary Fibrosis of Rats

2021 ◽  
Author(s):  
Hui Chen ◽  
Jinfeng Cui ◽  
Juan Wang ◽  
Yuan Wang ◽  
Fei Tong ◽  
...  
Keyword(s):  
Pulmonary Fibrosis ◽  
Signaling Pathway ◽  
Human Lung ◽  
Lung Fibroblasts ◽  
Control Group ◽  
Peroxisome Proliferator ◽  
Aminosalicylic Acid ◽  
Human Lung Fibroblasts ◽  
Group 5 ◽  
Tgf Β1

Abstract Pulmonary fibrosis is one of the most common complications of paraquat (PQ) poisoning, which becomes the focus of treatment. More and more studies have found that 5-Aminosalicylic acid (5-ASA) may be a prospective therapy against fibrotic diseases. In the present study, we observed whether 5-ASA could attenuate the pulmonary fibrosis in PQ-treated rats and human lung fibroblasts (WI38VA13) cells, and subsequently explored the possible underlying mechanisms. Wistar rats were divided into control group, 5-ASA group, PQ group and PQ + 5-ASA group. Rats were sacrificed on 3, 7, 14, and 28 days after PQ treatment. We observed pulmonary histopathological changes and fibrosis formation among different groups through hematoxylin and eosin (H&E) and Masson staining and TGF-β1, p-Smad3 and the peroxisome proliferator activated receptor γ (PPARγ) pulmonary content via immunohistochemical staining and Western blot. In addition, human lung fibroblasts WI38VA13 were also divided into control group, PQ group, 5-ASA group and PQ + 5-ASA group. And the role of TGF-β1 signaling pathway regulated factors (TGF-β1, p-Smad3 and PPARγ) were explored. Treatment with 5-ASA significantly inhibited the PQ-induced activation of TGF-β1 signaling pathway in human lung fibroblasts WI38VA13 cells. In conclusion, the results of this study suggested that 5-ASA has potential value in the treatment of PQ-induced pulmonary fibrosis via suppressing the activation of TGF-β1 signaling pathway.

scholarly journals Expression of TGF-β1/mTOR signaling pathway in pathological scar fibroblasts

2017 ◽  
Vol 15 (6) ◽  
pp. 3467-3472 ◽  
Author(s):  
Xiao-Xiang Zhai ◽  
Zhi-Ming Tang ◽  
Ji-Cun Ding ◽  
Xiao-Lan Lu
Keyword(s):  
Signaling Pathway ◽  
Mtor Signaling ◽  
Mtor Signaling Pathway ◽  
Pathological Scar ◽  
Tgf Β1

scholarly journals AMP-Activated Protein Kinase Alleviates Extracellular Matrix Accumulation in High Glucose-Induced Renal Fibroblasts through mTOR Signaling Pathway

2015 ◽  
Vol 35 (1) ◽  
pp. 191-200 ◽  
Author(s):  
Xia Luo ◽  
Lingyan Deng ◽  
Laxmi Pangeni Lamsal ◽  
Wenjuan Xu ◽  
Cheng Xiang ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Protein Kinase ◽  
Signaling Pathway ◽  
High Glucose ◽  
Mtor Signaling ◽  
Dependent Manner ◽  
Mtor Signaling Pathway ◽  
Matrix Synthesis ◽  
Matrix Accumulation ◽  
Amp Activated Protein Kinase

Background/Aims: Extracellular matrix accumulation contributes significantly to the pathogenesis of diabetic nephropathy. Although AMP-activated protein kinase (AMPK) has been found to inhibit extracellular matrix synthesis by experiments in vivo and vitro, its role in alleviating the deposition of extracellular matrix in renal interstitial fibroblasts has not been well defined. Methods: Currently, we conducted this study to investigate the effects of AMPK on high glucose-induced extracellular matrix synthesis and involved intracellular signaling pathway by using western blot in the kidney fibroblast cell line (NRK-49f). Results: Collagen IV protein levels were significantly increased by high glucose in a time-dependent manner. This was associated with a decrease in Thr72 phosphorylation of AMPK and an increase in phosphorylation of mTOR on Ser2448. High glucose-induced extracellular matrix accumulation and mTOR activation were significantly inhibited by the co-treatment of rAAV-AMPKα1312 (encoding constitutively active AMPKα1) whereas activated by r-AAV-AMPKα1D157A (encoding dominant negative AMPKα1). In cultured renal fibroblasts, overexpression of AMPKα1D157A upregulated mTOR signaling and matrix synthesis, which were ameliorated by co-treatment with the inhibitor of mTOR, rapamycin. Conclusion: Collectively, these findings indicate that AMPK exerts renoprotective effects by inhibiting the accumulation of extracellular matrix through mTOR signaling pathway.

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