Acute respiratory distress syndrome(ARDS)is one of the most fatal diseases worldwide.Pulmonary fibrosis occurs early in ARDS,and its severity plays a crucial role in ARDS mortality rate.Some studies suggested that fib...Acute respiratory distress syndrome(ARDS)is one of the most fatal diseases worldwide.Pulmonary fibrosis occurs early in ARDS,and its severity plays a crucial role in ARDS mortality rate.Some studies suggested that fibroproliferation is an essential mechanism in ARDS.Mitofusion2(Mfn2)overexpression plays a role in inhibiting cell proliferation.However,the role and potential mechanism of Mfn2 on the proliferation of fibroblasts is still unknown.In this study,we aimed at exploring the effect of Mfn2 on the human embryonic lung fibroblasts(HELF)and discussed its related mechanism.The HELF were treated with the Mfn2 overexpressing lentivirus(adv-Mfn2).The cell cycle was detected by flow cytometry.MTT,PCR and Western blotting were used to investigate the effect of Mfn2 on the proliferation of the HELF,collagen expression,the RAS-RAF-1-ERK1/2 pathway and the expression of cycle-related proteins(p21,p27,Rb,Raf-1,p-Raf-1,Erk1/2 and p-Erk1/2).The co-immunoprecipitation assay was used to explore the interaction between Mfn2 and Ras.The results showed that the overexpression of Mfn2 inhibited the proliferation of the HELF and induced the cell cycle arrest at the G0/G1 phase.Meanwhile,Mfn2 also inhibited the expression of collagen I,p-Erk and p-Raf-1.In addition,an interaction between Mfn2 and Ras existed in the HELF.This study suggests that the overexpression of Mfn2 can decrease the proliferation of HELF in ARDS,which was associated with the inhibition of the RAS-RAF-1-ERK1/2 pathway.The results may offer a potential therapeutic intervention for patients with ARDS.展开更多
Background:Intestinal barrier defect is an essential inflammatory bowel disease(IBD)pathogenesis.Mitochondrial dysfunction results in energy deficiency and oxidative stress,which contribute to the pathogenesis of IBD....Background:Intestinal barrier defect is an essential inflammatory bowel disease(IBD)pathogenesis.Mitochondrial dysfunction results in energy deficiency and oxidative stress,which contribute to the pathogenesis of IBD.β-arrestin1(ARRB1)is a negative regu-lator that promotes G protein-coupled receptors desensitization,endocytosis,and degradation.However,its role in maintaining the intestinal barrier remains unclear.Methods:Dextran sulfate sodium-induced colitis was performed in ARRB1 knockout and wild-type mice.Intestinal permeability and tight junction proteins were measured to evaluate the intestinal barrier.Mitochondria function and mitophagic flux in mice and cell lines were detected.Finally,the interaction between ARRB1 and mitofusin 2 was investigated by co-immunoprecipitation and dual luciferase assay.Results:We identified that ARRB1 protected the intestinal tight junction barrier against experimental colitis in vivo.ARRB1 deficiency was accompanied by abnormal mitochondrial morphology,lower adenosine triphosphate(ATP)production,and severe oxidative stress.In vitro,the knockdown of ARRB1 reduced ATP levels and mitochondrial membrane potential while increasing reactive oxygen species levels and oxidative stress.Upon ARRB1 ablation,mitophagy was inhibited,accompanied by decreased LC3BII,phosphatase and tension homologue-induced protein kinase1(PINK1),and parkin,but increased p62 expression.Mitophagy inhibition via PINK1 siRNA or mitochondrial division inhibitor 1 impaired ARRB1-mediated tight junction protection.The interaction of ARRB1 with E2F1 activated mitophagy by enhancing the transcription of mitofusin 2.Conclusions:Our results suggest that ARRB1 is critical to maintaining the intestinal tight junction barrier by promoting mitophagy.These results reveal a novel link between ARRB1 and the intestinal tight junction barrier,which provides theoretical support for coli-tis treatment.展开更多
基金This project was supported by Wuhan Medical Science Foundation of China(No.WX17B07,No.WX19A09,and No.WJ2019H324).
文摘Acute respiratory distress syndrome(ARDS)is one of the most fatal diseases worldwide.Pulmonary fibrosis occurs early in ARDS,and its severity plays a crucial role in ARDS mortality rate.Some studies suggested that fibroproliferation is an essential mechanism in ARDS.Mitofusion2(Mfn2)overexpression plays a role in inhibiting cell proliferation.However,the role and potential mechanism of Mfn2 on the proliferation of fibroblasts is still unknown.In this study,we aimed at exploring the effect of Mfn2 on the human embryonic lung fibroblasts(HELF)and discussed its related mechanism.The HELF were treated with the Mfn2 overexpressing lentivirus(adv-Mfn2).The cell cycle was detected by flow cytometry.MTT,PCR and Western blotting were used to investigate the effect of Mfn2 on the proliferation of the HELF,collagen expression,the RAS-RAF-1-ERK1/2 pathway and the expression of cycle-related proteins(p21,p27,Rb,Raf-1,p-Raf-1,Erk1/2 and p-Erk1/2).The co-immunoprecipitation assay was used to explore the interaction between Mfn2 and Ras.The results showed that the overexpression of Mfn2 inhibited the proliferation of the HELF and induced the cell cycle arrest at the G0/G1 phase.Meanwhile,Mfn2 also inhibited the expression of collagen I,p-Erk and p-Raf-1.In addition,an interaction between Mfn2 and Ras existed in the HELF.This study suggests that the overexpression of Mfn2 can decrease the proliferation of HELF in ARDS,which was associated with the inhibition of the RAS-RAF-1-ERK1/2 pathway.The results may offer a potential therapeutic intervention for patients with ARDS.
基金supported by grants from the National Natural Science Foundation of China[82070574,818004580]the Nature Science Foundation Team Project of Guangdong Province[2018B03031200].
文摘Background:Intestinal barrier defect is an essential inflammatory bowel disease(IBD)pathogenesis.Mitochondrial dysfunction results in energy deficiency and oxidative stress,which contribute to the pathogenesis of IBD.β-arrestin1(ARRB1)is a negative regu-lator that promotes G protein-coupled receptors desensitization,endocytosis,and degradation.However,its role in maintaining the intestinal barrier remains unclear.Methods:Dextran sulfate sodium-induced colitis was performed in ARRB1 knockout and wild-type mice.Intestinal permeability and tight junction proteins were measured to evaluate the intestinal barrier.Mitochondria function and mitophagic flux in mice and cell lines were detected.Finally,the interaction between ARRB1 and mitofusin 2 was investigated by co-immunoprecipitation and dual luciferase assay.Results:We identified that ARRB1 protected the intestinal tight junction barrier against experimental colitis in vivo.ARRB1 deficiency was accompanied by abnormal mitochondrial morphology,lower adenosine triphosphate(ATP)production,and severe oxidative stress.In vitro,the knockdown of ARRB1 reduced ATP levels and mitochondrial membrane potential while increasing reactive oxygen species levels and oxidative stress.Upon ARRB1 ablation,mitophagy was inhibited,accompanied by decreased LC3BII,phosphatase and tension homologue-induced protein kinase1(PINK1),and parkin,but increased p62 expression.Mitophagy inhibition via PINK1 siRNA or mitochondrial division inhibitor 1 impaired ARRB1-mediated tight junction protection.The interaction of ARRB1 with E2F1 activated mitophagy by enhancing the transcription of mitofusin 2.Conclusions:Our results suggest that ARRB1 is critical to maintaining the intestinal tight junction barrier by promoting mitophagy.These results reveal a novel link between ARRB1 and the intestinal tight junction barrier,which provides theoretical support for coli-tis treatment.
