Spinal and bulbar muscular atrophy is a neurodegenerative disease caused by extended CAG trinucleotide repeats in the androgen receptor gene,which encodes a ligand-dependent transcription facto r.The mutant androgen r...Spinal and bulbar muscular atrophy is a neurodegenerative disease caused by extended CAG trinucleotide repeats in the androgen receptor gene,which encodes a ligand-dependent transcription facto r.The mutant androgen receptor protein,characterized by polyglutamine expansion,is prone to misfolding and forms aggregates in both the nucleus and cytoplasm in the brain in spinal and bulbar muscular atrophy patients.These aggregates alter protein-protein interactions and compromise transcriptional activity.In this study,we reported that in both cultured N2a cells and mouse brain,mutant androgen receptor with polyglutamine expansion causes reduced expression of mesencephalic astrocyte-de rived neurotrophic factor.Overexpressio n of mesencephalic astrocyte-derived neurotrophic factor amelio rated the neurotoxicity of mutant androgen receptor through the inhibition of mutant androgen receptor aggregation.Conversely.knocking down endogenous mesencephalic astrocyte-derived neurotrophic factor in the mouse brain exacerbated neuronal damage and mutant androgen receptor aggregation.Our findings suggest that inhibition of mesencephalic astrocyte-derived neurotrophic factor expression by mutant androgen receptor is a potential mechanism underlying neurodegeneration in spinal and bulbar muscular atrophy.展开更多
BACKGROUND:To determine the protective role of mesencephalic astrocyte-derived neurotrophic factor(MANF) in regulating sepsis-associated acute kidney injury(S-AKI).METHODS:A total of 96 mice were randomly divided into...BACKGROUND:To determine the protective role of mesencephalic astrocyte-derived neurotrophic factor(MANF) in regulating sepsis-associated acute kidney injury(S-AKI).METHODS:A total of 96 mice were randomly divided into the control group,control+MANF group,S-AKI group,and S-AKI+MANF group.The S-AKI model was established by injecting lipopolysaccharide(LPS) at 10 mg/kg intraperitoneally.MANF(200 μg/kg) was administered to the control+MANF and S-AKI+MANF groups.An equal dose of normal saline was administered daily intraperitoneally in the control and S-AKI groups.Serum and kidney tissue samples were obtained for biochemical analysis.Western blotting was used to detect the protein expression of MANF in the kidney,and enzyme-linked immunosorbent assay(ELISA) was used to determine expression of MANF in the serum,pro-inflammatory cytokines(tumor necrosis factor-α [TNF-α] and interleukin-6 [IL-6]).Serum creatinine(SCr),and blood urea nitrogen(BUN)were examined using an automatic biochemical analyzer.In addition,the kidney tissue was observed for pathological changes by hematoxylin-eosin staining.The comparison between two groups was performed by unpaired Student’s t-test,and statistics among multiple groups were carried out using Tukey’s post hoc test following one-way analysis of variance(ANOVA).A P-value <0.05 was considered statistically significant.RESULTS:At the early stage of S-AKI,MANF in the kidney tissue was up-regulated,but with the development of the disease,it was down-regulated.Renal function was worsened in the S-AKI group,and TNF-α and IL-6 were elevated.The administration of MANF significantly alleviated the elevated levels of SCr and BUN and inhibited the expression of TNF-α and IL-6 in the kidney.The pathological changes were more extensive in the S-AKI group than in the S-AKI+MANF group.CONCLUSION:MANF treatment may significantly alleviate renal injury,reduce the inflammatory response,and alleviate or reverse kidney tissue damage.MANF may have a protective effect on S-AKI,suggesting a potential treatment for S-AKI.展开更多
BACKGROUND Nonalcoholic fatty liver disease (NAFLD) is a global metabolism-associated liver disease.Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a newly discovered secreted protein that is involved in...BACKGROUND Nonalcoholic fatty liver disease (NAFLD) is a global metabolism-associated liver disease.Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a newly discovered secreted protein that is involved in metabolic homeostasis.However,much remains to be discovered about its function in hepatic lipid metabolism;thus,we assessed whether MANF could regulate hepatic metabolism.AIM To establish in vivo and in vitro NAFLD models to explore the role of MANF in hepatic lipid metabolism.METHODS HepG2 cells treated with free fatty acids (FFAs) and ob/ob mice were used as NAFLD models.Liver tissues collected from wild type and ob/ob mice were used to detect MANF expression.Cells were treated with FFAs for different durations.Moreover,we used lentiviral constructs to establish overexpression and knockdown cell models in order to interfere with MANF expression levels and observe whether MANF influences hepatic steatosis.Western blot analysis and quantitative real-time PCR were used to detect protein and gene expression,and oil red O staining was used to visualize intracellular lipid droplets.RESULTS Hepatic MANF protein and mRNA expression in wild type mice were 10-fold and 2-fold higher,respectively,than those in ob/ob mice.The MANF protein was temporarily increased by 1.3-fold after stimulation with FFAs for 24 h and gradually decreased to 0.66-fold that of the control at the 72 h time point in HepG2 cells.MANF deficiency upregulated the expression of genes involved infatty acid synthesis,cholesterol synthesis,and fatty acid uptake and aggravated HepG2 cell steatosis,while MANF overexpression inhibited fatty acid synthesis and uptake and cholesterol synthesis,and rescued HepG2 cells from FFAsinduced steatosis.Furthermore,a significant decrease in triglyceride levels was observed in the MANF overexpression group compared with the control group(0.4288±0.0081 mmol/g vs 0.3746±0.0121 mmol/g,P <0.05) upon FF As treatment.There was also a 17%decrease in intracellular total cholesterol levels between the MANF overexpression group and the control group (0.1301±0.0059mmol/g vs 0.1088±0.0009 mmol/g,P <0.05) upon FF As treatment.Moreover,MANF suppressed lipid deposition in HepG2 cells.CONCLUSION Our findings indicate that MANF improves the phenotype of liver cell steatosis and may be a potential therapeutic target in hepatic steatosis processes.展开更多
Neurodegenerative diseases are a leading cause of disability worldwide,and despite significant resources put toward the discovery of potential therapeutic targets,there are currently no effective treatments.The rise o...Neurodegenerative diseases are a leading cause of disability worldwide,and despite significant resources put toward the discovery of potential therapeutic targets,there are currently no effective treatments.The rise of methods to derive and propagate stem cells in vitro offered展开更多
BACKGROUND Bone marrow mesenchymal stem cells(BMSCs)are capable of shifting the microglia/macrophages phenotype from M1 to M2,contributing to BMSCsinduced brain repair.However,the regulatory mechanism of BMSCs on micr...BACKGROUND Bone marrow mesenchymal stem cells(BMSCs)are capable of shifting the microglia/macrophages phenotype from M1 to M2,contributing to BMSCsinduced brain repair.However,the regulatory mechanism of BMSCs on microglia/macrophages after ischemic stroke is unclear.Recent evidence suggests that mesencephalic astrocyte-derived neurotrophic factor(MANF)and plateletderived growth factor-AA(PDGF-AA)/MANF signaling regulate M1/M2 macrophage polarization.AIM To investigate whether and how MANF or PDGF-AA/MANF signaling influences BMSCs-mediated M2 polarization.METHODS We identified the secretion of MANF by BMSCs and developed transgenic BMSCs using a targeting small interfering RNA for knockdown of MANF expression.Using a rat middle cerebral artery occlusion(MCAO)model transplanted by BMSCs and BMSCs-microglia Transwell coculture system,the effect of BMSCsinduced downregulation of MANF expression on the phenotype of microglia/macrophages was tested by Western blot,quantitative reverse transcription-polymerase chain reaction,and immunofluorescence.Additionally,microglia were transfected with mimics of miR-30a*,which inuenced expression of X-box binding protein(XBP)1,a key transcription factor that synergized with activating transcription factor 6(ATF6)to govern MANF expression.We examined the levels of miR-30a*,ATF6,XBP1,and MANF after PDGF-AA treatment in the activated microglia.RESULTS Inhibition of MANF attenuated BMSCs-induced functional recovery and decreased M2 marker production,but increased M1 marker expression in vivo or in vitro.Furthermore,PDGF-AA treatment decreased miR-30a*expression,had no influence on the levels of ATF6,but enhanced expression of both XBP1 and MANF.CONCLUSION BMSCs-mediated MANF paracrine signaling,in particular the PDGF-AA/miR-30a*/XBP1/MANF pathway,synergistically mediates BMSCs-induced M2 polarization.展开更多
In advanced hepatocellular carcinoma(HCC)tissues,M2-like tumor-associated macrophages(TAMs)are in the majority and promotes HCC progression.Contrary to the pro-tumor effect of M2-like TAMs,M1-like TAMs account for a s...In advanced hepatocellular carcinoma(HCC)tissues,M2-like tumor-associated macrophages(TAMs)are in the majority and promotes HCC progression.Contrary to the pro-tumor effect of M2-like TAMs,M1-like TAMs account for a small proportion and have anti-tumor effects.Since TAMs can switch from one type to another,reprogramming TAMs may be an important treatment for HCC therapy.However,the mechanisms of phenotypic switch and reprogramming TAMs are still obscure.In this study,we analyzed differential genes in normal macrophages and TAMs,and found that loss of MANF in TAMs accompanied by high levels of downstream genes negatively regulated by MANF.MANF reprogrammed TAMs into M1 phenotype.Meanwhile,loss of MANF promoted HCC progression in HCC patients and mice HCC model,especially tumor neovascularization.Additionally,macrophages with MANF supplement suppressed HCC progression in mice,suggesting MANF supplement in macrophage was an effective treatment for HCC.Mechanistically,MANF enhanced the HSF1-HSP70-1 interaction,restricted HSF1 in the cytoplasm of macrophages,and decreased both mRNA and protein levels of HSP70-1,which in turn led to reprogramming TAMs,and suppressing neovascularization of HCC.Our study contributes to the exploration the mechanism of TAMs reprogramming,which may provide insights for future therapeutic exploitation of HCC neovascularization.展开更多
The mesencephalic astrocyte-derived neurotrophic factor(MANF)has been recently identified as a neurotrophic factor,but its role in hepatic fibrosis is unknown.Here,we found that MANF was upregulated in the fibrotic li...The mesencephalic astrocyte-derived neurotrophic factor(MANF)has been recently identified as a neurotrophic factor,but its role in hepatic fibrosis is unknown.Here,we found that MANF was upregulated in the fibrotic liver tissues of the patients with chronic liver diseases and of mice treated with CCl4.MANF deficiency in either hepatocytes or hepatic mono-macrophages,particularly in hepatic mono-macrophages,clearly exacerbated hepatic fibrosis.Myeloid-specific MANF knockout increased the population of hepatic Ly6C^(high)macrophages and promoted HSCs activation.Furthermore,MANF-sufficient macrophages(from WT mice)transfusion ameliorated CCl4-induced hepatic fibrosis in myeloid cells-specific MANF knockout(MKO)mice.Mechanistically,MANF interacted with S100A8 to competitively block S100A8/A9 heterodimer formation and inhibited S100A8/A9-mediated TLR4-NF-κB signal activation.Pharmacologically,systemic administration of recombinant human MANF significantly alleviated CCl_(4)-induced hepatic fibrosis in both WT and hepatocytes-specific MANF knockout(HKO)mice.This study reveals a mechanism by which MANF targets S100A8/A9-TLR4 as a“brake”on the upstream of NF-κB pathway,which exerts an impact on macrophage differentiation and shed light on hepatic fibrosis treatment.展开更多
基金supported by the National Key R&D Program of China,No.2021YFA0805200(to SY)the National Natural Science Foundation of China,No.31970954(to SY)two grants from the Department of Science and Technology of Guangdong Province,Nos.2021ZT09Y007,2020B121201006(both to XJL)。
文摘Spinal and bulbar muscular atrophy is a neurodegenerative disease caused by extended CAG trinucleotide repeats in the androgen receptor gene,which encodes a ligand-dependent transcription facto r.The mutant androgen receptor protein,characterized by polyglutamine expansion,is prone to misfolding and forms aggregates in both the nucleus and cytoplasm in the brain in spinal and bulbar muscular atrophy patients.These aggregates alter protein-protein interactions and compromise transcriptional activity.In this study,we reported that in both cultured N2a cells and mouse brain,mutant androgen receptor with polyglutamine expansion causes reduced expression of mesencephalic astrocyte-de rived neurotrophic factor.Overexpressio n of mesencephalic astrocyte-derived neurotrophic factor amelio rated the neurotoxicity of mutant androgen receptor through the inhibition of mutant androgen receptor aggregation.Conversely.knocking down endogenous mesencephalic astrocyte-derived neurotrophic factor in the mouse brain exacerbated neuronal damage and mutant androgen receptor aggregation.Our findings suggest that inhibition of mesencephalic astrocyte-derived neurotrophic factor expression by mutant androgen receptor is a potential mechanism underlying neurodegeneration in spinal and bulbar muscular atrophy.
基金supported by the Health Commission Clinical Characteristic Discipline Construction Program of Pudong New Area,Shanghai (PW Yts2021-17)Youth Science and Technology Project Health and Family Planning Commission of Pudong New Area,Shanghai (PWRq2020-35)。
文摘BACKGROUND:To determine the protective role of mesencephalic astrocyte-derived neurotrophic factor(MANF) in regulating sepsis-associated acute kidney injury(S-AKI).METHODS:A total of 96 mice were randomly divided into the control group,control+MANF group,S-AKI group,and S-AKI+MANF group.The S-AKI model was established by injecting lipopolysaccharide(LPS) at 10 mg/kg intraperitoneally.MANF(200 μg/kg) was administered to the control+MANF and S-AKI+MANF groups.An equal dose of normal saline was administered daily intraperitoneally in the control and S-AKI groups.Serum and kidney tissue samples were obtained for biochemical analysis.Western blotting was used to detect the protein expression of MANF in the kidney,and enzyme-linked immunosorbent assay(ELISA) was used to determine expression of MANF in the serum,pro-inflammatory cytokines(tumor necrosis factor-α [TNF-α] and interleukin-6 [IL-6]).Serum creatinine(SCr),and blood urea nitrogen(BUN)were examined using an automatic biochemical analyzer.In addition,the kidney tissue was observed for pathological changes by hematoxylin-eosin staining.The comparison between two groups was performed by unpaired Student’s t-test,and statistics among multiple groups were carried out using Tukey’s post hoc test following one-way analysis of variance(ANOVA).A P-value <0.05 was considered statistically significant.RESULTS:At the early stage of S-AKI,MANF in the kidney tissue was up-regulated,but with the development of the disease,it was down-regulated.Renal function was worsened in the S-AKI group,and TNF-α and IL-6 were elevated.The administration of MANF significantly alleviated the elevated levels of SCr and BUN and inhibited the expression of TNF-α and IL-6 in the kidney.The pathological changes were more extensive in the S-AKI group than in the S-AKI+MANF group.CONCLUSION:MANF treatment may significantly alleviate renal injury,reduce the inflammatory response,and alleviate or reverse kidney tissue damage.MANF may have a protective effect on S-AKI,suggesting a potential treatment for S-AKI.
基金Supported by National Natural Science Foundation of China,No.81300702 and No.81501199Natural Science Foundation Project of Chongqing CSTC,No.cstc2018jcyj AX0210 and No.cstc2017jcyj AX0016the Kuanren Talents Program of the Second Affiliated Hospital of Chongqing Medical University
文摘BACKGROUND Nonalcoholic fatty liver disease (NAFLD) is a global metabolism-associated liver disease.Mesencephalic astrocyte-derived neurotrophic factor (MANF) is a newly discovered secreted protein that is involved in metabolic homeostasis.However,much remains to be discovered about its function in hepatic lipid metabolism;thus,we assessed whether MANF could regulate hepatic metabolism.AIM To establish in vivo and in vitro NAFLD models to explore the role of MANF in hepatic lipid metabolism.METHODS HepG2 cells treated with free fatty acids (FFAs) and ob/ob mice were used as NAFLD models.Liver tissues collected from wild type and ob/ob mice were used to detect MANF expression.Cells were treated with FFAs for different durations.Moreover,we used lentiviral constructs to establish overexpression and knockdown cell models in order to interfere with MANF expression levels and observe whether MANF influences hepatic steatosis.Western blot analysis and quantitative real-time PCR were used to detect protein and gene expression,and oil red O staining was used to visualize intracellular lipid droplets.RESULTS Hepatic MANF protein and mRNA expression in wild type mice were 10-fold and 2-fold higher,respectively,than those in ob/ob mice.The MANF protein was temporarily increased by 1.3-fold after stimulation with FFAs for 24 h and gradually decreased to 0.66-fold that of the control at the 72 h time point in HepG2 cells.MANF deficiency upregulated the expression of genes involved infatty acid synthesis,cholesterol synthesis,and fatty acid uptake and aggravated HepG2 cell steatosis,while MANF overexpression inhibited fatty acid synthesis and uptake and cholesterol synthesis,and rescued HepG2 cells from FFAsinduced steatosis.Furthermore,a significant decrease in triglyceride levels was observed in the MANF overexpression group compared with the control group(0.4288±0.0081 mmol/g vs 0.3746±0.0121 mmol/g,P <0.05) upon FF As treatment.There was also a 17%decrease in intracellular total cholesterol levels between the MANF overexpression group and the control group (0.1301±0.0059mmol/g vs 0.1088±0.0009 mmol/g,P <0.05) upon FF As treatment.Moreover,MANF suppressed lipid deposition in HepG2 cells.CONCLUSION Our findings indicate that MANF improves the phenotype of liver cell steatosis and may be a potential therapeutic target in hepatic steatosis processes.
基金supported by the NIH (NEI grant RO1 EY 018132,Kirschstein-NRSA 4T32HD007505-20)
文摘Neurodegenerative diseases are a leading cause of disability worldwide,and despite significant resources put toward the discovery of potential therapeutic targets,there are currently no effective treatments.The rise of methods to derive and propagate stem cells in vitro offered
文摘BACKGROUND Bone marrow mesenchymal stem cells(BMSCs)are capable of shifting the microglia/macrophages phenotype from M1 to M2,contributing to BMSCsinduced brain repair.However,the regulatory mechanism of BMSCs on microglia/macrophages after ischemic stroke is unclear.Recent evidence suggests that mesencephalic astrocyte-derived neurotrophic factor(MANF)and plateletderived growth factor-AA(PDGF-AA)/MANF signaling regulate M1/M2 macrophage polarization.AIM To investigate whether and how MANF or PDGF-AA/MANF signaling influences BMSCs-mediated M2 polarization.METHODS We identified the secretion of MANF by BMSCs and developed transgenic BMSCs using a targeting small interfering RNA for knockdown of MANF expression.Using a rat middle cerebral artery occlusion(MCAO)model transplanted by BMSCs and BMSCs-microglia Transwell coculture system,the effect of BMSCsinduced downregulation of MANF expression on the phenotype of microglia/macrophages was tested by Western blot,quantitative reverse transcription-polymerase chain reaction,and immunofluorescence.Additionally,microglia were transfected with mimics of miR-30a*,which inuenced expression of X-box binding protein(XBP)1,a key transcription factor that synergized with activating transcription factor 6(ATF6)to govern MANF expression.We examined the levels of miR-30a*,ATF6,XBP1,and MANF after PDGF-AA treatment in the activated microglia.RESULTS Inhibition of MANF attenuated BMSCs-induced functional recovery and decreased M2 marker production,but increased M1 marker expression in vivo or in vitro.Furthermore,PDGF-AA treatment decreased miR-30a*expression,had no influence on the levels of ATF6,but enhanced expression of both XBP1 and MANF.CONCLUSION BMSCs-mediated MANF paracrine signaling,in particular the PDGF-AA/miR-30a*/XBP1/MANF pathway,synergistically mediates BMSCs-induced M2 polarization.
基金funded by support programs for Jun Liu,including the National Natural Science Foundation of China(82073862)Excellent Youth Talent Program of Anhui Province Natural Science Foundation(2108085Y27,China)funded by Anhui Province Natural Science Foundation(2208085MH284,China)for Xiangpeng Hu,and funded by the National Natural Science Foundation of China(U21A20345)for Yuxian Shen。
文摘In advanced hepatocellular carcinoma(HCC)tissues,M2-like tumor-associated macrophages(TAMs)are in the majority and promotes HCC progression.Contrary to the pro-tumor effect of M2-like TAMs,M1-like TAMs account for a small proportion and have anti-tumor effects.Since TAMs can switch from one type to another,reprogramming TAMs may be an important treatment for HCC therapy.However,the mechanisms of phenotypic switch and reprogramming TAMs are still obscure.In this study,we analyzed differential genes in normal macrophages and TAMs,and found that loss of MANF in TAMs accompanied by high levels of downstream genes negatively regulated by MANF.MANF reprogrammed TAMs into M1 phenotype.Meanwhile,loss of MANF promoted HCC progression in HCC patients and mice HCC model,especially tumor neovascularization.Additionally,macrophages with MANF supplement suppressed HCC progression in mice,suggesting MANF supplement in macrophage was an effective treatment for HCC.Mechanistically,MANF enhanced the HSF1-HSP70-1 interaction,restricted HSF1 in the cytoplasm of macrophages,and decreased both mRNA and protein levels of HSP70-1,which in turn led to reprogramming TAMs,and suppressing neovascularization of HCC.Our study contributes to the exploration the mechanism of TAMs reprogramming,which may provide insights for future therapeutic exploitation of HCC neovascularization.
基金supported by the National Natural Science Foundation of China(81973336)the Joint Fund of the National Natural Science Foundation of China(U21A20345)。
文摘The mesencephalic astrocyte-derived neurotrophic factor(MANF)has been recently identified as a neurotrophic factor,but its role in hepatic fibrosis is unknown.Here,we found that MANF was upregulated in the fibrotic liver tissues of the patients with chronic liver diseases and of mice treated with CCl4.MANF deficiency in either hepatocytes or hepatic mono-macrophages,particularly in hepatic mono-macrophages,clearly exacerbated hepatic fibrosis.Myeloid-specific MANF knockout increased the population of hepatic Ly6C^(high)macrophages and promoted HSCs activation.Furthermore,MANF-sufficient macrophages(from WT mice)transfusion ameliorated CCl4-induced hepatic fibrosis in myeloid cells-specific MANF knockout(MKO)mice.Mechanistically,MANF interacted with S100A8 to competitively block S100A8/A9 heterodimer formation and inhibited S100A8/A9-mediated TLR4-NF-κB signal activation.Pharmacologically,systemic administration of recombinant human MANF significantly alleviated CCl_(4)-induced hepatic fibrosis in both WT and hepatocytes-specific MANF knockout(HKO)mice.This study reveals a mechanism by which MANF targets S100A8/A9-TLR4 as a“brake”on the upstream of NF-κB pathway,which exerts an impact on macrophage differentiation and shed light on hepatic fibrosis treatment.
