BACKGROUND Human-derived mesenchymal stromal cells have been shown to improve cognitive function following experimental stroke.The activity of exosomes has been verified to be comparable to the therapeutic effects of ...BACKGROUND Human-derived mesenchymal stromal cells have been shown to improve cognitive function following experimental stroke.The activity of exosomes has been verified to be comparable to the therapeutic effects of mesenchymal stromal cells.However,the effects of exosomes derived from human umbilical cord mesenchymal stem cells(HUC-MSCs)(ExoCtrl)on post-stroke cognitive impairment(PSCI)have rarely been reported.Moreover,whether exosomes derived from C-C chemokine receptor type 2(CCR2)-overexpressing HUC-MSCs(ExoCCR2)can enhance the therapeutic effects on PSCI and the possible underlying mechanisms have not been studied.AIM To investigate the effects of ExoCtrl on PSCI and whether ExoCCR2 can enhance therapeutic effects on PSCI.METHODS Transmission electron microscopy,qNano®particles analyzer,and Western blotting were employed to determine the morphology and CCR2 expression of ExoCtrl or ExoCCR2.ELISA was used to study the binding capacity of exosomes to CC chemokine ligand 2(CCL2)in vivo.After the intravenous injection of ExoCtrl or ExoCCR2 into experimental rats,the effect of ExoCtrl and ExoCCR2 on PSCI was assessed by Morris water maze.Remyelination and oligodendrogenesis were analyzed by Western blotting and immunofluorescence microscopy.QRT-PCR and immunofluorescence microscopy were conducted to compare the microglia/macrophage polarization.The infiltration and activation of hematogenous macrophages were analyzed by Western blotting and transwell migration analysis.RESULTS CCR2-overexpressing HUC-MSCs loaded the CCR2 receptor into their exosomes.The morphology and diameter distribution between ExoCtrl and ExoCCR2 showed no significant difference.ExoCCR2 bound significantly to CCL2 but ExoCtrl showed little CCL2 binding.Although both ExoCCR2 and ExoCtrl showed beneficial effects on PSCI,oligodendrogenesis,remyelination,and microglia/macrophage polarization,ExoCCR2 exhibited a significantly superior beneficial effect.We also found that ExoCCR2 could suppress the CCL2-induced macrophage migration and activation in vivo and in vitro,compared with ExoCtrl treated group.CONCLUSION CCR2 over-expression enhanced the therapeutic effects of exosomes on the experimental PSCI by promoting M2 microglia/macrophage polarization,enhancing oligodendrogenesis and remyelination.These therapeutic effects are likely through suppressing the CCL2-induced hematogenous macrophage migration and activation.Key words:Cognitive impairment;Stroke;Exosomes;C-C chemokine receptor type 2;Microglia/macrophage polarization;Remyelination.展开更多
BACKGROUND Massive hepatocyte death is the core event in acute liver failure(ALF).Gasdermin D(GSDMD)-mediated pyroptosis is a type of highly inflammatory cell death.However,the role of hepatocyte pyroptosis and its me...BACKGROUND Massive hepatocyte death is the core event in acute liver failure(ALF).Gasdermin D(GSDMD)-mediated pyroptosis is a type of highly inflammatory cell death.However,the role of hepatocyte pyroptosis and its mechanisms of expanding inflammatory responses in ALF are unclear.AIM To investigate the role and mechanisms of GSDMD-mediated hepatocyte pyroptosis through in vitro and in vivo experiments.METHODS The expression of pyroptosis pathway-associated proteins in liver tissues from ALF patients and a hepatocyte injury model was examined by Western blot.GSDMD short hairpin RNA(shRNA)was used to investigate the effects of downregulation of GSDMD on monocyte chemotactic protein 1(MCP1)and its receptor CC chemokine receptor-2(CCR2)in vitro.For in vivo experiments,we used GSDMD knockout mice to investigate the role and mechanism of GSDMD in a D-galactose/lipopolysaccharide(D-Galn/LPS)-induced ALF mouse model.RESULTS The levels of pyroptosis pathway-associated proteins in liver tissue from ALF patients and a hepatocyte injury model increased significantly.The level of GSDMD-N protein increased most obviously(P<0.001).In vitro,downregulation of GSDMD by shRNA decreased the cell inhibition rate and the levels of MCP1/CCR2 proteins(P<0.01).In vivo,GSDMD knockout dramatically eliminated inflammatory damage in the liver and improved the survival of DGaln/LPS-induced ALF mice(P<0.001).Unlike the mechanism of immune cell pyroptosis that involves releasing interleukin(IL)-1βand IL-18,GSDMDmediated hepatocyte pyroptosis recruited macrophages via MCP1/CCR2 to aggravate hepatocyte death.However,this pathological process was inhibited after knocking down GSDMD.CONCLUSION GSDMD-mediated hepatocyte pyroptosis plays an important role in the pathogenesis of ALF,recruiting macrophages to release inflammatory mediators by upregulating MCP1/CCR2 and leading to expansion of the inflammatory responses.GSDMD knockout can reduce hepatocyte death and inflammatory responses,thus alleviating ALF.展开更多
Atypical chemokine receptors have recently emerged as important molecular players in health and diseases; they affect chemokine availability and function and impact a multitude of pathophysiological events, including ...Atypical chemokine receptors have recently emerged as important molecular players in health and diseases; they affect chemokine availability and function and impact a multitude of pathophysiological events, including the tumorigenesis process. This family of atypical receptors comprises five members: ACKR1/DARC, ACKR2/D6,ACKR3/CXCR7, ACKR4/CCRL1, and ACKR5/CCRL2. This work evaluated the differential expression of these receptors in prostate cancer using quantitative PCR. Further evaluation of CCRL2 at the protein level confirmed its overexpression in a metastatic cell line and in malignant prostatic tissues from patients. CCRL2, a presumed member of the atypical chemokine receptor family, plays a key role in lung dendritic cell trafficking to peripheral lymph nodes.Recent studies have reported the expression of CCRL2 in different human cancer cell lines and tissues. However, its function and expression in prostate cancer has not been previously addressed.展开更多
基金the National Natural Science Foundation of China,No.81871847 and No.81672261
文摘BACKGROUND Human-derived mesenchymal stromal cells have been shown to improve cognitive function following experimental stroke.The activity of exosomes has been verified to be comparable to the therapeutic effects of mesenchymal stromal cells.However,the effects of exosomes derived from human umbilical cord mesenchymal stem cells(HUC-MSCs)(ExoCtrl)on post-stroke cognitive impairment(PSCI)have rarely been reported.Moreover,whether exosomes derived from C-C chemokine receptor type 2(CCR2)-overexpressing HUC-MSCs(ExoCCR2)can enhance the therapeutic effects on PSCI and the possible underlying mechanisms have not been studied.AIM To investigate the effects of ExoCtrl on PSCI and whether ExoCCR2 can enhance therapeutic effects on PSCI.METHODS Transmission electron microscopy,qNano®particles analyzer,and Western blotting were employed to determine the morphology and CCR2 expression of ExoCtrl or ExoCCR2.ELISA was used to study the binding capacity of exosomes to CC chemokine ligand 2(CCL2)in vivo.After the intravenous injection of ExoCtrl or ExoCCR2 into experimental rats,the effect of ExoCtrl and ExoCCR2 on PSCI was assessed by Morris water maze.Remyelination and oligodendrogenesis were analyzed by Western blotting and immunofluorescence microscopy.QRT-PCR and immunofluorescence microscopy were conducted to compare the microglia/macrophage polarization.The infiltration and activation of hematogenous macrophages were analyzed by Western blotting and transwell migration analysis.RESULTS CCR2-overexpressing HUC-MSCs loaded the CCR2 receptor into their exosomes.The morphology and diameter distribution between ExoCtrl and ExoCCR2 showed no significant difference.ExoCCR2 bound significantly to CCL2 but ExoCtrl showed little CCL2 binding.Although both ExoCCR2 and ExoCtrl showed beneficial effects on PSCI,oligodendrogenesis,remyelination,and microglia/macrophage polarization,ExoCCR2 exhibited a significantly superior beneficial effect.We also found that ExoCCR2 could suppress the CCL2-induced macrophage migration and activation in vivo and in vitro,compared with ExoCtrl treated group.CONCLUSION CCR2 over-expression enhanced the therapeutic effects of exosomes on the experimental PSCI by promoting M2 microglia/macrophage polarization,enhancing oligodendrogenesis and remyelination.These therapeutic effects are likely through suppressing the CCL2-induced hematogenous macrophage migration and activation.Key words:Cognitive impairment;Stroke;Exosomes;C-C chemokine receptor type 2;Microglia/macrophage polarization;Remyelination.
基金Supported by the National Natural Science Foundation of China,No.81570543 and No.81560104
文摘BACKGROUND Massive hepatocyte death is the core event in acute liver failure(ALF).Gasdermin D(GSDMD)-mediated pyroptosis is a type of highly inflammatory cell death.However,the role of hepatocyte pyroptosis and its mechanisms of expanding inflammatory responses in ALF are unclear.AIM To investigate the role and mechanisms of GSDMD-mediated hepatocyte pyroptosis through in vitro and in vivo experiments.METHODS The expression of pyroptosis pathway-associated proteins in liver tissues from ALF patients and a hepatocyte injury model was examined by Western blot.GSDMD short hairpin RNA(shRNA)was used to investigate the effects of downregulation of GSDMD on monocyte chemotactic protein 1(MCP1)and its receptor CC chemokine receptor-2(CCR2)in vitro.For in vivo experiments,we used GSDMD knockout mice to investigate the role and mechanism of GSDMD in a D-galactose/lipopolysaccharide(D-Galn/LPS)-induced ALF mouse model.RESULTS The levels of pyroptosis pathway-associated proteins in liver tissue from ALF patients and a hepatocyte injury model increased significantly.The level of GSDMD-N protein increased most obviously(P<0.001).In vitro,downregulation of GSDMD by shRNA decreased the cell inhibition rate and the levels of MCP1/CCR2 proteins(P<0.01).In vivo,GSDMD knockout dramatically eliminated inflammatory damage in the liver and improved the survival of DGaln/LPS-induced ALF mice(P<0.001).Unlike the mechanism of immune cell pyroptosis that involves releasing interleukin(IL)-1βand IL-18,GSDMDmediated hepatocyte pyroptosis recruited macrophages via MCP1/CCR2 to aggravate hepatocyte death.However,this pathological process was inhibited after knocking down GSDMD.CONCLUSION GSDMD-mediated hepatocyte pyroptosis plays an important role in the pathogenesis of ALF,recruiting macrophages to release inflammatory mediators by upregulating MCP1/CCR2 and leading to expansion of the inflammatory responses.GSDMD knockout can reduce hepatocyte death and inflammatory responses,thus alleviating ALF.
文摘Atypical chemokine receptors have recently emerged as important molecular players in health and diseases; they affect chemokine availability and function and impact a multitude of pathophysiological events, including the tumorigenesis process. This family of atypical receptors comprises five members: ACKR1/DARC, ACKR2/D6,ACKR3/CXCR7, ACKR4/CCRL1, and ACKR5/CCRL2. This work evaluated the differential expression of these receptors in prostate cancer using quantitative PCR. Further evaluation of CCRL2 at the protein level confirmed its overexpression in a metastatic cell line and in malignant prostatic tissues from patients. CCRL2, a presumed member of the atypical chemokine receptor family, plays a key role in lung dendritic cell trafficking to peripheral lymph nodes.Recent studies have reported the expression of CCRL2 in different human cancer cell lines and tissues. However, its function and expression in prostate cancer has not been previously addressed.