AIM To determine how a normal human colon cell line reacts to microbial challenge as a way to study oxidative stress-induced responses associated with inflammatory bowel disease.METHODS Normal human colon epithelial c...AIM To determine how a normal human colon cell line reacts to microbial challenge as a way to study oxidative stress-induced responses associated with inflammatory bowel disease.METHODS Normal human colon epithelial cells(ATCC?CRL.1790?)were stimulated with either heat killed E.coli or heat killed murine cecal contents(HKC)and examined for several relevant biomarkers associated with inflammation and oxidative stress including cytokine production,mitochondrial autophagy and oxidant status.TNFα,IL-1βand IL-8 protein concentrations were measured within the supernatants.Fluorescent microscopy was performed to quantify the production of reactive oxygen species(ROS)using an oxidation responsive fluorogenic probe.Mitochondrial morphology and mitochondrial membrane potential was assessed by dual staining using COXIV antibody and a dye concentrating in active mitochondria.Mitochondrial ROS scavenger was used to determine the source of ROS in stimulated cells.Autophagy was detected by staining for the presence of autophagic vesicles.Positive controls for autophagy and ROS/RNS experiments were treated with rapamycin and chloroquine.Mitochondrial morphology,ROS production and autophagy microscopy experiments were analyzed using a custom acquisition and analysis microscopy software(Image J).RESULTS Exposing CRL.1790 cells to microbial challenge stimulated cells to produce several relevant biomarkers associated with inflammation and oxidative stress.Heat killed cecal contents treatment induced a 10-12fold increase in IL-8 production by CRL.1790 cells compared to unstimulated controls at 6 and 12 h(P<0.001).Heat killed E.coli stimulation resulted in a4-5 fold increase in IL-8 compared to the unstimulated control cells at each time point(P<0.001).Both heat killed E.coli and HKC stimulated robust ROS production at 6(P<0.001),and 12 h(P<0.01).Mitochondrial morphologic abnormalities were detected at 6 and12 h based on reduced mitochondrial circularity and decreased mitochondrial membrane potential,P<0.01.Microbial stimulation also induced significant autophagy at 6 and 12 h,P<0.01.Lastly,blocking mitochondrial ROS generation using mitochondrial specific ROS scavenger reversed microbial challenge induced mitochondrial morphologic abnormalities and autophagy.CONCLUSION The findings from this study suggest that CRL.1790cells may be a useful alternative to other colon cancer cell lines in studying the mechanisms of oxidative stress events associated with intestinal inflammatory disorders.展开更多
The study on the factors influencing the biogenic gas production of low rank coal beds is of great significance to the search of the favorable biogenic gas enrichment areas of similar coal beds and the guidance of the...The study on the factors influencing the biogenic gas production of low rank coal beds is of great significance to the search of the favorable biogenic gas enrichment areas of similar coal beds and the guidance of the microbial stimulation technologies.In this paper,the lignite samples taken from the Jiergalangtu Sag in the Erlian Basin were selected to carry out the simulation experiment of biogenic coalbed methane under different temperatures,pH values,oxidation-reduction potentials(Eh)and trace element concentrations.Then,the influences on biogenic gas production were discussed and the optimal biogenic gas production condition was determined.And the following research results were obtained.First,in the simulation experiments,the biogenic gas production of coal beds increases first and then decreases with the increase of temperature,pH and trace element concentration and the decrease of Eh.Second,in the case of temperature 30-35℃,pH 7.0-7.5 and Eh 225 mV,the metabolism of methanogen is the most active and the most favorable for the generation of biogenic gas,so biogenic gas production can be promoted by adding the trace elements(Fe^(2+)and Ni^(2+))of proper concentration.It is concluded that a proper burial depth(300-600 m),a weak runoff hydrodynamic condition and an anaerobic environment constitute the favorable conditions for the generation of biogenic gas in the Jiergalangtu Sag,Erlian Basin.Moreover,the addition of appropriate trace elements can be an effective means of microbial stimulation.展开更多
基金Supported by an endowment from the Matilda R.Wilson Fund in Detroit,Michigan
文摘AIM To determine how a normal human colon cell line reacts to microbial challenge as a way to study oxidative stress-induced responses associated with inflammatory bowel disease.METHODS Normal human colon epithelial cells(ATCC?CRL.1790?)were stimulated with either heat killed E.coli or heat killed murine cecal contents(HKC)and examined for several relevant biomarkers associated with inflammation and oxidative stress including cytokine production,mitochondrial autophagy and oxidant status.TNFα,IL-1βand IL-8 protein concentrations were measured within the supernatants.Fluorescent microscopy was performed to quantify the production of reactive oxygen species(ROS)using an oxidation responsive fluorogenic probe.Mitochondrial morphology and mitochondrial membrane potential was assessed by dual staining using COXIV antibody and a dye concentrating in active mitochondria.Mitochondrial ROS scavenger was used to determine the source of ROS in stimulated cells.Autophagy was detected by staining for the presence of autophagic vesicles.Positive controls for autophagy and ROS/RNS experiments were treated with rapamycin and chloroquine.Mitochondrial morphology,ROS production and autophagy microscopy experiments were analyzed using a custom acquisition and analysis microscopy software(Image J).RESULTS Exposing CRL.1790 cells to microbial challenge stimulated cells to produce several relevant biomarkers associated with inflammation and oxidative stress.Heat killed cecal contents treatment induced a 10-12fold increase in IL-8 production by CRL.1790 cells compared to unstimulated controls at 6 and 12 h(P<0.001).Heat killed E.coli stimulation resulted in a4-5 fold increase in IL-8 compared to the unstimulated control cells at each time point(P<0.001).Both heat killed E.coli and HKC stimulated robust ROS production at 6(P<0.001),and 12 h(P<0.01).Mitochondrial morphologic abnormalities were detected at 6 and12 h based on reduced mitochondrial circularity and decreased mitochondrial membrane potential,P<0.01.Microbial stimulation also induced significant autophagy at 6 and 12 h,P<0.01.Lastly,blocking mitochondrial ROS generation using mitochondrial specific ROS scavenger reversed microbial challenge induced mitochondrial morphologic abnormalities and autophagy.CONCLUSION The findings from this study suggest that CRL.1790cells may be a useful alternative to other colon cancer cell lines in studying the mechanisms of oxidative stress events associated with intestinal inflammatory disorders.
基金supported by the National Science and Technology Major Project“Optimization and Evaluation of Blocks for Large-scale Development of Medium-and Low-rank Coalbed Methane(CBM)”(No.:2016ZX05041)PetroChina Major Science and Technology Project“Study and Application of Key Technologies for Exploration and Development of Coalbed Methane”(No.:2017E14).
文摘The study on the factors influencing the biogenic gas production of low rank coal beds is of great significance to the search of the favorable biogenic gas enrichment areas of similar coal beds and the guidance of the microbial stimulation technologies.In this paper,the lignite samples taken from the Jiergalangtu Sag in the Erlian Basin were selected to carry out the simulation experiment of biogenic coalbed methane under different temperatures,pH values,oxidation-reduction potentials(Eh)and trace element concentrations.Then,the influences on biogenic gas production were discussed and the optimal biogenic gas production condition was determined.And the following research results were obtained.First,in the simulation experiments,the biogenic gas production of coal beds increases first and then decreases with the increase of temperature,pH and trace element concentration and the decrease of Eh.Second,in the case of temperature 30-35℃,pH 7.0-7.5 and Eh 225 mV,the metabolism of methanogen is the most active and the most favorable for the generation of biogenic gas,so biogenic gas production can be promoted by adding the trace elements(Fe^(2+)and Ni^(2+))of proper concentration.It is concluded that a proper burial depth(300-600 m),a weak runoff hydrodynamic condition and an anaerobic environment constitute the favorable conditions for the generation of biogenic gas in the Jiergalangtu Sag,Erlian Basin.Moreover,the addition of appropriate trace elements can be an effective means of microbial stimulation.
