OBJECTIVE Na+/K+-ATPase(NKA)is large membrane protein expressed uni⁃versally which is indispensable for the mainte⁃nance of ionic gradient as well as neuronal excit⁃ability.The role of NKA in inflammatory regula⁃tion ...OBJECTIVE Na+/K+-ATPase(NKA)is large membrane protein expressed uni⁃versally which is indispensable for the mainte⁃nance of ionic gradient as well as neuronal excit⁃ability.The role of NKA in inflammatory regula⁃tion is still unclear.Inflammatory responses are initiated upon the activation of inflammasomes.In order to investigate the crosslink between NKA and inflammasome,NKAα1 knockout(KO)N2a cells were generated using CRISPR/Cas9 system.METHODS AND RESULTS qPCR results showed that NLRP1 and NLRP3 were upregulated in response to NKAα1 loss while both NLRC4 and AIM2 remained unaffected.Meanwhile,consistent with the change in NLRP1 and NLRP3,both the mRNA level of ASC and IL-1βwere significantly increased in NKAα1 KO cells.These data indicated that NKAα1 interfer⁃ence might influence the level of NLRP1 and NLRP3 inflammasomes in neuronal cells.Further evidence indicating the potential link between NKA and inflammasome pathway were provided using cytokine array assay where all the differen⁃tiated protein detected were closely linked to NLRP1 and NLRP3.To confirm this effect,we also observed the transcriptional levels of inflam⁃masome proteins in the brain cortex from both NKAα1+/+and NKAα1+/-mice.In line with the observation gained in NKAα1 KO cells,the mRNA level of NLRP1,NLRP3 and IL-1βwere significantly upregulated in NKAα1+/-mice brain.Interestingly,in the primary cultured astrocytes,treatment with LPS/ATP significantly reduced the mRNA and protein levels of NKAα1 expression.These data imply that a negative regulation loop between NKAα1 and inflammation may exist in the central nervous system.Since neuroinflam⁃matory mechanism is currently considered the most potential of interventions to target anxiety,we therefore perform behavioural experiments to investigate the role of NKAα1 in anxiety.Chronic restraint stress(CRS)for 10 d significantly reduced the time and frequency of entering the open arm and prolonged the retention time in the closed arm in the elevated plus-maze test.In the open field test,CRS also reduced both duration and frequency of entering into the central region.Although NKAα1 loss itself did not alter the behaviour performance in the normal condition,it exacerbated CRS-induced above behaviour abnormalities.CONCLUSION NKAα1 is regulat⁃ed upon inflammatory challenger and may be a novel target to treat anxiety.展开更多
BACKGROUND:Hepatic ischemia-reperfusion injury is a common phenomenon in hepatic surgical procedures and can result in further severe damage.This study aimed to investigate the protective effects of glutamine precondi...BACKGROUND:Hepatic ischemia-reperfusion injury is a common phenomenon in hepatic surgical procedures and can result in further severe damage.This study aimed to investigate the protective effects of glutamine preconditioning on hepatic ischemia-reperfusion injury in rats and its dose-dependency. METHODS:Thirty-two healthy male Wistar rats were randomly divided into four groups(n=8 per group).One group received 0.9%NaCl(control)and the other three received glutamine(Gln groups)4 hours before ischemia.The Gln groups were named GL,GM,and GH according to the glutamine dose.The liver was subjected to 1 hour of ischemia and 2 hours of reperfusion. Two hours later,the levels of alanine aminotransferase(ALT), intracellular free calcium(Ca 2+ ),and activity of Na + /K + adenosine triphosphatase(ATPase)and superoxide dismutase (SOD)were assessed,and liver tissue sections were examined under a microscope. RESULTS:The Gln and control groups differed in the concentration of intracellular free calcium(P<0.05),and the activity of Na + /K + ATPase and SOD in the Gln groups was higher than in the control group(P<0.05).The ALT level was lower in the GM and GH groups than in the control group(P<0.05).The levels of Na + /K + ATPase and SOD rose gradually with increasing glutamine dose(P<0.05),and the concentration of Ca 2+ declined gradually with increasing glutamine dose(P<0.05).The degree of hepatocyte injury was milder in the Gln groups than in the control group. CONCLUSIONS:Glutamine preconditioning protected effectively against hepatic ischemia-reperfusion injury.These protective effects were related to the dose of glutamine and due to the reduction of intracellular calcium overload and the improvements in the activity of Na + /K + ATPase and SOD.展开更多
A growing number of researches have shown that ouabain can regulate mammalian sperm function and male reproduction by modulating the sperm motility, capacitation and acrosome reaction in vitro. This study further exam...A growing number of researches have shown that ouabain can regulate mammalian sperm function and male reproduction by modulating the sperm motility, capacitation and acrosome reaction in vitro. This study further examined the relationship between ouabain and asthenozoospermia. In this study, the rat was intraperitoneally injected with ouabain at different concentrations（low-dose ouabain group： 12.5 μg/kg body weight per day, and high-dose ouabain group： 25 μg/kg body weight per day） for 30 days to establish the asthenozoospermia model. The sperms from 60 males with normal fertility were incubated with ouabain of gradient concentrations（10-7–10-2mol/L） for 4 h. The sperm motility was evaluated under a microscope. Moreover, the endogenous ouabain（EO） level was determined in seminal plasma of mild or severe asthenozoospermia patients and males with normal fertility by competitive inhibition ELISA. The results showed that the sperm motility was significantly diminished in the rats treated with different concentrations of ouabain. The number of motile sperms（grades a and b） was decreased greatly in a time- and dose-dependent manner in 10-5–10-2mol/L ouabain groups（P0.01）, while no obvious change in sperm motility was observed in 10-7–10-6mol/L groups even for 4-h incubation（P0.05）. Furthermore, the EO level was significantly increased in asthenozoospermia patients as compared with that in males with normal fertility（25.27±1.71 μg/L in mild asthenozoospermia patients, 26.52±1.82 μg/L in severe asthenozoospermia patients, 19.31±1.45 μg/L in normal fertility men）（P0.01）. In conclusion, rat asthenozoospermia was successfully established by intraperitoneal injection of ouabain, and 10-5mol/L ouabain was sufficient enough to inhibit sperm motility in vitro. Moreover, EO, a normal constituent of seminal plasma, was highly expressed in asthenozoospermia males as compared with normal fertility ones.展开更多
文摘OBJECTIVE Na+/K+-ATPase(NKA)is large membrane protein expressed uni⁃versally which is indispensable for the mainte⁃nance of ionic gradient as well as neuronal excit⁃ability.The role of NKA in inflammatory regula⁃tion is still unclear.Inflammatory responses are initiated upon the activation of inflammasomes.In order to investigate the crosslink between NKA and inflammasome,NKAα1 knockout(KO)N2a cells were generated using CRISPR/Cas9 system.METHODS AND RESULTS qPCR results showed that NLRP1 and NLRP3 were upregulated in response to NKAα1 loss while both NLRC4 and AIM2 remained unaffected.Meanwhile,consistent with the change in NLRP1 and NLRP3,both the mRNA level of ASC and IL-1βwere significantly increased in NKAα1 KO cells.These data indicated that NKAα1 interfer⁃ence might influence the level of NLRP1 and NLRP3 inflammasomes in neuronal cells.Further evidence indicating the potential link between NKA and inflammasome pathway were provided using cytokine array assay where all the differen⁃tiated protein detected were closely linked to NLRP1 and NLRP3.To confirm this effect,we also observed the transcriptional levels of inflam⁃masome proteins in the brain cortex from both NKAα1+/+and NKAα1+/-mice.In line with the observation gained in NKAα1 KO cells,the mRNA level of NLRP1,NLRP3 and IL-1βwere significantly upregulated in NKAα1+/-mice brain.Interestingly,in the primary cultured astrocytes,treatment with LPS/ATP significantly reduced the mRNA and protein levels of NKAα1 expression.These data imply that a negative regulation loop between NKAα1 and inflammation may exist in the central nervous system.Since neuroinflam⁃matory mechanism is currently considered the most potential of interventions to target anxiety,we therefore perform behavioural experiments to investigate the role of NKAα1 in anxiety.Chronic restraint stress(CRS)for 10 d significantly reduced the time and frequency of entering the open arm and prolonged the retention time in the closed arm in the elevated plus-maze test.In the open field test,CRS also reduced both duration and frequency of entering into the central region.Although NKAα1 loss itself did not alter the behaviour performance in the normal condition,it exacerbated CRS-induced above behaviour abnormalities.CONCLUSION NKAα1 is regulat⁃ed upon inflammatory challenger and may be a novel target to treat anxiety.
文摘BACKGROUND:Hepatic ischemia-reperfusion injury is a common phenomenon in hepatic surgical procedures and can result in further severe damage.This study aimed to investigate the protective effects of glutamine preconditioning on hepatic ischemia-reperfusion injury in rats and its dose-dependency. METHODS:Thirty-two healthy male Wistar rats were randomly divided into four groups(n=8 per group).One group received 0.9%NaCl(control)and the other three received glutamine(Gln groups)4 hours before ischemia.The Gln groups were named GL,GM,and GH according to the glutamine dose.The liver was subjected to 1 hour of ischemia and 2 hours of reperfusion. Two hours later,the levels of alanine aminotransferase(ALT), intracellular free calcium(Ca 2+ ),and activity of Na + /K + adenosine triphosphatase(ATPase)and superoxide dismutase (SOD)were assessed,and liver tissue sections were examined under a microscope. RESULTS:The Gln and control groups differed in the concentration of intracellular free calcium(P<0.05),and the activity of Na + /K + ATPase and SOD in the Gln groups was higher than in the control group(P<0.05).The ALT level was lower in the GM and GH groups than in the control group(P<0.05).The levels of Na + /K + ATPase and SOD rose gradually with increasing glutamine dose(P<0.05),and the concentration of Ca 2+ declined gradually with increasing glutamine dose(P<0.05).The degree of hepatocyte injury was milder in the Gln groups than in the control group. CONCLUSIONS:Glutamine preconditioning protected effectively against hepatic ischemia-reperfusion injury.These protective effects were related to the dose of glutamine and due to the reduction of intracellular calcium overload and the improvements in the activity of Na + /K + ATPase and SOD.
基金supported by 2012 Independence Innovation Foundation of Huazhong University of Science and Technology(No.01-18-519003)
文摘A growing number of researches have shown that ouabain can regulate mammalian sperm function and male reproduction by modulating the sperm motility, capacitation and acrosome reaction in vitro. This study further examined the relationship between ouabain and asthenozoospermia. In this study, the rat was intraperitoneally injected with ouabain at different concentrations（low-dose ouabain group： 12.5 μg/kg body weight per day, and high-dose ouabain group： 25 μg/kg body weight per day） for 30 days to establish the asthenozoospermia model. The sperms from 60 males with normal fertility were incubated with ouabain of gradient concentrations（10-7–10-2mol/L） for 4 h. The sperm motility was evaluated under a microscope. Moreover, the endogenous ouabain（EO） level was determined in seminal plasma of mild or severe asthenozoospermia patients and males with normal fertility by competitive inhibition ELISA. The results showed that the sperm motility was significantly diminished in the rats treated with different concentrations of ouabain. The number of motile sperms（grades a and b） was decreased greatly in a time- and dose-dependent manner in 10-5–10-2mol/L ouabain groups（P0.01）, while no obvious change in sperm motility was observed in 10-7–10-6mol/L groups even for 4-h incubation（P0.05）. Furthermore, the EO level was significantly increased in asthenozoospermia patients as compared with that in males with normal fertility（25.27±1.71 μg/L in mild asthenozoospermia patients, 26.52±1.82 μg/L in severe asthenozoospermia patients, 19.31±1.45 μg/L in normal fertility men）（P0.01）. In conclusion, rat asthenozoospermia was successfully established by intraperitoneal injection of ouabain, and 10-5mol/L ouabain was sufficient enough to inhibit sperm motility in vitro. Moreover, EO, a normal constituent of seminal plasma, was highly expressed in asthenozoospermia males as compared with normal fertility ones.
