Transient receptor potential melastatin 2(TRPM2) is an important ion channel that represents a potential target for treating injury caused by cerebral ischemia. However, it is unclear whether reducing TRPM2 expression...Transient receptor potential melastatin 2(TRPM2) is an important ion channel that represents a potential target for treating injury caused by cerebral ischemia. However, it is unclear whether reducing TRPM2 expression can help repair cerebral injury, and if so what the mechanism underlying this process involves. This study investigated the protective effect of reducing TRPM2 expression on pheochromocytoma(PC12) cells injured by oxygen-glucose deprivation(OGD). PC12 cells were transfected with plasmid encoding TRPM2 shRNAS, then subjected to OGD by incubation in glucose-free medium under hypoxic conditions for 8 hours, after which the cells were allowed to reoxygenate for 24 hours. Apoptotic cells, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels were detected using flow cytometry. The relative expression of C-X-C motif chemokine ligand 2(CXCL2), NACHT, LRR, and PYD domain–containing protein 3(NALP3), and caspase-1 were detected using fluorescence-based quantitative reverse transcription-polymerase chain reaction and western blotting. The rates of apoptosis, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels in the TRPM2-shRNA + OGD group were lower than those observed in the OGD group. Taken together, these results suggest that TRPM2 knockdown reduces OGD-induced neuronal injury, potentially by inhibiting apoptosis and reducing oxidative stress levels, mitochondrial membrane potentials, intracellular calcium concentrations, and NLRP3 inflammasome activation.展开更多
OBJECTIVE To investigate the role of transient receptor potential melastatin 2(TRPM2),a calcium-permeable non-selective cation channel which acts as an oxidative stress sensor,in mediating the production of pro-inflam...OBJECTIVE To investigate the role of transient receptor potential melastatin 2(TRPM2),a calcium-permeable non-selective cation channel which acts as an oxidative stress sensor,in mediating the production of pro-inflammatory IL-1βin high glucose condition.METHODS Human pro-monocytic leukemia cell U937 was purchased from ATCC and cultured in RPMI 1640(Life Technologies).Prior to high glucose(HG)stimulation,U937 cells were cultured in medium with glucose 5.5mmol·L-1 for 48 h.The cells were then incubated in high glucose concentration(30mmol·L-1)or mannitol(30mmol·L-1)for 48 h.The protein expression of TRPM2 and the production of human IL-1β were evaluated by ELISA.TRPM2 inhibitors(DPQ and AMP)and TRPM2 siRNAs were employed to further investigate the role of TRPM2 in HG-induced IL-1β production.RESULTS The TRPM2 protein expression was significantly up-regulated by 2-folds in U937 cells after the treatment of high glucose(30mmol·L-1 for 48h)(P<0.01).The production of IL-1β in U937 was also significantly increased by HG treatment and was time-and dose-dependent(10,20 or 30mmol·L-1 glucose for 24,48 or 72h)(P<0.01).The HG-induced IL-1β production in U937 could be abolished by using TRPM2 inhibitors DPQ(100μmol·L-1 for 45min)and AMP(100μmol·L-1 for 45 min)as well as by the transfection of TRPM2siRNAs(60nmol·L-1).CONCLUSION High glucose condition(such as in diabetes)might mediate pro-inflammatory environments via the modulation of TRPM2 channels on immune cells.展开更多
The wound is induced by several mechanical and metabolic factors.In the etiology of the wound recovery,excessive oxidative stress,calcium ion(Ca^(2+))influx,and apoptosis have important roles.Ca^(2+)-permeable TRPM2 c...The wound is induced by several mechanical and metabolic factors.In the etiology of the wound recovery,excessive oxidative stress,calcium ion(Ca^(2+))influx,and apoptosis have important roles.Ca^(2+)-permeable TRPM2 channel is activated by oxidative stress.Protective roles of Hypericum perforatum extract(HP)on the mechanical nerve injury-induced apoptosis and oxidative toxicity through regulation of TRPM2 in the experimental animals were recently reported.The potential protective roles in HP treatment were evaluated on the TRPM2-mediated cellular oxidative toxicity in the renal epithelium(MPK)cells.The cells were divided into three groups as control,wound,and wound+HP treatment(75μM for 72 h).Wound diameters were more importantly decreased in the wound+HP group than in the wound group.In addition,the results of laser confocal microscopy analyses indicated protective roles of HP and TRPM2 antagonists(N-(p-Amylcinnamoyl)anthranilic acid and 2-aminoethyl diphenylborinate)against the wound-induced increase of Ca^(2+) influx and mitochondrial ROS production.The wound-induced increase of early(annexin V-FITC)apoptosis and late(propidium iodide)apoptosis were also decreased in the cells by the HP treatment.In conclusion,HP treatment acted protective effects against wound-mediated oxidative cell toxicity and apoptosis through TRPM2 inhibition.These effects may be attributed to their potent antioxidant effect.展开更多
Hepatic ischemia–reperfusion(IR)injury is a serious clinical problem that complicates liver resection and transplantation.Despite recent advances in understanding of the pathophysiology of hepatic IR injury,effective...Hepatic ischemia–reperfusion(IR)injury is a serious clinical problem that complicates liver resection and transplantation.Despite recent advances in understanding of the pathophysiology of hepatic IR injury,effective interventions and therapeutics are still lacking.Here,we examined the role of transient receptor potential melastatin 2(TRPM2),a Ca^(2+)-permeable,non-selective cation channel,in mediating hepatic IR injury.Our data showed that TRPM2 deficiency attenuated IR-induced liver dysfunction,inflammation,and cell death in mice.Moreover,RNA sequencing analysis indicated that TRPM2-induced IR injury occurs via ferroptosis-related pathways.Consistently,as a ferroptosis inducer,(1S,3R)-RSL3 treatment induced mitochondrial dysfunction in hepatocytes and a TRPM2 inhibitor suppressed this.Interestingly,TRPM2-mediated calcium influx caused mitochondrial calcium accumulation via the mitochondrial Ca^(2+)-selective uniporter and increased the expression level of arachidonate 12-lipoxygenase(ALOX12),which results in mitochondrial lipid peroxidation during hepatic IR injury.Furthermore,hepatic IR injury-induced ferroptosis was obviously relieved by a TRPM2 inhibitor or calcium depletion,both in vitro and in vivo.Collectively,these findings demonstrate a crucial role for TRPM2-mediated ferroptosis in hepatic IR injury via increased Ca^(2+)-induced ALOX12 expression,indicating that pharmacological inhibition of TRPM2 may provide an effective therapeutic strategy for hepatic IR injury-related diseases,such as during liver resection and transplantation.展开更多
基金supported by the National Natural Science Foundation of China,Nos.81671532,81771625(to XF)the Jiangsu Provincial Key Medical Discipline of China,No.ZDXKA2016013(to XF)+3 种基金the Jiangsu Provincial Medical Youth Talent of China,No.QNRC2016758(to XF)the Jiangsu Province Women and Children Health Research Project of China,No.F201750(to XF)the Public Health Technology Project of Suzhou City of China,No.SYS201765(to XF)a grant from the Department of Pediatrics Clinical Center of Suzhou City of China,No.Szzx201504(to XF)。
文摘Transient receptor potential melastatin 2(TRPM2) is an important ion channel that represents a potential target for treating injury caused by cerebral ischemia. However, it is unclear whether reducing TRPM2 expression can help repair cerebral injury, and if so what the mechanism underlying this process involves. This study investigated the protective effect of reducing TRPM2 expression on pheochromocytoma(PC12) cells injured by oxygen-glucose deprivation(OGD). PC12 cells were transfected with plasmid encoding TRPM2 shRNAS, then subjected to OGD by incubation in glucose-free medium under hypoxic conditions for 8 hours, after which the cells were allowed to reoxygenate for 24 hours. Apoptotic cells, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels were detected using flow cytometry. The relative expression of C-X-C motif chemokine ligand 2(CXCL2), NACHT, LRR, and PYD domain–containing protein 3(NALP3), and caspase-1 were detected using fluorescence-based quantitative reverse transcription-polymerase chain reaction and western blotting. The rates of apoptosis, mitochondrial membrane potentials, reactive oxygen species levels, and cellular calcium levels in the TRPM2-shRNA + OGD group were lower than those observed in the OGD group. Taken together, these results suggest that TRPM2 knockdown reduces OGD-induced neuronal injury, potentially by inhibiting apoptosis and reducing oxidative stress levels, mitochondrial membrane potentials, intracellular calcium concentrations, and NLRP3 inflammasome activation.
基金The project supported by Science and Technology Development Fund of Macao SAR(118/2012/A)Research Committee of the University of Macao〔MYRG124(Y1-L3)-ICMS12-HPM〕
文摘OBJECTIVE To investigate the role of transient receptor potential melastatin 2(TRPM2),a calcium-permeable non-selective cation channel which acts as an oxidative stress sensor,in mediating the production of pro-inflammatory IL-1βin high glucose condition.METHODS Human pro-monocytic leukemia cell U937 was purchased from ATCC and cultured in RPMI 1640(Life Technologies).Prior to high glucose(HG)stimulation,U937 cells were cultured in medium with glucose 5.5mmol·L-1 for 48 h.The cells were then incubated in high glucose concentration(30mmol·L-1)or mannitol(30mmol·L-1)for 48 h.The protein expression of TRPM2 and the production of human IL-1β were evaluated by ELISA.TRPM2 inhibitors(DPQ and AMP)and TRPM2 siRNAs were employed to further investigate the role of TRPM2 in HG-induced IL-1β production.RESULTS The TRPM2 protein expression was significantly up-regulated by 2-folds in U937 cells after the treatment of high glucose(30mmol·L-1 for 48h)(P<0.01).The production of IL-1β in U937 was also significantly increased by HG treatment and was time-and dose-dependent(10,20 or 30mmol·L-1 glucose for 24,48 or 72h)(P<0.01).The HG-induced IL-1β production in U937 could be abolished by using TRPM2 inhibitors DPQ(100μmol·L-1 for 45min)and AMP(100μmol·L-1 for 45 min)as well as by the transfection of TRPM2siRNAs(60nmol·L-1).CONCLUSION High glucose condition(such as in diabetes)might mediate pro-inflammatory environments via the modulation of TRPM2 channels on immune cells.
文摘The wound is induced by several mechanical and metabolic factors.In the etiology of the wound recovery,excessive oxidative stress,calcium ion(Ca^(2+))influx,and apoptosis have important roles.Ca^(2+)-permeable TRPM2 channel is activated by oxidative stress.Protective roles of Hypericum perforatum extract(HP)on the mechanical nerve injury-induced apoptosis and oxidative toxicity through regulation of TRPM2 in the experimental animals were recently reported.The potential protective roles in HP treatment were evaluated on the TRPM2-mediated cellular oxidative toxicity in the renal epithelium(MPK)cells.The cells were divided into three groups as control,wound,and wound+HP treatment(75μM for 72 h).Wound diameters were more importantly decreased in the wound+HP group than in the wound group.In addition,the results of laser confocal microscopy analyses indicated protective roles of HP and TRPM2 antagonists(N-(p-Amylcinnamoyl)anthranilic acid and 2-aminoethyl diphenylborinate)against the wound-induced increase of Ca^(2+) influx and mitochondrial ROS production.The wound-induced increase of early(annexin V-FITC)apoptosis and late(propidium iodide)apoptosis were also decreased in the cells by the HP treatment.In conclusion,HP treatment acted protective effects against wound-mediated oxidative cell toxicity and apoptosis through TRPM2 inhibition.These effects may be attributed to their potent antioxidant effect.
基金the National Key Research and Development Project[2017YFC0110802]the Zhejiang Province Key Research and Development Project[2020C01059,2020]+4 种基金the Natural Science Foundation of China[82030108,31872796,81874059,32071102,and 82102105]the Natural Science Foundation of Zhejiang Province[LQ22H160017]the Zhejiang Engineering Research Center of Cognitive Healthcare[2017E10011]the National Key Scientific Instrument and Equipment Development Project[81827804]the China Postdoctoral Science Foundation[2021M702825].
文摘Hepatic ischemia–reperfusion(IR)injury is a serious clinical problem that complicates liver resection and transplantation.Despite recent advances in understanding of the pathophysiology of hepatic IR injury,effective interventions and therapeutics are still lacking.Here,we examined the role of transient receptor potential melastatin 2(TRPM2),a Ca^(2+)-permeable,non-selective cation channel,in mediating hepatic IR injury.Our data showed that TRPM2 deficiency attenuated IR-induced liver dysfunction,inflammation,and cell death in mice.Moreover,RNA sequencing analysis indicated that TRPM2-induced IR injury occurs via ferroptosis-related pathways.Consistently,as a ferroptosis inducer,(1S,3R)-RSL3 treatment induced mitochondrial dysfunction in hepatocytes and a TRPM2 inhibitor suppressed this.Interestingly,TRPM2-mediated calcium influx caused mitochondrial calcium accumulation via the mitochondrial Ca^(2+)-selective uniporter and increased the expression level of arachidonate 12-lipoxygenase(ALOX12),which results in mitochondrial lipid peroxidation during hepatic IR injury.Furthermore,hepatic IR injury-induced ferroptosis was obviously relieved by a TRPM2 inhibitor or calcium depletion,both in vitro and in vivo.Collectively,these findings demonstrate a crucial role for TRPM2-mediated ferroptosis in hepatic IR injury via increased Ca^(2+)-induced ALOX12 expression,indicating that pharmacological inhibition of TRPM2 may provide an effective therapeutic strategy for hepatic IR injury-related diseases,such as during liver resection and transplantation.
