Obejective:To explore the establishment of an oxygen glucose deprivation/reperfusion model of senescent SH-SY5Y cells.Methods:SH-SY5Y cells were randomly divided into control(D-galactose 0 mmol/L group),D-galactose(25...Obejective:To explore the establishment of an oxygen glucose deprivation/reperfusion model of senescent SH-SY5Y cells.Methods:SH-SY5Y cells were randomly divided into control(D-galactose 0 mmol/L group),D-galactose(25 mmol/L,50 mmol/L,100 mmol/L,200 mmol/L,400 mmol/L)groups,and treated with corresponding concentrations of D-galactose for 48 h.The changes of cell morphology,β-galactosidase,the cell morphology,β-galactosidase activity by microscopic observation,cell proliferation rate by EdU kit and cell survival rate by CCK-8 assay were used to determine the decaying concentration of D-galactose and to establish the senescence model.The senescent SH-SY5Y cells were randomly divided into control group(oxygen glucose deprivation without treatment group),oxygen glucose deprivation treatment(0.5 h,1 h,1.5 h,2 h)group,followed by re-glucose reoxygenation for 24 h,and CCK-8 assay for the survival rate of senescent SH-SY5Y cells.Results:There were no significant changes in cell morphology and β-gal activity in the 25 mmol/L and 50 mmol/L groups compared with the control group(P>0.05),cytosolic hypertrophy was seen in the cells of the 100 mmol/L group,chromatin fixation in the cells of the 200 mmol/L group,and massive vacuolization in the cells of the 400 mmol/L group;the positive rate ofβ-galactosidase staining in the cells of the(100-400 mmol/L)group was significantly higher compared with the control group(P<0.05),with little difference between the 100 mmol/L and 200 mmol/L groups(P>0.05);the cell proliferation ability of the(100-400 mmol/L)group was significantly decreased in a concentration-dependent manner(P<0.05);the cell survival rate was decreased in a concentration-dependent manner(P<0.05),with IC_(50) between 100 mmol/L and 200 mmol/L.The survival of senescent SH-SY5Y cells showed a time-dependent decrease in oxygen-glucose deprivation(P<0.05),with an IC_(50) close to 1 h.Conclusion:D-gal concentration of 100 mmoL/L and 48 h of cell action could establish a survival rate of about 50%of senescent SH-SY5Y cells,and oxygen glucose deprivation of senescent SH-SY5Y cells for 1 h and reperfusion for 24 h could establish an oxygen glucose deprivation/reperfusion model of senescent SH-SY5Y cells with a survival rate close to 50%.展开更多
Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ische...Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ischemic encephalopathy.The therapeutic potential of human placental chorionic plate-derived mesenchymal stem cells for various diseases has been explored.However,the potential use of human placental chorionic plate-derived mesenchymal stem cells for the treatment of neonatal hypoxic-ischemic encephalopathy has not yet been investigated.In this study,we injected human placental chorionic plate-derived mesenchymal stem cells into the lateral ventricle of a neonatal hypoxic-ischemic encephalopathy rat model and observed significant improvements in both cognitive and motor function.Protein chip analysis showed that interleukin-3 expression was significantly elevated in neonatal hypoxic-ischemic encephalopathy model rats.Following transplantation of human placental chorionic plate-derived mesenchymal stem cells,interleukin-3 expression was downregulated.To further investigate the role of interleukin-3 in neonatal hypoxic-ischemic encephalopathy,we established an in vitro SH-SY5Y cell model of hypoxic-ischemic injury through oxygen-glucose deprivation and silenced interleukin-3 expression using small interfering RNA.We found that the activity and proliferation of SH-SY5Y cells subjected to oxygen-glucose deprivation were further suppressed by interleukin-3 knockdown.Furthermore,interleukin-3 knockout exacerbated neuronal damage and cognitive and motor function impairment in rat models of hypoxic-ischemic encephalopathy.The findings suggest that transplantation of hpcMSCs ameliorated behavioral impairments in a rat model of hypoxic-ischemic encephalopathy,and this effect was mediated by interleukin-3-dependent neurological function.展开更多
A plasmid for cytoglobin expression, pAcGFP1-Cl-cytoglobin, was transfected into SH-SY5Y cells. Cobalt chloride was used to establish a model of hypoxia. Western blotting indicated that cytoglobin was overexpressed an...A plasmid for cytoglobin expression, pAcGFP1-Cl-cytoglobin, was transfected into SH-SY5Y cells. Cobalt chloride was used to establish a model of hypoxia. Western blotting indicated that cytoglobin was overexpressed and there was low expression of hypoxia-inducible factor-la in SH-SY5Y cells after transfection. Following cobalt chloride-induced hypoxia, cytoglobin and hypoxia-inducible fac- tor-la expression gradually increased in SH-SY5Y cells. Flow cytometry showed that with increas- ing duration of hypoxia, the proportion of normal cells significantly diminished in the transfected and non-transfected groups. The proportion of cells in the early stages of apoptosis increased. However, the proportion of apoptotic cells was significantly lower in the transfected group compared with the non-transfected group. These results demonstrate that cytoglobin and hypoxia-inducible factor-la are strongly up-regulated by hypoxia, and that there is a strong relationship between hy- poxia-inducible factor-la and cytoglobin during hypoxic injury.展开更多
In this study, we examined the effects of neuroglobin gene （Ngb） transfection into SH-SY5Y cells, using ultrasound-targeted microbubble destruction （UTMD）, on cobalt chloride-induced hypoxia. With an ultrasound in...In this study, we examined the effects of neuroglobin gene （Ngb） transfection into SH-SY5Y cells, using ultrasound-targeted microbubble destruction （UTMD）, on cobalt chloride-induced hypoxia. With an ultrasound intensity of 0.8 W/cm2, a 60-second exposure duration, 50% duty cycle, and 20% microbubble concentration, pAcGFP1-C1-Ngb-transfected cells exhibited the highest cell viability and transfection efficiency. The efficiency of plasmid delivery was significantly higher with UTMD than transfection with plasmid alone, transfection with plasmid using microbubbles, or transfection of plasmid by ultrasound. In addition, during cobalt chloride-induced hypoxia, caspase-3 activity in pAcGFP1-C1-Ngb-transfected cells was significantly lower than in untransfected cells. Ngb protein and mRNA expression were significantly higher in cells transfected by UTMD than in cells transfected with the other methods. These results demonstrate that UTMD can very efficiently mediate exogenous gene delivery, and that Ngb overexpression protects cells against cobalt chloride-induced hypoxia.展开更多
In vitro cultured human neuroblastoma SH-SY5Y cells were pretreated with 50 or 5 ug/mL geniposide for 12 hours and exposed to 400 umol/L corticosterone. Corticosterone exposure in cultures not pretreated with geniposi...In vitro cultured human neuroblastoma SH-SY5Y cells were pretreated with 50 or 5 ug/mL geniposide for 12 hours and exposed to 400 umol/L corticosterone. Corticosterone exposure in cultures not pretreated with geniposide resulted in inhibited cell growth, reduced cell survival, and increased P53 and P21 protein expression. However, in geniposide pretreated SH-SY5Y cells, cell viability and the number of cells in the G2 phase of the cell cycle were significantly increased, P21 and P53 protein expression was reduced, and cell apoptosis was inhibited following corticosterone exposure. These results indicate that geniposide can protect SH-SY5Y cells against high-dose corticosterone-induced injury.展开更多
The histone deacetylase inhibitor, trichostatin A, is used to treat Alzheimer’s disease and can improve learning and memory but its underlying mechanism of action is unknown. To determine whether the therapeutic effe...The histone deacetylase inhibitor, trichostatin A, is used to treat Alzheimer’s disease and can improve learning and memory but its underlying mechanism of action is unknown. To determine whether the therapeutic effect of trichostatin A on Alzheimer’s disease is associated with the nuclear factor erythroid 2-related factor 2(Nrf2) and Kelch-like epichlorohydrin-related protein-1(Keap1) signaling pathway, amyloid β-peptide 25–35(Aβ25–35) was used to induce Alzheimer’s disease-like pathological changes in SH-SY5 Y neuroblastoma cells. Cells were then treated with trichostatin A. The effects of trichostatin A on the expression of Keap1 and Nrf2 were detected by real-time quantitative polymerase chain reaction, western blot assays and immunofluorescence. Total antioxidant capacity and autophagy activity were evaluated by total antioxidant capacity assay kit and light chain 3-I/II levels, respectively. We found that trichostatin A increased cell viability and Nrf2 expression, and decreased Keap1 expression in SH-SY5 Y cells. Furthermore, trichostatin A increased the expression of Nrf2-related target genes, such as superoxide dismutase, NAD(P)H quinone dehydrogenase 1 and glutathione S-transferase, thereby increasing the total antioxidant capacity of SH-SY5 Y cells and inhibiting amyloid β-peptide-induced autophagy. Knockdown of Keap1 in SH-SY5 Y cells further increased trichostatin A-induced Nrf2 expression. These results indicate that the therapeutic effect of trichostatin A on Alzheimer’s disease is associated with the Keap1-Nrf2 pathway. The mechanism for this action may be that trichostatin A increases cell viability and the antioxidant capacity of SH-SY5 Y cells by alleviating Keap1-mediated inhibition Nrf2 signaling, thereby alleviating amyloid β-peptide-induced cell damage.展开更多
To investigate the molecular mechanism underlying the neuroprotective effect of lithium on cells, in this study, we exposed SH-SY5Y cells to 0.5 mmol/L lithium carbonate （Li2CO2） for 25-50 weeks and then detected th...To investigate the molecular mechanism underlying the neuroprotective effect of lithium on cells, in this study, we exposed SH-SY5Y cells to 0.5 mmol/L lithium carbonate （Li2CO2） for 25-50 weeks and then detected the expression levels of some neurobiology related genes and post-translational modifications of stress proteins in SH-SYSY cells, cDNA arrays showed that pyruvate kinase 2 （PKM2） and calmodulin 3 （CaM 3） expression levels were significantly down-regulated, phosphatase protein PP2A expression was lightly down-regulated, and casein kinase II （CK2）, threonine/tyrosine phosphatase 7 （PYST2）, and dopamine beta-hydroxylase （DBH） expression levels were significantly up-regulated. Besides, western blot analysis of stress proteins （HSP27, HSP70, GRP78 and GRP94） showed an over-expression of two proteins： a 105 kDa protein which is a hyper-phosphorylated isoform of GRP94, and a 108 kDa protein which is a phosphorylated tetramer of HSP27. These results suggest that the neuroprotective effects of lithium are likely related to gene expressions and post-translational modifications of proteins cited above.展开更多
The Chinese herb Shuyusan, whose main constituent is jasminoidin, has been shown to protect SH-SY5Y cells against corticosterone-induced damage. SH-SY5Y cells injured by 400 μmol/L cor- ticosterone were treated with ...The Chinese herb Shuyusan, whose main constituent is jasminoidin, has been shown to protect SH-SY5Y cells against corticosterone-induced damage. SH-SY5Y cells injured by 400 μmol/L cor- ticosterone were treated with 5 and 30 μg/mL Shuyusan-containing serum. Results revealed that Shuyusan-containing serum elevated the survival rate of SH-SY5Y cells, reduced Bax expression, increased Bcl-2 expression, markedly elevated brain-derived neurotrophic factor mRNA expression, and blocked cell apoptosis. Moreover, the effect of high-dose （30 μg/mL） Shuyusan-containing se- rum was more remarkable. Therefore, Shuyusan-containing serum appears to protect SH-SY5Y cells against corticosterone-induced impairment by adjusting the expression of apoptosis-associ- ated proteins and brain-derived neurotrophic factor. Moreover, high-dose Shuyusan-containing se- rum has a protective effect on high-dose corticosterone-induced impairment.展开更多
基金This is supported by the Youth Science Foundation of Guangxi Medical University(GXMUYSF202127)。
文摘Obejective:To explore the establishment of an oxygen glucose deprivation/reperfusion model of senescent SH-SY5Y cells.Methods:SH-SY5Y cells were randomly divided into control(D-galactose 0 mmol/L group),D-galactose(25 mmol/L,50 mmol/L,100 mmol/L,200 mmol/L,400 mmol/L)groups,and treated with corresponding concentrations of D-galactose for 48 h.The changes of cell morphology,β-galactosidase,the cell morphology,β-galactosidase activity by microscopic observation,cell proliferation rate by EdU kit and cell survival rate by CCK-8 assay were used to determine the decaying concentration of D-galactose and to establish the senescence model.The senescent SH-SY5Y cells were randomly divided into control group(oxygen glucose deprivation without treatment group),oxygen glucose deprivation treatment(0.5 h,1 h,1.5 h,2 h)group,followed by re-glucose reoxygenation for 24 h,and CCK-8 assay for the survival rate of senescent SH-SY5Y cells.Results:There were no significant changes in cell morphology and β-gal activity in the 25 mmol/L and 50 mmol/L groups compared with the control group(P>0.05),cytosolic hypertrophy was seen in the cells of the 100 mmol/L group,chromatin fixation in the cells of the 200 mmol/L group,and massive vacuolization in the cells of the 400 mmol/L group;the positive rate ofβ-galactosidase staining in the cells of the(100-400 mmol/L)group was significantly higher compared with the control group(P<0.05),with little difference between the 100 mmol/L and 200 mmol/L groups(P>0.05);the cell proliferation ability of the(100-400 mmol/L)group was significantly decreased in a concentration-dependent manner(P<0.05);the cell survival rate was decreased in a concentration-dependent manner(P<0.05),with IC_(50) between 100 mmol/L and 200 mmol/L.The survival of senescent SH-SY5Y cells showed a time-dependent decrease in oxygen-glucose deprivation(P<0.05),with an IC_(50) close to 1 h.Conclusion:D-gal concentration of 100 mmoL/L and 48 h of cell action could establish a survival rate of about 50%of senescent SH-SY5Y cells,and oxygen glucose deprivation of senescent SH-SY5Y cells for 1 h and reperfusion for 24 h could establish an oxygen glucose deprivation/reperfusion model of senescent SH-SY5Y cells with a survival rate close to 50%.
基金supported by the National Natural Science Foundation of China,No.82001604Guizhou Provincial Higher Education Science and Technology Innovation Team,No.[2023]072+1 种基金Guizhou Province Distinguished Young Scientific and Technological Talent Program,No.YQK[2023]040Guizhou Provincial Basic Research Program(Natural Science),No.ZK[2021]-368(all to LXiong),and Zunyi City Innovative Talent Team Training Plan,No.[2022]-2.
文摘Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ischemic encephalopathy.The therapeutic potential of human placental chorionic plate-derived mesenchymal stem cells for various diseases has been explored.However,the potential use of human placental chorionic plate-derived mesenchymal stem cells for the treatment of neonatal hypoxic-ischemic encephalopathy has not yet been investigated.In this study,we injected human placental chorionic plate-derived mesenchymal stem cells into the lateral ventricle of a neonatal hypoxic-ischemic encephalopathy rat model and observed significant improvements in both cognitive and motor function.Protein chip analysis showed that interleukin-3 expression was significantly elevated in neonatal hypoxic-ischemic encephalopathy model rats.Following transplantation of human placental chorionic plate-derived mesenchymal stem cells,interleukin-3 expression was downregulated.To further investigate the role of interleukin-3 in neonatal hypoxic-ischemic encephalopathy,we established an in vitro SH-SY5Y cell model of hypoxic-ischemic injury through oxygen-glucose deprivation and silenced interleukin-3 expression using small interfering RNA.We found that the activity and proliferation of SH-SY5Y cells subjected to oxygen-glucose deprivation were further suppressed by interleukin-3 knockdown.Furthermore,interleukin-3 knockout exacerbated neuronal damage and cognitive and motor function impairment in rat models of hypoxic-ischemic encephalopathy.The findings suggest that transplantation of hpcMSCs ameliorated behavioral impairments in a rat model of hypoxic-ischemic encephalopathy,and this effect was mediated by interleukin-3-dependent neurological function.
文摘A plasmid for cytoglobin expression, pAcGFP1-Cl-cytoglobin, was transfected into SH-SY5Y cells. Cobalt chloride was used to establish a model of hypoxia. Western blotting indicated that cytoglobin was overexpressed and there was low expression of hypoxia-inducible factor-la in SH-SY5Y cells after transfection. Following cobalt chloride-induced hypoxia, cytoglobin and hypoxia-inducible fac- tor-la expression gradually increased in SH-SY5Y cells. Flow cytometry showed that with increas- ing duration of hypoxia, the proportion of normal cells significantly diminished in the transfected and non-transfected groups. The proportion of cells in the early stages of apoptosis increased. However, the proportion of apoptotic cells was significantly lower in the transfected group compared with the non-transfected group. These results demonstrate that cytoglobin and hypoxia-inducible factor-la are strongly up-regulated by hypoxia, and that there is a strong relationship between hy- poxia-inducible factor-la and cytoglobin during hypoxic injury.
文摘In this study, we examined the effects of neuroglobin gene （Ngb） transfection into SH-SY5Y cells, using ultrasound-targeted microbubble destruction （UTMD）, on cobalt chloride-induced hypoxia. With an ultrasound intensity of 0.8 W/cm2, a 60-second exposure duration, 50% duty cycle, and 20% microbubble concentration, pAcGFP1-C1-Ngb-transfected cells exhibited the highest cell viability and transfection efficiency. The efficiency of plasmid delivery was significantly higher with UTMD than transfection with plasmid alone, transfection with plasmid using microbubbles, or transfection of plasmid by ultrasound. In addition, during cobalt chloride-induced hypoxia, caspase-3 activity in pAcGFP1-C1-Ngb-transfected cells was significantly lower than in untransfected cells. Ngb protein and mRNA expression were significantly higher in cells transfected by UTMD than in cells transfected with the other methods. These results demonstrate that UTMD can very efficiently mediate exogenous gene delivery, and that Ngb overexpression protects cells against cobalt chloride-induced hypoxia.
基金the Capital Specific Clinical Medical Subject of Beijing Science and Technology Commission,No.Z090507017709030
文摘In vitro cultured human neuroblastoma SH-SY5Y cells were pretreated with 50 or 5 ug/mL geniposide for 12 hours and exposed to 400 umol/L corticosterone. Corticosterone exposure in cultures not pretreated with geniposide resulted in inhibited cell growth, reduced cell survival, and increased P53 and P21 protein expression. However, in geniposide pretreated SH-SY5Y cells, cell viability and the number of cells in the G2 phase of the cell cycle were significantly increased, P21 and P53 protein expression was reduced, and cell apoptosis was inhibited following corticosterone exposure. These results indicate that geniposide can protect SH-SY5Y cells against high-dose corticosterone-induced injury.
文摘The histone deacetylase inhibitor, trichostatin A, is used to treat Alzheimer’s disease and can improve learning and memory but its underlying mechanism of action is unknown. To determine whether the therapeutic effect of trichostatin A on Alzheimer’s disease is associated with the nuclear factor erythroid 2-related factor 2(Nrf2) and Kelch-like epichlorohydrin-related protein-1(Keap1) signaling pathway, amyloid β-peptide 25–35(Aβ25–35) was used to induce Alzheimer’s disease-like pathological changes in SH-SY5 Y neuroblastoma cells. Cells were then treated with trichostatin A. The effects of trichostatin A on the expression of Keap1 and Nrf2 were detected by real-time quantitative polymerase chain reaction, western blot assays and immunofluorescence. Total antioxidant capacity and autophagy activity were evaluated by total antioxidant capacity assay kit and light chain 3-I/II levels, respectively. We found that trichostatin A increased cell viability and Nrf2 expression, and decreased Keap1 expression in SH-SY5 Y cells. Furthermore, trichostatin A increased the expression of Nrf2-related target genes, such as superoxide dismutase, NAD(P)H quinone dehydrogenase 1 and glutathione S-transferase, thereby increasing the total antioxidant capacity of SH-SY5 Y cells and inhibiting amyloid β-peptide-induced autophagy. Knockdown of Keap1 in SH-SY5 Y cells further increased trichostatin A-induced Nrf2 expression. These results indicate that the therapeutic effect of trichostatin A on Alzheimer’s disease is associated with the Keap1-Nrf2 pathway. The mechanism for this action may be that trichostatin A increases cell viability and the antioxidant capacity of SH-SY5 Y cells by alleviating Keap1-mediated inhibition Nrf2 signaling, thereby alleviating amyloid β-peptide-induced cell damage.
文摘To investigate the molecular mechanism underlying the neuroprotective effect of lithium on cells, in this study, we exposed SH-SY5Y cells to 0.5 mmol/L lithium carbonate （Li2CO2） for 25-50 weeks and then detected the expression levels of some neurobiology related genes and post-translational modifications of stress proteins in SH-SYSY cells, cDNA arrays showed that pyruvate kinase 2 （PKM2） and calmodulin 3 （CaM 3） expression levels were significantly down-regulated, phosphatase protein PP2A expression was lightly down-regulated, and casein kinase II （CK2）, threonine/tyrosine phosphatase 7 （PYST2）, and dopamine beta-hydroxylase （DBH） expression levels were significantly up-regulated. Besides, western blot analysis of stress proteins （HSP27, HSP70, GRP78 and GRP94） showed an over-expression of two proteins： a 105 kDa protein which is a hyper-phosphorylated isoform of GRP94, and a 108 kDa protein which is a phosphorylated tetramer of HSP27. These results suggest that the neuroprotective effects of lithium are likely related to gene expressions and post-translational modifications of proteins cited above.
基金funded by a grant from the Scientific Research Key Project of Armed Forces during the "12 th Five-Year Plan" Period,No.CWS12J129
文摘The Chinese herb Shuyusan, whose main constituent is jasminoidin, has been shown to protect SH-SY5Y cells against corticosterone-induced damage. SH-SY5Y cells injured by 400 μmol/L cor- ticosterone were treated with 5 and 30 μg/mL Shuyusan-containing serum. Results revealed that Shuyusan-containing serum elevated the survival rate of SH-SY5Y cells, reduced Bax expression, increased Bcl-2 expression, markedly elevated brain-derived neurotrophic factor mRNA expression, and blocked cell apoptosis. Moreover, the effect of high-dose （30 μg/mL） Shuyusan-containing se- rum was more remarkable. Therefore, Shuyusan-containing serum appears to protect SH-SY5Y cells against corticosterone-induced impairment by adjusting the expression of apoptosis-associ- ated proteins and brain-derived neurotrophic factor. Moreover, high-dose Shuyusan-containing se- rum has a protective effect on high-dose corticosterone-induced impairment.
