BACKGROUND:Hydroxy safflower yellow A has anti-ischemia,anti-oxidation,anti-thrombotic and anti-inflammatory effects.Whether it affects neuronal pyroptosis after glucose-oxygen deprivation/reglucose-reoxygenation is still unclear. OBJECTIVE:To investigate the protective effect of hydroxy safflower yellow A on neuronal pyroptosis and its mechanism. METHODS:HT22 cells in logarithmic growth phase were randomly divided into five groups:normal group,model group,hydroxy safflower yellow A group,colivelin group,and colivelin+hydroxy safflower yellow A group.HT22 cells were treated with glucose-oxygen deprivation/reglucose-reoxygenation to establish neuronal pyroptosis model,and then treated with STAT3 agonist Colivelin and hydroxy safflower yellow A.JC-1 probe was employed to assess changes in mitochondrial membrane potential.Reactive oxygen species kit was used to determine the content of reactive oxygen species in cells.GSDMD/TUNEL staining was conducted to observe cell pyroptosis.Immunofluorescence analysis was performed to detect STAT3 and GSDMD protein expression.RT-PCR was utilized for assessing mRNA expression levels of STAT3,NLRP3,and Caspase-1.Western blot assay was utilized to measure the protein expression levels of p-STAT3,NLRP3,GSDMD,Cleaved-caspase-1,and interleukin-1β. RESULTS AND CONCLUSION:(1)Compared with the normal group,the number of pyroptotic cells increased in HT22 cells in the model group along with a significant increase in protein expression levels of p-STAT3,NLRP3,Cleaved-caspase-1,GSDMD,and interleukin-1β.Compared with the model group,the number of pyroptotic cells reduced,and the expression of pyroptosis-related proteins significantly decreased in the hydroxy safflower yellow A group.(2)In comparison with the model group,pyroptosis worsened in the colivelin group where mitochondrial membrane potential decreased along with elevated reactive oxygen species content and increased mRNA expression levels of STAT3,NLRP3,and Caspase-1,as well as increased protein expression levels of p-STAT3,NLRP3,GSDMD,Cleaved-caspase-1,and interleukin-1β.Compared with the Colivelin group,above indexes were improved in the colivelin+hydroxy safflower yellow A group.These results suggest that hydroxy safflower yellow A plays a neuroprotective role through STAT3 signaling pathway to inhibit HT22 pyroptosis after glucose-oxygen deprivation/reglucose-reoxygenation treatment.

Objective:To explore effects and mechanism of Hydroxylsafflower Yellow A(HSYA)on expression of NLRP3 in glial cells after cerebral ischemic injury.Methods:A middle cerebral artery occlusion/reperfusion(MCAO/R)model was established in male SD rats.After successfully modeling for 24 h,Longa scoring and corner test were used to evaluate degree of neurological dys-function.Western blot and immunofluorescence were used to detect expressions of JAK2/STAT3 molecules and NLRP3,ELISA was used to measure IL-1β,IL-6 and TNF-α levels.A glucose-oxygen deprivation/reperfusion(OGD/R)model was established in microg-lia,and JAK2 and STAT3 inhibitor AG490 was used to further verify action of HSYA on NLRP3.Results:Compared with sham group,neurological dysfunction aggravated in MCAO/R group(P<0.01),HSYA treatment improved neurological function(P<0.01).Expres-sions of p-JAK2,p-STAT3 and NLRP3 in MCAO/R group were higher than those in the sham group(P<0.01);and HSYA treatment reduced expressions of p-JAK2,p-STAT3 and NLRP3(P<0.01).Levels of inflammatory factors IL-1β,IL-6 and TNF-α were higher in MCAO/R group than sham group(P<0.01),and HSYA inhibited expressions of IL-1β,IL-6 and TNF-α(P<0.01 or P<0.05).In vi-tro experiments showed expressions of p-JAK2,p-STAT3 and NLRP3 in OGD/R group were significantly higher than normal control group(P<0.01),after adding AG490,phosphorylation of JAK2 and STAT3 decreased,NLRP3 expression was inhibited(P<0.01).Inflammatory cytokines IL-1β,IL-6 and TNF-α levels were higher in OGD/R group than normal control group(P<0.01),and HSYA inhibited expressions of IL-1β,IL-6 and TNF-α(P<0.01 or P<0.05).Conclusion:HSYA alleviates brain damage,probably by regu-lating JAK2/STAT3 signaling pathway and inhibiting NLRP3 expression in microglia after cerebral ischemia and hypoxia.

Objective:To explore effects and mechanism of Hydroxylsafflower Yellow A(HSYA)on expression of NLRP3 in glial cells after cerebral ischemic injury.Methods:A middle cerebral artery occlusion/reperfusion(MCAO/R)model was established in male SD rats.After successfully modeling for 24 h,Longa scoring and corner test were used to evaluate degree of neurological dys-function.Western blot and immunofluorescence were used to detect expressions of JAK2/STAT3 molecules and NLRP3,ELISA was used to measure IL-1β,IL-6 and TNF-α levels.A glucose-oxygen deprivation/reperfusion(OGD/R)model was established in microg-lia,and JAK2 and STAT3 inhibitor AG490 was used to further verify action of HSYA on NLRP3.Results:Compared with sham group,neurological dysfunction aggravated in MCAO/R group(P<0.01),HSYA treatment improved neurological function(P<0.01).Expres-sions of p-JAK2,p-STAT3 and NLRP3 in MCAO/R group were higher than those in the sham group(P<0.01);and HSYA treatment reduced expressions of p-JAK2,p-STAT3 and NLRP3(P<0.01).Levels of inflammatory factors IL-1β,IL-6 and TNF-α were higher in MCAO/R group than sham group(P<0.01),and HSYA inhibited expressions of IL-1β,IL-6 and TNF-α(P<0.01 or P<0.05).In vi-tro experiments showed expressions of p-JAK2,p-STAT3 and NLRP3 in OGD/R group were significantly higher than normal control group(P<0.01),after adding AG490,phosphorylation of JAK2 and STAT3 decreased,NLRP3 expression was inhibited(P<0.01).Inflammatory cytokines IL-1β,IL-6 and TNF-α levels were higher in OGD/R group than normal control group(P<0.01),and HSYA inhibited expressions of IL-1β,IL-6 and TNF-α(P<0.01 or P<0.05).Conclusion:HSYA alleviates brain damage,probably by regu-lating JAK2/STAT3 signaling pathway and inhibiting NLRP3 expression in microglia after cerebral ischemia and hypoxia.

BACKGROUND:Ischemic stroke is a serious threat to human health.After ischemia and hypoxia,astrocyte expresses lipocalin-2 in large amounts to aggravate brain injury,but the specific mechanism is not clear.Hydroxysafflor yellow A has anti-ischemia,anti-oxidation,anti-thrombosis and anti-inflammatory effects.However,whether hydroxysafflor yellow A affects the expression of lipocalin-2 in astrocytes after cerebral ischemia and hypoxia and its mechanism are not clear. OBJECTIVE:To investigate the effect and mechanism of hydroxysafflor yellow A on the expression of lipocalin-2 in astrocytes after cerebral ischemia and reperfusion. METHODS:(1)Thirty adult SD rats were randomly divided into three groups:sham operation group,middle cerebral artery occlusion and reperfusion group,and hydroxysafflor yellow A group.The middle cerebral artery occlusion and reperfusion model was established in the latter two groups,and hydroxysafflor yellow A group was intraperitoneally injected with 12 mg/kg hydroxysafflor yellow A after reperfusion.Longa score was used to evaluate the degree of neurological impairment.Infarct volume was determined by TTC staining.JAK2/STAT3 pathway and lipocalin-2 expression were detected by western blot assay and immunofluorescence.Levels of interleukin 1β,interleukin 6 and tumor necrosis factor α were detected by ELISA.(2)Astrocytes were divided into four groups:Normal group,glucose-oxygen deprivation group,hydroxysafflor yellow A group and AG490 group.In the latter three groups,glucose-oxygen deprivation and glucose-oxygen recovery models were established.Astrocytes were treated with 75 μmol/L hydroxysafflor yellow A and 10 μmol/L tyrosine phosphorylation inhibitor AG490 for 8 hours during glucose-oxygen deprivation,respectively.The mechanism of hydroxysafflor yellow A on lipocalin-2 was further verified. RESULTS AND CONCLUSION:(1)Compared with the sham operation group,cerebral infarction was significantly increased in the middle cerebral artery occlusion and reperfusion group,accompanied by aggravated neurological impairment(P<0.01).Hydroxysafflor yellow A treatment could reduce cerebral infarction volume and improve neurological function(P<0.01).(2)The expressions of p-JAK2,p-STAT3 and lipocalin-2 in the middle cerebral artery occlusion and reperfusion group were higher than those in the sham operation group(P<0.01).Hydroxysafflor yellow A treatment reduced the expressions of JAK2,STAT3 and lipocalin-2(P<0.01).(3)The expression levels of interleukin 1β,interleukin-6 and tumor necrosis factor α in the middle cerebral artery occlusion and reperfusion group were higher than those in the sham operation group(P<0.01).Hydroxysafflor yellow A inhibited the expressions of interleukin 1β,interleukin-6 and tumor necrosis factor α(P<0.01).(4)In vitro,the expressions of p-JAK2,p-STAT3 and lipocalin-2 in the glucose-oxygen deprivation group were significantly higher than those in the normal group(P<0.01).After adding AG490,the phosphorylation of JAK2 and STAT3 decreased,and the expression of lipocalin-2 was inhibited(P<0.01).The results suggest that hydroxysafflor yellow A may inhibit the expression of lipocalin-2 in astrocytes after ischemia and hypoxia by regulating the JAK2/STAT3 signaling pathway,thereby reducing brain injury.

Objective:To quantify any correlation between serum levels of omega-3 polyunsaturated fatty acids (ω3-PUFAs) and post-stroke cognitive impairment (PSCI).Methods:The clinical data of 77 patients hospitalized after a first stroke were analyzed. The Minimum Mental State Examination (MMSE) was used to divide them into impaired (PSCI) and unimpaired (non-PSCI) cohorts. The serum levels of ω3-PUFAs, α-linolenic acid (ALA), eieosapentaenoic acid (EPA) and dueosahexenoie acid (DHA) were compared between the two groups and correlated with the individuals′ MMSE scores.Results:The average ALA, EPA, DHA and total ω3-PUFAs levels of PSCI group were in most cases significantly lower than those of the non-PSCI group. Spearman correlation analysis showed that serum DHA level was a weak positive predictor of the MMSE scores (R=0.32, P≤0.05). Logistic regression analysis indicated that low serum DHA level was an independent risk factor for PSCI ( P≤0.01). Conclusions:Cognitively impaired stroke survivors tend to have lower serum ω3-PUFAs levels than those without cognitive impairment. There is a weak positive correlation between serum DHA levels and MMSE scores. Low serum DHA level is an independent risk factor for PSCI. The serum level of ω3-PUFAs is of high value in the auxiliary diagnosis and evaluation of PSCI.