Effects of dexmedetomidine on the oncological behavior of hepatocellular carcinoma and role of Nrf2 / 安徽医科大学学报

Objective @# To investigate the impact of dexmedetomidine on the oncological behavior of hepatocellular carcinoma and explore the role of NF-E2-related factor 2 (Nrf2) at both in vitro and in vivo levels．@*Methods @# In vivo experiment，Male C57BL/6J mice were randomly divided into a control group ( Ctrl group) ，a hepatocellular carcinoma group ( HCC group) ，and a hepatocellular carcinoma + dexmedetomidine group ( HCC + Dex group) . Hepatocellular carcinoma was induced in mice by combining N-Nitrosodiethylamine ( DEN) / carbon tetrachloride ( CCl4 ) ，followed by daily intraperitoneal injection of 10% dexmedetomidine for two weeks．After feeding the mice for one month，the mice were assessed for the quantity and size of liver tumors．The proliferation ability of liver cancer was evaluated using Ki67 immunohistochemistry．Additionally，the expression level of Nrf2 protein in tumor tissue was measured through immunofluorescence．In vitro experiment，Hepa1-6 cells were incubated with different concentrations of dexmedetomidine (0. 1，1，5 nmol /L) for 48 hours to examine their effects．The proliferation， migration and invasion abilities of Hepa1-6 cells were evaluated using the MTT and Transwell methods．The expres- sion level of Nrf2 protein in the Hepa1-6 cells was measured using Western blot and immunofluorescence．Addition- ally，the proliferation ，migration and invasion abilities of cells were assessed after Nrf2 knockdown via si-RNA transfection，in combination with incubation with 1 nmol /L dexmedetomidine for 48 hours. @*Results @#ompared to the HCC group，the anatomical examination results revealed an increase in the number of liver tumors and the lon- gest diameter in the HCC + Dex group (P ＜0. 05) ． Ki67 immunohistochemistry results indicated the number of Ki67 positive cells in liver cancer tissue increased in the HCC + Dex group (P＜0. 01) ．The immunofluorescence assay demonstrated an upregulation of Nrf2 expression level in the HCC + Dex group (P ＜0. 05 ) ． MTT results showed that 1 nmol /L of dexmedetomidine increased the cell viability of Hepa1-6 cells (P＜0. 05) ．Transwell re- sults indicated that 0. 1 ，1 ，and 5 nmol /L of dexmedetomidine enhanced the invasive ability of Hepa1-6 cells， while 0. 1 and 1 nmol /L of dexmedetomidine enhanced the migration ability (P＜0. 05) ．Western blot and immu- nofluorescence results showed an upregulation of Nrf2 expression level in cells after treatment with 1 nmol /L dexme- detomidine (P＜0. 01) ．The Nrf2 expression level of cells was reduced using si-RNA，followed by treatment with 1 nmol /L dexmedetomidine．The results from MTT and Transwell assays revealed a decrease in the viability，invasion and migration ability of Hepa1-6 cells (P＜0. 01) ．@*Conclusion @# Dexmedetomidine may enhance the proliferation， invasion and migration capacity of hepatocellular carcinoma by upregulating the expression of Nrf2 .

Effect and mechanism of dioscin on renal injury in septic rats / 中国药房

OBJECTIVE To investigate the effect of dioscin on renal injury in septic rats and its possible mechanism. METHODS The septic rat model was induced by using cecal ligation and puncture. Sixty model rats were randomly divided into model group （0.5% sodium carboxymethyl cellulose solution）， dioscin low-dose， medium-dose and high-dose groups （30， 60， 120 mg/kg） and dexamethasone group （positive control， 10 mg/kg）， with 12 rats per group； another 12 rats were selected as the sham operation group （0.5% sodium carboxymethyl cellulose solution）. After 15 minutes of modeling， rats in each group were injected with medicine/0.5% sodium carboxymethyl cellulose solution via the tail vein. Twenty-four hours after administration， the levels of creatinine （Cr）， blood urea nitrogen （BUN）， neutrophil gelatinase associated lipocalin （NGAL）， kidney injury molecule-1 （KIM- 1）， interleukin 6 （IL-6）， IL-1β and tumor necrosis factor-α （TNF-α） in serum and malondialdehyde （MDA） in renal tissue， superoxide dismutase （SOD） activity and the protein expressions of nuclear factor E2-related factor 2 （Nrf2）， heme oxygenase-1 （HO-1）， NOD-like receptor protein 3 （NLRP3） were detected； renal histomorphology was observed. RESULTS Compared with model group， pathological injury of renal tissue was improved significantly in dioscin low-dose， medium-dose and high-dose groups； the levels of Cr， BUN， NGAL， KIM-1， IL-6， IL-1β and TNF-α in serum， MDA level and protein expression of NLRP3 in renal tissue were decreased significantly （P＜0.05）； SOD activity in renal tissue， protein expressions of Nrf2 and HO-1 were increased significantly （P＜0.05）， in a dose-dependent manner （P＜0.05）. The pathological damage of renal tissue in the dioscin high-dose group was similar to dexamethasone group， and there was no statistically significant difference in the levels of the above indicators （P＞0.05）. CONCLUSIONS Dioscin can activate the Nrf2/HO-1 signaling pathway to inhibit NLRP3 inflammasome， and realize the inhibition of inflammatory factors and oxidative stress， so as to protect the kidney injury in sepsis.

Effect and mechanism of dioscin on renal injury in septic rats / 中国药房

OBJECTIVE To investigate the effect of dioscin on renal injury in septic rats and its possible mechanism. METHODS The septic rat model was induced by using cecal ligation and puncture. Sixty model rats were randomly divided into model group （0.5% sodium carboxymethyl cellulose solution）， dioscin low-dose， medium-dose and high-dose groups （30， 60， 120 mg/kg） and dexamethasone group （positive control， 10 mg/kg）， with 12 rats per group； another 12 rats were selected as the sham operation group （0.5% sodium carboxymethyl cellulose solution）. After 15 minutes of modeling， rats in each group were injected with medicine/0.5% sodium carboxymethyl cellulose solution via the tail vein. Twenty-four hours after administration， the levels of creatinine （Cr）， blood urea nitrogen （BUN）， neutrophil gelatinase associated lipocalin （NGAL）， kidney injury molecule-1 （KIM- 1）， interleukin 6 （IL-6）， IL-1β and tumor necrosis factor-α （TNF-α） in serum and malondialdehyde （MDA） in renal tissue， superoxide dismutase （SOD） activity and the protein expressions of nuclear factor E2-related factor 2 （Nrf2）， heme oxygenase-1 （HO-1）， NOD-like receptor protein 3 （NLRP3） were detected； renal histomorphology was observed. RESULTS Compared with model group， pathological injury of renal tissue was improved significantly in dioscin low-dose， medium-dose and high-dose groups； the levels of Cr， BUN， NGAL， KIM-1， IL-6， IL-1β and TNF-α in serum， MDA level and protein expression of NLRP3 in renal tissue were decreased significantly （P＜0.05）； SOD activity in renal tissue， protein expressions of Nrf2 and HO-1 were increased significantly （P＜0.05）， in a dose-dependent manner （P＜0.05）. The pathological damage of renal tissue in the dioscin high-dose group was similar to dexamethasone group， and there was no statistically significant difference in the levels of the above indicators （P＞0.05）. CONCLUSIONS Dioscin can activate the Nrf2/HO-1 signaling pathway to inhibit NLRP3 inflammasome， and realize the inhibition of inflammatory factors and oxidative stress， so as to protect the kidney injury in sepsis.

Effect of Sargassum and Glycyrrhizae Radix et Rhizoma Incompatible Pair with Haizao Yuhutang on Oxidative Stress in Liver of Goiter Rats / 中国实验方剂学杂志

ObjectiveTo observe the effects of Sargassum and Glycyrrhizae Radix et Rhizoma incompatible pair with the Haizao Yuhutang （HYT） on oxidative stress in the liver of goiter rats under the condition of 2 times the dose limit of the Pharmacopoeia of the People's Republic of China 2020. MethodA total of 128 male Wistar rats were randomly divided into a blank group， a model group， a euthyrox group （20 μg·kg-1）， a HYT group （12.06 g·kg-1）， a HYT without Sargassum （HYT-H） group （9.90 g·kg-1）， a HYT without Glycyrrhizae Radix et Rhizoma （HYT-G） group （10.26 g·kg-1）， a HYT without Sargassum and Glycyrrhizae Radix et Rhizoma （HYT-HG） group （8.10 g·kg-1）， and a Sargassum and Glycyrrhizae Radix et Rhizoma （HG） group （3.96 g·kg-1）. The blank group was given deionized water by gavage， and the others were given propylthiouracil （PTU） to replicate the goiter pathological model. Euthyrox was taken as a positive control drug， and the rest of the Chinese medicine groups were given the corresponding decoction by gavage， the material was collected 12 hours after the last dose. The serum levels of aspartate aminotransferase （AST）， alanine aminotransferase （ALT）， alkaline phosphatase （ALP）， and the contents of superoxide dismutase （SOD）， glutathione peroxidase （GSH-Px）， malondialdehyde （MDA）， reactive oxygen species （ROS） in liver tissue were detected in each group. The pathological changes in the liver were observed via hematoxylin-eosin （HE） staining. Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was utilized to detect the mRNA expressions of Kelch-like Ech-associated protein 1 （Keap1）， nuclear factor E2-related factor 2 （Nrf2）， heme oxygenase-1 （HO-1）， p53 and Caspase-3 in liver tissues. Western blot was adopted to detect the protein expressions of Nrf2 and HO-1 in liver tissues in oxidative stress-related signaling pathways. ResultCompared with control group， the model group showed significantly increased serum ALT level and contents of MDA and ROS in liver tissues （P<0.05， P<0.01）， significantly reduced activities of SOD and GSH-Px in the liver （P<0.01）， significantly increased mRNA expression of Keap1 （P<0.01）， and significantly decreased mRNA and protein expressions of Nrf2 and HO-1 （P<0.05， P<0.01）. Compared with the model group， the HYT group manifested significantly reduced serum levels of AST， ALT， and ALP （P<0.05， P<0.01）， significantly reduced contents of MDA and ROS in liver tissue （P<0.01）， significantly increased the activities of SOD and GSH-Px （P<0.01）， significantly decreased mRNA expressions of Keap1， p53， and Caspase-3 （P<0.01）， and significantly increased mRNA and protein expressions of Nrf2 and HO-1 （P<0.05， P<0.01）. ConclusionUnder the condition of 2 times the dose limit of the Pharmacopoeia of the People's Republic of China 2020， Sargassum and Glycyrrhizae Radix et Rhizoma incompatible pair with the HYT on oxidative stress in the liver of goiter rats had different effects. The HYT that contains Sargassum and Glycyrrhizae Radix et Rhizoma has a protective effect on the liver of goiter rats， and the effect is better than that of the HG group， the euthyrox group， and the incomplete groups. Its mechanism may be related to activating the Nrf2/HO-1 signaling pathway to alleviate liver oxidative stress and inhibiting the p53/Caspase-3 signaling pathway to reduce hepatocyte apoptosis.

Experimental Study on Regulation of Nrf2/HO-1 by Linalool to Inhibit Hepatic Injury Induced by Aflatoxin B1 / 中国实验方剂学杂志

ObjectiveTo investigate the effect of linalool against acute liver injury induced by aflatoxin B1（AFB1） in rats and explore its protective mechanism. MethodTwenty male SPF SD rats were randomly divided into three groups： Control （n=6）， AFB1 （n=7）， and linalool （n=7） groups. Linalool solution （200 mg·kg-1） was administered preventatively for 14 days， while the control and AFB1 groups intragastrically received an equivalent volume of double distilled water. After preventative administration of linalool， AFB1 solution （1 mg·kg-1， dissolved in saline） was intraperitoneally injected for two consecutive days to induce acute liver injury in rats. Samples were collected and processed 14 days after model establishment. Pathological changes in liver tissue of rats were observed using Hematoxylin-eosin（HE） staining and Masson staining. Biochemical detection was performed to measure the levels of alanine transaminase（ALT）， aspartate transaminase（AST）， γ-glutamyl transferase（GGT）， lactate dehydrogenase（LDH）， alkaline phosphatase（ALP）， total bilirubin（TBil）， direct bilirubin（DBil）， indirect bilirubin（IBil）， malondialdehyde（MDA）， superoxidedismutase（SOD）， catalase（CAT） ， glutathione（GSH）， Fe3+， and Fe2+ in the liver tissue. Western blot was adopted to assess protein expression levels of nuclear factor-erythroid 2-related factor 2（Nrf2） and heme oxygenase-1（HO-1）. Molecular docking was performed to verify the binding between linalool and key proteins of the Nrf2/HO-1 signaling pathway. Molecular dynamics techniques were used to confirm the stability and affinity of linalool binding with key proteins of the Nrf2/HO-1 signaling pathway. ResultPathological results showed that compared to that in the AFB1 group， the liver structure in the linalool group tended to be normal， with a significant decrease in blue collagen fibers. The linalool group exhibited significantly reduced levels of ALT， AST， GGT， LDH， ALP， TBil， DBil， and IBil （P<0.01）， Fe3+ and Fe2+ content， and oxidative stress marker MDA （P<0.01）. The levels of antioxidants SOD， CAT， and GSH significantly increased （P<0.01）. Molecular docking showed a molecular docking energy between linalool and Nrf2 and HO-1 targets of -5.495 6 and -5.199 4 kcal·mol-1（1 cal≈4.186 J）， respectively. Molecular dynamics results indicated strong affinity in the binding of linalool with Nrf2 and HO-1. Western blot revealed a significant increase in Nrf2 protein expression （P<0.05） and a decrease in HO-1 protein expression （P<0.01） in the linalool group. ConclusionLinalool may protect against AFB1-induced acute liver injury by modulating the Nrf2/HO-1 ferroptosis signaling pathway to inhibit liver cell ferroptosis and regulate hepatic oxidative stress levels.

The Role of Nrf2 in Exercise Improving of NAFLD / 生物化学与生物物理进展

In cardiovascular disorders, neurological diseases, and chronic metabolic diseases, the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway is essential for maintaining cell homeostasis. According to studies, boosting Nrf2 expression can be used to cure or prevent chronic diseases that are characterized by oxidative stress, inflammation, and mitochondrial dysfunction. Nonalcoholic fatty liver disease (NAFLD) is a chronic metabolic liver disease characterized by hepatic steatosis brought on by a number of causes other than alcohol. In recent years, its incidence has gradually risen across the globe. According to relevant studies, NAFLD and the Nrf2 signaling pathway are tightly connected. Inhibiting lipid production and metabolism-related enzymes, repairing impaired liver metabolism, and lowering hepatic lipid storage are all possible with Nrf2 activation. Exercise is a powerful tool for treating and preventing NAFLD. However, exercise type, exercise intensity, environment, and exhaustion all have an impact on the Nrf2 signaling pathway. By activating Nrf2, exercise can lessen liver inflammation, oxidative stress, endoplasmic reticulum stress, and insulin resistance, and ameliorate liver damage to improve NAFLD. The activation of Nrf2 signaling pathway, its associated mechanism of controlling antioxidation, and the impact of exercise on the Nrf2 signaling pathway are all explained in this work. Based on the pathogenesis of NAFLD, this article examines the connection between exercise, Nrf2, and NAFLD, and the current state of knowledge regarding Nrf2’s role in the amelioration of NAFLD through exercise. It offers a theoretical frame of reference for future research into how Nrf2 might be used to improve NAFLD.

Saponin from Panax japonicus IVa alleviates lipopolysaccharide-induced acute lung injury in rats via its anti-inflammatory and antioxidative ef-fects / 中国病理生理杂志

AIM:To investigate the effects of saponin from Panax japonicus IVa(SPJ IVa)on acute lung inju-ry in rats and to explore its possible protective mechanism.METHODS:Sixty SD rats were randomly divided into four groups,15 rats in each group:the control group,model group,low-dose SPJ IVa group,and high-dose SPJ IVa group.A rat model of ALI was established via intratracheal instillation of lipopolysaccharide(LPS,2 mg/kg).Rats in the low-and high-dose SPJ IVa groups were intraperitoneally injected with 15 and 45 mg/kg SPJ IVa,respectively,30 min after model-ing.Serum,bronchoalveolar lavage fluid(BALF),and lungs were collected 24 h after modeling.Pathomorphological changes in lung tissues were assessed using HE staining.The wet weight/dry weight ratio of lung tissues was measured us-ing the weighing method,whereas ELISA was used to measure the levels of interleukin-1β(IL-1β),IL-6,and tumor ne-crosis factor-α(TNF-α)in the serum and BALF.The levels of malondialdehyde(MDA),superoxide dismutase(SOD),and glutathione(GSH)were assessed using the kit method.Cell apoptosis in lung tissues was evaluated by immunohisto-chemical staining of cleaved caspase-3 and TUNEL.Western blot was used to measure the expression of nuclear factor E2-related factor 2(Nrf2),heme oxygenase-1(HO-1),nuclear factor-κB(NF-κB)p65,and Toll-like receptor 4(TLR4)in lung tissues.RESULTS:Compared with control group,the lung tissues of the model group were significantly damaged,and the lung injury scores(0.21±0.22 vs 2.98±0.46)and lung wet/dry weight ratios(3.09±0.41 vs 6.36±0.61)were significantly increased(P<0.01).Compared with model group,the lung injury scores(1.80±0.31 and 1.05±0.25 vs 2.98±0.46)and lung wet/dry weight ratios(5.25±0.44 and 3.89±0.35 vs 6.36±0.61)in low-and high-dose SPJ IVa groups were significantly reduced(P<0.01).The administration of LPS resulted in elevated levels of pro-inflammatory cy-tokines(IL-1β,IL-6,and TNF-α)as well as the oxidative marker MDA in both serum and BALF(P<0.01).Additional-ly,it led to a decrease in antioxidant markers SOD and GSH(P<0.01).However,treatment with both low and high doses of SPJ IVa effectively attenuated the LPS-induced production of pro-inflammatory factors and oxidative markers MDA(P<0.01),while also increasing SOD and GSH levels(P<0.05 or P<0.01).In the model group,evident apoptosis was ob-served in lung tissues,whereas treatment with low and high doses of SPJ IVa significantly suppressed TUNEL-positive cells and the expression of cleaved caspase-3(P<0.01).The expression levels of Nrf2,HO-1,NF-κB p65,and TLR4 in lung tissues were significantly higher in the model group than in the control group(P<0.01);in turn,after treatment with low and high doses of SPJ IVa,Nrf2 and HO-1 were further upregulated(P<0.01),whereas NF-κB p65 and TLR4 were downregulated(P<0.01).CONCLUSION:The inhibitory effect of SPJ IVa on LPS-induced ALI in rats may be attribut-ed to its ability to suppress the TLR4/NF-κB-and Nrf2/HO-1-mediated inflammatory response and oxidative stress.

Effects of total flavonoids of Pterocarya hupehensis Skan on migration,invasion and ferroptosis of non-small-cell lung cancer A549 cells / 中国病理生理杂志

AIM:To investigate the impact of total flavonoids of Pterocarya hupehensis Skan(PHSTF)on the migration,invasion,and ferroptosis of non-small-cell lung cancer A549 cells.METHODS:The A549 cells were divided into control group,low-,medium-and high-dose(100,150 and 200 μg/mL)PHSTF groups,ferroptosis inhibitor liprox-statin-1(Lip-1)group,and high-dose PHSTF combined with Lip-1 group,each cultured in corresponding media.Cell via-bility was assessed using the CCK-8 assay,while cell migration and invasion abilities were determined through scratch and Transwell assays.Cell lipid peroxidation levels were measured using the glutathione(GSH)assay kit.RT-qPCR was em-ployed to assess the mRNA expression of solute carrier family 7 member 11(SLC7A11)and glutathione peroxidase 4(GPX4),while Western blot was utilized to examine the protein expression of SLC7A11,GPX4,Kelch-like epichlorohy-drin-associated protein-1(Keap-1),nuclear factor E2-related factor 2(Nrf2)and heme oxygenase-1(HO-1).RE-SULTS:Compared with control group,PHSTF significantly diminished the viability of A549 cells in a time-and dose-de-pendent manner(P＜0.01),and the cell migration and invasion were also reduced(P＜0.01),along with a significant de-crease in GSH level(P＜0.01).Treatment with PHSTF inhibited the mRNA and protein expression levels of ferroptosis-re-lated proteins,including SLC7A11 and GPX4(P＜0.01),suppressed the protein expression of Nrf2 and HO-1(P＜0.01),and enhanced the expression of Keap-1(P＜0.01).The Lip-1 partially restored the decrease in cell viability in-duced by PHSTF(P＜0.01),significantly up-regulated the protein expression levels of SLC7A11,GPX4,Nrf2 and HO-1,and suppressed the protein expression of Keap-1(P＜0.01).CONCLUSION:Total flavonoids of Pterocarya hupehen-sis Skan can inhibit the migration and invasion of non-small-cell lung cancer A549 cells,and induce the cell ferroptosis by regulating the Keap-1/Nrf2/HO-1 pathway.

Acacetin ameliorates alveolar epithelial cell damage caused by Streptococcus pneumoniae infection by regulating Sirt1-mediated AMPK/Nrf2 signaling pathway / 中国免疫学杂志

Objective:To investigate effect of acacetin on alveolar epithelial cell damage caused by Streptococcus pneumoniae(SP)infection by regulating sirtuin 1(Sirt1)-mediated 5'-AMP activated protein kinase(AMPK)/nuclear factor erythroid-2 related factor 2(Nrf2)signaling pathway.Methods:Alveolar epithelial cells A549 cultured in vitro were infected with SP to establish a cell damage model.After treatment with acacetin at final concentrations of 0,5,25,50,100,150,200 μmol/L,CCK-8 was performed to detect cell viability of each treatment group and optimal concentration of acacetin was screened.A549 cells cultured in vitro were ran-domly separated into five groups:control group,model group,acacetin(150 μmol/L)group,EX527(Sirt1 inhibitor,40 μmol/L)group,acacetin(150 μmol/L)+EX527(40 μmol/L)group,control group was not treated,other groups were infected with SP to establish a cell damage model,and then treated with 150 μmol/L acacetin and 40 μmol/L EX527,CCK-8 and flow cytometry were performed to measure cell viability and apoptosis rate in each group;kits were performed to measure levels of reactive oxygen species(ROS),superoxide dismutase(SOD),malondialdehyde(MDA),lactate dehydrogenase(LDH)and IL-10,IL-1β,TNF-α levels of cells in each group;Western blot was performed to measure proliferation-related proteins Ki-67,proliferating cell nuclear antigen(PCNA),apoptosis-related proteins caspase-9,Bax,Sirt1 and AMPK/Nrf2 signaling pathway proteins p-AMPK/AMPK,Nrf2 expres-sions of cells in each group.Results:Model group had decreased A549 cell viability,SOD and IL-10 levels,p-AMPK/AMPK,Sirt1,Nrf2,Ki-67 and PCNA protein expressions(P<0.05),and increased apoptosis rate,MDA,LDH,ROS,IL-1β and TNF-α levels than control group(P<0.05).Compared with model group and acacetin+EX527 group,acacetin group had increased A549 cell viability,SOD and IL-10 levels,p-AMPK/AMPK,Sirt1,Nrf2,Ki-67 and PCNA protein expressions(P<0.05),and decreased apoptosis rate,MDA,LDH,ROS,IL-1β and TNF-α levels(P<0.05);EX527 group had decreased A549 cell viability,SOD and IL-10 levels,p-AMPK/AMPK,Sirt1,Nrf2,Ki-67 and PCNA protein expressions(P<0.05),and increased apoptosis rate,MDA,LDH,ROS,IL-1β and TNF-α levels(P<0.05).Conclusion:Acnestin can activate AMPK/Nrf2 signaling by up-regulating Sirt1 expression,thereby promoting secretion of anti-inflammatory factors,reducing production of ROS and pro-inflammatory factors,reducing inflammation and oxidative stress,and finally alleviating neuronal damage.

Effects of casticin on lipopolysaccharide-induced BEAS-2B cell damage and NF-κB-Keap1-Nrf2/ARE pathway / 中国免疫学杂志

Objective:To investigate the effects of casticin(CAS)on lipopolysaccharide-induced injury of human normal lung epithelial cells(BEAS-2B)and NF-κB-Keap1-Nrf2/ARE pathway.Methods:BEAS-2B cells were pretreated with different concentra-tions of casticin for 1 h,2 h and 4 h,and then treated with 1 μg/ml liposolysaccharide to construct cell damage model.The contents of inflammatory factors in cell supernatant was detected by ELISA to screen the optimal concentration and time of casticin.The cells were divided into normal group,model group,Casticin group,ML385 group,Casticin+ML385 group and dexamethasone group.Cell apoptosis was detected by flow cytometry,the concentrations of inflammatory factors were detected by ELISA,and the levels of NF-κB p65,p-NF-κB p65,Keap1,Nrf2 and Nrf2 protein in cell nucleus were detected by Western blot.Results:The optimal concentration of CAS was 10 μmol/L and the optimal time was 2 h.Compared with normal group,the apoptosis rate,the contents of inflammatory fac-tors,p-NF-κB p65 and Keap1 protein expression levels in model group were significantly increased(P<0.01),and Nrf2 protein expression levels in cells and nuclei were significantly decreased(P<0.01).Compared with model group,apoptosis rate and contents of inflammatory factors in casticin group and dexamethasone group were significantly decreased(P<0.01),protein expression levels of p-NF-κB p65 and Keap1 in Casticin group were significantly decreased(P<0.01).The expression level of Nrf2 protein in cells and nuclei was significantly increased(P<0.01).Compared with ML385 group,apoptosis rate,inflammatory factors contents,p-NF-κB p65 and Keap1 protein expression levels in Casticin+ML385 group were significantly decreased(P<0.01),the expression level of Nrf2 protein in cells and nuclei were significantly increased(P<0.01).Conclusion:CAS can inhibit cell inflammation induced by lipo-polysaccharide by regulating NF-κB-Keap1-Nrf2/ARE signaling pathway.

Exploring the therapeutic effect of licorice zinc on chloasma mice via the Nrf-2/HO-1 pathway / 中国比较医学杂志

Objective To investigate the therapeutic effect of licorice zinc on melasma.Methods Thirty-six BALB/c mice were equally divided into blank group,model group,licorzinc low-dose group,licorzine medium-dose group,licorzinc high-dose group and tranexamic acid group.Melasma was induced by 100 mJ/cm2 UVB irradiation combined with 15 mg/kg progesterone injection.Mice were treated with tranexamic acid(0.065 g/kg)and low(0.65 g/kg),medium(1.3 g/kg),or high(2.6 g/kg)doses of zinc licorice for 14 days.Skin was taken for HE and Masson-Fontana staining and measurement of SOD,MDA,GSP-Px,TNF-α,IL-1β,IL-6,plasma protein Nrf-2,nuclear protein Nrf-2 and HO-1 expression levels.Results Compared with model group,high-dose licorice zinc group showed decreased melanocyte formation,collagen cell necrosis,and inflammatory infiltration(P＜0.01);decreased MDA,IL-6,IL-1β,TNF-α and plasma protein Nrf-2 expression(P＜0.01);and increased GSP-Px,SOD and nuclear protein Nrf-2 and HO-1 expression(P＜0.01).Conclusions Zinc licorice activates the Nrf-2/HO-1 pathway to initiate high expression of HO-1,SOD and GSP-Px and fight oxidative stress,thereby reducing melanogenesis.

Mechanism on Banxia Xiexintang Inducing Ferroptosis in Gastric Cancer Cells Based on Nrf2/GPX4 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo observe the effect of Banxia Xiexintang （BXT） on the proliferation of human gastric cancer HGC-27， MKN-45， and AGS cells and its mechanism. MethodCell counting kit-8 （CCK-8） was used to detect the effects of different concentrations of BXT-containing serum （5%， 10%， and 20%） on the proliferation of HGC-27， MKN-45， and AGS cells. A mitochondrial membrane potential probe （TMRE） was used to detect the expression of mitochondrial membrane potential in cells. A kit was used to detect iron ion （Fe2+） content， lipid peroxide （LPO）， and superoxide dismutase （SOD） activity. Western blot was used to detect the protein expression levels of glycogen synthase3β （GSK3β）， phosphorylated GSK3β （p-GSK3β）， nuclear factor E2 related factor 2 （Nrf2）， and glutathione peroxidase 4 （GPX4）. The real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） was used to detect the mRNA expression of member 11 of the cystine/glutamic acid reverse transporter solute vector family 7 （SLC7A11）， member 2 of the heavy chain solute vector family 3 （SLC3A2）， transferrin receptor 3 （TFRC）， and tumor protein （TP）53. ResultCCK-8 results showed that BXT and capecitabine could significantly reduce the survival rate of three kinds of gastric cancer cells after treatment with drug-containing serum for 24 h （P<0.01）. After 48 h of intervention with drug-containing serum， the survival rate of three kinds of gastric cancer cells was significantly decreased in both the capecitabine group and the BXT group compared with the blank group. The BXT group was dose-dependent， with 20% BXT having the most significant effect （P<0.01）. In terms of biochemical indicators of ferroptosis， compared with the blank group， BXT and capecitabine significantly decreased the expression of mitochondrial membrane potential （P<0.01） and SOD activity （P<0.01） and significantly increased the contents of LPO and Fe2+ （P<0.01）， so as to improve the sensitivity of gastric cancer cells to ferroptosis. In terms of the Nrf2/GPX4 pathway， compared with the blank group， the BXT group could reduce the protein expressions of p-GSK3β， Nrf2， and GPX4 （P<0.01） in gastric cancer cells and increase mRNA expressions of SLC7A11 and SLC3A2 （P<0.05）. It could also increase the protein expression of GSK3β （P<0.01） and mRNA expression of TP53 and TFRC （P<0.05， P<0.01） in gastric cancer cells. Inhibition of the Nrf2/GPX4 pathway induces ferroptosis in gastric cancer cells. Compared with the capecitabine group， the 20% BXT group showed a more obvious effect. ConclusionBanxia Xiexintang can induce ferroptosis in gastric cancer cells HGC-27， MKN-45， and AGS by inhibiting the Nrf2/GPX4 pathway.

Mechanism of Modified Tianwang Buxindan on Skin of Sleep-deprived Mice Through PI3K/Akt/Nrf2 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo observe the effect of modified Tianwang Buxindan （MTBD） on the skin of sleep-deprived （SD） mice and investigate its mechanism. MethodSixty 2-month-old female Kunming mice were randomly divided into a blank group， a model group， a vitamin C （VC， 0.08 g·kg-1）， and MTBD low-， medium-， and high-dose groups （6.5， 12.5， 25 g·kg-1）. Except for the blank group， the other groups were subjected to SD mouse model induction （using multiple platform water environment method for 18 hours of sleep deprivation daily from 15：00 to next day 9：00）， continuously for 14 days， and caffeine （CAF， 7.5 mg·kg-1） was injected intraperitoneally from the 2nd week onwards， continuously for 7 days. While modeling， the blank group and the model group were administered with normal saline （0.01 mL·g-1）， and the other groups received corresponding drugs for treatment. On the day of the experiment， general observations were recorded （such as body weight， spirit， fur， and skin）. After sampling， skin tissue pathological changes were observed under an optical microscope using hematoxylin-eosin （HE） and Masson staining methods. Skin thickness and skin moisture content were measured. Biochemical assay kits were used to detect skin hydroxyproline （HYP） content， skin and serum superoxide dismutase （SOD） activity， and malondialdehyde （MDA） content. Enzyme-linked immunosorbent assay （ELISA） was used to detect serum interleukin （IL）-6， tumor necrosis factor （TNF）-α， and IL-1β levels in mice. Western blot was used to detect skin tissue type Ⅰ collagen （ColⅠ）， type Ⅲ collagen （ColⅢ）， phosphatidylinositol 3-kinase （PI3K）， phosphorylated （p）-PI3K， protein kinase B （Akt）， p-Akt， nuclear factor E2-related factor 2 （Nrf2）， heme oxygenase （HO）-1， and nuclear factor （NF）-κB protein expression. ResultCompared with the blank group， the model group showed varying degrees of changes. In general， signs of aging such as reduced body weight （P<0.01）， listlessness， dull fur color， and formation of wrinkles on the skin appeared. Tissue specimen testing revealed skin thinning， flattening of the dermoepidermal junction （DEJ）， and reduced collagen fibers under the optical microscope. Skin thickness and moisture content decreased， skin tissue HYP content significantly decreased （P<0.01）， skin and serum SOD activity significantly decreased （P<0.01）， and MDA content significantly increased （P<0.01）. Serum IL-6， TNF-α， and IL-1β levels significantly increased （P<0.01）. Skin ColⅠ， ColⅢ， p-PI3K/PI3K， p-Akt/Akt， Nrf2， and HO-1 protein expression significantly decreased （P<0.05， P<0.01）， and NF-κB expression increased （P<0.01）. Compared with the model group， the VC group and the MTBD low-dose group showed increased skin moisture content， HYP content， SOD activity， and ColⅠ， ColⅢ， p-PI3K/PI3K protein expression （P<0.05， P<0.01）， and decreased serum MDA content （P<0.05）. In addition， a decrease in serum IL-6 and IL-1β levels was detected in the MTBD low-dose group （P<0.05）， while the above indicators in the MTBD medium- and high-dose groups improved （P<0.05， P<0.01）. ConclusionSleep deprivation accelerates the aging process of the skin in SD model mice. MTBD can improve this phenomenon， exerting anti-inflammatory and antioxidant effects， and its mechanism of action may be related to the activation of the PI3K/Akt/Nrf2 signaling pathway.

Effect of Sinisan on Oxidative Stress in Cholestatic Hepatitis Rats Based on Nrf2/HO-1 Signaling Pathway / 中国实验方剂学杂志

ObjectiveBased on the nuclear factor erythroid 2 related factor 2 （Nrf2）/heme oxygenase-1 （HO-1） signaling pathway， this paper explores the effect of Sinisan （SNS） on liver oxidative stress injury in cholestatic hepatitis rats and its mechanism. MethodThirty 6-week-old male SD rats were randomly divided into a control group， model group， low and high dose groups of SNS （2.5 and 5 g·kg-1） and ursodeoxycholic acid group （UDCA， 63 mg·kg-1）， with six rats in each group. Rats were administrated for seven consecutive days. On the 5th day， the control group was given olive oil of 10 mL·kg-1， and the other groups were given alpha-naphthalene isothiocyanate （ANIT） of 80 mg·kg-1. The serum biochemical indicator levels of cholestasis and the content of antioxidant factors in rat liver were detected by enzyme-linked immunosorbent assay （ELISA）. Hematoxylin-eosin （HE） staining was used to observe the pathological changes in liver tissue. The relative mRNA and protein expressions of Nrf2， HO-1， and quinone oxidoreductase 1 （NQO1） in liver tissue were detected by real-time fluorescence quantitative polymerase chain reaction （Real-time PCR） and Western blot. ResultCompared with the control group， the model group showed a significant increase in the serum biochemical indicator levels of cholestasis and the content of antioxidant factors in liver tissue （P<0.01）. There were obvious pathological changes in the model group such as the disordered arrangement of hepatocytes， obvious congestion and necrosis in the portal area， infiltration of inflammatory cells， and destruction of the interlobular bile duct. The relative mRNA and protein expressions of Nrf2， HO-1， and NQO1 in liver tissue were significantly down-regulated in the model group （P<0.05， P<0.01）. Compared with the model group， the groups of SNS showed a significant decrease in the serum biochemical indicator levels of cholestasis and the content of antioxidant factors in liver tissue （P<0.01）， and the pathological liver injury was obviously improved. The necrotic area was reduced， and the infiltration of inflammatory cells was decreased. In addition， there was a small amount of extravasated blood in the interlobular vein. The relative mRNA and protein expressions of Nrf2， HO-1， and NQO1 in liver tissue were significantly up-regulated （P<0.05， P<0.01）. ConclusionSNS can significantly improve liver injury in cholestatic hepatitis rats， and its mechanism may be related to the inhibition of oxidative stress response mediated by the Nrf2/HO-1 signaling pathway.

Exploring the mechanism of traditional Chinese medicine indirubin derivative E804 inhibiting the proliferation and migration of lung cancer A549 cells based on the Nrf2-HO-1/GPX4 pathway / 安徽医科大学学报

Objective To investigate the effects of indirubatin derivative E804 on proliferation and migration of non-small cell lung cancer(NSCLC)A549 cells,and to elucidate the possible mechanism of Nrf2-HO-1/GPX4 pathway.Methods Lung cancer A549 cells were used as the cell model.The proliferation and migration of differ-ent specific inhibitors(Nec-1,CQ,Z-VAD,DFO,Fer-1 and Lip-1)in 0,10 μmol/L E804 and 10 μmol/L E804+groups were observed by MTT and cell scratch assay.The contents of reactive oxygen species(ROS)were de-tected by DCFH-DA fluorescence probe method,the contents of Fe2+were detected by colorimetric method,the contents of reduced glutathione(GSH)were detected by spectrophotometry,and the contents of malondialdehyde(MDA)were detected by micromethod.The expression levels of SLC7A11,Transferrin,GPX4,SLC40A1,Nrf2 and HO-1 were detected by Western blot in cells of 0,2.5,5 and 10 μmol/L E804 groups.Results Compared with the control group(0 μmol/L E804),2.5,5 and 10 μmol/L E804 significantly increased intracellular ROS,Fe2+and MDA levels,and decreased intracellular GSH content(P＜0.01).Meanwhile,the expression levels of SLC7A11,GPX4,SLC40A1,Nrf2 and HO-1 significantly decreased(P＜0.01),and the expression level of Transferrin increased(P＜0.05).Compared with the 10 μmol/L E804 group alone,the apoptosis inhibitor(Z-VAD)group and the ferroptosis inhibitor(DFO,Fer-1 and Lip-1)group could significantly reverse the inhibition of proliferation and migration of A549 cells by 10 μmol/L E804(P＜0.01).Conclution E804 can induce ferrop-tosis and inhibit the proliferation and migration of A549 cells,which may be related to the inhibition of Nrf2-HO-1/GPX4 pathway.

Sodium hydrosulfide increases glutathione synthesis in cardiac muscle cell line HL-1 under hyperglycemia and hyperlipidemia / 基础医学与临床

Objective To investigate sodium hydrogen sulfide(NaHS)with function of regulating glutathione(GSH)synthesis to reduce reactive oxygen species(ROS)production in type 2 diabetic cardiomyopathy(DCM).Methods Mouse cardiomyocyte cell line HL-1 was incubated with high concentration of glucose(HG:40 mmol/L)and palmitate(Pal:500 μmol/L)as a cell model of type 2 DCM.HL-1 cells were incubated with NaHS(100 μmol/L),DL-propargylglycin(PPG,1 mmol/L)and N-acetyl-l-cysteine(NAC,5 mmol/L),respectively for 72 hours.The expression of cystathionine-γ-lyase(CSE)and the key enzymes of glutathione production was tested by Western blot.Dihydroethidium(DHE)and dichlorofluoromethane(DCFH)were used to detect the content of ROS in HL-1 cells.Cell viability was detected by CCK8 kit.The content of total GSH was detected.The interaction between muscle specific ring finger protein 1(Murf1)and nuclear factor erythroid-derived 2-related factor 2(Nrf2)and Nrf2 ubiquitylation was determined by co-immunoprecipitation(co-IP).Results Compared with control group,the expression level of CSE,solute carrier family 7 members 11(SLC7A11),glutamate cysteine ligase C(GCLC),glutamate cysteine ligase M(GCLM)and glutathione synthetase(GSS)in HL-1 cells treated incubated with high glucose and palmitate was decreased,however,NaHS was found to restore it.NaHS reduced the content of ROS in HL-1 cells treated with high glucose and palmitate.The interaction between murf1 and Nrf2 was confirmed by co-immunoprecipitation(Co-IP).Compared with NaHS group,the ubiquitylation level of Nrf2 was enhanced in high glucose and palmitate group.Conclusions Sodium hydrosulfide may reduce the ubiquitylation level of Nrf2 and promote the expression of key enzymes of GSH synthesis.

Effects of intervention in autophagy regulation of p62-Keap1/Nrf2-GPX4 pathway on ferroptosis and oxaliplatin resistance in colorectal cancer cells / 临床与实验病理学杂志

Purpose To investigate the effect of autophagy intervention on ferroptosis and drug resistance of colorectal canc-er cells and its molecular mechanism.Methods The human colorectal cancer cell lines HCT-8,COLO205,HCT-116,SW620,and SW480 were cultured.HCT-116 cells with moder-ate expression of LC3 were screened,and the expression differ-ences of LC3,p62,Keap1,Nrf2,GPX4 proteins,Fe2+,GSH,and MDA between them and OXA-resistant HCT-116/OXA cell lines were detected.The expression levels of LC3,p62,Keap1,Nrf2,GPX4,Fe2+,GSH and MDA were assessed in HCT-116/OXA cells through the intervention of autophagy and ferroptosis intervention agent combined with oxaliplatin.The proliferative activity and sensitivity to oxaliplatin in each group were detected by CCK-8 assay.Cell growth and invasion ability of each group were detected by plate cloning and Trans well assay.Results LC3,p62 and GPX4 expression levels of HCT-116 cells in the 5 groups were moderate.Compared with HCT-116 cells,HCT-116/OXA was less sensitive to oxaliplatin,and the proteins of p62,Nrf2 and GPX4 were highly expressed,LC3 and Keap1 were lowly expressed,and the expression of Fe2+,GSH and MDA were increased(P＜0.05).The levels of LC3,Keap1 protein,Fe2+and MDA in Rapa and Rapa+Fer-1 groups were higher than those in Fer-1 and control groups,while p62,Nrf2,GPX4 and GSH levels were lower.The expressions of GPX4 pro-tein and GSH in Rapa+Fer-1 group were lower than those in Rapa group(P＜0.05).In the autophagy inhibitor group,LC3,p62,Nrf2,GPX4 and GSH were highly expressed in the CQ and CQ+Erastin groups compared with the control and Eras-tin groups,while Keap1 protein,Fe2+and MDA were low.The levels of GPX4 protein and GSH in Erastin group were lower than those in the other three groups,and the levels of Fe2+and MDA were higher than those in the other three groups(P＜0.05).The combination of autophagy activator OXA showed that Rapa intervention group had higher chemical sensitivity to OXA,less number of migrating cells and lower cell proliferation activity than the other three groups.The sensitivity of Rapa+Fer-1 group to oxaliplatin was lower than that of Rapa group,but higher than that of Fer-1 group and control group(P＜0.05).There was no significant difference between Fer-1 group and con-trol group(P＜0.05).Compared with the control group,the cell activity,migration capacity and clonogenesis capacity of Erastin,CQ+Erastin and CQ groups were decreased when auto-phagy inhibitor was combined with OXA,and the Erastin group was the lowest,while the CQ+Erastin group was higher than the Erastin group,and lower than the CQ group(P＜0.05).Con-clusion In colorectal cancer,autophagy is involved in the regu-lation of ferroptosis,and intervention in autophagy can regulate ferroptosis in colorectal cancer cells through the p62-Keap1/Nrf2-GPX4 pathway,thereby reversing oxaliplatin resistance.

The relationship between EGCG targeted regulation of Nrf2-Keap1 signaling pathway and neuroprotective effect in cerebral infarction / 实用医学杂志

Objective The preventive effect of epigallocatechin gallate(EGCG)on hyperglycemia-induced hemorrhagic transformation(HT)was analyzed,and the underlying mechanisms were further explored.Methods Male SD rats were randomly divided into sham operation group(Sham,n = 20),model group(n = 27),hyperglycemia model group(HG,n = 43),and EGCG group(n = 43).In the model group,only the electrocoagulation cerebral ischemia model was established,and the HG group and the EGCG group were used to establish the HT model with acute hyperglycemia combined with electrocoagulation cerebral ischemia model.In addition,EGCG was adminis-tered by gavage for 5 days before cerebral ischemia at a dose of 50 mg/kg/d.Further studies confirmed the relevant targets by using network pharmacology to predict the potential targets and pathways of EGCG in the occurrence of HT.Results Compared with the model group,the mortality rate of the rats in the HG group was significantly increased[21.2%(6/27)vs.51.2%(22/43),P＜0.05].The mortality of rats in the EGCG group was significantly lower than that in the HG group[30.20%(13/43)vs.51.2%(22/43),P＜0.05].Second,mNSS,Longa score and infarct volume in the EGCG group were significantly lower than those in the HG group(P＜0.05).The incidence of HT in the HG group was higher than that in the model group(59.3%vs.90.7%).EGCG significantly reduced the incidence of hyperglycemia-induced HT to 69.8%.Compared with the HG group,EGCG decreased the hemoglobin content from(53.42±5.11)mg/dL to(37.04±2.39)mg/dL respectively(P＜0.05).Network pharmacology revealed that Nrf2-Keap1-mediated neuroinflammation may be associated with hyperglycemia-induced HT.The expression of Nrf2 and Keap1 was significantly decreased and the expression of TLR4 and phosphorylation of NF-κB was significantly increased in the HG group,but EGCG reversed this process.Conclusion EGCG pretreatment prevents the occurrence of HT,which may be related to the neuroprotection mediated by activation of the Nrf2-Keap1 signaling pathway.

Mechanism of black phosphorus regulating oxidative stress-inflammation cascade in retarding intervertebral disc degeneration / 中国组织工程研究

BACKGROUND:Oxidative stress plays a critical role in intervertebral disc degeneration.As a reducing material with good biocompatibility,black phosphorus quantum dots have the potential to resist oxidative stress and retard intervertebral disc degeneration.OBJECTIVE:To evaluate the effect of black phosphorus quantum dots on scavenging reactive oxygen species in the microenvironment of an intervertebral disc through in vivo and in vitro experiments,and further explore the role of black phosphorus quantum dots in Nrf2/ARE pathway and intervertebral disc inflammation.METHODS:Black phosphorus quantum dots were prepared by a liquid exfoliation technique.(1)In vitro experiment:The nucleus pulposus cells of SD rats were isolated and extracted,and the passages 2-4 nucleus pulposus cells were cocultured with different solutions,including F12-DMEM medium(blank group),black phosphorus quantum dot solution,hydrogen peroxide solution,hydrogen peroxide+black phosphorus quantum dot solution,hydrogen peroxide+black phosphorus quantum dot+Nrf2 specific inhibitor ML385 solution.Cell live/dead staining and intracellular reactive oxygen species,mitochondrial membrane potential and western blot assay were performed respectively.(2)In vivo experiment:Thirty SD rats were randomly divided into sham operation,puncture and puncture + black phosphorus groups,with 10 rats in each group.A Co7-10 intervertebral disc degeneration model was established using intervertebral disc puncture in the puncture group and the puncture+black phosphorus group.Black phosphorus quantum dot solution was injected in the intervertebral disc after a puncture in the puncture+black phosphorus group.The intervertebral disc tissue imaging and histological staining were evaluated at 4 and 8 weeks after surgery.RESULTS AND CONCLUSION:(1)In vitro experiment:Live/dead staining revealed that the black phosphorus quantum dots had good biocompatibility,were non-toxic to cells,and had a protective effect on nucleus pulposus cells under oxidative stress.Intracellular reactive oxygen species and JC-1 fluorescent probes showed that black phosphorus quantum dots could regulate the reduction of mitochondrial membrane potential caused by oxidative stress in nucleus pulposus cells and protected cells from hydrogen peroxidation-induced intracellular oxidative stress.Western blot analysis showed that compared with the blank group,the protein expressions of Nrf2,heme oxygenase 1,quinone oxidoreductase and type Ⅱ collagen were decreased in the hydrogen peroxide group(P<0.05),while the protein expressions of tumor necrosis factor α,interleukin 1β,matrix metalloproteinase 13 and p65 were increased(P<0.05).The addition of black phosphorus quantum dots could reverse the inhibitory effect of hydrogen peroxide on the Nrf2 pathway and reduce the inflammatory response caused by oxidative stress,but NrF2-specific inhibitors could cancel this effect.(2)In vivo experiment:X-ray and MRI demonstrated that at 4 and 8 weeks after surgery,the intervertebral disc height and water content of nucleus pulposus in the puncture group were lower than those in the sham operation group(P<0.05),and the intervertebral disc height and water content of nucleus pulposus in the puncture+black phosphorus group were higher than those in the puncture group(P<0.05).Histological staining exhibited that the degree of intervertebral disc degeneration in the puncture+black phosphorus group was less than that in the puncture group,and the expression of heme oxygenase 1 protein was higher than that in the puncture+black phosphorus group.(3)Our results have indicated that black phosphorus quantum dots can exert an antioxidant effect and delay intervertebral disc degeneration by regulating Nrf2/ARE pathway.

Oxymatrine hydrogel promotes wound healing by activating Nrf2/HO-1 pathway in keratinocytes / 中国组织工程研究

BACKGROUND:Inflammation and oxidative stress contribute to the barriers of regeneration in chronic wound.Oxymatrine has various biological activities,such as anti-oxidation,anti-inflammation and so on,which may have the potential effect of promoting wound healing. OBJECTIVE:To investigate the effect of oxymatrine on wound healing and the protective effect on H2O2-induced oxidative stress injury in human keratinoid cell line HaCaT cells. METHODS:(1)In vivo experiment:Hyaluronic acid methacryloyl hydrogels containing 0,0.05,0.1,0.2 g/L oxymatrine were prepared.A full-layer skin defect model with a diameter of 12 mm was made in the back of 75 diabetic mice and randomly divided into five groups for intervention,with 15 mice in each group.The wounds of the model group were bandaged and fixed.The wounds of the hydrogel group were covered with hyaluronic acid methacryloyl hydrogel.The wounds of the low-dose,moderate-dose and high-dose oxymatrine groups were covered with hyaluronic acid methacryloyl hydrogel containing 0.05,0.1,and 0.2 g/L oxymatrine,respectively,and then bandaged and fixed after light curing.Relevant indicators were detected within 14 days.(2)In vitro experiment:Human keratinocyte line HaCaT was divided into five groups.The normal group was cultured conventionally.H2O2 group and low-,moderate-and high-concentration oxymatrine groups were treated with H2O2 for 4 hours,and then the medium was replaced with medium containing 0,0.05,0.1,and 0.2 g/L oxymatrine,respectively,and the relevant indexes were detected after 24 hours of culture. RESULTS AND CONCLUSION:(1)In vivo experiment:Compared with the model group,the wound healing rate of mice in the hydrogel group had no significant change.The wound healing rate of mice in the low-,moderate-and high-dose oxymatrine group was increased at 7 and 14 days after treatment(P<0.05).Pathological observation of wound section 14 days after treatment showed that compared with the model group,the thickness of regenerated epidermal layer,the number of microvessels,and collagen deposition in the moderate-and high-dose oxymatrine groups were increased(P<0.05).Western blot assay analysis of wound samples 7 days after surgery showed that compared with the model group,the protein expressions of tumor necrosis factor α and interleukin 6 in the moderate-and high-dose oxymatrine groups were decreased(P<0.05).(2)In vitro experiment:CCK8 assay,EdU and Ki67 staining showed that compared with the H2O2 group,the cell proliferation ability of the moderate-and high-concentration oxymatrine groups was significantly increased(P<0.05).Compared with the H2O2 group,mitochondrial membrane potential was increased(P<0.05)and reactive oxygen species content was decreased(P<0.05)in the moderate-and high-concentration oxymatrine groups.Western blot assay results showed that compared with the H2O2 group,the expression levels of Nrf2 nuclear protein,Nrf2 total protein,HO-1 protein,and superoxide dismutase 1 protein were increased in the high-concentration oxymatrine group(P<0.05).(3)These findings confirm that oxymatrine can alleviate oxidative stress damage in HaCat cells and accelerate wound healing by upregulating the levels of Nrf2 and HO-1 protein.