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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.
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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.
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OBJECTIVE To investigate the intervention effect and potential mechanism of breviscapine on hepatic fibrosis （HF） in rats based on the transforming growth factor-β（1 TGF-β1）/Smad2/extracellular signal-regulated protein kinase 1（ERK1） and Kelch-like epichlorohydrin-associated protein 1（Keap1）/nuclear factor-erythroid 2-related factor 2（Nrf2）/heme oxygenase-1（HO-1） pathways. METHODS Totally 60 rats were randomly divided into normal control group， model group， breviscapine low-dose， medium-dose and high-dose groups （5.4， 10.8， 21.6 mg/kg）， and colchicine group （positive control， 0.45 mg/kg）， with 10 rats in each group， half male and half female. Except for the normal control group， HF model of the other groups was induced by carbon tetrachloride. Subsequently， each drug group was given corresponding medicine by gavage once a day for 28 days. The liver appearance of rats in each group was observed and their liver coefficients were calculated. The levels of alanineaminotransferase （ALT） and aspartate aminotransferase （AST）in serum， those of ALT， AST， superoxide dismutase （SOD），malondialdehyde （MDA） and glutathione peroxidase （GSH- Px） in liver tissue were detected. The liver tissue inflammatory and fibrotic changes were observed. The protein and mRNA expressions of TGF-β1， Smad2， ERK1， Nrf2， Keap1 and HO-in liver tissue were detected. RESULTS Compared with the normal control group， the model group showed large areas of white nodular lesions in the liver， obvious inflammatory cell infiltration and collagen fiber deposition. The body weight， the levels of SOD and GSH-Px in liver tissue， the protein and mRNA expressions of Nrf2 and HO-1 were significantly lowered in the model group （P＜0.05）； the liver coefficient， the percentage of Masson staining positive area， ALT and AST levels of serum and liver tissue， MDA level of liver tissue， the protein and mRNA expressions of TGF-β1， Smad2， ERK1 and Keap1 were significantly increased （P＜0.05）. Compared with the model group， the liver lesions of rats in each drug group were improved， and the above quantitative indexes were generally reversed （P＜0.05）. CONCLUSIONS Breviscapine has a good intervention effect on HF rats， which may be related to inhibiting TGF-β1/Smad2/ERK1 pathway for anti-fibrosis and regulating Keap1/Nrf2/HO-1 pathway to inhibit oxidative stress.
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Objective To investigate the effect of dioscin on uric acid(UA)-induced oxidative stress injury of human renal tubular epithelial cells(HK-2)and its molecular mechanism.Methods HK-2 cells were cultured and divided into four groups:blank group(normal group),model group(uric acid-stimulation modeling),condition control group(UA+DMSO)and dioscin group(UA+dioscin).Oxidative stress injury model was induced by UA in HK-2 cells.Cells viability was detected by CCK-8.ROS level was detected by flow cytometry.Real-time PCR was used to detect the expressions of glycogen synthase kinase 3β(GSK3β),nuclear factor erythroid 2-related factor 2(Nrf2)and heme oxygenase 1(HO-1)at mRNA level,and Western Blot was used to detect the expressions of phosphorylated glycogen synthesis kinase 3β(p-GSK3β),GSK3β,Nrf2 and HO-1 at protein level.Results After stimulation by UA,HK-2 cells viability was obviously decreased,and ROS level was significantly increased(all P＜0.001).When treated with dioscin,HK-2 cells viability was obviously increased,and the ROS level of HK-2 cells was significantly decreased(all P＜0.001).The expressions of Nrf2 and HO-1 decreased at the protein and mRNA levels after stimulation with UA.But the expressions of Nrf2 and HO-1 significantly increased after treated with dioscin(all P＜0.001).Compared with the blank group,the p-GSK3β/GSK3β ratio in the model group decreased significantly at the protein level,but the p-GSK3β/GSK3β ratio increased after treated with dioscin(all P＜0.001).Conclusion Dioscin can alleviate UA-induced oxidative stress injury in HK-2 cells.The mechanism might be that dioscin can promote phosphorylation of GSK3β,and activate Nrf2/HO-1 pathway.
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AIM: To investigate the protective effect and mechanism of Danlou tablet on retinal ischemia-reperfusion injury(RIRI)in mice.METHODS: A total of 40 ApoE-/- mice were fed with high fat diet for 6 wk, and the RIRI model was established by anterior chamber infusion of pressurized saline. The mice were divided into control group(normal saline for 8 wk), RIRI model group(normal saline for 8 wk), and low-, medium-, and high-dose Danlou tablets groups [1, 2, and 4 g/(kg·d), respectively, for 8 wk]. The morphological changes of retina were observed by hematoxylin-eosin(HE)staining, retinal cell apoptosis was detected by terminal-deoxynucleoitidyl transferase mediated Nick-End Labeling(TUNEL)staining. The Western-blot assay was used to detect the expression of retinal tissue sample Kelch-like ech-associated protein 1(Keap1), nuclear factor E2 related factor 2(Nrf2), heme oxygenase 1(HO-1), and superoxide dismutase(Sod2)proteins.RESULTS: Compared with the control group, the mouse retina was atrophic with thinning thickness and increasing cell apoptosis, down-regulation of Sod2 protein expression, and up-regulation of Keap1 protein expression in the RIRI model group(all P<0.01). Compared with the RIRI model group, the retinal thickness increased in the medium- and high-dose of Danlou tablets groups(all P<0.01), and the cell apoptosis of retina decreased in the low-, medium- and high-dose of Danlou tablets groups(all P<0.05). There were no significant differences in the expression of Keap1 and HO-1 proteins of mouse retina tissue in the low-dose of Danlou tablets group(P>0.05). The expression of Sod2, Nrf2 and HO-1 proteins up regulated, and the expression of Keap1 protein down regulated in the medium- and high-dose of Danlou tablets groups(all P<0.05).CONCLUSION: Danlou tablet can alleviate RIRI-induced atrophy and thinning of retina and retinal cell apoptosis by regulating Keap1-Nrf2/HO-1 signal pathway and reducing oxidative stress.
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Objective To investigate the effects of Polygonatum kingianum on wound healing and nuclear factor-erythroid 2-related factor 2(Nrf2)/heme oxygenase-1(HO-1)signaling pathway in diabetic skin injury rats.Methods Diabetic rat models were established and 48 rats were randomly divided into model group,sitagliptin group(10 mg·kg-1),Polygonatum kingianum water extract low dose group(2 g·kg-1),Polygonatum kingianum water extract high dose group(8 g·kg-1)and Polygonatum Kingianum alcohol extract low and high dose group(2 g·kg-1 and 8 g·kg-1).8 in each group;Another control group was set up.After 4 weeks of intragastric administration,all rats established back skin wounds,and continued to be administered for 14 days after operation.At the end of the experiment,the rats were killed,and blood glucose,glycosylated hemoglobin(GHb),plasma levels of H2O2,MDA,SOD,GSH and wound margin skin tissue T-AOC,SOD,MDA levels were measured.The relative expression of Nrf2 and HO-1 mRNA in wound margin skin tissue was measured by fluorescence quantitative method.Results Compared with the model group,the levels of SOD and GSH in plasma and the relative expressions of Nrf2 and HO-1 mRNA in wound margin skin tissue in the low and high dose groups of ethanol extract of Polygonatum kingianum,and the low and high dose groups of water extract of Polygonatum kingianum were increased(P<0.01,P<0.001).The levels of glycosylated hemoglobin(GHb)in plasma and MDA in wound margin tissue were decreased(P<0.05,P<0.01,P<0.001).The level of T-AOC in the high dose group was increased(P<0.01).The SOD level of skin tissue in the low dose group of Polygonatum kingianum water extract was increased(P<0.01),and the level of H2O2 in the low dose group was decreased(P<0.01).Conclusion Polygonatum canaliculatum can up-regulate Nrf2/HO-1 signaling pathway,regulate excessive oxidative stress,and promote wound healing.
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AIM: To explore schisandrin B (Sch B) pretreatment reduces intestinal ischemia reperfusion injury (IIRI) through inhibiting apoptosis by activation of Nrf2/HO-1 signing pathway in mice by network pharmacology and in vivo experiment. METHODS: (1) The targets of Sch B and IIRI were searched from online databases, Drawing Venn diagram to obtain the common target of them. Cytoscape software was imported to construct the protein-protein interaction (PPI) network to establish the "Drugs-Disease-core target gene" network. The mechanism of Sch B against IIRI was predicted through GO and KEGG enrichment analysis. (2) Thirty-six C57BL/6J mice were randomly divided into six groups (n = 6). The model of IIRI was established in four groups except the sham operation group. Three of the groups were pretreated with Sch B, Nrf2 inhibitor ML385, and Sch B + ML385, respectively. After the experiment, intestinal tissue samples were taken for HE staining, Chiu ' s score, apoptosis staining, immunohistochemistry (IHC), and immunoblotting (Western blot). RESULTS: A total of 412 Sch B related tar- gets, 2 166 IIRI related targets and 153 common targets were screened out through network pharmacology. There were 88 "Sch B-IIRI-core target gene" included NFE2L2 (Nrf2), HMOX1 (HO-1), BCL2, CASP3 (caspase 3), and so on. KEGG enrichment analysis screened 163 related pathways, apoptosis pathway ranked high showing that the pathway may play a key role in the treatment of IIRI by Sch B. The animal experiment had shown that Sch B reduced the Chiu's score and apoptotic while upregulating Nrf2, HO-1, Bcl-2 protein expression levels and Bcl-2/Bax, downregulating Bax, and cleaved caspase-3 expression levels, thereby reducing IIRI in mice, and that Nrf2 inhibitor ML385 reversed this process (P < 0.05). CONCLUSION: This study reveals that Sch B has the characteristics of multi-target and multi-pathway in the reduction of IIRI, and Sch B can reduce IIRI through inhibiting apoptosis by activation of Nrf2/ HO-1 pathway.
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To investigate the antidepressant mechanism of Shenling Kaixin Granules(SLKX) in treating chronic unpredictable mild stress(CUMS) model rats. Ninety male SD rats were randomly divided into control group, model group, Shugan Jieyu Capsules(110 mg·kg~(-1)) group and SLKX low-(90 mg·kg~(-1)), medium-(180 mg·kg~(-1)), and high-dose(360 mg·kg~(-1)) groups. Depression rat model was replicated by CUMS method. After treatment, the behavioral changes of rats were evaluated by sugar preference, open field, elevated cross maze and forced swimming experiments. The contents of interleukin 1 beta(IL-1β), tumor necrosis factor α(TNF-α), brain-derived neurotrophic factor(BDNF) and 5-hydroxytryptamine(5-HT) in serum were determined by enzyme linked immunosorbent assay(ELISA), and the activities of superoxide dismutase(SOD) and catalase(CAT) in hippocampal CA1 region were also detected. Pathological changes in hippocampal CA1 region were detected by hematoxylin-eosin(HE) staining, and Western blot was used to determine the expression of nerve growth factor(NGF), BDNF, phospho-tyrosine kinase receptor(p-TrkB)/TrkB, phospho-cAMP-response element binding protein(p-CREB)/CREB, nuclear factor E2 related factor 2(Nrf2), heme oxygenase 1(HO-1), B-cell lymphoma-2(Bcl-2)/Bcl-2 associated X protein(Bax) and caspase-3 in hippocampal CA1 region. RESULTS:: showed that compared with the control group, the model group had decreased sugar preference, reduced number of entries and time spent in the center of open field and shortened total distance of movement, reduced number of entries and proportion of time spent in open arm, and increased number and time of immobility in forced swimming experiment. Additionally, the serum contents of IL-1β and TNF-α and the expression of caspase-3 were higher, while the contents of BDNF and 5-HT, the activities of SOD and CAT in hippocampal CA1 region, the expressions of NGF, BDNF, p-TrkB/TrkB, p-CREB/CREB, HO-1 and Bcl-2/Bax, and the Nrf2 nuclear translocation were lower in model group than in control group. Compared with the conditions in model group, the sugar preference, the number of entries and time spent in the center of open, total distance of movement, and the number of entries and proportion of time spent in open arm in treatment groups were increased while the number and time of immobility in forced swimming experiment were decreased; the serum contents of IL-1β and TNF-α and the expression of caspase-3 were down regulated, while the contents of BDNF and 5-HT, the activities of SOD and CAT in hippocampal CA1 region, the expressions of NGF, BDNF, p-TrkB/TrkB, p-CREB/CREB, HO-1, Bcl-2/Bax, and Nrf2 nuclear translocation were enhanced. In conclusion, SLKX might regulate the Nrf2 nucleus translocation by activating BDNF/TrkB/CREB pathway, lower oxidative stress damage in hippocampus, inhibit caspase-3 activity, and reduce apoptosis of hippocampal nerve cells, thereby playing an antidepressant role.
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Rats , Male , Animals , bcl-2-Associated X Protein/metabolism , Caspase 3/metabolism , Nerve Growth Factor/metabolism , Brain-Derived Neurotrophic Factor/metabolism , Signal Transduction , Tumor Necrosis Factor-alpha/metabolism , Serotonin/metabolism , NF-E2-Related Factor 2/metabolism , Rats, Sprague-Dawley , Antidepressive Agents/pharmacology , Hippocampus/metabolism , Superoxide Dismutase/metabolism , Sugars/pharmacology , Depression/genetics , Stress, Psychological/metabolismABSTRACT
Objective To investigate protective effects of polysaccharides from Platycarya strobilacea Sieb.et Zucc on cerebral ischemia-reperfusion injury in rats.MethodsForty Wistar rats were randomly divided into four groups:sham group,model group,Rehmannia glutinosa polysaecharide low,and high dose groups.Each group was given oral administration for 7 days.After 1 h of last administration, the CIRI (cerebral ischemia-reperfusion injury) group was produced inserting 5-0 line into the internal carotid artery.Neurological functional score was evaluated according to the method of Zea longa's score, meanwhile,infarct size was detected by TTC staining.Levels of superoxide dismutase (SOD) activity,malondialdehyde (MDA), IL-10, IL-1β and TNF-α contents in cerebral tissues were measured.TUNEL staining was used to assess the number of TUNEL-positive cells of the ischemic cortex.Western blot was used to analyze the expressions of Nrf2 and HO-1 in cerebral tissues.ResultsCompared to model group, Platycarya strobilacea a polysaccharides could significantly improve neurogical functions,greatly decrease the contents of MDA, IL-10 and IL-1β in cerebral tissues,improve SOD activity and IL-10 content in cerebral tissues,and effectively reduce cerebral infarct range.Meanwhile Platycarya strobilacea polysaccharides could enhance the expressions of Nrf2 and HO-1 in cerebral tissues.ConclusionPlatycarya strobilacea polysaccharides have beneficial effects on rats with cerebral ischemia-reperfusion injury, which may be mediated at least in part by the Nrf2/HO-1 pathway.
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OBJECTIVE: To study the effect mechanism of tempol against hypobaric hypoxia-induced heart damage in mice. METHODS: One hundred and ten BALB/c mice were randomly divided into normal control group, hypoxia model group, acetazolamide group and tempol group. After single intraperitoneal injection for 30 min, the mice were exposed to a simulated high altitude of 8 000 m for 12 h. After hypoxic exposure, blood was collected from the eye sockets and separated into serum to measure the activities of lactic dehydrogenase (LDH)and creatine kinase (CK). Then the mice were sacrificed and the content of H2O2 and malondialdehyde (MDA) as well as ATPase and antioxidant enzyme activity in heart were determined. HIF-1, VEGF, Nrf2, and HO-1 were detected by immunohistochemistry. RESULTS: Compared with normal control group, the activities of plasma CK and LDH in hypoxia model group significantly increased. In addition, the content of H2O2 and MDA in hypoxia model group significantly increased while ATPase and antioxidant enzymes activity markedly decreased compared with the normal control group. Moreover, the expression of HIF-1α, VEGF, Nrf2 and HO-1 increased. Prior administration of tempol effectively decreased the activities of plasma CK and LDH as well as the content of H2O2 and MDA in heart tissue. Tempol could increase ATPase and antioxidant enzyme activities and decreased the expression of HIF-1α and VEGF compared with hypoxia model, while it could further increase the expression of Nrf2 and HO-1. CONCLUSION: Tempol has protective effect on heart injury induced by hypobaric hypoxia in mice. Its mechanism may be attributed to the amelioration of energy metabolism, scavenging free radical, improvement of antioxidant enzyme activity the activation of the Nrf2/HO-1 pathway as well as alleviation of oxidative stress.