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1.
Article in Chinese | WPRIM | ID: wpr-912063

ABSTRACT

Objective:To investigate the effects and regulation mechanism of lipid-associated membrane proteins (LAMPs) derived from Mycoplasma pneumoniae( Mp) on the expression of quinine oxidoreductase 1 (NQO-1) in human monocyte cell line THP-1 cells, and to know the effect of NQO-1 to interleukin 8 secretion in LAMPs stimulated cells, so as to better understand the regulation mechanism upon Mp infection. Methods:Mp were cultivated and the precipitate was collected to extract LAMPs. The cytotoxicity of LAMPs to THP-1 cells was analyzed by using CCK8 test. THP-1 cells were cultured in vitro with different concentrations of LAMPs for different times, and the expression of NQO-1 protein was detected by Western blot. Nrf2 siRNA was used to investigate the role of Nrf2 in NQO-1 expression in LAMPs induced cells, and NQO-1 inhibitor Diminutol was performed to test whether they blocked interleukin 8 (IL-8) secretion when treated with LAMPs in THP-1 cells. Results:LAMPs extracted from Mp had no cytotoxicity to THP-1 cells. The expression of NQO-1 protein in LAMPs-stimulated THP-1 cells showed a dose-dependent and time-dependent manner. The production of NQO-1 protein reached peaks when treated with 5.0 μg/ml or 7.5 μg/ml of LAMPs for 12 h. Silencing of Nrf2 by siRNA significantly decreased NQO-1 production, and blocking NQO-1 by Dim increased the level of IL-8 in LAMPs-stimulated cells. Conclusions:LAMPs derived from Mp induced the expression of NQO-1 protein in THP-1 cells via Nrf2, and NQO-1 can inhibit IL-8 secretion in LAMPs stimulated monocytes.

2.
Chinese Journal of Endemiology ; (12): 627-634, 2021.
Article in Chinese | WPRIM | ID: wpr-909066

ABSTRACT

Objective:To observe the role of nuclear factor erythroid 2-related factor 2 (Nrf2) in regulating apoptosis during malignant transformation of human bronchial epithelial cells (HBE cells) induced by sodium arsenite (NaAsO 2). Methods:HBE cells were treated with 0.0 and 1.0 μmol/L NaAsO 2, which were control group and arsenic exposed group respectively. HBE cells were treated with 1.0 μmol/L NaAsO 2 for 43 passages to establish a malignant transformation model. The dynamic changes of indexes in different passages (0, 1st, 8th, 15th, 22nd, 29th, 36th, and 43rd) after exposure to NaAsO 2 were monitored, including the apoptosis rate detected by flow cytometry and apoptosis-related proteins and Nrf2 protein detected by Western blotting. Nrf2 siRNA was transfected into malignant transformed HBE cells (T-HBE cells) to silence Nrf2. The silencing effect of Nrf2 protein was verified. And, the apoptosis rate and apoptosis-related proteins were detected. Results:With the increase of arsenic exposure, the apoptosis rates of HBE cells decreased (0, 1, 8, 15, 22, 29, 36 and 43 passages were 0.370 ± 0.029, 0.443 ± 0.069, 0.357 ± 0.046, 0.330 ± 0.016, 0.273 ± 0.050, 0.160 ± 0.024, 0.110 ± 0.022, 0.097 ± 0.012, respectively, Ftrend = 22.981, P < 0.05). Compared with the 0 passage cells, the apoptosis rates of the 22nd, 29th, 36th and 43rd passages in the arsenite group were lower. The differences between them were statistically significant ( P < 0.05). With the increase of arsenic exposure, the expressions of pro-apoptotic proteins caspase-3, cleaved-caspase-3, C/EBP-homologous protein (CHOP) and B-cell lymphoma-2 (Bcl-2) associated X protein (Bax) showed downward trends ( Ftrend = 22.356, 3.738, 6.130, 8.061, P < 0.05), while the anti-apoptotic proteins myeloid cell leukemia 1 protein (Mcl-1) and Bcl-2 showed upward trends ( Ftrend = 58.201, 7.691, P < 0.05). Compared with the 0 passage and the control group of the same passage, from the 22nd passage of caspase-3, cleaved-caspase-3, from the 15th passage of CHOP, Mcl-1, and Bcl-2, from the 29th passage of Bax in the arsenite group, the differences of protein were statistically significant ( P < 0.05). However, there were no significant differences in caspase-8, cleaved-caspase-8, caspase-12 and cleaved-caspase-12 protein expressions in the arsenic group ( P > 0.05). Compared with the 0 passage and the control group of the same passage, from the 8th passage of Nrf2 proteins in the arsenite group, the differences of expressions were statistically significant ( P < 0.05). Compared with T-HBE cells transfected with Con siRNA (control), the apoptosis rate of T-HBE cells transfected with Nrf2 siRNA was higher ( P < 0.05). Compared with T-HBE cells transfected with Con siRNA, the expression levels of Nrf2, Bcl-2 and Mcl-1 in T-HBE cells transfected with Nrf2 siRNA were lower ( P < 0.05), while the expression levels of cleaved-caspase-3/caspase-3, caspase-3, cleaved-caspase-3, CHOP, and Bax were higher ( P < 0.05). Conclusion:Nrf2 may regulate mitochondrial apoptotic pathway through Bcl-2, Mcl-1 and Bax, and endoplasmic reticulum apoptotic pathway through CHOP, so as to inhibit the apoptosis of HBE cells and participate in the process of malignant transformation of HBE cells induced by NaAsO 2.

3.
Article in Chinese | WPRIM | ID: wpr-908530

ABSTRACT

Objective:To study the gene expression of nuclear factor erythroid-2-related factor 2 (Nrf2), glutathione-S-transferase (GST) and interleukin-1 β (IL-1β) in A549 cells exposed to hyperoxia and cell apoptosis after siRNA interference with Nrf2.Method:Normal A549 cell lines were assigned into normoxia+siRNA group, normoxia+control group, hyperoxia+siRNA group and hyperoxia+control group according to whether siRNA interference was used and the exposure environment (normoxia/hyperoxia). The hyperoxia environment contained 95%O 2 and 5%CO 2. The levels of mRNA expression of Nrf2, GST and IL-1β were detected using quantitative real-time polymerase chain reaction (qRT-PCR). Flow cytometry was used to examine cell apoptosis of the hyperoxia+control group and hyperoxia+siRNA group at different time points. Analysis of variance (ANOVA) was used to test the relative gene expression and apoptosis of A549 cells. Result:(1) Compared with the normoxia+control group, the expression of Nrf2 and GST in the hyperoxia+control group was significantly increased ( P<0.05), and the expression of IL-1β was significantly decreased ( P<0.05); the expression of Nrf2 and GST in the normoxia+siRNA group decreased significantly ( P<0.05), while the expression of IL-1β increased significantly ( P<0.05). (2) Compared with the normoxia+siRNA group, Nrf2 expression in the hyperoxia+siRNA group showed no significant changes ( P=0.230), GST expression increased slightly ( P=0.057), and IL-1β expression decreased slightly ( P=0.112). (3) Compared with the hyperoxia+control group, the expression of Nrf2 and GST in the hyperoxia+siRNA group decreased significantly ( P<0.05), and the expression of IL-1β increased significantly ( P=0.042). (4) Compared with the hyperoxia+control group, the apoptosis of A549 cells in the hyperoxia+siRNA group increased significantly at 24 h, 48 h and 72 h ( P<0.05). Conclusion:After interfering with Nrf2, siRNA may regulate the expression of GST and IL-1β, preventing oxidative stress, reducing inflammatory response and inhibiting apoptosis.

4.
Article in Chinese | WPRIM | ID: wpr-906078

ABSTRACT

Objective:To explore the mechanism of Banxia Xiexintang (BXXX) in preventing and treating chronic atrophic gastritis (CAG) through Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant response element (ARE) signaling pathway. Method:SD rats were divided into a normal group (<italic>n</italic>=12) and an experimental group for CAG model induction. The model rats were then randomly divided into a model group, a vatacoenayme (VG) group (60 mg·kg<sup>-1</sup>), and high- (280 mg·kg<sup>-1</sup>), medium- (140 mg·kg<sup>-1</sup>), and low-dose (70 mg·kg<sup>-1</sup>) BXXX groups. The doses in the BXXX groups were equivalent to 28, 14, and 7 g·kg<sup>-1</sup> crude drugs. The rats in the normal group and the model group received distilled water at an equal volume, and those in the VG group and the BXXX groups were treated correspondingly by gavage. After 12 weeks of treatment, hematoxylin-eosin (HE) staining was carried out to observe pathological changes in the gastric mucosa of CAG rats. Western blot and real-time fluorescence-based quantitative PCR was used to detect the protein and mRNA expression levels of Nrf2, glutathione S-transferase (GST), and NAD (P)H:quinone oxidoreductase 1 (NQO1) in the gastric mucosa of CAG rats. Result:Compared with the normal group, the model group showed increased protein and mRNA expression levels of Nrf2, NQO1, and GST in the gastric mucosa of the rats (<italic>P</italic><0.05), atrophic gastric mucosa, and even intestinal metaplasia. The protein and mRNA expression levels of Nrf2, NQO1, and GST in the VG group and the high- and medium-dose BXXX groups were lower than those in the model group (<italic>P</italic><0.05), and gastric mucosa atrophy and intestinal metaplasia were significantly improved, especially in the high-dose BXXX group. However, the effect in the low-dose BXXX group was not significant. Conclusion:BXXX can blunt the transcriptional activity of Nrf2, shut down Nrf2 signaling pathway, and reduce the expression levels of NQO1 and GST to achieve normal oxidation-anti-oxidation balance, which may be one of its action mechanisms in the treatment of CAG.

5.
Article in Chinese | WPRIM | ID: wpr-906019

ABSTRACT

Objective:To investigate the renoprotective effects that Sanjiao Qushi prescription ameliorates cationic bovine serum albumin (C-BSA) induced membranous nephropathy(MN) in mice model and its influence on nuclear factor erythroid-2-related factor-2(Nrf2)/heme oxygenase-1(HO-1) signaling pathway. Method:Sixty female BALB/c mice were randomly divided into the normal group(<italic>n</italic>=10) and the model group(<italic>n</italic>=50). The mice in the model group received C-BSA injection via tail vein (6.5 mg·kg<sup>-1</sup>). The mice that were successfully modeled were randomized into the model group, the low dose Sanjiao Qushi prescription group(3.71 g·kg<sup>-1</sup>), the high dose Sanjiao Qushi prescription group(7.42 g·kg<sup>-1</sup>) and benazepril hydrochloride group(1.3 mg·kg<sup>-1</sup>). And they were administered with the corresponding medicine by gavage once a day for four consecutive weeks. 24 hour-urine protein quantitation were performed before C-BSA injection and after C-BSA injection as well as the medicine gavage. When the treatment was finished, all of the mice were sacrificed and the biochemical indicators such as serum creatinine(SCr), blood urea nitrogen(BUN), triglyceride(TG), total cholesterol(TC), total protein(TP) and albumin(Alb) were measured. And the renal pathological morphology changes were observed by light microscope with hematoxylineosin(HE), Masson and periodic acid-silver metheramine(PASM) staining. The deposition of immunoglobulin G(IgG) in the glomerulus was detected by fluorescence microscope. The expression of reactive oxygen species (ROS) of kidney was detected by fluorescence immunoassay. The protein expression levels of Nrf2 in cell nucleus and cytoplasm and the downstream protein factors HO-1 and NADH quinone acceptor oxidoreductase 1(NQO1) were detected by Western blot. Result:Compare to normal group, the levels of 24 hour-urine protein quantitation, TG and TC significantly increased in model group(<italic>P</italic><0.01), while TP and Alb levels significantly decreased(<italic>P</italic><0.01). The model group exhibited enlarged volume of glomerular, significantly thickened glomerular basement membrane(GBM), fuchsinophilic protein deposition and spike formation through light microscope. Immunofluorescence staining for the model group exhibited granular deposition of IgG along the capillary wall. The expression of ROS in kidney significantly increased(<italic>P</italic><0.01). The protein expression levels of Nrf2 in cell nucleus significantly increased(<italic>P</italic><0.01), while Nrf2 in cytoplasm significantly decreased(<italic>P</italic><0.01).The protein expression levels of HO-1 and NQO1 significantly increased(<italic>P</italic><0.01). Compared to model group, the levels of 24 hour-urine protein quantitation, TG and TC significantly decreased in each treated group(<italic>P</italic><0.01), TP and Alb levels significantly increased(<italic>P</italic><0.05,<italic>P</italic><0.01). The pathological damages alleviated obviously. The expression of ROS in kidney significantly decreased(<italic>P</italic><0.01). The protein expression levels of Nrf2 in cell nucleus significantly decreased(<italic>P</italic><0.01), while Nrf2 in cytoplasm significantly increased(<italic>P</italic><0.05,<italic>P</italic><0.01). The protein expression levels of HO-1 and NQO1 significantly increased(<italic>P</italic><0.01). Conclusion:Sanjiao Qushi prescription worked on MN mice possibly by regulating related proteins in the Nrf2/HO-1 signaling pathway and relieving oxidative stress, thus decreasing 24 hour-urine protein and blood lipid, increasing serum protein, and alleviating the pathological damages to protect renal function and delay progress of the disease.

6.
Article in Chinese | WPRIM | ID: wpr-905061

ABSTRACT

Objective:To observe the effect of Shaofu Zhuyutang on nuclear factor erythroid-2-related factor 2 (Nrf2) /antioxidant response element (ARE) signaling pathway in blood vessels by establishing the model of rats with cold coagulation and blood stasis syndrome, and to explore the protective effect and mechanism of Shaofu Zhuyutang on vascular endothelial injury. Method:The 50 SPF rats were randomly divided into high dose group (4.8 g·kg-1), middle dose group (2.4 g·kg-1), low dose group (1.2 g·kg-1), model group and normal group (ten of each group). The rat model of cold coagulation and blood stasis syndrome was established by subcutaneous injection of epinephrine hydrochloride combined with ice bath. At the same time of modeling, the drug was administered by gavage. After 28 days of continuous administration, the hemorheology indexes were detected by automatic hemorheology instrument. Levels of nitric oxide (NO), endothelin (ET)-1, superoxide dismutase (SOD), glutathione (GSH-Px), intercellular adhesion molecule-1(ICAM-1), vascular cell adhesion molecule-1(VCAM-1), von Willebrand factor (vWF) in serum were determined by ELISA. Hematoxylin and eosin (HE) staining was used to observe the endothelial injury of vascular tissue of thoracic aorta. The protein expression of Nrf2 and HO-1 in vascular tissue of thoracic aorta was detected by Western blot. Real-time fluorescence quantitative polymerase chain reaction(Real-time PCR)was used to observe the expression of Nrf2 and heme oxygenase-1 (HO-1) mRNA in vascular tissue of thoracic aorta. Result:Compared with the blank group, model group rats whole blood viscosity and plasma viscosity were significantly increased (P<0.05,P<0.01), vWF, ICAM 1, VCAM 1 content increased significantly (P<0.01), NO, SOD, gsh-px levels decreased significantly (P<0.01), significantly increased the content of ET-1(P<0.01), thoracic aorta vascular tissue Nrf2, HO-1 mRNA expression was significantly increased (P<0.01), Nrf2 protein expression in the cell nucleus increased significantly (P<0.05), The protein expression level of Nrf2 in cytoplasm was significantly decreased (P<0.05), while the protein expression level of HO-1 was significantly increased (P<0.01). Compared with model group, the whole blood viscosity (high and middle cut), plasma viscosity, were significantly reduced in high and meduim-dose Shaofu Zhuyutang groups(P<0.05,P<0.01). The levels of vWF, ICAM-1, VCAM-1 and ET-1 in serum were significantly reduced (P<0.05,P<0.01), NO, SOD and GSH-Px increased significantly (P<0.05,P<0.01). The pathological changes such as hyperplasia, swelling and shedding of endothelial cells of thoracic aorta, rupture of internal elastic membrane and disorder of smooth muscle arrangement were improved. The expression levels of Nrf2, HO-1 protein and gene were significantly increased in vascular tissue of thoracic aorta (P<0.01). Conclusion:Shaofu Zhuyutang has a protective effect on vascular endothelial injury in rats with cold coagulation and blood stasis syndrome. The mechanism of action is related to the activation of Nrf2/ARE signaling pathway, which leading to the increased expression of antioxidant enzymes and decreased the expression of adhesion factors.

7.
Acta Pharmaceutica Sinica B ; (6): 3791-3805, 2021.
Article in English | WPRIM | ID: wpr-922441

ABSTRACT

Acetaminophen (APAP) overdose can induce liver injury and is the most frequent cause of acute liver failure in the United States. We investigated the role of p62/SQSTM1 (referred to as p62) in APAP-induced liver injury (AILI) in mice. We found that the hepatic protein levels of p62 dramatically increased at 24 h after APAP treatment, which was inversely correlated with the hepatic levels of APAP-adducts. APAP also activated mTOR at 24 h, which is associated with increased cell proliferation. In contrast, p62 knockout (KO) mice showed increased hepatic levels of APAP-adducts detected by a specific antibody using Western blot analysis but decreased mTOR activation and cell proliferation with aggravated liver injury at 24 h after APAP treatment. Surprisingly, p62 KO mice recovered from AILI whereas the wild-type mice still sustained liver injury at 48 h. We found increased number of infiltrated macrophages in p62 KO mice that were accompanied with decreased hepatic von Willebrand factor (VWF) and platelet aggregation, which are associated with increased cell proliferation and improved liver injury at 48 h after APAP treatment. Our data indicate that p62 inhibits the late injury phase of AILI by increasing autophagic selective removal of APAP-adducts and mitochondria but impairs the recovery phase of AILI likely by enhancing hepatic blood coagulation.

8.
Acta Pharmaceutica Sinica B ; (6): 3740-3755, 2021.
Article in English | WPRIM | ID: wpr-922437

ABSTRACT

Acetaminophen (APAP) is a widely used analgesic and antipyretic drug, which is safe at therapeutic doses but can cause severe liver injury and even liver failure after overdoses. The mouse model of APAP hepatotoxicity recapitulates closely the human pathophysiology. As a result, this clinically relevant model is frequently used to study mechanisms of drug-induced liver injury and even more so to test potential therapeutic interventions. However, the complexity of the model requires a thorough understanding of the pathophysiology to obtain valid results and mechanistic information that is translatable to the clinic. However, many studies using this model are flawed, which jeopardizes the scientific and clinical relevance. The purpose of this review is to provide a framework of the model where mechanistically sound and clinically relevant data can be obtained. The discussion provides insight into the injury mechanisms and how to study it including the critical roles of drug metabolism, mitochondrial dysfunction, necrotic cell death, autophagy and the sterile inflammatory response. In addition, the most frequently made mistakes when using this model are discussed. Thus, considering these recommendations when studying APAP hepatotoxicity will facilitate the discovery of more clinically relevant interventions.

9.
Acta Pharmaceutica Sinica B ; (6): 1148-1157, 2021.
Article in English | WPRIM | ID: wpr-881190

ABSTRACT

As one of the most lethal diseases, pancreatic cancer shows a dismal overall prognosis and high resistance to most treatment modalities. Furthermore, pancreatic cancer escapes early detection during the curable period because early symptoms rarely emerge and specific markers for this disease have not been found. Although combinations of new drugs, multimodal therapies, and adjuvants prolong survival, most patients still relapse after surgery and eventually die. Consequently, the search for more effective treatments for pancreatic cancer is highly relevant and justified. As a newly re-discovered mediator of gasotransmission, hydrogen sulfide (H

10.
Acta Pharmaceutica Sinica B ; (6): 89-99, 2021.
Article in English | WPRIM | ID: wpr-881126

ABSTRACT

Acetaminophen (APAP) overdose is the leading cause of drug-induced liver injury, and its prognosis depends on the balance between hepatocyte death and regeneration. Sirtuin 6 (SIRT6) has been reported to protect against oxidative stress-associated DNA damage. But whether SIRT6 regulates APAP-induced hepatotoxicity remains unclear. In this study, the protein expression of nuclear and total SIRT6 was up-regulated in mice liver at 6 and 48 h following APAP treatment, respectively.

11.
International Eye Science ; (12): 21-26, 2021.
Article in Chinese | WPRIM | ID: wpr-837709

ABSTRACT

@#AIM: To investigate the protective effect and mechanism of luteolin on H2O2-induced oxidative damage of retinal pigment epithelium(RPE)cells. <p>METHODS:ARPE-19 cells were divided into the control group, H2O2 group, different doses of luteolin groups and Nrf2 inhibitor group, and the oxidative damage model of RPE was prepared by 100μmol/L H2O2, except for the control group. Cell activity was detected by Methyl thiazolyl tetrazolium(MTT)assay and proper experimental concentration of luteolin was determined. The cell morphology and activity was observed in each group. Cell apoptosis rate and reactive oxygen species(ROS)were detected by flow cytometry, malondialdehyde(MDA)and superoxide dismutase(SOD)were detected by kit method, and the expression of caspase-3, poly adeno-sine diphosphate ribose polymerase(PARP), B cell lymphoma-2(Bcl-2), nuclear factor erythroid 2-related factor 2(Nrf2)and heme oxygenase-1(HO-1)proteins were detected by Western blot. <p>RESULTS: 100μmol/L luteolin has toxic effects on ARPE-19 cells, so 25μmol/L and 50μmol/L luteolin were selected for subsequent experiments. The cell activity, SOD activity and the protein expression levels of Bcl-2, Nrf2, HO-1 in 25μmol/L and 50μmol/L luteolin groups were significantly higher than the H2O2 group(<i>P</i><0.05). The apoptosis rate, ROS, MDA content and the protein expression levels of Caspase-3 and PARP in 25μmol/L and 50μmol/L luteolin groups were significantly lower than the H2O2 group(<i>P</i><0.05). The cell activity, SOD activity \〖(13.83±1.49)U/mL <i>vs</i>(22.69±1.83)U/mL\〗 and the protein expression levels of Bcl-2, Nrf2 and HO-1 protein expression in the Nrf2 inhibitor group were significantly lower than the 50μmol/L luteolin group(<i>P</i><0.05). The apoptosis rate, ROS, MDA content \〖(654.96±26.99)<i>vs</i>(446.52±29.42),(3.89±0.29)nmol/mL <i>vs</i>(2.06±0.19)nmol/mL\〗 and the protein expression levels of Caspase-3 and PARP in the Nrf2 inhibitor group were significantly higher than the 50μmol/L luteolin group(<i>P</i><0.05).<p>CONCLUSION: Luteolin can improve the oxidative damage of RPE cells induced by H2O2, and its mechanism may be related to the activation of the Nrf2/HO-1 pathway.

12.
Article in English | WPRIM | ID: wpr-888500

ABSTRACT

Atherosclerosis is a common pathological change in cardiovascular disease. Vascular smooth muscle cell is the main source of plaque cell and extracellular matrix, and nuclear factor-erythroid 2-related factor 2 (Nrf2) is a key transcription factor regulating the function of vascular smooth muscle cell. In the process of atherosclerosis, Nrf2 signaling pathway has the following regulatory effects on vascular smooth muscle cell: regulating the phenotype of vascular smooth muscle cell to change to the direction conducive to the alleviation of disease progression; inhibiting the proliferation and migration of vascular smooth muscle cell; mitigating the level of blood lipid; alleviating vascular smooth muscle cell calcification, aging and apoptosis process. This article reviews the specific mechanisms of Nrf2 regulating atherosclerosis, such as phenotypic transformation, proliferation and migration, lipid metabolism, calcification, aging and apoptosis in atherosclerosis, in order to provide a basis for understanding the molecular mechanism of atherosclerosis development and finding therapeutic targets.


Subject(s)
Atherosclerosis , Cell Movement , Cell Proliferation , Cells, Cultured , Humans , Muscle, Smooth, Vascular , Myocytes, Smooth Muscle , NF-E2-Related Factor 2/metabolism , Signal Transduction
13.
International Eye Science ; (12): 1156-1161, 2021.
Article in Chinese | WPRIM | ID: wpr-877371

ABSTRACT

@#AIM:To investigate the protective effects of naringin(Nar)phospholipid complex(NPC)on oxidative injury in retinal pigment epithelium cells(ARPE-19 cells)induced by tert-butyl hydroperoxide(t-BHP)and elucidate the underlying mechanism.<p>METHODS:The NPC was prepared by solvent method. Experimental cells are divided into seven groups: control group \〖cultured with dimethylsulfoxide(DMSO)\〗, model group(intervention with 200μmol/L t-BHP), nuclear factor erythroid 2-related factor 2(Nrf2)-siRNA group(cell transfection for Nrf2 gene), naringin group(add 200μmol/L t-BHP after pretreatment with 200μmol/L naringin medium), NPC group(add 200μmol/L t-BHP after pretreatment with 200μmol/L NPC medium), Nrf2-siRNA+ naringin group(after 200μmol/L naringin pretreatment, Nrf2 gene interference, then add 200μmol/L t-BHP)and Nrf2-siRNA+ NPC group(after 200μmol/L NPC pretreatment, Nrf2 gene interference, then add 200μmol/L t-BHP). The intracellular levels of superoxide dismutase(SOD), malondialdehyde(MDA)and total antioxidant capacity(T-AOC)were detected, intracellular level of reactive oxygen species(ROS)was detected by DCFH-DA staining method. The mRNA and protein expressions of HO-1, NQO-1, GCL and Nrf2 were detected by real-time PCR and western blot, respectively. <p>RESULTS:NPC more significantly increased the levels of SOD and T-AOC, reduced the contents of ROS and MDA than naringin in t-BHP-treated ARPE-19 cells. After naringin and NPC pre-protected ARPE-19 cells, the relative expression and protein expression of Nrf2, HO-1, NQO-1 and GCL mRNA were higher than those of the model group and Nrf2-siRNA group. There were statistically significant differences in the relative expression of 4 genes and the expression levels of 4 proteins in the naringin group and the NPC group, the Nrf2-siRNA+naringin group and the Nrf2-siRNA+NPC group. The expression of Nrf2, HO-1 and NQO-1 protein in the Nrf2-siRNA+naringin group was not significantly different than that in the Nrf2-siRNA group. Compared with the Nrf2-siRNA group, the expression of 4 proteins in the Nrf2-siRNA+NPC group was statistically significant, and the effect of NPC was significantly stronger than that of naringin.<p>CONCLUSION: After naringin forms a phospholipid complex, it can significantly increase the antioxidant capacity in cells and reduce the oxidation level. It up-regulates the expression of Nrf2 and its downstream antioxidant enzymes and phase Ⅱ detoxification enzymes by activating the Nrf2/ARE antioxidative stress pathway to better protect ARPE-19 cells from oxidative damage.

14.
International Eye Science ; (12): 1051-1055, 2021.
Article in Chinese | WPRIM | ID: wpr-876754

ABSTRACT

@#AIM: To detect the relative expression levels of microRNA-27a(miR-27a)and nuclear factor erythroid-2-related factor 2(NRF2)in serum of patients with age-related macular degeneration(ARMD)bleeding, and to explore the correlation between the expression levels and the prognosis of ARMD bleeding. <p>METHODS: A retrospective case series observation was carried out.From June 2018 to October 2019, 80 patients with ARMD bleeding who were treated in our hospital were selected as ARMD bleeding group, and 80 healthy people who had routine examination in our hospital were selected as control group. The relative expression levels of miR-27a and NRF2 were detected by real-time fluorescent quantitative PCR(qRT-PCR), the diagnostic value of serum miR-27a and NRF2 expression for ARMD bleeding was evaluated by receiver operating characteristic curve(ROC). The incidence of poor prognosis was analyzed; in addition, Logistic regression was used to analyze the influencing factors of poor prognosis in patients with ARMD bleeding. <p>RESULTS: The relative expression level of miR-27a in serum of ARMD bleeding group was significantly higher than that of control group(<i>P</i><0.01), and the relative expression level of NRF2 mRNA in serum was significantly lower than that in control group(<i>P</i><0.01). ROC results showed that the AUC of serum miR-27a and NRF2 in the diagnosis of ARMD bleeding was 0.867 and 0.820 respectively, and the cutoff value was 1.10 and 1.08 respectively, at this time, the corresponding sensitivity was 71.3% and 91.3%, and the specificity was 90.0% and 63.7%, respectively. The AUC of serum miR-27a combined with NRF2 in the diagnosis of ARMD bleeding was 0.912, and the corresponding sensitivity and specificity were 86.3% and 85.0%, respectively. The incidence of poor prognosis in high miR-27a group was significantly higher than that in low miR-27a group(<i>P</i><0.05); and the incidence of poor prognosis in high NRF2 group was significantly lower than that in low NRF2 group(<i>P</i><0.05). Logistic analysis showed that the high expression of serum miR-27a was an independent risk factor for poor prognosis in patients with ARMD bleeding, and the high relative expression of NRF2 in serum was a protective factor for the poor prognosis of patients with ARMD bleeding. <p>CONCLUSION: The relative expression level of miR-27a in the serum of patients with ARMD hemorrhage is significantly increased, and the relative expression level of NRF2 is significantly decreased, both of them have certain diagnostic value for ARMD bleeding, and their relative expressions are closely related to the prognosis of patients, which is suggested that miR-27a and NRF2 can be used as potential biological indexes for early diagnosis and prognosis evaluation of ARMD bleeding.

15.
China Occupational Medicine ; (6): 660-665, 2020.
Article in Chinese | WPRIM | ID: wpr-881949

ABSTRACT

OBJECTIVE: To investigate the role of nuclear factor erythroid-2-related factor 2(NRF2) signaling pathway in trichloroethylene(TCE) induced oxidative stress in human liver cancer HepG2 cells. METHODS: HepG2 cells in the exponential growth phase were randomly divided into control group and low-, medium-and high-dose groups, and the cells were stimulated with TCE at the final concentrations of 0, 2, 4 or 8 mmol/L respectively for 24 hours. After TCE exposure, the cells were collected. The activity of superoxide dismutase(SOD) was measured by 2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulphenyl)-2 h-tetrazole monosodium salt method.The activities of catalase(CAT) and glutathione peroxidase(GSH-Px) were detected by enzymatic method. The level of malondialdehyde(MDA) was detected by thiobarbituric acid method. The level of 8-hydroxydeoxyguanosine(8-OHDG) was detected by enzyme-linked immunosorbent assay. The mRNA expression of NRF2, heme oxygenase(HO1), glutamate-cysteine ligase catalytic subunit(GCLC), NAD(P) quinone oxidoreductase 1(NQO1) were detected by real-time polymerase chain reaction and the protein expression of NRF2, HO1, GCLC, NQO1 were detected by Western blotting. RESULTS: The activity of CAT in HepG2 cells decreased in the 3 doses drug exposure groups(all P<0.05). The activity of GSH-Px increased(all P<0.05), while the level of MDA in HepG2 cells decreased in low-dose group compared with the control group(P<0.05). There was no statistical significance in SOD activity and 8-OHDG level of HepG2 cells among these 4 groups(all P>0.05). The mRNA and protein relative expression of NRF2 and GCLC in HepG2 cells decreased in the low-and medium-dose groups(all P<0.05) compared with the control group. The mRNA relative expression of HO1 decreased(all P<0.05). The HO1 protein relative expression of HepG2 cells increased in low-dose group compared with the control group(P<0.05). The mRNA and protein relative expression of NQO1 in low-dose group increased(both P<0.05). The protein relative expression of HO1 in HepG2 cells in medium-dose group was lower than that in control group(P<0.05).The mRNA and protein relative expression levels of NRF2 and NQO1 increased in HepG2 cells of high-dose group(all P<0.05) and the mRNA relative expression of HO1 increased in HepG2 cells of high-dose group(all P<0.05) compared with the other 3 groups. CONCLUSION: The low and medium dose TCE stimulation can cause oxidative stress in HepG2 cells and decrease the antioxidant enzyme activity. The high dose TCE stimulation to HepG2 cells can activate NRF2 signaling pathway, thus upregulating the expression of downstream antioxidant enzymes HO1, GCLC and NQO1, and relieving the oxidative damage caused by TCE.

16.
China Occupational Medicine ; (6): 650-655, 2020.
Article in Chinese | WPRIM | ID: wpr-881947

ABSTRACT

OBJECTIVE: To explore the role of N~6-methyladenosine(m~6A) catalytic enzymes(methyltransferases and demethylases) in cadmium-induced oxidative damage in human renal epithelial cells(HK-2 cells), and to analyze the correlation between nuclear factor-erythroid 2-related factor 2(NRF2) and m~6A catalytic enzymes. METHODS: i) HK-2 cells in logarithmic growth phase were randomly divided into control group and 6 cadmium sulfate treatment groups, then treated with 0, 2, 4, 8, 16, 32 and 64 μmol/L cadmium sulfate solution for 24 hours. The cell survival rates were detected by CCK-8 assay, and the appropriate doses of cadmium sulfate were selected for subsequent experiments. ii) HK-2 cells in logarithmic growth phase were randomly divided into control group and low-, medium-, and high-dose groups, and treated with 0, 4, 8, and 16 μmol/L cadmium sulfate solution respectively for 24 hours. Subsequently, the levels of reactive oxygen species(ROS) were detected by fluorescence probe. The mRNA expression of NRF2, the m~6A methyltransferases such as methyltransferase like proteins(METTL) 3, METTL14, METTL16 and the m~6A demethylases such as fat mass and obesity associated protein(FTO), AlkB family of nonheme Fe(Ⅱ)/α-ketoglutarate(α-KG)-dependent dioxygenases 5(ALKBH5) were determined by real-time polymerase chain reaction. RESULTS: i) The survival rate of HK-2 cells was more than 60.00% and lower than that of the control group(P<0.05) after the cells were stimulated with 16 μmol/L of cadmium sulfate. Therefore, 4, 8 and 16 μmol/L of cadmium sulfate were selected as the stimulation concentrations in the follow-up experiments. ii) The relative expression of NRF2, METTL3, METTL14 and METTL16 in HK-2 cells in low-dose group increased(all P<0.05), while the levels of ROS and the relative mRNA expression of NRF2, METTL3, METTL14, METTL16 and FTO in HK-2 cells in medium and high-dose groups increased(all P<0.05) when compared with the control group. There was no significant difference in the expression of ALKBH5 mRNA among these 4 groups(P>0.05). In the correlation analysis, NRF2 mRNA expression was positively correlated with the mRNA expression of METTL3 and METTL16 [Pearson correlation coefficient(r) = 0.61 and 0.66, respectively, all P<0.05]. There was no correlation between NRF2 mRNA expression and METTL14, FTO and ALKBH5(r=0.53, 0.48, and 0.01 respectively, all P>0.05). CONCLUSION: Cadmium sulfate may increase intracellular ROS level, up-regulate NRF2 expression and activate NRF2 signaling pathway as well as enhance the expression of METTL3 and METTL16 in HK-2 cells, thus increasing intracellular oxidative damage and decreasing the cell survival rate.

17.
Article in English | WPRIM | ID: wpr-846971

ABSTRACT

The organosulfur compound sulforaphane (SFN; C6H11NOS2) is a potent cytoprotective agent promoting antioxidant, anti-inflammatory, antiglycative, and antimicrobial effects in in vitro and in vivo experimental models. Mitochondria are the major site of adenosine triphosphate (ATP) production due to the work of the oxidative phosphorylation (OXPHOS) system. They are also the main site of reactive oxygen species (ROS) production in nucleated human cells. Mitochondrial impairment is central in several human diseases, including neurodegeneration and metabolic disorders. In this paper, we describe and discuss the effects and mechanisms of action by which SFN modulates mitochondrial function and dynamics in mammalian cells. Mitochondria-related pro-apoptotic effects promoted by SFN in tumor cells are also discussed. SFN may be considered a cytoprotective agent, at least in part, because of the effects this organosulfur agent induces in mitochondria. Nonetheless, there are certain points that should be addressed in further experiments, indicated here as future directions, which may help researchers in this field of research.

18.
Article in Chinese | WPRIM | ID: wpr-873273

ABSTRACT

Cardiovascular disease is the leading cause of death in China. Due to its great individual differences in genetic background, pathogenesis and disease development trend, the survival risk rate after standardized western medicine treatment under the guidance of the current guidelines remains high. Traditional Chinese medicine(TCM) has unique multiple-target, multiple-pathway and multiple-layer advantages, which can effectively make up for shortcomings of western medicine. Therefore, it has been widely used in the treatment of cardiovascular diseases. Oxidative stress is one of the important causes of cardiovascular diseases. Nuclear factor erythroid-2-related factor 2(Nrf2) is the central regulator of this reaction. When being activated, it can transfer to the nucleus and initiate signaling in the downstream pathway, thus playing an anti-oxidative stress role. As one of the most important endogenous protection systems in the body, the Nrf2/heme oxygenase-1 (Nrf2/HO-1) signaling pathway is the most classical approach for Nrf2 in playing roles. There have been certain achievements in studying and clarifying TCM by regulating this pathway to treat cardiovascular diseases using modern molecular biology and other methods. Based on this, this paper summarized the relationship between Nrf2/HO-1 signaling pathway and cardiovascular diseases, then concluded and analyzed the mechanism and pharmacological effects of TCM and its active ingredients in Nrf2/HO-1 signaling pathway on different cardiovascular diseases, involving active ingredients of TCM, TCM pairs active ingredients, TCM extracts and TCM formula. This paper provides a theoretical reference for the development and utilization of anti-cardiovascular drugs.

19.
China Pharmacy ; (12): 2386-2391, 2020.
Article in Chinese | WPRIM | ID: wpr-825896

ABSTRACT

OBJECTIVE:To study the improvement effects of icariside Ⅱ(ICS Ⅱ)on neurological function of focal cerebral ischemia model rats by regulating miR- 141-3p/Notch/nuclear factor erythroid- 2-related factor 2(Nrf2)axis(miR-141-3p/Notch/ Nrf2). METHODS :The rats were divided into sham operation group ,model group ,nimodipine group (20 mg/kg)and ICS Ⅱ low-dose,medium-dose and high-dose groups (4,8,16 mg/kg),with 20 rats in each group. Twenty-four hours after establishing focal cerebral ischemia model ,model rats were given re levant medicine or normal saline intragastrically ,twice a day ,for consecutive 3 d. The neurological deficit of rats in each group was scored ;the volume of cerebral infarction was measured by 2,3, 5-triphenyltetrazolium chloride (TTC)staining;water content of cerebral tissue and the permeability of blood-brain barrier were measured;HE staining was performed to observe the pathological change of cerebral tissue of rats ;the expression of miR- 141-3p in cerebral tissue of rats was measured by qRT-PCR ;the protein expression of Notch and Nrf 2 in cerebral tissue of rats were measured by Western blotting assay. RESULTS :Compared with sham operation group ,the neurological deficit score ,expression of Notch-1 and Nrf 2 in model group were significantly lowered (P<0.05);infarction volume ,brain water content ,the permeability of blood-brain barrier and the expression of miR- 141-3p in cerebral tissue were increased significantly (P<0.05);the distribution of cortical cells was disordered ,and inflammatory infiltration and necrosis were observed in a large number of nerve cells. Compared with model group ,the neurological deficit score ,the protein expression of Notch- 1 and Nrf 2 in cerebral tissue were significantly increased in ICS Ⅱgroups(P<0.05);infarction volume ,brain water content ,the permeability of blood-brain barrier and the expression of miR- 141-3p in cerebral tissue were decreased significantly (P<0.05);the arrangement of cortical cells was regular,and the inflammatory infiltration and necrosis of nerve cells were decreased significantly. CONCLUSIONS :ICS Ⅱ can promote the recovery of neurological function in focal cerebral ischemic model rats ,which may be related to down-regulation of miR-141-3p and activation of Notch/Nrf 2 axis.

20.
Article in Chinese | WPRIM | ID: wpr-804817

ABSTRACT

Objective@#To observe the changes of LC3, lc3-Ⅱ/lc3-Ⅰ ratio, Nrf2 and Bcl2 in myocarditis induced by coxsackievirus group B type 3 (CV-B3) infection and myocardial damage in SD rats caused by particulate matter of four different pollution sources, and to further explore the mechanism of autophagy and apoptosis of myocardial cells and myocardial damage.@*Methods@#Adult SD rats were randomly divided into CV-B3 infection group (20 rats), automobile exhaust group (20 rats), coal smoke group (20 rats), burning straw group (20 rats), atmosphere group (20 rats) and control group (20 rats). The expressions of LC3, Bcl2 and Nrf2 in rats were detected by Western blot at 12 hours, 48 hours, 5 days and 10 days.@*Results@#In the first three groups of rats expression of LC3, Bcl2 and Nrf2 was upregulated, this was seen early in CV-B3 group, the peak was high, and recovery was fast; while in automobile exhaust group the above changes appeared later, the amplitude was low; in the coal smoke group rats the above changes appeared even later, but the amplitude of change was higher than that in automobile exhaust group, but lower than that of CV-B3 group. In automobile exhaust and coal smoke groups Bcl2 and Nrf2 expression was still slightly increased at day 10. After 48 hours, the above measurements in rats in the atmosphere group were temporarily up-regulated, and returned to normal on day 5. The above measurements of rats in the straw smoke and the control group did not show significant change.@*Conclusions@#In the SD rats with acute viral myocarditis induced by CV-B3 and myocardial damage induced by automobile exhaust, coal smoke and atmospheric particulate matter, the whole process of metabolism, renewal, repair and anti-damage activity of myocardial cells can be accomplished through autophagy activation, apoptosis inhibition and antioxidant mechanism.

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