Programmed cell death, antioxidant response and oxidative stress in wheat f lag leaves induced by chemical hybridization agent SQ-1

Male sterility induced by a chemical hybridization agent （CHA） is an important tool for utilizing crop heterosis. Leaves, especially the flag leaves, as CHA initial recipients play a decisive role in inducing male sterility. To investigate effects of different treatment times of CHA-SQ-1 used, morphological, biochemical and physiological responses of wheat flag leaves were detected in thistudy. CHA induced programmed cell death （PCD） as shown in terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling （TUNEL） and DNA laddering analysis. In the early phase, CHA-SQ-1 trig- gered organelle changes arid PCD in wheat leaves accompanied by excess production of reactive oxygen species （O2- and H202） and down-regulation of the activities of superoxide dismutase （SOD）, catalase （CAT） and guaiacol peroxidase （POD）. Meanwhile, leaf cell DNAs showed ladder-like patterns on agarose gel, indicating that CHA-SQ-1 led to the activation of the responsible endonuclease. The oxidative stress assays showed that lipid peroxidation was strongly activated and photosynthesis was obviously inhibited in SQ-l-induced leaves. However, CHA contents in wheat leaves gradually reduced along with the time CHA-SQ-1 applied. Young flags returned to an oxidative/antioxidative balance and ultimately developed into mature green leaves. These results provide explanation of the relations between PCD and anther abortion and practical application of CHA for hybrid breeding.

Hepatic Histopathological Characteristics and Antioxidant Response of Phytoplanktivorous Silver Carp Intraperitoneally Injected with Extracted Microcystins

Objective To investigate the hispathological characteristics and antioxidant responses in liver of silver carp after intraperitoneal administration of microcystins （MCs） for further understanding hepatic intoxication and antioxidation mechanism in fish. Methods Phytoplanktivorous silver carp was injected intraperitoneally （i.p.） with extracted hepatotoxic microcystins （mainly MC-RR and -LR） at a dose of 1000 μg MC-LReq./kg body weight, and liver histopathological changes and antioxidant responses were studied at 1, 3, 12, 24, and 48 h, respectively, after injection. Results The damage to liver structure and the activities of hepatic antioxidant enzymes including catalase （CAT）, superoxide dismutase （SOD）, and glutathione peroxide （GPX） were increased in a time-dependent manner. Conclusion In terms of clinical and histological signs of intoxication and LD50 （i.p.） dose of MC-LR, silver carp appears rather resistant to MCs exposure than other fishes. Also the significantly increased SOD activity in the liver of silver carp suggests a higher degree of response to MCs exposure than CAT and GPX.

An Ultimate Approach to Retarding the Aging Process by Integrating Traditional Chinese Medicine and Western Medicine

Aging and death are unavoidable in life. While immortality may be impossible, many people dream of living a long and healthy life. Throughout history, humans have searched for ways to stay young, but have not found an effective way. This may be because the methods used do not target the causes of aging directly. To address this, we investigated how to delay aging using traditional Chinese medicine (TCM) and Western medicine approaches. In this article, we will explain the causes of aging in the context of TCM and Western medicine and suggest methods to delay it. By integrating TCM and Western medicine, I hope to help everyone age healthily and enjoy a long life.

Physiological Changes in Leaves of Medicago falcata Under Low-Temperature Stress

Low temperature significantly restricts crop yield and quality.Medicago falcata(M.falcata)is a typical legume species that exhibits great capacity of tolerance to low temperature.To understand the low-temperature responses in M.falcata,the electrolyte leakage and lipid peroxidation level,and activities of superoxide dismutase(SOD),catalase(CAT)and peroxidase(POD),and contents of reduced glutathione(GSH),soluble protein,soluble sugar and proline were investigated in low-temperature-stressed M.falcata leaves.The electrolyte leakage and malondialdehyde(MDA)content increased,and could be used to quantify low-temperature damage at cellular level.And then,the significant change of SOD,POD and CAT activities,and GSH content reflected the higher reactive oxygen species(ROS)scavenging capacity in M.falcata.In addition,the significant change of soluble protein,soluble sugar and proline contents helped to maintain osmotic equilibrium,energy supply and protein functions.These nine physiological traits were analyzed by gray relational grade analysis and ranked from the highest to the lowest as follows:electrolyte leakage,GSH,proline,soluble protein,MDA,soluble sugar,SOD,CAT and POD,and illustrated that the electrolyte leakage level,GSH and proline contents should be selected and measured priority in M.falcata low-temperature tolerance to improve measurement efficiency.

Sulforaphane protects liver injury induced by intestinal ischemia reperfusion through Nrf2-ARE pathway

AIM: To investigate the effect of sulforaphane (SFN) on regulation of NF-E2-related factor-2 (Nrf2)-antiox-idant response element (ARE) pathway in liver injury induced by intestinal ischemia/reperfusion (I/R). METHODS: Rats were divided randomly into four ex-perimental groups: control, SFN control, intestinal I/R and SFN pretreatment groups (n = 8 in each group). The intestinal I/R model was established by clamping the superior mesenteric artery for 1 h and 2 h reperfu-sion. In the SFN pretreatment group, surgery was performed as in the intestinal I/R group, with intraperitoneal administration of 3 mg/kg SFN 1 h before the op-eration. Intestine and liver histology was investigated. Serum levels of aspartate aminotransferase (AST), and alanine aminotransferase (ALT) were measured. Liver tissue superoxide dismutase (SOD), myeloperoxidase (MPO), glutathione (GSH) and glutathione peroxidase (GSH-Px) activity were assayed. The liver transcription factor Nrf2 and heme oxygenase-1 (HO-1) were determined by immunohistochemical analysis and Western blotting analysis.RESULTS: Intestinal I/R induced intestinal and liver injury, characterized by histological changes as well as a signif icant increase in serum AST and ALT levels (AST: 260.13 ± 40.17 U/L vs 186.00 ± 24.21 U/L, P < 0.01; ALT: 139.63 ± 11.35 U/L vs 48.38 ± 10.73 U/L, P < 0.01), all of which were reduced by pretreatment with SFN, respectively (AST: 260.13 ± 40.17 U/L vs 216.63 ± 22.65 U/L, P < 0.05; ALT: 139.63 ± 11.35 U/L vs 97.63 ± 15.56 U/L, P < 0.01). The activity of SOD in the liver tissue decreased after intestinal I/R (P < 0.01), which was enhanced by SFN pretreatment (P < 0.05). In ad-dition, compared with the control group, SFN markedly reduced liver tissue MPO activity (P < 0.05) and elevat-ed liver tissue GSH and GSH-Px activity (P < 0.05, P < 0.05), which was in parallel with the increased level of liver Nrf2 and HO-1 expression.CONCLUSION: SFN pretreatment attenuates liver injury induced by intestinal I/R in rats, attributable to the antioxidant effect through Nrf2-ARE pathway.

Pachymic Acid Ameliorates Pulmonary Hypertension by Regulating Nrf2-Keap1-ARE Pathway

Objective:Pulmonary hypertension(PH)is a severe pulmonary vascular disease that eventually leads to right ventricular failure and death.The purpose of this study was to investigate the mechanism by which pachymic acid(PA)pretreatment affects PH and pulmonary vascular remodeling in rats.Methods:PH was induced via hypoxia exposure and administration of PA(5 mg/kg per day)in male Sprague-Dawley rats.Hemodynamic parameters were measured using a right ventricular floating catheter and pulmonary vascular morphometry was measured by hematoxylin-eosin(HE),a-SMA and Masson staining.MTT assays and EdU staining were used to detect cell proliferation,and apoptosis was analyzed by TUNEL staining.Western blotting and immunohistochemistry were used to detect the expression of proteins related to the Nrf2-Keapl-ARE pathway.

Induction of Antioxidant and Heat Shock Protein Responses During Torpor in the Gray Mouse Lemur, Microcebus murinus

A natural tolerance of various environmental stresses is typically supported by various cytoprotective mechanisms that protect macromolecules and promote extended viability. Among these are antioxidant defenses that help to limit damage from reactive oxygen species and chaperones that help to minimize protein misfolding or unfolding under stress conditions. To understand the molecular mechanisms that act to protect cells during primate torpor, the present study characterizes antioxidant and heat shock protein（HSP） responses in various organs of control（aroused）and torpid gray mouse lemurs, Microcebus murinus. Protein expression of HSP70 and HSP90 a was elevated to 1.26 and 1.49 fold, respectively, in brown adipose tissue during torpor as compared with control animals, whereas HSP60 in liver of torpid animals was 1.15 fold of that in control（P 〈 0.05）. Among antioxidant enzymes, protein levels of thioredoxin 1 were elevated to 2.19 fold in white adipose tissue during torpor, whereas Cu–Zn superoxide dismutase 1 levels rose to 1.1 fold in skeletal muscle（P 〈 0.05）. Additionally, total antioxidant capacity was increased to 1.6 fold in liver during torpor（P 〈 0.05）, while remaining unchanged in the five other tissues. Overall, our data suggest that antioxidant and HSP responses are modified in a tissue-specific manner during daily torpor in gray mouse lemurs. Furthermore, our data also show that cytoprotective strategies employed during primate torpor are distinct from the strategies in rodent hibernation as reported in previous studies.

Metformin activates Nrf2 signaling and induces the expression of antioxidant genes in skeletal muscle and C2C12 myoblasts

As a first line anti-diabetes drug, the molecular mechanisms by which metformin exerts its pharmacological activities are still under extensive investigations. The Nrf2 signaling plays a crucial role in protecting cells from oxidative damages, and has emerged as a promising target for treatment of diabetes and related complexes in recent years. In the present study, the effect of metformin on Nrf2 signaling was tested in vitro and in vivo, and the possible mechanism was explored. Metformin activated AMPK and Nrf2 signaling and induced the expression of antioxidant genes NQO1 and y-GCSm in C2C12 mouse myoblast cells in a similar concentration- and time-dependent manner. Moreover, overexpression of AMPK significantly elevated the basal and metformin-induced ARE-driven luciferase reporter activities, suggesting the involvement of AMPK in metformin-activated Nrf2 signaling. Finally, metformin activated Nrf2 signaling and induced the expression of antioxidant genes such as HO-1 and SOD, and resulted in increased GSH level in mouse liver and skeletal muscle tissues. Take together, our results clearly demonstrated that metformin activated Nrf2 signaling and enhanced the tissue antioxidant capacity, and provide a new molecular mechanism of action of metformin.

Ectomycorrhizal fungi and dark septate endophyte inoculation improve growth and tolerance of Pinus tabulaeformis under cadmium stress

Forest trees can establish symbiotic associations with dark septate endophytes(DSEs)and ectomycorrhizal fungi(ECMF)simultaneously.However,the combined effects of these two fungi on the growth and cadmium(Cd)tolerance of host plants remain largely unexplored.To address this knowledge gap,a pot experiment was conducted to examine the effects of the interaction between an ECMF strain(Suillus granulatus)and a DSE strain(Pseudopyrenochaeta sp.)on Pinus tabulaeformis under Cd stress,by assessing plant growth and physiological parameters,nutrient uptake,and soil properties.Notably,the colonization rates of both fungal strains were found to increase in response to Cd stress,with the extent of this increase being influenced by the specific fungal species and the Cd level in the soil.Compared to the non-inoculation treatment,single inoculation with fungal strain resulted in enhanced biomass,root development,and nutrient contents in P.tabulaeformis seedlings under Cd stress.Furthermore,a synergistic effect was observed when these seedlings were co-inoculated with S.granulatus and Pseudopyrenochaeta sp.,as indicated by significantly greater measurements in various indicators compared to both the single and non-inoculation treatments.Fungal inoculation effectively regulated the antioxidant defense responses and photosynthesis of P.tabulaeformis seedlings subjected to Cd stress,particularly in the co-inoculation treatment.In addition,fungal inoculation facilitated the Cd accumulation in P.tabulaeformis,suggesting a promising potential for the implementation of bioremediation strategies in the areas contaminated with heavy metals.The findings from this study indicate that the utilization of root symbiotic fungi obtained from stress environments could potentially enhance the growth performance and tolerance of P.tabulaeformis towards heavy metals,and co-inoculation of both fungal groups may result in even more pronounced synergistic effects on the overall fitness of the plant.

Effects of extracts from soothing-liver and invigorating-spleen formulas on the injury induced by oxidative stress in the hepatocytes of rats with non-alcoholic fatty liver disease induced by high-fat diet

OBJECTIVE:To investigate effects of the extracts from soothing-liver and invigorating-spleen formulas on the injury due to oxidative stress,mediated by the Nuclear factor-like 2(Nrf2)-Antioxidant response element(ARE) pathway,in the hepatocytes of rats with non-alcoholic fatty liver disease(NAFLD) induced by high-fat diet.METHODS:Soothing-liver and invigorating-spleen formula mixtures were prepared for five groups:normal,model,soothing-liver formula group(SLG),invigorating-spleen formula group(ISG),integrated formula group(IG).The rat model of NAFLD was induced by feeding rats a high-fat diet(HFD).After 16 weeks,the hepatic tissue was examined following Haematoxylin-Eosin(H&E) staining and with Transmission electron microscopy(TEM).Levels of hepatic lipids,serum lipids and hepatic functions were measured using a biochemical analyser.Hepatocytes were isolated from the livers of rats and were identified by cellar immunohistochemistry,cellular immunofluorescence and flow cytometry.The expression levels of Nrf2,Kelch-like epichlorohydrin-associated protein 1(Keap-1),haeme oxygenase-1(HO-1) and NAD(P)H:quinone oxidoreductase 1(NQO1) m RNAs were assessed by real-time fluorescence quantitative PCR.Nrf2,Keap-1,HO-1 and NQO1 proteins were measured by Western blotting.Finally,the levels of oxidative stress factors Superoxide Dismutase(SOD),malonaldehyde(MDA) and Glutathione peroxidase(GSH-Px) in hepatocytes were measured by WST-1,TBA and colorimetry.RESULTS:The H & E and TEM results showed that the NAFLD model rats successfully reproduced typical pathogenetic and histopathological features.The liver function and levels of hepatic lipids and serum lipids from the model rats were dramatically increased.Compared with the model group,the levels of hepatic lipids,serum lipids and hepatic function in the treatment groups were ameliorated to different degrees.The yields of purified hepatocytes in each rat were 4-5 × 108.The viability of the isolated hepatocytes was higher than 95%,with a purity over 93.2%.Cellular immunohistochemistry analysis showed that the hepatocytes were brown,while in the cellular immunofluorescence analysis,the hepatocytes showed green fluorescence.The expression levels of Nrf2,Keap-1,HO-1 and NQO1 m RNA and protein in the hepatocytes were significantly higher in the model group than in the normal group(P < 0.05,P < 0.01).Compared with the model group,the expression of Nrf2,Keap-1,HO-1 and NQO-1 m RNAs and proteins in all treatment groups increased,especially in the IG(P < 0.01).CONCLUSION:The extracts from soothing-liver and invigorating-spleen formulas may protect the liver against the injury induced by oxidative stress in hepatocytes by influencing the Nrf2-ARE pathway,which may be the mechanism having the potential for prevention and treatment of NAFLD.

Growth inhibition and oxidative damage of Microcystis aeruginosa induced by crude extract of Sagittaria trifolia tubers

Aquatic macrophytes are considered to be promising in controlling harmful cyanobacterial blooms. In this research, an aqueous extract of Sagittaria trifolia tubers was prepared to study its inhibitory effect on Microcystis aeruginosa in the laboratory. Several physiological indices of M. aeruginosa, in response to the environmental stress, were analyzed. Results showed that S. trifolia tuber aqueous extract significantly inhibited the growth of M. aeruginosa in a concentration-dependent way. The highest inhibition rate reached 90% after 6 day treatment. The Chlorophyll-a concentration of M. aeruginosa cells decreased from 343.1 to314.2 μg/L in the treatment group. The activities of superoxide dismutase and peroxidase and the content of reduced glutathione in M. aeruginosa cells initially increased as a response to the oxidative stress posed by S. trifolia tuber aqueous extract, but then decreased as time prolonged. The lipid peroxidation damage of the cyanobacterial cell membranes was reflected by the malondialdehyde level, which was notably higher in the treatment group compared with the controls. It was concluded that the oxidative damage of M. aeruginosa induced by S.trifolia tuber aqueous extract might be one of the mechanisms for the inhibitory effects.

Low- and high-dose hydrogen peroxide regulation of transcription factor NF-E2-related factor 2

Background Reactive oxygen species （ROS） may play both physiological and pathophysiological roles. Transcription factor NF-E2-related factor 2 （Nrf2） regulates antioxidant response element （ARE）-mediated genes expression and coordinates induction of chemoprotective proteins in response to physical and chemical stresses. The exact role of Nrf2 in cellular responses to different levels of oxidative stresses remains unknown. Methods Rat pulmonary microvascular endothelial cells were cultured and treated with 0 mmol/L, 0.125 mmol/L, 0.25 mmol/L, 0.5 mmol/L, 1.0 mmol/L and 2.0 mmol/L hydrogen peroxide solution for 2 hours. Nrf2 gene expression was assayed by reverse transcription-PCR, Nrf2-ARE binding activity was assayed with electrophoretic mobility shift assay （EMSA）, and localization of Nrf2 was detected with immunohistochemistry. Results Low and moderate （0.125 mmol/L, 0.25 mmol/L and 0.5 mmol/L） doses hydrogen peroxide exposure of rat pulmonary microvascular endothelial cells led to the nuclear accumulation of Nrf2, increased activity of transcription regulation and up-regulation of ARE-medicated gene expression. In contrast, high doses of hydrogen peroxide （1 mmol/L 2 mmol/L） exposure of the cells led to the nuclear exclusion of Nrf2, decreased activity transcription regulation and down-regulation of ARE-mediated gene expression. Conclusion Low and moderate doses of hydrogen peroxide play protective roles by increasing transcription activity of Nrf2, whereas high- dose hydrogen peroxide plays a deleterious role by decreasing transcription activity of Nrf2.