Hydrogen sulfide protects retinal pigment epithelium cells against ferroptosis through the AMPK- and p62-dependent non-canonical NRF2-KEAP1 pathway.

Oxidative stress-induced ferroptosis of retinal pigment epithelium (RPE) cells contributes to retinal degenerative diseases. The antioxidant molecule hydrogen sulfide (H2S) regulates oxidative stress response, but its effect on the ferroptosis of RPE cells is unclear. In this study, sodium hydrosulfide (NaHS) was used as an exogenous H2S donor to intervene tert-butyl hydroperoxide (t-BHP)-induced ferroptosis of APRE-19 cells. We found that NaHS pretreatment attenuates t-BHP-induced oxidative stress and ferroptosis. Analysis of mRNA-sequencing coupled with FerrDb database identified nuclear factor erythroid-2-related factor 2 (NRF2) as a primary target for the cytoprotective role of H2S. NRF2 inhibitor ML385 reverses the effects of H2S on ferroptosis. Biochemical analysis revealed that H2S stabilizes NRF2. H2S decreases the interaction between NRF2 and KEAP1, but enhances the interaction between KEAP1 and p62. These results suggest that H2S activates the non-canonical NRF2-KEAP1 pathway. Further study demonstrated that H2S stimulates AMPK to interact and phosphorylate p62. Additionally, inhibiting AMPK or knocking down p62 blocks the effects of H2S. We speculate that targeting the non-canonical NRF2-KEAP1 pathway by H2S-based drug may benefit the treatment of retinal degenerative diseases.

Melatonin increases susceptibility to atrial fibrillation in obesity via Akt signaling impairment in response to lipid overload.

Melatonin has been proven to have antiarrhythmic potential; however, several studies have recently challenged this view. Herein, using a mouse model of obesity-induced atrial fibrillation (AF), we tentatively explored whether exogenous melatonin supplementation could increase AF susceptibility in the context of obesity. We observed that an 8-week drinking administration of melatonin (60 µg/ml in water) induced a greater susceptibility to AF in obese mice, although obesity-induced structural remodeling was alleviated. An investigation of systemic insulin sensitivity showed that melatonin treatment improved insulin sensitivity in obese mice, whereas it inhibited glucose-stimulated insulin secretion. Notably, melatonin treatment inhibited protein kinase B (Akt) signaling in the atria of obese mice and palmitate-treated neonatal rat cardiomyocytes, thereby providing an AF substrate. Melatonin increased lipid stress in obesity, as evidenced by elevated lipid accumulation and lipolysis-related gene expression, thus contributing to the impairment in atrial Akt signaling. Taken together, our results demonstrated that melatonin could increase AF susceptibility in obesity, probably due to increased lipid stress and resultant impairment of atrial Akt signaling. Our findings suggest that special precautions should be taken when administering melatonin to obese subjects.

Enhancing wuyiencin productivity of Streptomyces albulus (CK15) by mutagenesis breeding with atmospheric and room temperature plasma.

Streptomyces species are known for their ability to efficiently produce secondary metabolites, including various antibiotics. Wuyiencin, an antibiotic produced by Streptomyces albulus CK15, is commonly used in agriculture to control fungal diseases in crops and vegetables. In this study, we utilized atmospheric and room temperature plasma (ARTP) mutagenesis to generate mutant S. albulus strains with improved fermentation capabilities for wuyiencin production. After mutagenizing the wild-type S. albulus CK15 strain once and conducting two rounds of antimicrobial screening, three genetically stable mutants (M19, M26, and M28) were identified. These mutants showed increased wuyiencin production by 17.4%, 13.6%, and 18.5% in comparison to the CK15 strain in flask culture, respectively. The M28 mutant exhibited the highest wuyiencin activity, producing 1443.0 ± 134.6 U/mL in flask culture and 1673.8 ± 127.4 U/mL in a 5 L fermenter. These results demonstrate that ARTP is an efficient tool for microbial mutation breeding and improving wuyiencin production.

The mechanism of immune related signal pathway Egr2-FasL-Fas in transcription regulation and methylated modification of Paralichthys olivaceus under acute hypoxia stress.

Apoptosis genes Egr2, Fas and FasL are related to immune responses. However, the mechanism of these genes inducing apoptosis in fish are still not very clear. An acute hypoxia treatment (1.73 ± 0.06 mg/L) for 24 h was carried out on Japanese flounder (Paralichthys olivaceus). The increasingly dense apoptotic signals at 3 h, 6 h, 12 h by TUNEL in skeletal muscle indicated that hypoxia could quickly affect muscle growth and development. Furthermore, we concluded that the Egr2-FasL-Fas signal pathway, which was located at the upstream of apoptotic executor protein caspases, was related to the apoptosis by quantitative real-time PCR, protein concentration detection in ELISA and double gene in situ hybridization methods. The mechanism of the pathway was researched in transcription regulation and epigenetic modification by dual-luciferase reporter assay and bisulfite modified method, respectively. Egr2, as a transcription factor, could up-regulate the expression of FasL gene. And its binding site was mainly between -479 to -1 of FasL gene promoter. The 5th CpG dinucleotides (-514) methylation levels in FasL gene were significantly affected by hypoxia, and they were negatively correlated with its expressions. These suggested that the -514 site may be a very important site to regulate the FasL gene expression. Above results, we concluded that hypoxia activated the immune related signal pathway Egr2-FasL-Fas to induced skeletal muscle apoptosis to affect growth and development of Japanese flounder. The study revealed the mechanism of hypoxia induced apoptosis, which could provide a reference for fish immunity and aquaculture management.

Effect of toyF on wuyiencin and toyocamycin production by Streptomyces albulus CK-15.

Streptomyces albulus CK-15 produces various secondary metabolites, including the antibiotics wuyiencin and toyocamycin, which can reportedly control a broad range of plant fungal diseases. The production of these nucleoside antibiotics in CK-15 is regulated by two biosynthesis gene clusters. To investigate the potential effect of toyocamycin biosynthesis on wuyiencin production, we herein generated S. albulus strains in which a key gene in the toyocamycin biosynthesis gene cluster, namely toyF, was either deleted or overexpressed. The toyF deletion mutant ∆toyF did not produce toyocamycin, while the production of wuyiencin increased by 23.06% in comparison with that in the wild-type (WT) strain. In addition, ΔtoyF reached the highest production level of wuyiencin 4 h faster than the WT strain (60 h vs. and 64 h). Further, toyocamycin production by the toyF overexpression strain was two-fold higher than by the WT strain, while wuyiencin production was reduced by 29.10%. qRT-PCR showed that most genes in the toyocamycin biosynthesis gene cluster were expressed at lower levels in ∆toyF as compared with those in the WT strain, while the expression levels of genes in the wuyiencin biosynthesis gene cluster were upregulated. Finally, the growth rate of ∆toyF was much faster than that of the WT strain when cultured on solid or liquid medium. Based on our findings, we report that in industrial fermentation processes, ∆toyF has the potential to increase the production of wuyiencin and reduce the timeframe of fermentation.

Hyper-O-GlcNAcylation impairs insulin response against reperfusion-induced myocardial injury and arrhythmias in obesity.

Myocardial ischemia/reperfusion (I/R) injury is a major determinant of morbidity and mortality in patients undergoing treatment for cardiac disease. A variety of treatments are reported to have benefits against reperfusion injury, yet their cardioprotective effects seem to be diminished in obesity, and the underlying mechanism remains elusive. In this study, we found that db/db mice exhibit cardiac hyper-O-GlcNAcylation. In parallel, palmitate treatment (200 mM; 12 h) in H9c2 cells showed an increase in global protein O-GlcNAcylation, along with an impaired insulin response against reperfusion injury. To investigate whether O-GlcNAcylation underlies this phenomenon, glucosamine was used to increase global protein O-GlcNAc levels. Interestingly, histological staining, electrophysiological studies, serum cardiac markers and oxidative stress biomarker assays showed that preischemic treatment with glucosamine attenuated insulin cardioprotection against myocardial infarction, arrhythmia and oxidative stress. Mechanistically, glucosamine treatment decreased insulin-stimulated Akt phosphorylation, a key modulator of cell survival. Furthermore, inhibition of O-GlcNAcylation via 6-diazo-5-oxo-l-norleucine (DON) apparently increased insulin-induced Akt phosphorylation and restored its cardioprotective response against reperfusion injury in palmitate-induced insulin-resistant H9c2 cells. Our findings demonstrated that obesity-induced hyper-O-GlcNAcylation might contribute to the attenuation of insulin cardioprotection against I/R injury.

Immune correlates of NF-κB and TNFα promoter DNA methylation in Japanese flounder (Paralichthys olivaceus) muscle and immune parameters change response to vibrio anguillarum infection.

Vibrio anguillarum infection can activate NF-κB/TNFα pathway in the immune organs of fish. Fish muscle is also an important immune organ, but the research on its immune function is few. Our aim was to study regulating mechanism of NF-κB and TNFα gene expressions in the muscle of Japanese flounder (Paralichthys olivaceus) which was under Vibrio anguillarum infection (0, 24, 48, 72 and 96 h). The results showed that the expressions of NF-κB and TNFα increased significantly at 48 h, and there was a significant positive correlation between them. In situ hybridization confirmed the co-existence of NF-κB and TNFα genes in Japanese flounder muscle. Interestingly, the expression of the TNFα gene was regulated by the DNA methylation and its methylation level was negatively correlated with the expression. The lowest methylation level of TNFα occurred at 48 h under Vibrio anguillarum infection (P < 0.05). And more, when the fragment (-2122 âˆ¼ -730) was deleted on TNFα gene promoter, double luciferase activity was the highest, indicating that fragment (-730-0) was the transcription factor binding region. The site (-78 ~ -69) on the fragment (-730-0) binding NF-κB was mutated, and double luciferase activity decreased significantly. The results confirmed that the site (-78 ~ -69) was indeed an important binding site for NF-κB. In addition, the activity of TNFα in the serum of Japanese flounder changed with the prolongation of vibrio anguillarum infection, and the concentration of other immune factors such as ALP, ALT, AST and LDH also changed in the muscle under vibrio anguillarum infection. They all showed a trend of first increasing and then decreasing. Above studies implied that Japanese flounder responded to Vibrio anguillarum infection at the immune level with the change of its methylation status and the activation of transcription factor. By studying the mechanism of immune pathways, understanding the response to immune stress is great significant to the research of fish breeding for disease resistance.

Serendipita indica alleviates drought stress responses in walnut (Juglans regia L.) seedlings by stimulating osmotic adjustment and antioxidant defense system.

Juglans regia L. is a good host for Serendipita indica. Under drought condition, seedlings colonized with S. indica showed higher values in plant height, total fresh biomass, root/shoot ratio, relative growth rate, leaf relative water content and chlorophyll content, gas exchange parameters, maximal photochemical efficiency, photochemical quenching, and effective photosystem II quantum yield than the uncolonized seedlings. It suggested beneficial effects of S. indica on host plants' growth and physiological parameters in response to drought. In comparison with the uncolonized seedlings, S. indica-colonized seedlings showed lower levels in hydrogen peroxide, superoxide anion, malondialdehyde, and relative electrical conductivity under drought condition, suggesting the ability of S. indica to prevent or retard the accumulation of reactive oxygen species and to diminish the oxidative injure. Furthermore, walnut seedlings responded to drought by actively accumulating osmotic regulation substances including soluble protein, soluble sugar, and proline. Root colonization with S. indica was more conductive to the accumulation. Moreover, in response to drought stress, walnut seedlings, regardless of colonization, increased activities of superoxide dismutase, catalase, peroxidase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, and glutathione reductase, levels of ascorbate and glutathione, and ratios of reduced ascorbate/dehydroascorbic acid and reduced glutathione/oxidized glutathione in leaves and roots. S. indica colonization induced much more increase in the abovementioned indicators as compared to the uncolonized seedlings. Overall, S. indica colonization alleviated the detrimental effects of drought stress by altering root system, enhancing osmotic adjustment, and repressing the accumulation of reactive oxygen species via stimulating antioxidant system including enzymatic and nonenzymatic components. KEY POINTS: • S. indica stimulated root growth of walnut seedlings under drought condition. • S. indica accelerated osmotic adjustment under drought condition. • S. indica activated antioxidant defense mechanism under drought condition.

Pyroptosis in liver disease.

Pyroptosis is an inflammatory cell death process that is dependent on caspase. Pyroptosis is a specific form of programmed cell death with the morphological characteristics of formation of pores on the cell membrane, cell swelling, and rupture of the plasma membrane. Recent studies have demonstrated that pyroptosis plays an important role in the occurrence and development of liver diseases. Here, we focus on the mechanisms of pyroptosis, as well as on the relationship between pyroptosis and liver diseases.

Morphological and Compositional Analysis of Two Walnut (Juglans regia L.) Cultivars Growing in China.

Two selected walnut cultivars (Xiangling and Jizhaomian) growing in China were analyzed in terms of proximate and mineral composition, fatty acid and amino acid profile. According to the higher kernel rate (61.22%), thinner shell (1.03 mm), higher content of fat (66.93%), protein (20.97%), essential amino acids (29.31% of total amino acids) and minerals such as manganese, zinc and copper, Xiangling would be the more nutritive cultivar with more commercial value. However, Jizhaomian would be the more healthful cultivar based on the higher content of polyunsaturated fatty acids (79.39% of total fatty acids), and it would be more sensitive to rancidity. Jizhaomian also showed higher content in potassium, calcium, magnesium and iron. Genotype is the major source of variability in morphology and chemical compositions. Regardless of the cultivar, consumption of walnuts is desirably encouraged, since all of them contain compounds potentially beneficial to health.

iTRAQ-based proteomic analysis reveals the mechanisms of Botrytis cinerea controlled with Wuyiencin.

BACKGROUND: Grey mould is an important plant disease worldwide, caused by Botrytis cinerea, resulting in serious economic loss. Wuyiencin, a low toxicity, high efficiency, and broad-spectrum agricultural antibiotic, has been demonstrated effectiveness against B. cinerea. RESULTS: Wuyiencin treatment inhibited growth and sporulation of B. cinerea, specifically altering hypha morphology and intracellular structures. These changes were accompanied by differential expression (fold change > 2.0) of 316 proteins identified by iTRAQ-labelling LC-MS/MS analysis (P < 0.05). Up-regulation of 14 proteins, including carbohydrate metabolism proteins and cell wall stabilization proteins, was validated by parallel reaction monitoring (PRM). Down-regulation of 13 proteins was validated by PRM, including regulators of energy metabolism, nucleotide/protein synthesis, and the biosynthesis of mediators of plant stress and decay. CONCLUSION: Our results confirm the inhibitory biological effects of wuyiencin on B. cinereal and elaborate on the differentially expressed proteins and associated pathways implicated in the capacity of wuyiencin to debilitate the growth and pathogenicity of grey mould. This study provides validated candidates for further targeted exploration with the goal of optimizing wuyiencin as a safe, low-toxicity agent for biological control.

Indole-3-acetic acid production by Streptomyces fradiae NKZ-259 and its formulation to enhance plant growth.

BACKGROUND: Indole-3-acetic acid (IAA) is produced by microorganisms and plants via either tryptophan-dependent or tryptophan-independent pathways. Herein, we investigated the optimisation of IAA production by Streptomyces fradiae NKZ-259 and its formulation as a plant growth promoter to improve economic and agricultural development. RESULTS: The maximum IAA yield achieved using optimal conditions was 82.363 µg/mL in the presence of 2 g/L tryptophan after 6 days of incubation. Thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC) analysis of putative IAA revealed an RF value of 0.69 and a retention time of 11.842 min, comparable with the IAA standard. Regarding product formulation, kaolin-based powder achieved a suspension rate of 73.74% and a wetting time of 80 s. This carrier exhibited good shelf life stability for NKZ-259, and the cell population did not decrease obviously over 4 months of storage at 4 °C. In vivo analysis of plant growth promotion showed that tomato seedlings treated with kaolin powder containing NKZ-259 cells displayed a significant increase in root and shoot length of 7.97 cm and 32.77 cm, respectively, and an increase in fresh weight and dry weight of 6.72 g and 1.34 g. Compared to controls, plant growth parameters were increased almost it two-fold. CONCLUSION: Optimising the culture conditions resulted in an almost four-fold increase in IAA secretion by NKZ-259 cells. The results clearly demonstrate that S. fradiae NKZ-259 holds great potential for plant growth promotion and IAA production. Furthermore, kaolin-based powder is an effective carrier for NKZ-259 cells and may be useful for commercial applications.

Characterization of novel DeoR-family member from the Streptomyces ahygroscopicus strain CK-15 that acts as a repressor of morphological development.

Wuyiencin is produced by Streptomyces ahygroscopicus var. wuyiensis, which has been widely used in China as an industrially produced biopesticide to control various fungal diseases. Although its mechanism of action, breeding, and fermentation had been extensively characterized, less is known about the regulatory functions that affect its biosynthesis or morphological development. The wysR3 gene of S. ahygroscopicus strain CK-15, a novel member of the DeoR family of regulatory genes, was assessed to determine its function by gene knockdown. Herein, we demonstrate for the first time that DeoR family proteins derived from the same source are likely to be a single branch in a phylogenetic tree and show that wysR3 acts as a repressor for its morphological development without effecting wuyiencin production. We found that the ΔwysR3 strain can grow quickly to reach a plateau stage of maximum biomass at 60 h, which is ∼12 h faster than the wild-type strain. In the industrial fermentation production process, the ΔwysR3 strain can reduce consumption and save both time and money.

[Screening and identification of Bacillus pumilus producing double active protein of anticoagulation and thrombolysis].

OBJECTIVE: The aim of this study was to screen bacteria that can produce antithrombotic. METHODS: We screened the target bacteria on VY/4 plate and casein plate from more than 20 samples such as water, soil, rabbit manure, sheep manure and deadwood. We detected the antithrombotic activity by fibrin plate and fibrin tube. We identified the target bacteria by morphological characteristics, physical and chemical properties and 16S DNA sequence homology. RESULTS: We obtained 5 strains that can produce antithrombotic. We found that the extracellular protein of strain LDS33 shows both stronger fibrinolytic activity and stronger anticoagulation activity. According to the morphology, physiochemical properties, 16S DNA sequencing and phylogenetic tree, strain LDS33 is identified as Bacillus pumilus. CONCLUSION: Bacillus pumilus LDS33 can produce highly active anticoagulation and thrombolysis double active protein.

Effects of perspective taking on attention bias to body-related information among junior high school students with body image disturbance.

Using lexical judgment tasks, the present study explored whether perspective taking affected attention bias to body-related information among junior high school students with body image disturbance. Experiment 1 examined the junior high school students' attention bias to body schema-related words; the results showed the body image disturbance group responded significantly more quickly to negative body schema-related words than positive words, whereas the control group did not show a significant difference between positive and negative words. In Experiment 2, participants were asked to judge whether the positive or negative body schema-related words were suitable to describe themselves, when adopting their own perspective or that of another person. The results showed that reaction times to negative words were significantly shorter than to positive words when adopting a self-perspective. When taking another's perspective, there was no significant difference of reaction time between positive and negative words. This result demonstrated that perspective taking reduced attention bias to negative body schema-related information among junior high school students with body image disturbance. The present research suggests that guiding adolescents to view themselves from different perspectives can help them form a more accurate and objective body image.

Intermittent fasting attenuates obesity-related atrial fibrillation via SIRT3-mediated insulin resistance mitigation.

OBJECTIVE: Atrial fibrillation (AF) is the most common tachyarrhythmia in urgent need of therapeutic optimization. Obesity engenders AF, and its pathogenesis is closely intertwined with insulin resistance (IR), but mechanism-based management is still underinvestigated. Intermittent fasting (IF) is a novel lifestyle intervention that mitigates IR, a potential AF driver, yet whether IF can prevent obesity-related AF remains elusive. Here, we aimed to evaluate the impacts of short-term IF on AF and to uncover the underlying mechanism. METHODS: We subjected obese mice (high-fat diet for 8-week) to IF (alternative-day fasting for another 5-week) for AF vulnerability and substrate formation assessment, and similarly treated neonatal atrial cardiomyocytes (NRCMs) and fibroblasts (NRCFs) (palmitate, 200 µM) with IF (alternative-day short-term starvation for 8-day) for mechanism investigation. RESULTS: Obese mice were prone to AF and atrial remodeling. IF reduced AF inducibility, duration, and reversed atrial remodeling including channel disturbance, left atrial dilation, cardiac hypertrophy and fibrosis in obese mice independent of weight loss. Mechanistically, IF up-regulated the SIRT3 protein level both in vivo and in vitro, and pharmacologic inhibition (3-(1H-1,2,3-Triazol-4-yl) pyridine, 50 µM) and genetic suppression of SIRT3 could attenuate the IF-mediated benefits against hypertrophy and fibrosis. Furthermore, IF activated AMPK and Akt signaling, two positive downstream targets of SIRT3, and inactivated HIF1α signaling, a negative downstream target of SIRT3 in both obese mice atria and palmitate-treated cells, while inhibition of SIRT3 reversed these effects. CONCLUSION: IF prevents obesity-related AF via SIRT3-mediated IR mitigation, thus representing a feasible lifestyle intervention to improve AF management.

Influence of rootstock on drought response in young 'Gale Gala' apple (Malus domestica Borkh.) trees.

BACKGROUND: Drought is a major environmental stress limiting plant growth, productivity, and survival worldwide. Rootstocks are widely used to enhance plants resistance to drought stresses. This study determined influence of rootstock on drought responses in 1-year-old 'Gale Gala' apple trees grafted onto Malus sieversii or M. hupehensis. RESULTS: Choice of rootstock resulted in differential response to drought stress. Specifically, M. sieversii caused less drought-induced reduction in relative growth rate, biomass accumulation, leaf area, leaf chlorophyll content, relative water content, photosynthesis rate and maximum chlorophyll fluorescence yield but greater increase in whole-plant water use efficiency compared to M. hupehensis. Secondly, compared with M. hupehensis, M. sieversii caused less drought-induced accumulation of reactive oxygen species but more increase in activities of antioxidant enzymes. In addition, xylem sap abscisic acid concentration was greater in trees grafted onto M. hupehensis than in those grafted onto M. sieversii under drought stress. CONCLUSION: 'Gale Gala' trees' response to drought stress was associated with the rootstock's genotype onto which it was grafted. Trees with M. sieversii as rootstock are more drought resistant than trees with M. hupehensis as rootstock, which suggests that M. sieversii can be widely used as rootstock in arid and semi-arid regions.

Pharmacological inhibition of sphingolipid synthesis reduces ferroptosis by stimulating the HIF-1 pathway.

Ferroptosis is crucial to the pathology of many neurological diseases. Here, we found pre-treatment with myriocin, an inhibitor of de novo synthesis of sphingolipid, significantly decreased the erastin- or glutamate-induced ferroptosis of HT22 cells without requiring the recovery of intracellular glutathione. The transcriptome analysis of HT22 cells treated with or without myriocin identified the hypoxia-inducible factor 1 (HIF-1) pathway as a prime and novel drug target. Further study validated that HIF1α was required for the cytoprotective effects of myriocin. Myriocin treatment promoted the expression of HIF-1 pathway effectors including PDK1 and BNIP3 and altered the intracellular levels of glucose metabolites. Additionally, myriocin treatment stabilized HIF1α protein by decreasing its ubiquitination and proteasomal degradation. Similar effects of myriocin on HIF1α stabilization were also found in other mammalian cell lines indicating this is a common mechanism for the cytoprotective role of myriocin.

Gluconolactone Alleviates Myocardial Ischemia/Reperfusion Injury and Arrhythmias via Activating PKCε/Extracellular Signal-Regulated Kinase Signaling.

Gluconolactone (D-glucono-1,5-lactone or GDL) is a food additive which presents in dietary products such as tofu, yogurt, cheese, bread, wine, etc. GDL has long been considered as a free radical scavenger; however, its role in cardioprotection remains elusive. In this study, using a mouse model of myocardial ischemia/reperfusion (I/R) injury and a model of hypoxia/reoxygenation (H/R) in neonatal rat cardiomyocytes (NRCM), we explored the role of GDL in I/R injury. We found that GDL (5 mg/kg, i.p.) attenuated myocardial I/R injury as evidenced by decreased infarct size, release of cardiac injury markers and apoptosis. Additionally, GDL decreased reperfusion-induced arrhythmias and oxidative stress. These effects were also observed in parallel in vitro studies. Mechanistically, we found that GDL treatment was strongly associated with activation of pro-survival extracellular signal-regulated kinase (ERK) signaling both in vivo and in vitro, and pharmacological inhibition of ERK signaling via U0126 attenuated GDL-induced cardioprotection against H/R injury in NRCM cells. To reveal how GDL regulates ERK signaling, we predicted the putative targets of GDL by Swiss Target Prediction, and protein kinase C (PKC) emerged as the most promising target for GDL. By pharmacological intervention and immunofluorescence, we found that PKCε, an important member of the PKC family, was activated after GDL treatment in heart, thereby leading to ERK activation and cardioprotection against I/R injury. Taken together, our results demonstrated that GDL acts as a potent activator of PKCε and, thus, provides cardioprotection against I/R injury via activation of ERK signaling.

DNA methylation in promoter region of immune related genes STAT3 and VEGFA and biochemical parameters change in muscle of Japanese flounder under acute hypoxia.

Acute hypoxic stress can lead to immune response in fish, but the molecular mechanism of muscle immunity in fish under acute hypoxia are still unclear. In this study, we carried out the effect of signal transducer and activator of transcription3(STAT3) and vascular endothelial growth factor A(VEGFA) on muscle immune responses of Japanese flounder (Paralichthys olivaceus) during acute hypoxic stimulation (1.65 ± 0.28mg/L O2; 3h, 6h, 12h, 24h) and reoxygenation (7.30 ± 0.40mg/L O2; R12h, R24h, R48h). In situ hybridization (ISH) showed that STAT3 and VEGFA RNA were co-located in the skeletal muscle of Japanese flounder. Japanese flounder was seriously affected by hypoxia for 3h and 6h. The expression of STAT3 and VEGFA increased significantly. The methylation levels of STAT3 5'UTR region and VEGFA promoter region were significantly lower than those in normoxia group, which was negatively correlated with the expression levels of STAT3 and VEGFA. The enzyme activities (LDH, ALT, AST, ALP) changed significantly. In addition, enzyme-linked immunosorbent assay (ELISA) detected a positive correlation between serum VEGFA concentration and muscle VEGFA mRNA. The current study have shown that Japanese flounder responded to acute hypoxic stress at multiple metabolic levels by changing DNA methylation status and activating transcription factors such as HIF-1α, Nrf2 and STAT3. It is significant for the scientific development of aquaculture through analyzing the effects of hypoxia on biological immunity.