Glutamate-cysteine Ligase Modifier Subunit Research Articles

Transcriptome analysis of sheep follicular development during prerecruitment, dominant, and mature stages after FSH superstimulation

Sheep is usually a monovular animal; superovulation technology is used to increase the number of offspring per individual and shorten generation intervals. To date, mature FSH superstimulatory treatments have been successfully used in sheep breeding, but much remains unknown about genes, pathways, and biological functions involved in follicular development. Therefore, in this study, we performed transcriptome profiling of small follicles (SFs; 2–2.5 mm), medium follicles (MFs; 3.5–4.5 mm), and large follicles (LFs; > 6 mm) in Mongolian ewes after FSH superstimulation. Furthermore, we identified differentially expressed genes and performed Kyoto Encyclopedia of Genes and Genomes pathway and Gene Ontology enrichment analyses in 3 separate pairwise comparisons. We found that ovarian steroidogenesis was significantly enriched in the SFs versus MFs analysis; the associated genes, cytochrome P450 family 19 (CYP19) and Hydroxy-delta-5-steroid dehydrogenase 3 beta- and steroid delta-isomerase 1 (HSD3B1), were significantly upregulated. Moreover, proline metabolism, glutathione metabolism, and PPAR signaling pathways were significantly enriched in the LFs versus SFs analysis; the associated genes, glutamate-cysteine ligase modifier subunit (GCLM) and cystathionine gamma-lyase (CTH), were significantly upregulated, whereas peroxisome proliferator-activated receptor gamma (PPARγ) was significantly downregulated. In summary, our study provides basic data and possible biological direction to further explore the molecular mechanism of sheep follicular development after FSH superstimulation.

Protective Effect of Low Molecular Weight Peptides from Solenocera crassicornis Head against Cyclophosphamide-Induced Nephrotoxicity in Mice via the Keap1/Nrf2 Pathway.

The major component of the Solenocera crassicornis head protein hydrolysates-fraction 1 (SCHPs-F1) are low molecular weight peptides (MW < 1 kDa). In this study, we investigated the potential renoprotective effects of SCHPs-F1 in a cyclophosphamide (CTX) toxicity mouse model. In brief, 40 male mice were randomly divided into 5 groups and received either saline or 80 mg/kg body weight (BW) CTX by intraperitoneal injection for 5 days, followed by either saline or SCHPs-F1 (100, 200, and 400 mg/kg BW) by intragastric administration for 15 days. SCHPs-F1 treatment significantly reversed the CTX-induced decreases in the levels of blood urea nitrogen (BUN), creatinine (CRE), and cytochrome P450 (CYP450), as well as the renal histological lesions. Furthermore, the results indicated that SCHPs-F1 potentially alleviated CTX-induced nephrotoxicity through mitigating inflammatory responses, oxidative stress, and apoptosis status of the kidneys, as evidenced by decreased levels of malondialdehyde (MDA), interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ and increased levels of total antioxidant capacity (T-AOC), catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Moreover, overexpression of pro-apoptotic proteins pair B-cell lymphoma-2 (Bcl-2)-associated X (Bax)/Bcl-2, cysteinyl aspartate specific proteinase (caspase)-3 and caspase-9 in renal tissues were suppressed by treatment with SCHPs-F1. In addition, the protein levels of the antioxidant factor nuclear factor erythroid-2 related factor 2 (Nrf2) and the expression levels of its downstream target genes heme-oxygenase (HO-1), glutamate-cysteine ligase modifier subunit (GCLM) and NAD(P)H dehydrogenase (quinone) 1 (NQO-1) were stimulated by treatment with SCHPs-F1 in the CTX-induced renal injury model. Taken together, our data suggested that SCHPs-F1 could provide a novel potential strategy in mitigating the nephrotoxicity caused by CTX.

Vernonia patula (Dryand.) Merr. and Leucas chinensis (Retz.) R. Brown exert anti-inflammatory activities and relieve oxidative stress via Nrf2 activation

Ethnopharmacological relevanceVernonia patula (Dryand.) Merr. and Leucas chinensis (Retz.) R. Brown have anti-inflammatory properties and are popularly used as complementary and alternative medicine in Asia. Aim of the studyTo investigate the underlying molecular mechanism and active chemicals in the ethanol extracts of V. patula (VP) and L. chinensis (LC). Materials and methodsThe inhibitory activities of VP and LC on lipopolysaccharide (LPS)-stimulated nitric oxide (NO) and interleukin-6 (IL-6) production were investigated in RAW264.7 macrophages and BV2 microglia. Downregulation of pro-inflammatory genes and upregulation of Nrf2 (NF-E2 p45-related factor 2)-ARE (antioxidant response element) pathway were investigated using RT-Q-PCR and Western blotting. Direct antioxidant capacities were measured using free radical scavenging and Folin-Ciocalteu assays. The flavonoids and triterpenes in VP and LC were identified by HPLC-ESI-MS. ResultsVP and LC inhibited NO and IL-6 production and suppressed iNOS, IL-6, IL-1β and CCL2 gene expression. VP and LC were potent direct antioxidants and effective indirect antioxidants assayed by Nrf2 activation and induction of heme oxygenase (HO)-1, glutamate-cysteine ligase modifier subunit (GCLM) and NAD(P)H quinone oxidoreductase 1 (NQO1). Three flavonoids including apigenin (1), luteolin (2) and chryseriol (3), and one triterpene betulinic acid (4) were found in VP; while compounds 1–4 and oleanolic acid (5) were in LC. ConclusionAnti-inflammatory and antioxidant activities of VP and LC may be in great part attributed to the identified Nrf2 activating compounds, which induce expression of Phase II enzymes and attenuate the upregulation of pro-inflammatory genes.

Genetic variants in glutamate cysteine ligase confer protection against type 2 diabetes.

Oxidative stress contributes to the pathogenesis of type 2 diabetes (T2D). This study investigated whether single nucleotide polymorphisms (SNPs) at genes encoding glutamate cysteine ligase catalytic (rs12524494, rs17883901, rs606548, rs636933, rs648595, rs761142 at GCLC) and modifier (rs2301022, rs3827715, rs7517826, rs41303970 at GCLM) subunits are associated with susceptibility to type 2 diabetes. 2096 unrelated Russian subjects were enrolled for the study. Genotyping was done with the use of the MassArray System. Plasma levels of reactive oxygen species (ROS) and glutathione in the study subjects were analyzed by fluorometric and colorimetric assays, respectively.The present study found, for the first time, an association of SNP rs41303970 in the GCLM gene with a decreased risk of T2D (P = 0.034, Q = 0.17). Minor alleles such as rs12524494-G GCLC gene (P = 0.026, Q = 0.17) and rs3827715-C GCLM gene (P = 0.03, Q = 0.17) were also associated with reduced risk for T2D. Protective effects of variant alleles such as rs12524494-G at GCLC (P = 0.02, Q = 0.26) and rs41303970-A GCLM (P = 0.013, Q = 0.25) against the risk of T2D were seen solely in nonsmokers. As compared with healthy controls, diabetic patients had markedly increased levels of ROS and decreased levels of total GSH in plasma. Interestingly, fasting blood glucose level positively correlated with oxidized glutathione concentration (rs = 0.208, P = 0.01). Three SNPs rs17883901, rs636933, rs648595 at GCLC and one rs2301022 at GCLM were associated with decreased levels of ROS, while SNPs rs7517826, rs41303970 at GCLM were associated with increased levels of total GSH in plasma. Single nucleotide polymorphisms in genes encoding glutamate cysteine ligase subunits confer protection against type 2 diabetes and their effects are mediated through increased levels of glutathione.

Integrative transcriptome analysis and discovery of signaling pathways involved in the protective effects of curcumin against oxidative stress in tilapia hepatocytes

Summer outbreaks of the hepatobiliary syndrome in fish impose a heavy burden on aquaculture in China. Curcumin is a polyphenol with antioxidant activity that has been used to protect the health of fish livers, but the mechanism underlying its protective effect is unclear. In this study, an in vitro model of hepatocyte oxidative damage in Oreochromis niloticus was established using H2O2. Treatment with 5 mM H2O2 for 2.5 h markedly reduced cell viability and antioxidant activity and elevated lactate dehydrogenase (LDH) activity, indicating conditions that can be used to establish an oxidative stress model. Under H2O2 stress, curcumin pretreatment significantly maintained cell viability, reduced malondialdehyde (MDA) levels, and increased superoxide dismutase (SOD) activity. RNA-seq results showed that acute H2O2 treatment resulted in minor changes in gene expression, whereas curcumin changed the expression profile and affected cytochrome P450 (Cyp 450), glutathione (GSH) metabolism, and the peroxisome proliferator-activated receptor (PPAR) signaling pathway. Several critical antioxidant defense signaling pathways were identified, and altered expression was confirmed by q-PCR. These results indicate that curcumin might upregulate PPAR expression by increasing Cyp2J2 expression. Further experiments showed that curcumin can upregulate the Nrf2-Keap1 signaling pathway at the transcriptional level, and this upregulation can induce downstream defense genes, including glutamate cysteine ligase catalytic subunit(GCLC) and glutamate cysteine ligase modifier subunit (GCLM), and thereby promote GSH synthesis and the expression of related antioxidases. This study might shed light on the effects of curcumin on the prevention and alleviation of liver diseases in fish.

&lt;p&gt;Copper Oxide Nanoparticles Induce Oxidative DNA Damage and Cell Death via Copper Ion-Mediated P38 MAPK Activation in Vascular Endothelial Cells&lt;/p&gt;

BackgroundInhaled nanoparticles can cross pulmonary air–blood barrier into circulation and cause vascular endothelial injury and progression of cardiovascular disease. However, the molecular mechanism underlying the vascular toxicity of copper oxide nanoparticles (CuONPs) remains unclear. We have recently demonstrated that the release of copper ions and the accumulation of superoxide anions contributed to CuONPs-induced cell death in human umbilical vein endothelial cells (HUVECs). Herein, we further demonstrate the mechanism underlying copper ions-induced cell death in HUVECs.Methods and ResultsCuONPs were suspended in culture medium and vigorously vortexed for several seconds before exposure. After treatment with CuONPs, HUVECs were collected, and cell function assays were conducted to elucidate cellular processes including cell viability, oxidative stress, DNA damage and cell signaling pathways. We demonstrated that CuONPs uptake induced DNA damage in HUVECs as evidenced by γH2AX foci formation and increased phosphorylation levels of ATR, ATM, p53 and H2AX. Meanwhile, we showed that CuONPs exposure induced oxidative stress, indicated by the increase of cellular levels of superoxide anions, the upregulation of protein levels of heme oxygenase-1 (HO-1) and glutamate-cysteine ligase modifier subunit (GCLM), the elevation of the levels of malondialdehyde (MDA), but the reduction of glutathione to glutathione disulfide ratio. We also found that antioxidant N-acetyl-L-cysteine (NAC) could ameliorate CuONPs-induced oxidative stress and cell death. Interestingly, we demonstrated that p38 mitogen-activated protein kinase (MAPK) signaling pathway was activated in CuONPs-treated HUVECs, while p38α MAPK knockdown by siRNA significantly rescued HUVECs from CuONPs-induced DNA damage and cell death. Importantly, we showed that copper ions chelator tetrathiomolybdate (TTM) could alleviate CuONPs-induced oxidative stress, DNA damage, p38 MAPK pathway activation and cell death in HUVECs.ConclusionWe demonstrated that CuONPs induced oxidative DNA damage and cell death via copper ions-mediated p38 MAPK activation in HUVECs, suggesting that the release of copper ions was the upstream activator for CuONPs-induced vascular endothelial toxicity, and the copper ions chelator TTM can alleviate CuONPs-associated cardiovascular disease.

Activation of Farnesoid X Receptor by Schaftoside Ameliorates Acetaminophen-Induced Hepatotoxicity by Modulating Oxidative Stress and Inflammation.

Aims: Acetaminophen (APAP) overdose leads to acute liver injury by inducing hepatic mitochondrial oxidative stress and inflammation. However, the molecular mechanisms involved are still unclear. Farnesoid X receptor (FXR) serves as a therapeutic target for the treatment of liver disorders, whose activation has been proved to protect APAP-induced hepatotoxicity. In this study, we examined whether FXR activation by schaftoside (SS), a naturally occurring flavonoid from Desmodium styracifolium, could protect mice against APAP-induced hepatotoxicity via regulation of oxidative stress and inflammation. Results: We first found that SS exhibited potent protective effects against APAP-induced hepatotoxicity in mice. The study reveals that SS is a potential agonist of FXR, which protects mice from hepatotoxicity mostly via regulation of oxidative stress and inflammation. Mechanistically, the hepatoprotective SS is associated with the induction of the genes of phase II detoxifying enzymes (e.g., UGT1A1, GSTα1), phase III drug efflux transporters (e.g., bile salt export pump, organic solvent transporter protein β), and glutathione metabolism-related enzymes (e.g., glutamate-cysteine ligase modifier subunit [Gclm], glutamate-cysteine ligase catalytic subunit [Gclc]). More importantly, SS-mediated FXR activation could fine-tune the pro- and anti-inflammatory eicosanoids generation via altering eicosanoids metabolic pathway, thereby resulting in decrease of hepatic inflammation. In contrast, FXR deficiency can abrogate the above effects. Innovation and Conclusion: Our results provided the direct evidence that FXR activation by SS could attenuate APAP-induced hepatotoxicity via inhibition of nuclear factor kappa-B signaling and fine-tuning the generation of proinflammatory mediators' eicosanoids. Our findings indicate that strategies to activate FXR signaling in hepatocytes may provide a promising therapeutic approach to alleviate liver injury induced by APAP overdose.

FSHR ablation induces depression-like behaviors.

Alteration in reproductive hormones profile is associated with the increasing risk of menopausal depression in women. Serum follicle-stimulating hormone (FSH) level is changed during the menopause transition, while the effect of FSH on menopausal depression has remained undefined. In this study we investigated whether or how FSH affected menopausal depression in postmenopausal (ovariectomized) FSHR knockout mice (Fshr−/−). We found that Fshr−/− mice displayed aggravated depression-like behaviors, accompanied by severe oxidative stress in the whole brain, resulted from significantly reduced glutamate cysteine ligase modifier subunit (GCLm) in glutathione synthesis and glucose-6-phosphate dehydrogenase (G6PD) in NADP/NADPH transition. Importantly, administration of ROS scavenger N-acetyl cysteine (NAC, 150 mg · kg−1 · d−1, i.p. for 12 weeks) attenuated the depression-like behaviors of Fshr−/− mice. Consistent with these in vivo experiment results, we found that pretreatment with FSH (50, 100 ng/mL) dose-dependently increased protein levels of GCLm and G6PD, and decreased the ROS production in N2a mouse neuroblastoma cells. These findings demonstrate that FSH signaling is involved in pathogenesis of menopausal depression, and likely to maintain the redox-optimized ROS balance in neurons.

Rosmarinic acid protects against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced dopaminergic neurotoxicity in zebrafish embryos

Rosmarinic acid (RA) is an extract that can be obtained from Lamiaceae herbs and the Boraginaceae family. This study aimed to evaluate the effect of RA on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced dopaminergic neurotoxicity in zebrafish embryos. Embryos were challenged with MPTP and then were treated with RA or brusatol (a Nrf2 inhibitor). Locomotor activity of zebrafish was recorded using a video camera. The swimming distance was analyzed with SMART 3.0 software. Tyrosine hydroxylase (TH) immunohistochemistry, reactive oxygen species (ROS), glutathione (GSH), and malondialdehyde (MDA) contents were evaluated. The expressions of proteins in the DJ-1/Akt/Nrf2 signaling pathway were measured. The results showed that RA not only prevented MPTP-induced dopaminergic neuron loss, but also attenuated the deficit in locomotor behavior. RA attenuated the increases of ROS and MDA induced by MPTP. Treatment with RA augmented expression of glutamate cysteine ligase catalytic subunit, glutamate cysteine ligase modifier subunit, and GSH. Furthermore, RA increased the expression of DJ-1, p-Akt, Nuclear-Nrf2, HO-1 and inhibited the expression of PTEN. Brusatol partially abolished the neuroprotective effect of RA in MPTP-induced Parkinson's disease (PD) model of zebrafish embryos. The results of this study indicate that RA exerts neuroprotective effects on MPTP-induced neurotoxicity in dopaminergic neurons of a zebrafish PD model. The mechanism underlying the effects of RA is associated with promotion of antioxidant gene expression via regulation of the DJ-1/Akt/Nrf2 signaling pathway.

DJ-1 protects retinal pericytes against high glucose-induced oxidative stress through the Nrf2 signaling pathway

Oxidative stress has been associated with the etipathogenesis of Diabetic retinopathy (DR). Studies have shown that DJ-1 plays an important role in regulating the reactive oxygen species (ROS) production and resistance to oxidative stress-induced apoptosis. This study aimed to investigate whether DJ-1 upregulates oxidative stress and prevents damage to retinal capillary pericytes by increasing antioxidant capacity through the Nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway. Nrf2 is a redox-sensitive transcription factor that encode antioxidant enzymes and phase II metabolic enzymes, activation of Nrf2 functions is one of the critical defensive mechanisms against oxidative stress in many tissues. Our results showed after DJ-1 overexpression, apoptosis of rat retinal pericytes (RRPs) decreased, the ratio of B-cell lymphoma-2 (Bcl-2) to BCL2-Associated X Protein (BAX) increased, the production of ROS decreased, and the protein expression and activity of manganese superoxide dismutase (MnSOD, also called SOD2) and catalase (CAT) increased. DJ-1 overexpression activated Nrf2 expression, however, after Nrf2 silencing, apoptosis of RRPs increased, the ratio of Bcl-2 to BAX decreased, the production of ROS increased, the protein expression of MnSOD and CAT decreased, and the expression of heme oxygenase-1 (HO-1), NADP(H) quinone oxidoreductase (NQO1), glutamate-cysteine ligase catalytic subunit (GCLC) and modifier subunit (GCLM) decreased. These data suggest that enhancement of the Nrf2 pathway is a potential protective strategy for the treatment of DR. Therefore, DJ-1 may prevent high glucose-induced oxidative stress and RRPs apoptosis through the Nrf2 signaling pathway, thereby preventing the early onset and progression of DR.

Homologous-targeting biomimetic nanoparticles for photothermal therapy and Nrf2-siRNA amplified photodynamic therapy against oral tongue squamous cell carcinoma

Photothermal therapy (PTT) and photodynamic therapy (PDT) have some advantages such as site selectivity, non- or little-invasive property and negligible drug resistance for treatment of oral tongue squamous cell carcinoma (OTSCC). However, the activation of intracellular antioxidant responses will partly counteract anticancer effects of PDT. As a well-known redox-regulated transcription factor, Nrf2 has been found to participate in the resistance of tumor cells against PDT. Hereon, a biomimetic nanoparticle system was designed for co-loading and targeted delivery of photosensitizer indocyanine green (ICG) and Nrf2-siRNA, thus hope to achieve synergistic effects of PTT and amplified PDT in OTSCC. Poly(β-amino ester) (PBAE)/poly lactic-co-glycolic acid (PLGA) blended nanoparticles co-loaded with ICG and Nrf2-siRNA were prepared and then encapsulated within the vesicles of cancer cell membrane (CCM) specifically derived from homologous OTSCC, thus obtained biomimetic nanoparticles named as M@PPI-siRNA. Both in vitro and in vivo, M@PPI-siRNA nanoparticles exhibited strong OTSCC-targeting ability due to tumor-homing effect of homologous CCM. Upon laser irradiation at 808 nm, M@PPI-siRNA nanoparticles significantly down-regulated the expressions of Nrf2 and its regulated genes, glutamate-cysteine ligase catalytic subunit (GCLC) and modifier subunit (GCLM) that play important roles in ROS detoxification, and subsequently indirectly enhanced PDT anticancer effects by maintaining the accumulation of intracellular ROS. Through cooperation of PTT and Nrf2-siRNA amplified PDT, M@PPI-siRNA nanoparticles significantly inhibited the proliferation and induced the apoptosis of OTSCC cells, and also notably suppressed tumor growth and angiogenesis in OTSCC tumor-bearing mice. In summary, this study provides a promising OTSCC-targeted delivery system for both photosensitizer and gene, and also confirms that Nrf2-siRNA is an efficient synergist for amplifying PDT.

Potential neuroprotection of wheat alkylresorcinols in hippocampal neurons via Nrf2/ARE pathway.

5-n-Alkylresorcinols (ARs) are abundant in wheat bran and potentially antioxidative, although the neuroprotective mechanism is not fully understood. The neuroprotective effect of wheat bran ARs on H2O2-induced neuronal cells and the relationship between neuroprotection and the nuclear factor erythroid 2-related factor 2 (Nrf2)/antioxidant-response element (ARE) pathway were investigated in this study. Seven homologs were identified in the purified ARs by high-performance liquid chromatography-tandem mass spectrometry. Pretreatment with 80 μg mL-1 ARs alleviated 23% HT22 cell death and the up-regulation of intracellular reactive oxygen species level and malondialdehyde under H2O2 stimulation. The neuroprotection effect was proved by the increase in the Nrf2 nuclear location and up-regulation of mRNA and protein expressions of heme oxygenase-1, NAD(P)H quinone dehydrogenase 1, glutamate-cysteine ligase catalytic subunit, and glutamate-cysteine ligase modifier subunit l. Wheat bran ARs displayed a neuroprotective function, possibly by promoting the endogenous antioxidant defense system. ARs may be regarded as a functional food ingredient for preventing neurodegenerative diseases in the future.

Ferroptosis is controlled by the coordinated transcriptional regulation of glutathione and labile iron metabolism by the transcription factor BACH1

Ferroptosis is an iron-dependent programmed cell death event, whose regulation and physiological significance remain to be elucidated. Analyzing transcriptional responses of mouse embryonic fibroblasts exposed to the ferroptosis inducer erastin, here we found that a set of genes related to oxidative stress protection is induced upon ferroptosis. We considered that up-regulation of these genes attenuates ferroptosis induction and found that the transcription factor BTB domain and CNC homolog 1 (BACH1), a regulator in heme and iron metabolism, promotes ferroptosis by repressing the transcription of a subset of the erastin-induced protective genes. We noted that these genes are involved in the synthesis of GSH or metabolism of intracellular labile iron and include glutamate-cysteine ligase modifier subunit (Gclm), solute carrier family 7 member 11 (Slc7a11), ferritin heavy chain 1 (Fth1), ferritin light chain 1 (Ftl1), and solute carrier family 40 member 1 (Slc40a1). Ferroptosis has also been previously shown to induce cardiomyopathy, and here we observed that Bach1-/- mice are more resistant to myocardial infarction than WT mice and that the severity of ischemic injury is decreased by the iron-chelator deferasirox, which suppressed ferroptosis. Our findings suggest that BACH1 represses genes that combat labile iron-induced oxidative stress, and ferroptosis is stimulated at the transcriptional level by BACH1 upon disruption of the balance between the transcriptional induction of protective genes and accumulation of iron-mediated damage. We propose that BACH1 controls the threshold of ferroptosis induction and may represent a therapeutic target for alleviating ferroptosis-related diseases, including myocardial infarction.

Cardamonin protects against doxorubicin-induced cardiotoxicity in mice by restraining oxidative stress and inflammation associated with Nrf2 signaling

The clinical application of doxorubicin (DOX) for cancer treatment is limited due to its cardiotoxicity. However, the basic pathophysiological molecular mechanisms underlying DOX-induced cardiomyopathy have not yet been completely clarified, and the disease-specific therapeutic strategies are lacking. The aim of the present study was to investigate the potential cardioprotective effect of cardamonin (CAR), a flavone found in Alpinia plant, on DOX-induced cardiotoxicity in a mouse model. At first, in DOX-treated mouse cardiomyocytes, CAR showed significantly cytoprotective effects through elevating nuclear factor erythroid-2 related factor 2 (Nrf2) signaling, and reducing the degradation of Nrf2. This process then improved the anti-oxidant system, as evidenced by the up-regulated expression levels of haem oxygenase-1 (HO1), NAD(P)H:quinone oxidoreductase 1 (NQO1), glutamate-cysteine ligase modifier subunit (GCLM), superoxide dismutase (SOD), glutathione (GSH) and catalase (CAT). In contrast, DOX-induced increases in malondialdehyde (MDA) and reactive oxygen species (ROS) were highly inhibited by CAR treatments. Additionally, DOX-induced apoptosis and inflammatory response in cardiomyocytes were diminished by CAR through reducing the Caspase-3 and nuclear factor-κB (NF-κB) signaling pathways, respectively. Then, in the DOX-induced animal model with cardiotoxicity, we confirmed that through improving Nrf2 signaling, CAR markedly suppressed oxidative stress, apoptosis and inflammatory response in hearts of mice, improving cardiac function eventually. Together, our findings demonstrated that CAR activated Nrf2-related cytoprotective system, and protected the heart from oxidative damage, apoptosis and inflammatory injury, suggesting that CAR might be a potential therapeutic strategy in the prevention of DOX-associated myocardiopathy.

Curcumin Alleviates IUGR Jejunum Damage by Increasing Antioxidant Capacity through Nrf2/Keap1 Pathway in Growing Pigs.

Simple SummaryIntrauterine growth retardation (IUGR) is usually defined as fetal growth below the 10th percentile for gestational age and results in impaired growth and development of the fetus and/or its organs during gestation. IUGR not only has a permanent effect on postnatal growth and long-term health, but also results in high fetal mortality and morbidity. Recent results have demonstrated that IUGR can cause jejunum damage in piglets. The jejunum is not only the main organ for the digestion and absorption of nutrients, but also an immune organ in the body. However, few effective methods to alleviate jejunum damage and oxidative stress in IUGR pigs have been found. In recent years, studies have found that curcumin (CUR) may be an effective and safe feed additive for regulating oxidative stress in the body. Our results indicated that dietary added 200 mg/kg curcumin to the basal diet can increase the antioxidant capacity of the IUGR growing pigs, jejunum and alleviate the damage in jejunum of the IUGR growing pigs. Therefore, the use of curcumin as a feed additive has certain economic value.The purpose of this study was to explore the effects of curcumin on IUGR jejunum damage. A total of 24 IUGR and 12 normal-birth weight (NBW) female crossbred (Duroc × Landrace × Large White) piglets were randomly assigned into three groups at weaning (26 days): IUGR group, NBW group, and IUGR + CUR group, which were fed diets containing 0 mg/kg (NBW), 0 mg/kg (IUGR) and 200 mg/kg (IUGR + CUR) curcumin from 26 to 115 days of age. Results showed that dietary supplementation with 200 mg/kg curcumin significantly increased the total superoxide dismutase (T-SOD) activity and decreased the malondialdehyde (MDA) content in the jejunum of IUGR pigs (p < 0.05). Results of real-time PCR showed that the IUGR + CUR group significantly increased the gene expression of NF-E2-related factor 2 (Nrf2) (p < 0.05), and increased the glutamate-cysteine ligase catalytic subunit (GCLC), superoxide dismutase 1 (SOD1), glutamate-cysteine ligase modifier subunit (GCLM), and NAD(P)H quinone dehydrogenase 1 (NQO1) mRNA expression compared with the IUGR group (p < 0.05). Western blot results showed that dietary supplementation with 200 mg/kg curcumin significantly increased the protein levels of Nrf2 and NQO1. Compared with the IUGR group, pigs in IUGR + CUR group showed significantly decreased the levels of tumor necrosis factor-α (TNFα), interleukin-6 (IL-6), and interferon gamma (IFNγ) (p < 0.05), and increased the interleukin-2 (IL-2) level (p < 0.05). Dietary supplementation with 200 mg/kg curcumin significantly reduced cysteinyl aspartate specific proteinase 3 (caspase3), BCL2-associated X protein (bax), B-cellCLL/lymphoma 2 (bcl2), and heat-shock protein 70 (hsp70) mRNA expression, and increased occludin (ocln) mRNA expression (p < 0.05). In conclusion, dietary supplementation with 200 mg/kg curcumin can alleviate jejunum damage in IUGR growing pigs, through Nrf2/Keap1 pathway.

Unification of Opposites between Two Antioxidant Transcription Factors Nrf1 and Nrf2 in Mediating Distinct Cellular Responses to the Endoplasmic Reticulum Stressor Tunicamycin.

The water-soluble Nrf2 (nuclear factor, erythroid 2-like 2, also called Nfe2l2) is accepted as a master regulator of antioxidant responses to cellular stress, and it was also identified as a direct target of the endoplasmic reticulum (ER)-anchored PERK (protein kinase RNA-like endoplasmic reticulum kinase). However, the membrane-bound Nrf1 (nuclear factor, erythroid 2-like 1, also called Nfe2l1) response to ER stress remains elusive. Herein, we report a unity of opposites between these two antioxidant transcription factors, Nrf1 and Nrf2, in coordinating distinct cellular responses to the ER stressor tunicamycin (TU). The TU-inducible transcription of Nrf1 and Nrf2, as well as GCLM (glutamate cysteine ligase modifier subunit) and HO-1 (heme oxygenase 1), was accompanied by activation of ER stress signaling networks. Notably, the unfolded protein response (UPR) mediated by ATF6 (activating transcription factor 6), IRE1 (inositol requiring enzyme 1) and PERK was significantly suppressed by Nrf1α-specific knockout, but hyper-expression of Nrf2 and its target genes GCLM and HO-1 has retained in Nrf1α−/− cells. By contrast, Nrf2−/−ΔTA cells with genomic deletion of its transactivation (TA) domain resulted in significant decreases of GCLM, HO-1 and Nrf1; this was accompanied by partial decreases of IRE1 and ATF6, rather than PERK, but with an increase of ATF4 (activating transcription factor 4). Interestingly, Nrf1 glycosylation and its trans-activity to mediate the transcriptional expression of the 26S proteasomal subunits, were repressed by TU. This inhibitory effect was enhanced by Nrf1α−/− and Nrf2−/−ΔTA, but not by a constitutive activator caNrf2ΔN (that increased abundances of the non-glycosylated and processed Nrf1). Furthermore, caNrf2ΔN also enhanced induction of PERK and IRE1 by TU, but reduced expression of ATF4 and HO-1. Thus, it is inferred that such distinct roles of Nrf1 and Nrf2 are unified to maintain cell homeostasis by a series of coordinated ER-to-nuclear signaling responses to TU. Nrf1α (i.e., a full-length form) acts in a cell-autonomous manner to determine the transcription of most of UPR-target genes, albeit Nrf2 is also partially involved in this process. Consistently, transactivation of ARE (antioxidant response element)-driven BIP (binding immunoglobulin protein)-, PERK- and XBP1 (X-box binding protein 1)-Luc reporter genes was mediated directly by Nrf1 and/or Nrf2. Interestingly, Nrf1α is more potent than Nrf2 at mediating the cytoprotective responses against the cytotoxicity of TU alone or plus tBHQ (tert-butylhydroquinone). This is also further supported by the evidence that the intracellular reactive oxygen species (ROS) levels are increased in Nrf1α−/− cells, but rather are, to our surprise, decreased in Nrf2−/−ΔTA cells.

Intrarenal Transplantation of Hypoxic Preconditioned Mesenchymal Stem Cells Improves Glomerulonephritis through Anti-Oxidation, Anti-ER Stress, Anti-Inflammation, Anti-Apoptosis, and Anti-Autophagy.

To confer further therapeutic potential and prevent some adverse effects by the mesenchymal stem cells (MSCs) transplantation, we explored the effects of locally intrarenal arterial administration of hypoxic preconditioned MSCs in the anti-Thy1.1 induced rat glomerulonephritis. Proteinuria, histochemical staining, and western blotting were used to explore the therapeutic effects and mechanisms. Locally intrarenal arterial MSCs transplantation successfully implanted the fluorescent or CD44 labeled MSCs in the nephritic glomeruli, ameliorated proteinuria, and glomerulosclerosis in nephritic rats. Hypoxic preconditioning significantly upregulated hypoxic inducible factor-1α/VEGF (HIF-1α/VEGF) in the MSCs and was more efficient than normoxic MSCs in reducing the degree of urinary protein, glomerulosclerosis, fibrosis, macrophage/monocyte infiltration, GRP78 mediated endoplasmic reticulum stress, Beclin-1/LC3-II mediated autophagy, and Bax/Bcl-2/caspase 3 mediated apoptosis. Hypoxic MSCs could further promote intranuclear nuclear factor (erythroid-derived 2, Nrf2) and reduce nuclear factor kappa B expression in nephritic kidneys. As compared to normoxic MSCs, hypoxic MSCs transplantation significantly upregulated the renal expression of anti-oxidative response elements/enzymes including glutamate-cysteine ligase catalytic subunit, glutamate-cysteine ligase modifier subunit, glutathione peroxidase, catalase, Mn, and Cu/Zn superoxide dismutase. In summary, intrarenal hypoxic preconditioning MSCs transplantation was more effective to activate hypoxic inducible factor-1α/VEGF/Nrf2 (HIF-1α/VEGF/Nrf2) signaling, preserve anti-oxidant proteins and anti-oxidative responsive element proteins, and subsequently reduce glomerular apoptosis, autophagy, and inflammation.

Effects of zearalenone-induced oxidative stress and Keap1–Nrf2 signaling pathway-related gene expression in the ileum and mesenteric lymph nodes of post-weaning gilts

Zearalenone (ZEA) contamination of feed affects animal husbandry and the human health. Currently, the molecular mechanism underlying small intestine-related diseases caused by ZEA-induced oxidative stress is not well understood. In this study, we aimed to identify the mechanisms involved in ZEA (0.5–1.5 mg/kg)-induced oxidative stress in the ileum and mesenteric lymph nodes (MLNs) and the role of the Kelch-like erythroid cell-derived protein with CNC homology-associated protein 1 (Keap1)–nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway in post-weaning gilts. Forty post-weaning gilts (Landrace × Yorkshire × Duroc) with an average body weight of 14.01 ± 0.86 kg were randomly allocated to four groups and fed a corn–soybean meal basal diet supplemented with < 0.1, 0.5, 1.0, or 1.5 mg/kg ZEA. The results showed that the activity of total superoxide dismutase and glutathione peroxidase decreased (p < 0.05) linearly and quadratically and that the content of malondialdehyde increased (p < 0.05) quadratically in the ileum and MLNs with increasing ZEA in the diet. Immunohistochemical analysis showed that the expression of Nrf2 and glutathione peroxidase 1 (Gpx1) immunoreactive proteins in the ileum and MLNs were significantly enhanced with increasing ZEA. The relative mRNA and protein expression of Nrf2, Gpx1, quinone oxidoreductase 1 (Nqo1), hemeoxygenase 1 (Ho1), modifier subunit of glutamate-cysteine ligase (Gclm), and catalytic subunit of glutamate-cysteine ligase (Gclc) increased (p < 0.05) linearly and quadratically, and the relative mRNA and protein expression of Keap1 decreased (p < 0.05) linearly and quadratically in the ileum with increasing ZEA concentrations in the diet. Further, the relative mRNA and protein expression of Nrf2 and Gpx1 increased (p < 0.05) linearly and quadratically, and the relative mRNA and protein expression of Nqo1, Ho1, and Gclm decreased (p < 0.05) quadratically in the MLNs as ZEA concentrations increased in the diet. Our results provide valuable genetic information on ZEA-induced oxidative stress in the ileum and MLNs of post-weaning gilts and have elucidated the key regulatory genes involved in the Keap1–Nrf2 signaling pathway. Results indicated that the Keap1–Nrf2 signaling pathway might be a key target to further prevent and treat ZEA-induced injury to the ileum in post-weaning gilts.

Automatic feature extraction in X-ray image based on deep learning approach for determination of bone age

Aim:Determination of bone age is an important method to skeletal maturity and growth potential. This paper proposes a determination bone age via X-ray image recognition to obtain a better identification effect by comparison with state-of-the-art techniques. Methods: The proposed approach comprises two steps: the feature extraction and classification method. The feature extraction utilizes depth neural network to study the features of X-ray image, and the Local Binary Patterns (LBP) features and Glutamate cysteine ligase modifier subunit (GCLM) features in the image are extracted. Then, the classification method base on support vector machine is used to classify the features. Results:The experimental results show that the average absolute error of bone age assessment model based on multi-dimensional data feature fusion is 0.455, which is superior to the traditional method and support vector machine method. Because the model is based on feature extraction of deep neural network, it shows that the feature extraction method based on deep neural network can extract feature information better than traditional image analysis method. Conclusion:Compared with the traditional feature extraction method, the feature extraction based on deep convolution neural network has better performance in the bone age regression model. Combining population and gender information, the accuracy of bone age prediction based on image can be further improved.

Hydrangenol Isolated from the Leaves of Hydrangea serrata Attenuates Wrinkle Formation and Repairs Skin Moisture in UVB-Irradiated Hairless Mice.

Our previous study showed that hydrangenol isolated from Hydrangea serrata leaves exerts antiphotoaging activity in vitro. In this study, we determined its antiphotoaging effect in UVB-irradiated HR-1 hairless mice. We evaluated wrinkle formation, skin thickness, histological characteristics, and mRNA and protein expression using qRT-PCR and Western blot analysis in dorsal skins. Hydrangenol mitigated wrinkle formation, dorsal thickness, dehydration, and collagen degradation. Hydrangenol increased the expression of involucrin, filaggrin, and aquaporin-3 (AQP3) as well as hyaluronic acid (HA) production via hyaluronidase (HYAL)-1/-2 downregulation. Consistent with the recovery of collagen composition, the expression of Pro-COL1A1 was increased by hydrangenol. Matrix metalloproteinase (MMP)-1/-3, cyclooxygenase-2 (COX-2), and interleukin-6 (IL-6) expression was reduced by hydrangenol. Hydrangenol attenuated the phosphorylation of mitogen-activated protein kinases (MAPKs) including ERK and p38, activator protein 1 (AP-1) subunit, and signal transduction and activation of transcription 1 (STAT1). Hydrangenol upregulated the expression of nuclear factor-E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NAD(P)H quinone dehydrogenase 1 (NQO-1), glutamate cysteine ligase modifier subunit (GCLM), and glutamate cysteine ligase catalysis subunit (GCLC). Taken together, our data suggest that hydrangenol can prevent wrinkle formation by reducing MMP and inflammatory cytokine levels and increasing the expression of moisturizing factors and antioxidant genes.