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N-acetyl cysteine (NAC) is a versatile drug used in various conditions, but the limitations and toxicities are not clear. The acute toxicity and toxicological mechanisms of an intraperitoneal injection of NAC in normal mice were deciphered. The LD50 for male and female BALB/cByJNarl mice were 800 mg/kg and 933 mg/kg. The toxicological mechanisms of 800 mg/kg NAC (N800) were investigated. The serum biomarkers of hepatic and renal indices dramatically increased, followed by hepatic microvesicular steatosis, renal tubular injury and necrosis, and splenic red pulp atrophy and loss. Thus, N800 resulted in mouse mortality mainly due to acute liver, kidney, and spleen damages. The safe dose (275 mg/kg) of NAC (N275) increased hepatic antioxidant capacity by increasing glutathione levels and catalase activity. N275 elevated the hepatic gene expressions of lipid transporter, lipid synthesis, ß-oxidation, and ketogenesis, suggesting a balance between lipid production and consumption, and finally, increased ATP production. In contrast, N800 increased hepatic oxidative stress by decreasing glutathione levels through suppressing Gclc, and reducing catalase activity. N800 decreased the hepatic gene expressions of lipid transporter, lipid synthesis, and interferred ß-oxidation, leading to lipid accumulation and increasing Cyp2E1 expression, and finally, decreased ATP production. Therefore, NAC doses are limited for normal individuals, especially via intraperitoneal injection or similar means.
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Background and aim: In traditional medicine, Machilus zuihoensis Hayata bark (MZ) is used in combination with other medicines to treat gastric cancer, gastric ulcer (GU), and liver and cardiovascular diseases. This study aims to evaluate the gastroprotective effects and possible mechanism(s) of MZ powder against acidic ethanol (AE)-induced GU and its toxicity in mice. Experimental procedure: The gastroprotective effect of MZ powder was analyzed by orally administering MZ for 14 consecutive days before AE-inducing GU. Ulcer index (UI) and protection percentage were calculated, hematoxylin and eosin staining and periodic acid-Schiff staining were performed, and gastric mucus weights were measured. The antioxidative, anti-inflammatory, and anti-apoptotic mechanisms, and possible signaling pathway(s) were studied. Results and conclusion: Pretreatment with MZ (100 and 200 mg/kg) significantly decreased 10 µL/g AE-induced mucosal hemorrhage, edema, inflammation, and UI, resulted in protection percentages of 88.9% and 93.4%, respectively. MZ pretreatment reduced AE-induced oxidative stress by decreasing malondialdehyde level and restoring superoxide dismutase activity. MZ pretreatment demonstrated anti-inflammatory effects by reducing both serum and gastric tumor necrosis factor-α, interleukin (IL)-6, and IL-1ß levels. Furthermore, MZ pretreatment exhibited anti-apoptotic effect by decreasing Bcl-2 associated X protein/B-cell lymphoma 2 ratio. The gastroprotective mechanisms of MZ involved inactivations of nuclear factor kappa-light-chain enhancer of activated B cells (NF-κB) and mitogen activated protein kinase (MAPK) signaling pathways. Otherwise, 200 mg/kg MZ didn't induce liver or kidney toxicity. In conclusion, MZ protects AE-induced GU through mucus secreting, antioxidative, anti-inflammatory, and anti-apoptotic mechanisms, and inhibitions of NF-κB and MAPK signaling pathways.
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The nucleolar rRNA 2'-O-methyltransferase fibrillarin (FBL) contains a highly conserved methyltransferase domain at the C-terminus and a diverse glycine arginine-rich (GAR) domain at the N-terminus in eukaryotes. We found that a nine-exon configuration of fbl and exon 2-3 encoded GAR domain are conserved and specific in vertebrates. All internal exons except exon 2 and 3 are of the same lengths in different vertebrate lineages. The lengths of exon 2 and 3 vary in different vertebrate species but the ones with longer exon 2 usually have shorter exon 3 complementarily, limiting lengths of the GAR domain within a certain range. In tetrapods except for reptiles, exon 2 appears to be longer than exon 3. We specifically analyzed different lineages of reptiles for their GAR sequences and exon lengths. The lengths of exon 2 in reptiles are around 80-130-nt shorter and the lengths of exon 3 in reptiles are around 50-90 nt longer than those in other tetrapods, all in the GAR-coding regions. An FSPR sequence is present at the beginning of the GAR domain encoded by exon 2 in all vertebrates, and a specific FXSP/G element (X can be K, R, Q, N, and H) exist in the middle of GAR with phenylalanine as the 3rd exon 3-encoded amino acid residue starting from jawfish. Snakes, turtles, and songbirds contain shorter exon 2 compared with lizards, indicating continuous deletions in exon 2 and insertions/duplications in exon 3 in these lineages. Specifically, we confirmed the presence the fbl gene in chicken and validated the RNA expression. Our analyses of the GAR-encoding exons of fbl in vertebrates and reptiles should provide the basis for further evolutionary analyses of more GAR domain encoding proteins.
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Arginina , Glicina , Animais , Vertebrados/genética , Éxons/genética , Metiltransferases , Répteis/genética , AvesRESUMO
Cordyceps militaris (CM) is a popular medicinal fungus; however, few studies have focused on its impact on the male reproductive system. We evaluated the effects of CM fermentation products on the reproductive development of juvenile male (JM) mice. Mice were divided into four experimental groups, each fed 5% CM products (weight per weight (w/w) in normal diet): extracellular polysaccharides (EPS), fermentation broth (FB), mycelia (MY), and whole fermentation products (FB plus MY, FBMY) for 28 days, while mice in the control group (CT) were fed a normal diet. Basic body parameters, testicular structure, sperm parameters, and sex hormones concentrations were analyzed. Compared to the CT group, mice in the EPS, MY, and FBMY groups showed a significantly increased mean seminiferous tubule area (p < 0.05), mice in the FB and MY groups had significantly higher sperm concentrations (p < 0.05), and mice in the EPS, FB, and FBMY groups showed significantly increased ratios of motile sperm (p < 0.05). Meanwhile, EPS significantly promoted the ability of JM mice to synthesize testosterone (p < 0.05). Furthermore, all CM products significantly increased the food intake of JM mice (p < 0.05) but did not significantly change their water intake and body weight gain (p > 0.05). In conclusion, CM products, especially EPS, exhibit strong androgen-like activities that can promote male reproductive development.
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Cordyceps , Animais , Cordyceps/química , Fermentação , Masculino , Camundongos , Micélio , Polissacarídeos/análise , SementesRESUMO
T helper (Th)2 cytokines such as interleukin (IL)-4 and IL-13 control immune function by acting on leukocytes. They also regulate multiple responses in non-hematopoietic cells. During pregnancy, IL-4 and IL-13 facilitate alveologenesis of mammary glands. This particular morphogenesis generates alveoli from existing ducts and requires substantial cell proliferation. Using 3D cultures of primary mouse mammary epithelial cells, we demonstrate that IL-4 and IL-13 promote cell proliferation, leading to enlargement of mammary acini with partially filled lumens. The mitogenic effects of IL-4 and IL-13 are mediated by STAT6 as inhibition of STAT6 suppresses cell proliferation and improves lumen formation. In addition, IL-4 and IL-13 stimulate tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1). Prolonged treatment with these cytokines leads to increased IRS-1 abundance, which, in turn, amplifies IL-4- and IL-13-stimulated IRS-1 tyrosine phosphorylation. Through signaling crosstalk between IL-4/IL-13 and insulin, a hormone routinely included in mammary cultures, IRS-1 tyrosine phosphorylation is further enhanced. Lowering IRS-1 expression reduces cell proliferation, suggesting that IRS-1 is involved in IL-4- and IL-13-stimulated cell proliferation. Thus, a Th2-dominant cytokine milieu during pregnancy confers mammary gland development by promoting cell proliferation.
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Técnicas de Cultura de Células em Três Dimensões/métodos , Células Epiteliais/citologia , Proteínas Substratos do Receptor de Insulina/metabolismo , Interleucina-13/metabolismo , Interleucina-4/metabolismo , Glândulas Mamárias Animais/citologia , Fator de Transcrição STAT6/metabolismo , Animais , Proliferação de Células , Células Epiteliais/metabolismo , Feminino , Glândulas Mamárias Animais/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Modelos Animais , Fosforilação , Gravidez , Transdução de SinaisRESUMO
Acetaminophen (APAP) overdose induces acute liver damage and even death. The standard therapeutic dose of N-acetyl cysteine (NAC) cannot be applied to every patient, especially those with high-dose APAP poisoning. There is insufficient evidence to prove that increasing NAC dose can treat patients who failed in standard treatment. This study explores the toxicity of NAC overdose in both APAP poisoning and normal mice. Two inbred mouse strains with different sensitivities to propacetamol-induced hepatotoxicity (PIH) were treated with different NAC doses. NAC therapy decreased PIH by reducing lipid oxidation, protein nitration and inflammation, and increasing glutathione (GSH) levels and antioxidative enzyme activities. However, the therapeutic effects of NAC on PIH were dose-dependent from 125 (N125) to 275 mg/kg (N275). Elevated doses of NAC (400 and 800 mg/kg, N400 and N800) caused additional deaths in both propacetamol-treated and normal mice. N800 treatments significantly decreased hepatic GSH levels and induced inflammatory cytokines and hepatic microvesicular steatosis in both propacetamol-treated and normal mice. Furthermore, both N275 and N400 treatments decreased serum triglyceride (TG) and induced hepatic TG, whereas N800 treatment significantly increased interleukin-6, hepatic TG, and total cholesterol levels. In conclusion, NAC overdose induces hepatic and systemic inflammations and interferes with fatty acid metabolism.
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This study was designed to evaluate the anti-inflammatory effects of Alpinia officinarum Hance extract (AOE) and identify its main active ingredients. AOE was obtained using a 95% ethanol extraction method. Lipopolysaccharide (LPS) were used to induce an inflammatory response in RAW264.7 cells. The results showed that AOE exerts anti-inflammatory effects via inhibition of prostaglandin E2 secretion and cyclooxygenase -2 (COX-2) production. We further analyzed the components of AOE using high-performance liquid chromatography and found that AOE is comprised of several bioactive flavonoids including quercetin (Q), kaempferol (K), galangin (G), and curcumin (C). These four flavonoids effectively inhibited nitric oxide (NO), interleukin (IL)-1ß, IL-6, and tumor necrosis factor-α production. Moreover, they reduced COX-2 and inducible NO synthase expressions via regulation of nuclear factor kappa-light-chain-enhancer of activated B cells and c-Jun N-terminal kinase signaling pathways. Furthermore, we compared and contrasted the anti-inflammatory effects and mechanisms of these four flavonoids at the same dose in the LPS-induced cell inflammation model. The results showed that C is the most effective inhibitor of LPS-induced NO production. However, only Q and K effectively attenuated LPS-induced extracellular signal-regulated kinase and p38 elevations. In conclusion, AOE and its major bioactive compounds exert anti-inflammatory effects on LPS-induced inflammation. As A. officinarum Hance is much cheaper than any of its four flavonoids, especially G, we suggest using AOE as an anti-inflammatory agent.
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Alpinia , NF-kappa B , Alpinia/metabolismo , Animais , Anti-Inflamatórios/farmacologia , Ciclo-Oxigenase 2 , Lipopolissacarídeos , Macrófagos , Camundongos , NF-kappa B/metabolismo , Óxido Nítrico , Óxido Nítrico Sintase Tipo IIRESUMO
O-acetyl-ADP-ribose (AAR) is a metabolic small molecule relevant in epigenetics that is generated by NAD-dependent histone deacetylases, such as Sir2. The formation of silent heterochromatin in yeast requires histone deacetylation by Sir2, structural rearrangement of SIR complexes, spreading of SIR complexes along the chromatin, and additional maturation processing. AAR affects the interactions of the SIR-nucleosome in vitro and enhances the chromatin epigenetic silencing effect in vivo. In this study, using isothermal titration calorimetry (ITC) and dot blotting methods, we showed the direct interaction of AAR with Sir3. Furthermore, through chromatin immunoprecipitation (ChIP)-on-chip and chromatin affinity purification (ChAP)-on chip assays, we discovered that AAR is capable of increasing the extended spreading of Sir3 along telomeres, but not Sir2. In addition, the findings of a quantitative real-time polymerase chain reaction (qRT-PCR) and examinations of an in vitro assembly system of SIR-nucleosome heterochromatin filament were consistent with these results. This study provides evidence indicating another important effect of AAR in vivo. AAR may play a specific modulating role in the formation of silent SIR-nucleosome heterochromatin in yeast.
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Cromatina/genética , O-Acetil-ADP-Ribose/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Telômero/genética , Epigênese Genética , Regulação Fúngica da Expressão Gênica , Código das Histonas , Ligação Proteica , Saccharomyces cerevisiaeRESUMO
In Saccharomyces cerevisiae, Sir proteins mediate heterochromatin epigenetic gene silencing. The assembly of silent heterochromatin requires histone deacetylation by Sir2, conformational change of SIR complexes, and followed by spreading of SIR complexes along the chromatin fiber to form extended silent heterochromatin domains. Sir2 couples histone deacetylation and NAD hydrolysis to generate an epigenetic metabolic small molecule, O-acetyl-ADP-ribose (AAR). Here, we demonstrate that AAR physically associates with Sir3 and that polySir3-AAR formation has a specific and essential role in the assembly of silent SIR-nucleosome pre-heterochromatin filaments. Furthermore, we show that AAR is capable of stabilizing binding of the Sir3 BAH domain to the Sir3 carboxyl-terminal region. Our data suggests that for the assembly of SIR-nucleosome pre-heterochromatin filament, the structural rearrangement of SIR-nucleosome is important and result in creating more stable interactions of Sir3, such as the inter-molecule Sir3-Sir3 interaction, and the Sir3-nucleosome interaction within the filaments. In conclusion, our results reveal the importance of AAR, indicating that it not only affects the conformational rearrangement of SIR complexes but also might function as a critical fine-tuning modulatory component of yeast silent SIR-nucleosome pre-heterochromatin by stabilizing the intermolecular interaction between Sir3 N- and C-terminal regions.
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Heterocromatina/metabolismo , Nucleossomos/metabolismo , O-Acetil-ADP-Ribose/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Epigênese Genética , Ligação Proteica , Conformação Proteica , Estabilidade Proteica , Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/química , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/genética , Sirtuína 2/genética , Sirtuína 2/metabolismoRESUMO
To study the epigenetic gene silencing, yeast is an excellent model organism. Sir proteins are required for the formation of silent heterochromatin. Sir2 couples histone deacetylation and NAD hydrolysis to generate an endogenous epigenetic metabolic small molecule, O-acetyl-ADP-ribose (AAR). AAR is involved in the conformational change of SIR complexes, modulates the formation of SIR-nucleosome preheterochromatin and contributes to the spreading of SIR complexes along the chromatin fiber to form extended silent heterochromatin regions. Here, we show that AAR is capable of enhancing the chromatin silencing effect under either an extra exogenous AAR or a defect AAR metabolic enzyme situation, but decreasing the chromatin silencing effect under a defect AAR synthetic enzyme state. Our results provide an evidence of biological function importance of AAR. It is indicated that AAR does not only function in vitro but also play a role in vivo to increase the effect of heterochromatin epigenetic gene silencing. However, further investigations of AAR are warranted to expand our knowledge of epigenetics and associated small molecules.
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Cromatina/genética , O-Acetil-ADP-Ribose/genética , O-Acetil-ADP-Ribose/metabolismo , Cromatina/fisiologia , Epigênese Genética/genética , Epigenômica/métodos , Inativação Gênica/fisiologia , Heterocromatina/metabolismo , Histonas/metabolismo , Nucleossomos/metabolismo , O-Acetil-ADP-Ribose/fisiologia , Processamento de Proteína Pós-Traducional/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuína 2/genética , Sirtuína 2/metabolismo , Sirtuínas/genética , Sirtuínas/metabolismoRESUMO
INTRODUCTION: The development of tyrosinase inhibitors is a hot research topic. Recently, the Chinese herb Paeonia suffruticosa Andrews, commonly named as Cortex Moutan (CM), was reported as being capable of reducing melanogenesis. We developed an A2058 human melanoma cell model to test the safety and efficacy of tyrosinase inhibition. The aim was to further clarify the bioactivities of CM extracts and paeonol for the purpose of skin whitening. METHODS: The 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity, total polyphenol and flavonoid contents, and in vitro tyrosinase inhibitory effects of water and ethanol CM extracts were determined. Cellular inhibitions of tyrosinase and melanin production were also evaluated. RESULTS: Water and ethanol CM extracts were both shown to have strong DPPH scavenging abilities in a dose-dependent manner. The polyphenol content was higher in the ethanol CM extract compared to the water extract, while the flavanone content was comparable. Kinetic analyses revealed that the ethanol CM extract and paeonol are noncompetitive tyrosinase inhibitors. The cellular melanin content and l-DOPA oxidation assays demonstrated that the ethanol CM extract was an appropriate alternative whitening agent to paeonol and arbutin in ultraviolet-induced A2058 human melanoma cells. CONCLUSION: The results of this study suggest that a human cell model is more suitable for determining tyrosinase activity than mouse cell models for determining cellular tyrosinase activity and melanin production. The ethanol CM extract was also confirmed as a promising ingredient in sun protection and skin whitening cosmetics. Future work should focus on melanogenesis-related gene expressions.
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Medicamentos de Ervas Chinesas/uso terapêutico , Melanoma/tratamento farmacológico , Monofenol Mono-Oxigenase/antagonistas & inibidores , Fitoterapia , Extratos Vegetais/uso terapêutico , Neoplasias Cutâneas/tratamento farmacológico , Linhagem Celular Tumoral , Medicamentos de Ervas Chinesas/efeitos adversos , Humanos , Paeonia/efeitos adversos , Extratos Vegetais/efeitos adversos , Resultado do TratamentoRESUMO
Eucalyptus globulus possesses important pharmacological activities, including antioxidant and anti-inflammatory effects. We investigated the anti-fatigue, antioxidant, and anti-inflammatory effects of eucalyptus essential oil after swimming exercise using an animal model. Male Sprague Dawley rats were administered eucalyptus oil (200 µL/h) daily via inhalation (15 min), and anti-fatigue effects were assessed following eucalyptus essential oil administration for 2 or 4 weeks when forced to swim until exhaustion while carrying ~5% body weight-equivalent. To assess antioxidant and anti-inflammatory effects, control and oil-treated groups were subjected to swimming, which was intensified from 90 min to 120 min daily over 4 weeks, with non-swimming groups included as controls. The 2- and 4-week-treated rats increased their swimming-to-exhaustion time by 46 s and 111 s, respectively. Additionally, lactate (LA), creatine kinase (CK), and lactate dehydrogenase (LDH) activities increased significantly in the non-treated swimming relative to levels observed in the non-swimming groups (P < 0.05); however, no significant differences in these markers were observed between the treated groups. The anti-fatigue effects were related to LA clearance and reduced LDH and CK concentrations. Moreover, compared to the corresponding levels in the non-swimmers, the non-treated swimmers showed markedly elevated levels of liver malondialdehyde (MDA), xanthine oxidase (XO), and other factors, but significantly decreased (P < 0.05) glutathione (GSH) concentrations. However, compared with that of the non-swimmer group, the treated swimming group showed no significant changes in these levels (P > 0.05), suggesting stable XO and MDA production and maintenance of GSH levels. These results suggested that eucalyptus oil aromatherapy increased rat swimming performance and antioxidant capacity and decreased oxidative damage and inflammatory reactions in tissues, indicating good anti-fatigue, antioxidant, and anti-inflammatory effects after high-intensity endurance exercise.
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Aromaterapia , Animais , Anti-Inflamatórios , Antioxidantes , Óleo de Eucalipto , Fadiga , Masculino , Condicionamento Físico Animal , Ratos , Ratos Sprague-Dawley , NataçãoRESUMO
Acetaminophen (APAP) overdose can induce acute liver injury (ALI) with significant morbidity and mortality. Propacetamol is an APAP prodrug, which is clinically bioequivalent to APAP. Kaempferol, a dietary flavonoid, has antioxidant, anti-inflammatory, and anti-apoptotic effects. In this study, we investigated the protective effect of kaempferol on propacetamol-induced ALI and its underlying mechanism in mice. Kaempferol pretreatment (125â¯mg/kg) before propacetamol injection significantly decreased propacetamol-induced serum ALT and AST activities, and DNA fragmentation. Kaempferol administration also reduced propacetamol-induced oxidative stress by inhibiting thiobarbituric acid reactive substances (TBARS) and 3-nitrotyrosine (3-NT) formation partly through downregulation of cytochrome P450 2E1 (CYP2E1) expression, upregulation of UDP glucuronosyltransferase family 1 member A1 (UGT1A1) expression, restoration of the activities of antioxidant enzymes including SOD, GPx and catalase toward normal, recovery of propacetamol-suppressed Nrf2 and GCLC expressions, and maintenance of normal glutathione level. Furthermore, kaempferol markedly attenuated APAP-induced serum TNF-α and IL-6 productions, downregulated APAP-induced phosphorylations of JNK and ERK, and decreased early hepatic apoptosis via decreasing Bax/Bcl-2 ratio and caspase 3 activation. Furthermore, administration of N-acetylcysteine (NAC) and kaempferol significantly rescued more mice than a low dose of NAC only did when a lethal dose of propacetamol injected and therapized at a delayed time point. These data suggested that kaempferol protects the liver against propacetamol-induced injury through anti-oxidative, anti-inflammatory and anti-apoptotic activities.
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Acetaminofen/análogos & derivados , Lesão Pulmonar Aguda/prevenção & controle , Apoptose/efeitos dos fármacos , Citocromo P-450 CYP2E1/metabolismo , Glucuronosiltransferase/metabolismo , Inflamação/prevenção & controle , Quempferóis/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Acetaminofen/toxicidade , Acetilcisteína/farmacologia , Lesão Pulmonar Aguda/induzido quimicamente , Animais , Dano ao DNA , MAP Quinases Reguladas por Sinal Extracelular/antagonistas & inibidores , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Fator 2 Relacionado a NF-E2/fisiologia , Superóxido Dismutase/metabolismoRESUMO
Lipopolysaccharide (LPS) released from gram-negative bacteria stimulates immune responses in infected cells. Epigenetic modifications such as DNA methylation and protein methylation modulate LPS-induced innate immune gene expressions. Expression of the Klotho protein decreased with LPS treatment in rats. In a cellular model, information regarding the effect of LPS on Klotho expression was meager. In the present study, we demonstrated that LPS triggered global DNA and protein methylation in glomerular mesangial MES-13 cells. LPS upregulated protein expressions of enzymes central to cellular methylation reactions, especially protein arginine methyltransferase 6 (PRMT6) in MES-13 cells. Expression of the Klotho protein was diminished by LPS and was restored by 5-Aza-2'-deoxycytidine (5-Aza-2'-dc), AMI-1, and ammonium pyrrolidinedithiocarbamate (PDTC), but not adenosine aldehyde (AdOx). NF-κB was identified as a substrate for arginine methylation and interacted with PRMT6 in MES-13 cells. Inhibition of PRMT activity by AMI-1 blocked LPS-induced NF-κB nuclear translocation in MES-13 cells. Our data indicate that NF-κB negatively regulated Klotho expression with an interaction with PRMT6, which was upregulated by LPS in MES-13 cells.
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Glucuronidase/metabolismo , Lipopolissacarídeos/farmacologia , Células Mesangiais/citologia , NF-kappa B/metabolismo , Proteína-Arginina N-Metiltransferases/metabolismo , Animais , Células Cultivadas , Metilação de DNA , Epigênese Genética , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas Klotho , Células Mesangiais/efeitos dos fármacos , Células Mesangiais/metabolismo , Metilação , Camundongos , Regulação para CimaRESUMO
Cisplatin is a chemotherapeutic agent widely used in the treatment of various cancers. However, cisplatin can induce nephrotoxicity and neurotoxicity, limiting its dosage and usage. Galangin, a natural flavonol, has been found to exhibit anti-oxidant and anti-inflammatory effects in vivo. Here, we investigated the effects of galangin on cisplatin-induced acute kidney injury (AKI) and its molecular mechanisms in mice. Galangin administration reduced the cisplatin-induced oxidative stress by decreasing renal MDA and 3-NT formations. Galangin administration also increased renal anti-oxidative enzyme activities (SOD, GPx, and CAT) and GSH levels depleted by cisplatin. Furthermore, galangin administration inactivated stress-induced Nrf2 protein and its downstream products, HO-1 and GCLC. In terms of the inflammatory response, galangin administration reduced IκBα phosphorylation, NF-κB phosphorylation and nuclear translocation, and then inhibited cisplatin-induced secretions of pro-inflammatory TNF-α, IL-1ß and IL-6. In addition, cisplatin-induced ERK and p38 phosphorylations were inhibited by galangin administration. In terms of cell death, galangin administration reduced levels of p53, pro-apoptotic Bax and activated caspase-3 to inhibit the cisplatin-induced apoptosis. Galangin administration also reduced the expression levels of RIP1 and RIP3 to inhibit cisplatin-induced RIP1/RIP3-dependent necroptosis. Therefore, galangin administration significantly ameliorates cisplatin-induced nephrotoxicity by attenuating oxidative stress, inflammation, and cell death through inhibitions of ERK and NF-κB signaling pathways. Galangin might be a potential adjuvant for clinical cisplatin therapy.
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Injúria Renal Aguda/prevenção & controle , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Cisplatino , Citocinas/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Flavonoides/farmacologia , Mediadores da Inflamação/metabolismo , Rim/efeitos dos fármacos , NF-kappa B/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Injúria Renal Aguda/induzido quimicamente , Injúria Renal Aguda/enzimologia , Injúria Renal Aguda/patologia , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Citoproteção , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Glutamato-Cisteína Ligase/metabolismo , Heme Oxigenase-1/metabolismo , Rim/enzimologia , Rim/patologia , Masculino , Malondialdeído/metabolismo , Proteínas de Membrana/metabolismo , Camundongos Endogâmicos BALB C , Fator 2 Relacionado a NF-E2/metabolismo , Fosforilação , Transdução de Sinais/efeitos dos fármacos , Tirosina/análogos & derivados , Tirosina/metabolismoRESUMO
Yeast silent heterochromatin provides an excellent model with which to study epigenetic inheritance. Previously we developed an in vitro assembly system to demonstrate the formation of filament structures with requirements that mirror yeast epigenetic gene silencing in vivo. However, the properties of these filaments were not investigated in detail. Here we show that the assembly system requires Sir2, Sir3, Sir4, nucleosomes, and O-acetyl-ADP-ribose. We also demonstrate that all Sir proteins and nucleosomes are components of these filaments to prove that they are SIR-nucleosome filaments. Furthermore, we show that the individual localization patterns of Sir proteins on the SIR-nucleosome filament reflect those patterns on telomeres in vivo. In addition, we reveal that magnesium exists in the SIR-nucleosome filament, with a role similar to that for chromatin condensation. These results suggest that a small number of proteins and molecules are sufficient to mediate the formation of a minimal yeast silent pre-heterochromatin in vitro.
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Inativação Gênica/fisiologia , Nucleossomos/metabolismo , O-Acetil-ADP-Ribose/metabolismo , Sítios de Ligação , Cromatina/metabolismo , Montagem e Desmontagem da Cromatina , Epigenômica/métodos , Heterocromatina/metabolismo , Histonas/metabolismo , Magnésio , Processamento de Proteína Pós-Traducional , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Informação Silenciosa de Saccharomyces cerevisiae/metabolismo , Sirtuínas/metabolismo , Telômero/metabolismoRESUMO
Insulated isothermal PCR (iiPCR) method was recently available for rapid on-site detection of roundup ready soybean (RRS; event GTS40-3-2) in food materials and products. Performance of this method was evaluated in this study. The 100% detection endpoint for the RRS by iiPCR was found in samples containing 0.1% RRS, equivalent to the results of the reference real-time PCR (rtPCR). Analysis of nucleic acids of soybean-based processed food products indicated 95% agreement between the iiPCR and rtPCR for RRS detection. By testing soybean milk and tofu samples using simple pretreatment methods, we found that the agreements between iiPCR and rtPCR methods of the aforementioned samples were 80% and 90%, respectively. Replicated tests of all discrepant samples implied that these samples had trace amounts of RRS, suggesting that the iiPCR system is more sensitive than the rtPCR method. In conclusion, the iiPCR technology can be a useful point-of-need tool to help make a timely decision in the consumption of genetically modified organisms.
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Acetaminophen (APAP) overdose causes severe liver and kidney damage. APAP-induced liver injury (AILI) represents the most frequent cause of drug-induced liver failure. APAP is relatively insoluble and can only be taken orally; however, its prodrug, propacetamol, is water soluble and usually injected directly. In this study, we examined the time-dependent effects of AILI after propacetamol injection in mice. After analyses of alanine aminotransferase and aspartate aminotransferase activities and liver histopathology, we demonstrated that a novel AILI mouse model can be established by single propacetamol injection. Furthermore, we compared the protective and therapeutic effects of galangin with a known liver protective extract, silymarin, and the only clinical agent for treating APAP toxicity, N-acetylcysteine (NAC), at the same dose in the model mice. We observed that galangin and silymarin were more effective than NAC for protecting against AILI. However, only NAC greatly improved both the survival time and rate consequent to a lethal dose of propacetamol. To decipher the hepatic protective mechanism(s) of galangin, galangin pretreatment significantly decreased the hepatic oxidative stress, increased hepatic glutathione level, and decreased hepatic microsomal CYP2E1 levels induced by propacetamol injection. In addition, propacetamol injection also reproduced the probability of APAP-induced kidney injury (AIKI), appearing similar to a clinical APAP overdose. Only galangin pretreatment showed the protective effect of AIKI. Thus, we have established a novel mouse model for AILI and AIKI using a single propacetamol injection. We also demonstrated that galangin provides significant protection against AILI and AIKI in this mouse model.
Assuntos
Acetaminofen/efeitos adversos , Doença Hepática Induzida por Substâncias e Drogas/prevenção & controle , Flavonoides/uso terapêutico , Fígado/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Fitoterapia , Extratos Vegetais/uso terapêutico , Acetaminofen/administração & dosagem , Acetaminofen/análogos & derivados , Acetilcisteína/uso terapêutico , Alanina Transaminase/sangue , Alpinia/química , Animais , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Doença Hepática Induzida por Substâncias e Drogas/patologia , Citocromo P-450 CYP2E1/metabolismo , Modelos Animais de Doenças , Flavonoides/farmacologia , Glutationa/metabolismo , Helichrysum/química , Fígado/metabolismo , Fígado/patologia , Masculino , Camundongos Endogâmicos BALB C , Extratos Vegetais/farmacologia , Substâncias Protetoras/farmacologia , Substâncias Protetoras/uso terapêutico , Silimarina/uso terapêuticoRESUMO
Homeobox genes encode transcription factors that regulate embryonic development programs including organogenesis, axis formation and limb development. Previously, we identified and cloned a mouse double homeobox gene, Duxbl, whose homeodomain exhibits the highest identity (67 %) to human DUX4, a candidate gene of facioscapulohumeral muscular dystrophy (FSHD). Duxbl proteins have been shown to be expressed in elongated myocytes and myotubes of trunk and limb muscles during embryogenesis. In this study, we found that Duxbl maintained low expression levels in various adult muscles. Duxbl proteins were induced to express in activated satellite cells and colocalized with MyoG, a myogenic differentiating marker. Furthermore, Duxbl proteins were not detected in quiescent satellite cells but detected in regenerated myocytes and colocalized with MyoD and MyoG following cardiotoxin-induced muscle injury. Ectopic Duxbl overexpressions in C2C12 myoblast cells promoted cell proliferation through mainly enhancing cyclin D1 and hyper-phosphorylated retinoblastoma protein but reducing p21 expression. However, Duxbl overexpression in C2C12 cells inhibited myogenic differentiation by decreasing MyoD downstream gene expressions, including M-cadherin, MyoG, p21 and cyclin D3 but not MyoD itself. Duxbl overexpressions also promoted cell proliferation but blocked MyoD-induced myogenic conversion in multipotent mesenchymal C3H10T1/2 cells. In addition, results of a luciferase reporter assay suggest that Duxbl negatively regulated MyoG promoter activity through the proximal two E boxes. In conclusion, these results indicate that Duxbl may play a crucial role in myogenesis and postnatal muscle regeneration by activating and proliferating satellite and myoblast cells.
Assuntos
Diferenciação Celular , Proteínas de Homeodomínio/genética , Proteína MyoD/genética , Mioblastos/citologia , Mioblastos/metabolismo , Fatores de Transcrição/genética , Ativação Transcricional/genética , Envelhecimento/metabolismo , Animais , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/genética , Proliferação de Células , Imunofluorescência , Proteínas de Homeodomínio/metabolismo , Humanos , Camundongos , Desenvolvimento Muscular , Proteína MyoD/metabolismo , Miogenina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Regeneração , Células Satélites de Músculo Esquelético/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Aristolochic acid (AA) is a common cause of Chinese herb nephropathy. The mechanisms involved in the pathogenesis of AA nephropathy (AAN) are intricate. One well-documented effect of AA in the kidney is its pro-fibrotic activity. Nitric oxide (NO), a messenger gas generated from l-arginine, is the product of nitric oxide synthase (NOS). NO is involved in renal hemodynamics and exerts cytoprotective effects against renal injury. In the present study, the role of NO in AAN was investigated in MES-13 cells, a glomerular mesangial cell line. NO endogenously generated by the induction of inducible nitric oxide synthase (iNOS) with lipopolysaccharide (LPS)/interferon-γ (IFN-γ) significantly downregulated connective tissue growth factor (CTGF) protein expression in MES-13 cells. AA significantly suppressed LPS/IFN-γ-induced NO production and reversed CTGF expression that was downregulated by LPS/IFN-γ. AA decreased iNOS gene and protein expressions in a concentration-dependent manner. AA caused declines in LPS/IFN-γ-induced signal transducer and activator of transcription-1α (STAT-1α) phosphorylation and interferon response factor-1 (IRF-1) mRNA expression. Furthermore, AA attenuated IκB phosphorylation and reduced NF-κB translocation to the nuclear fraction. Taken together, our data indicate that AA reversed the CTGF expression inhibited by LPS/IFN-γ treatment via suppression of NO and iNOS expressions in MES-13 cells through inhibition of the JAK/STAT-1α and NF-κB signaling pathways. NO potentially exerts antifibrotic activity by down regulation of CTGF in MES-13 cells and inhibition of the iNOS gene by AA might partially account for the fibrotic effects of AA in nephropathy.