RESUMO
Staphylococcus aureus (S. aureus)forms biofilm that causes periprosthetic joint infections and osteomyelitis (OM) which are the intractable health problems in clinics. The silver-containing nanoparticles (AgNPs) are antibacterial nanomaterials with less cytotoxicity than the classic Ag compounds. Likewise, gold nanoparticles (AuNPs) have also been demonstrated as excellent nanomaterials for medical applications. Previous studies have showed that both AgNPs and AuNPs have anti-microbial or anti-inflammatory properties. We have developed a novel green chemistry that could generate the AuAg nanocomposites, through the reduction of tannic acid (TNA). The bioactivity of the nanocomposites was investigated inS. aureusbiofilm-exposed human osteoblast cells (hFOB1.19). The current synthesis method is a simple, low-cost, eco-friendly, and green chemistry approach. Our results showed that the AuAg nanocomposites were biocompatible with low cell toxicity, and did not induce cell apoptosis nor necrosis in hFOB1.19 cells. Moreover, AuAg nanocomposites could effectively inhibited the accumulation of reactive oxygen species (ROS) in mitochondria and in rest of cellular compartments after exposing to bacterial biofilm (by reducing 0.78, 0.77-fold in the cell and mitochondria, respectively). AuAg nanocomposites also suppressed ROS-triggered inflammatory protein expression via MAPKs and Akt pathways. The current data suggest that AuAg nanocomposites have the potential to be a good therapeutic agent in treating inflammation in bacteria-infected bone diseases.
Assuntos
Nanopartículas Metálicas , Nanocompostos , Humanos , Ouro/farmacologia , Nanopartículas Metálicas/química , Staphylococcus aureus , Espécies Reativas de Oxigênio/metabolismo , Antibacterianos/farmacologia , Antibacterianos/química , Bactérias , Nanocompostos/química , Biofilmes , Inflamação/tratamento farmacológico , Testes de Sensibilidade MicrobianaRESUMO
The burning incense (BI) behavior could be widely observed in Asia families. Incense sticks are often believed to be made from natural herbs and powders, and to have minimal impact on human health; however, there is limited research to support this claim. The current study aimed to identify the components of BI within the particulate matter 2.5 µm (PM2.5) range and explore if BI has bio-toxicity effects on rat astrocytes (CTX-TNA2). The study also examined the protective effects and underlying molecular mechanisms of tanshinone IIA, a primary lipid-soluble compound found in the herb danshen (Salvia miltiorrhiza Bunge), which has been shown to benefit the central nervous system. Results showed that despite the differences in BI components compared to the atmospheric particulate matter (PM) standards, BI still had a bio-toxicity on astrocytes. BI exposure caused early and late apoptosis, reactive oxygen species (ROS) production, MAPKs (JNK, p38, and ERK), and Akt signaling activation, and inflammation-related proteins (cPLA2, COX-2, HO-1, and MMP-9) increases. Our results further exhibit that the tanshinone IIA pre-treatment could significantly avoid the BI-induced apoptosis and inflammatory signals on rat astrocytes. These findings suggest that BI exposure may cause oxidative stress in rat astrocytes and increase inflammation-related proteins and support the potential of tanshinone IIA as a candidate for preventing BI-related adverse health effects.
Assuntos
Abietanos , Astrócitos , Ratos , Animais , Humanos , Abietanos/farmacologia , Estresse Oxidativo , Inflamação/induzido quimicamenteRESUMO
BACKGROUND: Although the powerful clinical effects of radiofrequency and microwave ablation have been established, such ablation is associated with several limitations, including a small ablation size, a long ablation time, the few treatment positioning, and biosafety risks. To overcome these limitations, biosafe and efficient magnetic ablation was achieved in this study by using biocompatible liquid gallium as an ablation medium and a contrast medium for imaging. RESULTS: Magnetic fields with a frequency (f) lower than 200 kHz and an amplitude (H) × f value lower than 5.0 × 109 Am-1 s-1 were generated using the proposed method. These fields could generate an ablation size of 3 cm in rat liver lobes under a temperature of approximately 300 °C and a time of 20 s. The results of this study indicate that biomedical gallium can be used as a contrast medium for the positioning of gallium injections and the evaluation of ablated tissue around a target site. Liquid gallium can be used as an ablation medium and imaging contrast medium because of its stable retention in normal tissue for at least 3 days. Besides, the high anticancer potential of gallium ions was inferred from the self-degradation of 100 µL of liquid gallium after around 21 days of immersion in acidic solutions. CONCLUSIONS: The rapid wireless ablation of large or multiple lesions was achieved through the simple multi-injection of liquid gallium. This approach can replace the currently favoured procedure involving the use of multiple ablation probes, which is associated with limited benefits and several side effects. METHODS: Magnetic ablation was confirmed to be highly efficient by the consistent results obtained in the simulation and in vitro tests of gallium and iron oxide as well as the electromagnetic specifics and thermotherapy performance comparison detailed in this study Ultrasound imaging, X-ray imaging, and magnetic resonance imaging were found to be compatible with the proposed magnetic ablation method. Self-degradation analysis was conducted by mixing liquid gallium in acidic solutions with a pH of approximately 5-7 (to imitate a tumour-containing microenvironment). X-ray diffraction was used to identify the gallium oxides produced by degraded gallium ions.
Assuntos
Técnicas de Ablação , Ablação por Cateter , Gálio , Animais , Gálio/farmacologia , Fígado/diagnóstico por imagem , Imageamento por Ressonância Magnética , Ratos , UltrassonografiaRESUMO
Osteosarcoma, a primary bone tumor, responds poorly to chemotherapy and radiation therapy in children and young adults; hence, as the basis for an alternative treatment, this study investigated the cytotoxic and antiproliferative effects of naringenin on osteosarcoma cell lines, HOS and U2OS, by using cell counting kit-8 and colony formation assays. DNA fragmentation and the increase in the G2/M phase in HOS and U2OS cells upon treatment with various naringenin concentrations were determined by using the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay and Annexin V/propidium iodide double staining, respectively. Flow cytometry was performed, and 2',7'-dichlorodihydrofluorescein diacetate, JC-1, and Fluo-4 AM ester probes were examined for reactive oxygen species (ROS) generation, mitochondrial membrane potential, and intracellular calcium levels, respectively. Caspase activation, cell cycle, cytosolic and mitochondrial, and autophagy-related proteins were determined using western blotting. The results indicated that naringenin significantly inhibited viability and proliferation of osteosarcoma cells in a dose-dependent manner. In addition, naringenin induced cell cycle arrest in osteosarcoma cells by inhibiting cyclin B1 and cyclin-dependent kinase 1 expression and upregulating p21 expression. Furthermore, naringenin significantly inhibited the growth of osteosarcoma cells by increasing the intracellular ROS level. Naringenin induced endoplasmic reticulum (ER) stress-mediated apoptosis through the upregulation of ER stress markers, GRP78 and GRP94. Naringenin caused acidic vesicular organelle formation and increased autophagolysosomes, microtubule-associated protein-light chain 3-II protein levels, and autophagy. The findings suggest that the induction of cell apoptosis, cell cycle arrest, and autophagy by naringenin through mitochondrial dysfunction, ROS production, and ER stress signaling pathways contribute to the antiproliferative effect of naringenin on osteosarcoma cells.
Assuntos
Espécies Reativas de OxigênioRESUMO
OBJECTIVE: To investigate protective effects of Taiwanese green propolis (TGP) against high glucose-induced inflammatory responses in human gingival fibroblasts (HGFs) through NLRP3 inflammasome signaling pathway. BACKGROUND: NLRP3 inflammasome has been implicated in the progression of both diabetes mellitus and periodontitis, suggesting a common potential therapeutic target for these diseases. Propolis is renowned for various biological activities, particularly anti-inflammation and antioxidant, representing a promising therapy for many conditions. However, underlying mechanisms remain unclear. METHODS: The cytotoxicity of TGP was evaluated by cell viability assay. The mRNA levels and protein expression or secretion of various inflammatory molecules and NLRP3 inflammasome-related molecules in high glucose-exposed HGFs with or without pretreatment of TGP (5 µg/ml) were determined by real-time PCR and western blot or specific kits, respectively. Intracellular and mitochondrial ROS measurements, NADPH oxidase activity determination, and subcellular fractions were performed to assess ROS generation. The transcriptional activity of NF-κB was measured by luciferase reporter kit. The signaling components were further differentiated using pharmacological inhibitors of ROS and small interfering RNAs of TLR2, TLR4, or NF-κB. RESULTS: High glucose could induce IL-1ß-driven inflammatory responses in HGFs via the activation of NLRP3 inflammasome regulated by TLR2/TLR4 coupled ROS in NF-κB-dependent manner. TGP had no adverse impact on the cell viability of HGFs at concentrations no greater than 10 µg/ml, and could exert inhibitory effects on high glucose-induced inflammatory responses via the interruption of NLRP3 inflammasome signaling pathway. CONCLUSION: Taiwanese green propolis could elicit protective effects against IL-1ß-driven inflammation in high glucose-exposed HGFs through TLR2/TLR4 combined ROS/NF-κB/NLRP3 inflammasome pathway.
Assuntos
Inflamassomos , Própole , Fibroblastos , Glucose/toxicidade , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Proteína 3 que Contém Domínio de Pirina da Família NLR , Própole/farmacologia , Transdução de SinaisRESUMO
Osteoarthritis (OA) is still a recalcitrant musculoskeletal disease on account of its complex biochemistry and mechanical stimulations. Apart from stimulation by external mechanical forces, the regulation of intracellular mechanics in chondrocytes has also been linked to OA development. Recently, visfatin has received significant attention because of the clinical finding of the positive correlation between its serum/synovial level and OA progression. However, the precise mechanism involved is still unclear. This study determined the effect of visfatin on intracellular mechanics and catabolism in human primary chondrocytes isolated from patients. The intracellular stiffness of chondrocytes was analyzed by the particle-tracking microrheology method. It was shown that visfatin damages the microtubule and microfilament networks to influence intracellular mechanics to decrease the intracellular elasticity and viscosity via glycogen synthase kinase 3ß (GSK3ß) inactivation induced by p38 signaling. Further, microtubule network destruction in human primary chondrocytes is predominantly responsible for the catabolic effect of visfatin on the cyclooxygenase 2 upregulation. The present study shows a more comprehensive interpretation of OA development induced by visfatin through biochemical and biophysical perspectives. Finally, the role of GSK3ß inactivation, and subsequent regulation of intracellular mechanics, might be considered as theranostic targets for future drug development for OA.
Assuntos
Condrócitos , Citocinas/fisiologia , Glicogênio Sintase Quinase 3 beta/metabolismo , Nicotinamida Fosforribosiltransferase/fisiologia , Osteoartrite , Citoesqueleto de Actina/metabolismo , Células Cultivadas , Condrócitos/metabolismo , Condrócitos/patologia , Humanos , Microtúbulos/metabolismo , Osteoartrite/metabolismo , Osteoartrite/patologia , Cultura Primária de CélulasRESUMO
Several diseases have been linked to particulate matter (PM) exposure. Outdoor activities, such as road running or jogging, are popular aerobic exercises due to few participatory limitations. Osteoarthritis (OA) is a progressive degenerative joint disease, usually observed at age 40, and not noticed before pain or diagnosis. Although exercise has health benefits, it is unclear whether outdoor jogging in higher PM (standard reference material 1649b, SRM 1649b) concentration environments could affect OA development or severity. Hence, a PM exposure monosodium iodoacetate (MIA)-induced OA animal jogged model was established for investigation. Results showed that high doses of PM (5 mg) significantly increased pro-inflammatory factors such as tumor necrosis factor α (TNF-α), interleukin (IL)-1ß, and IL-6, and M1 macrophages in the lung region, also obtained in systemic IL-6 and TNF-α expressions in this MIA-OA rat model. Moreover, levels of osteocalcin, cartilage oligomeric matrix protein (COMP), and N-telopeptides of type I collagen were especially influenced in MIA+PM groups. Morphological and structural changes of the knee joint were detected by micro-computed tomography images (micro-CT) and immunohistochemistry. MIA + PM rats exhibited severe bone density decrease, cartilage wear, and structure damages, accompanied by lower levels of physical activity, than the sham group and groups receiving MIA or PM alone. The findings suggest that the severity of OA could be promoted by PM exposure with a PM concentration effect via systemic inflammatory mechanisms. To the best of our knowledge, this is the first study to provide direct effects of PM exposure on OA severity.
Assuntos
Artrite Experimental/etiologia , Exposição por Inalação/efeitos adversos , Osteoartrite do Joelho/etiologia , Material Particulado/efeitos adversos , Animais , Artrite Experimental/sangue , Artrite Experimental/patologia , Biomarcadores/sangue , Citocinas/sangue , Ácido Iodoacético , Articulação do Joelho/patologia , Pulmão/metabolismo , Macrófagos/metabolismo , Masculino , Osteoartrite do Joelho/sangue , Osteoartrite do Joelho/patologia , Condicionamento Físico Animal , RatosRESUMO
Spreading depression (SD) is a pathophysiological phenomenon characterized by propagating waves of profound neuronal and glial depolarization in central nervous system gray matter. Although SD is primarily mediated by neurons with a subsequent astrocytic response, it remains unclear how astrocytic activity is modulated after SD and how altered astrocyte signaling contribute to neuronal excitability. Here, we report that after the concurrent Ca2+ wave, SD enhanced astrocytic activity by promoting a secondary period of Ca2+ oscillations. SD-induced Ca2+ oscillations did not require the activation of metabotropic glutamate receptors or purinergic receptors; instead, they were mediated by the activation of GABAB receptors and 1,4,5-trisphosphate (IP3) receptors. Furthermore, SD increased the number of NMDA receptor-mediated slow inward currents (SICs) in CA1 pyramidal neurons. The frequency of SD-induced SICs was reduced by blockade of GABAB receptors or by limiting Ca2+ efflux from the ER. Selective inhibition of astrocytic Ca2+ signals by dialysis of BAPTA into astrocytes or by knocking out the astrocytic type of IP3 receptors suppressed SICs after SD. These results demonstrated a causative link between the SD-induced Ca2+ oscillations and the enhanced glutamatergic astrocyte-neuron signaling. Therefore, we conclude that SD enhances the astrocyte Ca2+ signals and further promotes gliotransmission and neuronal excitability.
Assuntos
Astrócitos/fisiologia , Sinalização do Cálcio/fisiologia , Depressão Alastrante da Atividade Elétrica Cortical/fisiologia , Hipocampo/fisiologia , Neurônios/fisiologia , Transmissão Sináptica/fisiologia , Animais , Feminino , Masculino , Camundongos , Camundongos Endogâmicos ICRRESUMO
Particulate matter (PM), a widespread air pollutant, consists of a complex mixture of solid and liquid particles suspended in air. Many diseases have been linked to PM exposure, which induces an imbalance in reactive oxygen species (ROS) generated in cells, and might result in skin diseases (such as aging and atopic dermatitis). New techniques involving nanomedicine and nano-delivery systems are being rapidly developed in the medicinal field. Fullerene, a kind of nanomaterial, acts as a super radical scavenger. Lower water solubility levels limit the bio-applications of fullerene. Hence, to improve the water solubility of fullerene, while retaining its radical scavenger functions, a fullerene derivative, fullerenol C60(OH)36, was synthesized, to examine its biofunctions in PM-exposed human keratinocyte (HaCaT) cells. The PM-induced increase in ROS levels and expression of phosphorylated mitogen-activated protein kinase and Akt could be inhibited via fullerenol pre-treatment. Furthermore, the expression of inflammation-related proteins, cyclooxygenase-2, heme oxygenase-1, and prostaglandin E2 was also suppressed. Fullerenol could preserve the impaired state of skin barrier proteins (filaggrin, involucrin, repetin, and loricrin), which was attributable to PM exposure. These results suggest that fullerenol could act against PM-induced cytotoxicity via ROS scavenging and anti-inflammatory mechanisms, and the maintenance of expression of barrier proteins, and is a potential candidate compound for the treatment of skin diseases.
Assuntos
Poluição do Ar/prevenção & controle , Fulerenos/análise , Material Particulado/toxicidade , Água/química , Apoptose/efeitos dos fármacos , Linhagem Celular , Cidades , Proteínas Filagrinas , Fulerenos/química , Humanos , Inflamação/patologia , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Queratinócitos/ultraestrutura , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Mitocôndrias/ultraestrutura , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Modelos Biológicos , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Espécies Reativas de Oxigênio/metabolismo , SolubilidadeRESUMO
Malignant melanoma is developed from pigment-containing cells, melanocytes, and primarily found on the skin. Malignant melanoma still has a high mortality rate, which may imply a lack of therapeutic agents. Lakoochin A, a compound isolated from Artocarpus lakoocha and Artocarpus xanthocarpus, has an inhibitory function of tyrosinase activity and melanin production, but the anti-cancer effects are still unclear. In the current study, the therapeutic effects of lakoochin A with their apoptosis functions and possible mechanisms were investigated on A375.S2 melanoma cells. Several methods were applied, including 3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT), flow cytometry, and immunoblotting. Results suggest that lakoochin A attenuated the growth of A375.S2 melanoma cells through an apoptosis mechanism. Lakoochin A first increase the production of cellular and mitochondrial reactive oxygen species (ROSs); mitochondrial ROSs then promote mitogen-activated protein kinases (MAPKs) pathway activation and raise downstream apoptosis-related protein and caspase expression. This is the first study to demonstrate that lakoochin A, through ROS-MAPK, apoptosis-related proteins, caspases cascades, can induce melanoma cell apoptosis and may be a potential candidate compound for treating malignant melanoma.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Melanoma/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Estilbenos/farmacologia , Caspases/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Melanoma/tratamento farmacológico , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismoRESUMO
Methadone maintenance treatment (MMT) is the major tapering therapy for morphine addictive patients. There have gender differences reported in response to MMT. This study discovered that the estrogen-response element single nucleotide polymorphism (ERE-SNP; rs16974799, C/T) of cytochrome 2B6 gene (cyp2b6; methadone catabolic enzyme) responded differently to MMT dosing. Oestradiol was associated with high MMT dosing, high enantiomer (R- or S-) of 2-ethylidene-1,5-dimethyl-3,3-dipheny-pyrrolidine (EDDP; methadone metabolite) to methadone ratio and increased drug-seeking behaviour, implicating oestradiol-CYP-EDDP/methadone axis decreasing MMT efficacy. In mouse model, oestrogen mitigates methadone antinociceptive response, facilitates methadone catabolism and up-regulates methadone-associated metabolizing enzymes. Oestrogen also ablates chronic methadone administration-induced rewarding response. Mechanism dissection revealed the CC genotype of CYP2B6-ERE-SNP exerts higher ERE sequence alignment score, higher estrogenic response as compared to TT genotype. At last, preclinical study via targeting estrogen signal that tamoxifen (TMX; selective estrogen receptor modulator, SERM) could facilitate the tolerance phase rewarding response of methadone. Strikingly, TMX also reduces tapering/abstinence phases methadone liability in mice. In conclusion, this study demonstrates altering methadone metabolism through targeting estrogen signals might be able to free morphine addictive patients from the addiction of opioid replacement therapy. Therefore, the add-on therapy clinical trial introducing SERM in MMT regimen is suggested.
Assuntos
Citocromo P-450 CYP2B6/genética , Estradiol/metabolismo , Antagonistas de Estrogênios/farmacologia , Metadona/farmacologia , Dependência de Morfina/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Tamoxifeno/farmacologia , Adulto , Animais , Citocromo P-450 CYP2B6/metabolismo , Estradiol/farmacologia , Feminino , Regulação da Expressão Gênica , Humanos , Masculino , Metadona/farmacocinética , Camundongos , Camundongos Endogâmicos C57BL , Dependência de Morfina/genética , Dependência de Morfina/metabolismo , Dependência de Morfina/fisiopatologia , Tratamento de Substituição de Opiáceos , Ovariectomia , Polimorfismo de Nucleotídeo Único , Pirrolidinas/metabolismo , Elementos de Resposta , Fatores SexuaisRESUMO
Melanoma is the most malignant form of skin cancer and is associated with a very poor prognosis. The aim of this study was to evaluate the apoptotic effects of cudraflavone C on A375.S2 melanoma cells and to determine the underlying mechanisms involved in apoptosis. Cell viability was determined using the MTT and real-time cytotoxicity assays. Flow cytometric evaluation of apoptosis was performed after staining the cells with Annexin V-FITC and propidium iodide. The mitochondrial membrane potential was evaluated using the JC-1 assay. Cellular ROS production was measured using the CellROX assay, while mitochondrial ROS production was evaluated using the MitoSOX assay. It was observed that cudraflavone C inhibited growth in A375.S2 melanoma cells, and promoted apoptosis via the mitochondrial pathway mediated by increased mitochondrial ROS production. In addition, cudraflavone C induced phosphorylation of MAPKs (p38, ERK, and JNK) and up-regulated the expression of apoptotic proteins (Puma, Bax, Bad, Bid, Apaf-1, cytochrome C, caspase-9, and caspase-3/7) in A375.S2 cells. Pretreatment of A375.S2 cells with MitoTEMPOL (a mitochondria-targeted antioxidant) attenuated the phosphorylation of MAPKs, expression of apoptotic proteins, and the overall progression of apoptosis. In summary, cudraflavone C induced apoptosis in A375.S2 melanoma cells by increasing mitochondrial ROS production; thus, activating p38, ERK, and JNK; and increasing the expression of apoptotic proteins. Therefore, cudraflavone C may be regarded as a potential form of treatment for malignant melanoma.
Assuntos
Apoptose/efeitos dos fármacos , Flavonas/farmacologia , Melanoma/enzimologia , Melanoma/patologia , Mitocôndrias/metabolismo , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Caspases/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Ativação Enzimática/efeitos dos fármacos , Flavonas/química , Humanos , Melanoma/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Proteínas de Neoplasias/metabolismo , Fosforilação/efeitos dos fármacosRESUMO
BACKGROUND: Heroin use among young women of reproductive age has drawn much attention around the world. Although methadone is widely used in maintenance therapy for heroin/morphine addiction, the long-term effects of prenatal exposure to methadone and preventative therapy remain unclear. For revealing this question, female pregnant Sprague-Dawley rats were sub-grouped to receive (1) vehicle, (2) methadone 5 mg/kg at embryonic day 3 (E3) and then 7 mg/kg from E4 to E20, (3) dextromethorphan (DM) 3 mg/kg, and (4) methadone + DM (the rats received methadone followed by DM treatment), subcutaneously, twice a day from E3 to E20. The body weight, natural withdrawal, pain sensitivity, ED50, conditioned place preference and water maze were conducted at different postnatal stages (P1 to P79) of offspring. The quantitative real-time RT-PCR and electrophysiology were also used to measure the gene expression of opioid receptors in the spinal cord and changes of LTP/LTD in the hippocampus, separately. RESULTS: Prenatal exposure to methadone or DM did not affect survival rate, body weight, water maze and LTP or LTD of offspring. However, prenatal methadone significantly increased the withdrawal symptoms, pain sensitivity, addiction liability and decreased the mRNA expression of pain related opioid receptors. Co-administration of DM with methadone in the maternal rats effectively prevented these abnormalities of offspring induced by methadone. CONCLUSIONS: Our study clearly showed that co-administration of dextromethorphan with methadone in the maternal rats prevented the adverse effects induced by prenatal methadone exposure. It implies that dextromethorphan may have a potential to be used in combination with methadone for maintenance treatment in pregnant heroin-addicted women to prevent the adverse effects induced by methadone on offspring.
Assuntos
Antitussígenos/uso terapêutico , Dextrometorfano/uso terapêutico , Metadona/administração & dosagem , Dependência de Morfina/tratamento farmacológico , Entorpecentes/administração & dosagem , Efeitos Tardios da Exposição Pré-Natal/tratamento farmacológico , Animais , Feminino , Masculino , Metadona/toxicidade , Dependência de Morfina/etiologia , Dependência de Morfina/fisiopatologia , Entorpecentes/toxicidade , Gravidez , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Ratos , Ratos Sprague-DawleyRESUMO
Eupafolin, a major active component found in the methanol extracts of Phyla nodiflora, has been used to treat inflammation of skin. We examined its effects on cyclooxygenase-2 (COX-2) expression in LPS-treated human dermal fibroblasts. Lipopolysaccharide (LPS) significantly increased prostaglandin-E2 (PGE2) production associated with increased COX-2 expression in Hs68 cells. This effect was blocked by eupafolin, TLR-4 antibody, antioxidants (APO and NAC), as well as inhibitors, including U0126 (ERK1/2), SB202190 (p38), SP600125 (JNK1/2), and Tanshinone IIA (AP-1). In gene regulation level, qPCR and promoter assays revealed that COX-2 expression was attenuated by eupafolin. In addition, eupafolin also ameliorated LPS-induced p47 phox activation and decreased reactive oxygen species (ROS) generation and NADPH oxidase (Nox) activity. Moreover, pretreatment with eupafolin and APO led to reduced LPS-induced phosphorylation of ERK1/2, JNK, and p38. Further, eupafolin attenuated LPS-induced increase in AP-1 transcription factor binding activity as well as the increase in the phosphorylation of c-Jun and c-Fos. In vivo studies have shown that in dermal fibroblasts of LPS treated mice, eupafolin exerted anti-inflammation effects by decreasing COX-2 protein levels. Our results reveal a novel mechanism for anti-inflammatory and anti-oxidative effects of eupafolin that involved inhibition of LPS-induced ROS generation, suppression of MAPK phosphorylation, diminished DNA binding activity of AP-1 and attenuated COX-2 expression leading to reduced production of prostaglandin E2 (PGE2). Our results demonstrate that eupafolin may be used to treat inflammatory responses associated with dermatologic diseases.
Assuntos
Inibidores de Ciclo-Oxigenase 2/farmacologia , Ciclo-Oxigenase 2/metabolismo , Dinoprostona/metabolismo , Fibroblastos/efeitos dos fármacos , Flavonas/farmacologia , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Lipopolissacarídeos/farmacologia , Glicoproteínas de Membrana/metabolismo , NADPH Oxidases/metabolismo , Transdução de Sinais/efeitos dos fármacos , Pele/efeitos dos fármacos , Fator de Transcrição AP-1/metabolismo , Antioxidantes/farmacologia , Linhagem Celular , Ciclo-Oxigenase 2/genética , Relação Dose-Resposta a Droga , Regulação para Baixo , Fibroblastos/metabolismo , Genes Reporter , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/antagonistas & inibidores , Glicoproteínas de Membrana/genética , NADPH Oxidase 2 , NADPH Oxidases/genética , Estresse Oxidativo/efeitos dos fármacos , Fosforilação , Regiões Promotoras Genéticas , Inibidores de Proteínas Quinases/farmacologia , Interferência de RNA , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Pele/enzimologia , Fatores de Tempo , TransfecçãoRESUMO
Heroin use among young women of reproductive age has drawn much attention around the world. However, there is lack of information on the long-term effects of prenatal exposure to opioids on their offspring. Our previous study demonstrated that prenatally buprenorphine-exposed offspring showed a marked change in the cross-tolerance to morphine compared with other groups. In the current study, this animal model was used to study effects of methamphetamine (METH)-induced behavioral sensitization in the offspring at their adulthood. The results showed no differences in either basal or acute METH-induced locomotor activity in any of the groups of animals tested. When male offspring received METH injections of 2 mg/kg, i.p., once a day for 5 days, behavioral sensitization was induced, as determined by motor activity. Furthermore, the distance and rate of development (slope) of locomotor activity and conditioned place preference induced by METH were significantly increased in the prenatally buprenorphine-exposed animals compared with those in other groups. The dopamine D1 R in the nucleus accumbens of the prenatally buprenorphine-exposed offspring had lower mRNA expression; but no significant changes in the µ-, κ-opioid, nociceptin, D2 R and D3 R receptors were noted. Furthermore, significant alterations were observed in the basal level of cAMP and the D1 R agonist enhanced adenylyl cyclase activity in the prenatally buprenorphine-exposed group. Overall, the study demonstrates that D1 R and its downregulated cAMP signals are involved in enhancing METH-induced behavioral sensitization in prenatally buprenorphine-exposed offspring. The study reveals that prenatal exposure to buprenorphine caused long-term effects on offspring and affected the dopaminergic system-related reward mechanism.
Assuntos
Buprenorfina/farmacologia , Metadona/farmacologia , Metanfetamina/farmacologia , Morfina/farmacologia , Atividade Motora/efeitos dos fármacos , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Analgésicos Opioides/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Estimulantes do Sistema Nervoso Central/farmacologia , Feminino , Masculino , Modelos Animais , Gravidez , RNA Mensageiro/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , RecompensaRESUMO
Background: As lung cancer is the leading cause of cancer death worldwide, the development of new medicines is a crucial endeavor. Naringenin, a flavanone derivative, possesses anti-cancer and anti-inflammatory properties and has been reported to have cytotoxic effects on various cancer cells. The current study investigated the underlying molecular mechanism by which naringenin induces cell death in lung cancer. Methods: The expression of apoptosis, cell cycle arrest, and autophagy markers in H1299 and A459 lung cancer cells was evaluated using a terminal deoxynucleotidyl transferase dUTP nick end labeling assay (TUNEL), Western blot, Annexin V/PI stain, PI stain, acridine orange staining, and transmission electron microscopy (TEM). Using fluorescence microscopy, DALGreen was used to observe the degradation of p62, a GFP-LC3 plasmid was used to evaluate puncta formation, and a pcDNA3-GFP-LC3-RFP-LC3ΔG plasmid was used to evaluate autophagy flux. Furthermore, the anti-cancer effect of naringenin was evaluated in a subcutaneous H1299 cell xenograft model. Results: Naringenin treatment of lung cancer cells (H1299 and A459) reduced cell viability and induced cell cycle arrest. Pretreatment of cells with ROS scavengers (N-acetylcysteine or catalase) suppressed the naringenin-induced cleavage of apoptotic protein and restored cyclin-dependent kinase activity. Naringenin also triggered autophagy by mediating ROS generation, thereby activating AMP-activated protein kinase (AMPK) signaling. ROS inhibition not only inhibited naringenin-induced autophagic puncta formation but also decreased the ratio of microtubule-associated proteins 1A/1B light chain 3 II (LC3II)/LC3I and activity of the AMPK signaling pathway. Furthermore, naringenin suppressed tumor growth and promoted apoptosis in the xenograft mouse model. Conclusion: This study demonstrated the potent anti-cancer effects of naringenin on lung cancer cells, thereby providing valuable insights for developing small-molecule drugs that can induce cell cycle arrest, apoptosis, and autophagic cell death.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Flavanonas , Neoplasias Pulmonares , Humanos , Animais , Camundongos , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Apoptose , Neoplasias Pulmonares/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Linhagem Celular Tumoral , Pontos de Checagem da Fase G2 do Ciclo Celular , Autofagia , Flavanonas/farmacologiaRESUMO
Previously, we found that chronic methamphetamine treatment altered cannabinoid type 1 receptor (CB1R)-dependent cAMP/PKA/dopamine and cAMP-regulated phosphoprotein of Mr 32,000 (DARPP-32)/T34/PP2B signaling and decreased levels of CB1R protein and mRNA in the nucleus accumbens. These findings suggested the existence of signaling interplay between mesolimbic dopamine and CB1R. In this study, we further investigate interactions between CB1R and dopamine D2 receptor (D2R) signaling. Activation of either CB1R or D2R increased extracellular signal-regulated kinases 1 and 2 (ERK1/2) phosphorylation, while co-stimulation of CB1R and D2 R evoked an additive effect on the phospho-ERK1/2 signal. This effect was mediated through a pertussis toxin-sensitive Gαi/o pathway in primary striatal cells. Furthermore, the mRNA level of CB1R was increased via dopamine D2 receptor short form (D(2S)R) by treatment with D2R agonist quinpirole in D(2S)R/C6 glioma cells. This effect could be suppressed by co-treatment with the ERK1/2 inhibitor U0126. To test if D(2S)R could transcriptionally regulate CB1R, the 5'-untranslated region (5'-UTR) of the cannabinoid receptor 1 (CNR1) gene was sequenced from rat brain. Results showed that the CNR1 gene includes two exons, which contain 375 bp of 5'-UTR and are separated by a 17-kb intron. A luciferase reporter assay showed that the maximal D(2S)R-responsive promoter activity is located in the -1 to -222 region of CNR1 promoter. Overall, we demonstrate previously unidentified crosstalk between D2R and CB1R via ERK1/2 signaling that enhances the expression of CB1R by modulating its promoter activity.
Assuntos
Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Receptor Cross-Talk/efeitos dos fármacos , Receptor CB1 de Canabinoide/efeitos dos fármacos , Receptores de Dopamina D2/efeitos dos fármacos , Animais , Sequência de Bases , Western Blotting , Linhagem Celular Tumoral , Cicloexanóis/farmacologia , DNA/biossíntese , DNA/isolamento & purificação , Feminino , Genes Reporter/efeitos dos fármacos , Glioma/metabolismo , Dados de Sequência Molecular , Neostriado/citologia , Neostriado/metabolismo , Plasmídeos/genética , Regiões Promotoras Genéticas/efeitos dos fármacos , RNA/biossíntese , RNA/genética , RNA/isolamento & purificação , Ratos , Ratos Sprague-Dawley , Receptor CB1 de Canabinoide/agonistas , Receptor CB1 de Canabinoide/biossíntese , Receptor CB1 de Canabinoide/genética , TransfecçãoRESUMO
Background: Lung cancer is a malignant tumor with metastatic potential. Chemokine ligand 14 (CXCL14) has been reported to be associated with different cancer cell migration and invasion. However, few studies have explored the function of CXCL14 and its specific receptor in lung cancer metastasis. This study aims to determine the mechanism of CXCL14-promoted cancer metastasis. Methods: The expression of CXCL14, atypical chemokine receptor 2 (ACKR2), and epithelial mesenchymal transition (EMT) markers was evaluated by the public database of The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO), Western blot, enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qPCR), immunohistochemistry (IHC), and immunofluorescence (IF). Migration and wound healing assays were used to observe the motility of cancer cells. A luciferase reporter assay was performed to analyze transcription factor activity. The metastasis of lung cancer cells was evaluated in an orthotopic model. Results: We have presented that overexpression of CXCL14 and ACKR2 was observed in lung cancer datasets, human lung tumor sections, and lung cancer cells. Furthermore, the migration of CXCL14-promoted lung cancer cells was determined in vitro and in vivo. In particular, ACKR2 knockdown abolished CXCL14-induced cancer cell motility. Additionally, ACKR2 was involved in CXCL14-triggered phospholipase Cß3 (PLCß3), protein kinase Cα (PKCα), and proto-oncogene c-Src signaling pathway and subsequently upregulated nuclear factor κB (NF-κB) transcription activity leading to EMT and migration of lung cancer cells. These results indicated that the CXCL14/ACKR2 axis played an important role in lung cancer metastasis. Conclusion: This study is the first to reveal the function of CXCL14 in promoting EMT and metastasis in lung cancer. As a specific receptor for CXCL14 in lung cancer, ACKR2 mediates CXCL14-induced signaling that leads to cell motility. Our findings can be used as a prognostic biomarker of lung cancer metastasis.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Carcinoma Pulmonar de Células não Pequenas/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Linhagem Celular Tumoral , Transdução de Sinais/genética , Receptores de Quimiocinas , Quimiocinas CXC/genéticaRESUMO
Despite advances in therapeutic strategies, lung cancer remains the leading cause of cancer-related death worldwide. Acetylshikonin is a derivative of the traditional Chinese medicine Zicao and presents a variety of anticancer properties. However, the effects of acetylshikonin on lung cancer have not been fully understood yet. This study explored the mechanisms underlying acetylshikonin-induced cell death in non-small cell lung cancer (NSCLC). Treating NSCLC cells with acetylshikonin significantly reduced cell viability, as evidenced by chromatin condensation and the appearance of cell debris. Acetylshikonin has also been shown to increase cell membrane permeability and induce cell swelling, leading to an increase in the population of necrotic cells. When investigating the mechanisms underlying acetylshikonin-induced cell death, we discovered that acetylshikonin promoted oxidative stress, decreased mitochondrial membrane potential, and promoted G2/M phase arrest in lung cancer cells. The damage to NSCLC cells induced by acetylshikonin resembled results involving alterations in the cell membrane and mitochondrial morphology. Our analysis of oxidative stress revealed that acetylshikonin induced lipid oxidation and down-regulated the expression of glutathione peroxidase 4 (GPX4), which has been associated with necroptosis. We also determined that acetylshikonin induces the phosphorylation of receptor-interacting serine/threonine-protein kinase 1 (RIPK1)/RIPK3 and mixed lineage kinase domain-like kinase (MLKL). Treatment with RIPK1 inhibitors (necrostatin-1 or 7-Cl-O-Nec-1) significantly reversed acetylshikonin-induced MLKL phosphorylation and NSCLC cell death. These results indicate that acetylshikonin activated the RIPK1/RIPK3/MLKL cascade, leading to necroptosis in NSCLC cells. Our findings indicate that acetylshikonin reduces lung cancer cells by promoting G2/M phase arrest and necroptosis.