RESUMEN
In this study, we established a coculture model comprising human neuroblastoma SH-SY5Y cells and induced pluripotent stem cell-derived astrocytes, faithfully replicating the human brain environment for in vitro neurotoxicity assessment. We optimized the cell differentiation duration and cell ratios to obtain images conducive to neurite outgrowth evaluation. Subsequently, the neurotoxic effects in the coculture and monoculture of SH-SY5Y cells were confirmed using neurotoxic agents such as acrylamide (ACR) and hydrogen peroxide (H2O2). Disparities in the neurotoxic impacts of ACR and H2O2 within the coculture were mirrored in the expression of genes associated with early neuronal injury. Notably, the reduction in neurite outgrowth induced by neurotoxic agents revealed the coculture's lower sensitivity compared to monocultures. Furthermore, the coculture system exhibited distinct effects of test agents on nerve damage and manifested protective influences on nerve cells. The proposed methodology holds promise for large-scale chemical neurotoxicity screening through neurite change measurements. This in vitro coculture model, accounting for cell interactions, emerges as a valuable tool in toxicity testing, offering insights into the potential effects of chemicals within the human body.
Asunto(s)
Neuroblastoma , Síndromes de Neurotoxicidad , Humanos , Astrocitos , Técnicas de Cocultivo , Peróxido de Hidrógeno , Acrilamida/toxicidadRESUMEN
The genotoxicity, development toxicity, carcinogenicity, and acute or chronic toxic effects of glutaraldehyde (GA), particularly during occupational exposure through its use as a fixative, disinfectant, and preservative, are well-documented but its effects on neurotoxicity have not been investigated. We performed in vitro and in vivo studies to examine the developmental neurotoxicity (DNT) of GA. Neurite outgrowth was examined in an in vitro co-culture model consisting of SH-SY5Y human neuroblastoma cells and human astrocytes. Cell Counting Kit-8, lactate dehydrogenase assay, and high-content screening revealed that GA significantly inhibited neurite outgrowth at non-cytotoxic concentration. Further studies showed that GA upregulated the mRNA expression of the astrocyte markers GFAP and S100ß and downregulated the expression of the neurodevelopmental genes Nestin, ßIII-tubulin, GAP43, and MAP2. Furthermore, in vivo zebrafish embryo toxicity tests explored the effects of GA on neural morphogenesis. GA adversely affected the early development of zebrafish embryos, resulting in decreased survival, irregular hatching, and reduced heart rate in a time- and concentration-dependent manner. Furthermore, the width of the brain and spinal cord was reduced, and the myelination of Schwann cells and oligodendrocytes was decreased by GA in transgenic zebrafish lines. These data suggest that GAs have potential DNT in vitro and in vivo, highlighting the need for caution regarding the neurotoxicity of GA.
Asunto(s)
Neuroblastoma , Síndromes de Neurotoxicidad , Animales , Astrocitos , Células Cultivadas , Técnicas de Cocultivo , Glutaral/farmacología , Humanos , Neuronas , Pez CebraRESUMEN
High-quality antibody (Ab) production depends on the availability of immunologically relevant antigens. We present a potentially universal platform for generating soluble antigens from bacterial hosts, tailored to immunized animals for Ab production. A novel RNA-dependent chaperone, in which the target antigen is genetically fused with an RNA-interacting domain (RID) docking tag derived from the immunized host, promotes the solubility and robust folding of the target antigen. We selected the N-terminal tRNA-binding domain of lysyl-tRNA synthetase (LysRS) as the RID for fusion with viral proteins and demonstrated the expression of the RID fusion proteins in their soluble and native conformations; immunization predominantly elicited Ab responses to the target antigen, whereas the "self" RID tag remained nonimmunogenic. Differential immunogenicity of the fusion proteins greatly enriched and simplified the screening of hybridoma clones of monoclonal antibodies (mAbs), enabling specific and sensitive serodiagnosis of MERS-CoV infection. Moreover, mAbs against the consensus influenza hemagglutinin stalk domain enabled a novel assay for trivalent seasonal influenza vaccines. The Fc-mediated effector function was demonstrated, which could be harnessed for the design of next-generation "universal" influenza vaccines. The nonimmunogenic built-in antigen folding module tailored to a repertoire of immunized animal hosts will drive immunochemical diagnostics, therapeutics, and designer vaccines.
Asunto(s)
Anticuerpos Monoclonales/biosíntesis , Anticuerpos Antivirales/biosíntesis , Antígenos Virales/química , Infecciones por Coronavirus/diagnóstico , Hibridomas/inmunología , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Chaperonas Moleculares , Animales , Anticuerpos Monoclonales/inmunología , Anticuerpos Monoclonales/aislamiento & purificación , Anticuerpos Antivirales/inmunología , Anticuerpos Antivirales/aislamiento & purificación , Antígenos Virales/genética , Antígenos Virales/inmunología , Infecciones por Coronavirus/sangre , Ensayo de Inmunoadsorción Enzimática , Femenino , Glicoproteínas Hemaglutininas del Virus de la Influenza/química , Glicoproteínas Hemaglutininas del Virus de la Influenza/genética , Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Humanos , Inmunización , Vacunas contra la Influenza , Lisina-ARNt Ligasa/química , Lisina-ARNt Ligasa/genética , Ratones , Ratones Endogámicos BALB C , Conformación Proteica , Dominios Proteicos , Pliegue de Proteína , Proteínas Recombinantes de Fusión/química , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/inmunología , Pruebas Serológicas , SolubilidadRESUMEN
Esophageal cancer (EC) is one of the leading causes to cancer death in the worldwide and major population of EC is esophageal squamous cell carcinoma (ESCC). Still, ESCC-targeted therapy has not been covered yet. In the present study we have identified that Licochalcone B (Lico B) inhibited the ESCC growth by directly blocking the Janus kinase (JAK) 2 activity and its downstream signaling pathway. Lico B suppressed KYSE450 and KYSE510 ESCC cell growth, arrested cell cycle at G2/M phase and induced apoptosis. Direct target of Lico B was identified by kinase assay and verified with in vitro and ex vivo binding. Computational docking model predicted for Lico B interaction to ATP-binding pocket of JAK2. Furthermore, treatment of JAK2 clinical medicine AZD1480 to ESCC cells showed similar tendency with Lico B. Thus, JAK2 downstream signaling proteins phosphorylation of STAT3 at Y705 and S727 as well as STAT3 target protein Mcl-1 expression was decreased with treatment of Lico B. Our results suggest that Lico B inhibits ESCC cell growth, arrests cell cycle and induces apoptosis, revealing the underlying mechanism involved in JAK2/STAT3 signaling pathways after Lico B treatment. It might provide potential role of Lico B in the treatment of ESCC.
Asunto(s)
Chalconas/uso terapéutico , Carcinoma de Células Escamosas de Esófago/tratamiento farmacológico , Janus Quinasa 2/antagonistas & inhibidores , Apoptosis , Línea Celular Tumoral , Chalconas/farmacología , Carcinoma de Células Escamosas de Esófago/patología , HumanosRESUMEN
Patients with non-small-cell lung cancer (NSCLC) containing epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to tyrosine kinase inhibitor gefitinib; however, the treatment is less effective over time. Gefitinib resistance mechanisms include MET gene amplification. A therapeutic strategy targeting MET as well as EGFR can overcome resistance to gefitinib. In the present study we identified Echinatin (Ecn), a characteristic chalcone in licorice, which inhibited both EGFR and MET and strongly altered NSCLC cell growth. The antitumor efficacy of Ecn against gefitinib-sensitive or -resistant NSCLC cells with EGFR mutations and MET amplification was confirmed by suppressing cell proliferation and anchorage-independent colony growth. During the targeting of EGFR and MET, Ecn significantly blocked the kinase activity, which was validated with competitive ATP binding. Inhibition of EGFR and MET by Ecn decreases the phosphorylation of downstream target proteins ERBB3, AKT and ERK compared with total protein expression or control. Ecn induced the G2/M cell cycle arrest, and apoptosis via the intrinsic pathway of caspase-dependent activation. Ecn induced ROS production and GRP78, CHOP, DR5 and DR4 expression as well as depolarized the mitochondria membrane potential. Therefore, our results suggest that Ecn is a promising therapeutic agent in NSCLC therapy.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas/patología , Chalconas/farmacología , Gefitinib/farmacología , Neoplasias Pulmonares/patología , Proteínas Proto-Oncogénicas c-met/antagonistas & inhibidores , Apoptosis/efectos de los fármacos , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Resistencia a Antineoplásicos , Chaperón BiP del Retículo Endoplásmico , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/genética , Glycyrrhiza/química , Humanos , Neoplasias Pulmonares/tratamiento farmacológico , Simulación del Acoplamiento Molecular , Raíces de Plantas/química , Inhibidores de Proteínas Quinasas , Proteínas Proto-Oncogénicas c-met/genética , Quinazolinas/farmacologíaRESUMEN
Licochalcone (LC) families have been reported to have a wide range of biological function such as antioxidant, antibacterial, antiviral, and anticancer effects. Although various beneficial effects of LCD were revealed, its anticancer effect in human oral squamous cancer has not been identified. To examine the signaling pathway of LCD's anticancer effect, we determined whether LCD has physical interaction with Janus kinase (JAK2)/signal transducer and activator of transcription-3 (STAT3) signaling, which is critical in promoting cancer cell survival and proliferation. Our results demonstrated that LCD inhibited the kinase activity of JAK2, soft agar colony formation, and the proliferation of HN22 and HSC4 cells. LCD also induced mitochondrial apoptotic events such as altered mitochondrial membrane potential and reactive oxygen species production. LCD increased the expression of apoptosis-associated proteins in oral squamous cell carcinoma (OSCC) cells. Finally, the xenograft study showed that LCD significantly inhibited HN22 tumor growth. Immunohistochemical data supported that LCD suppressed p-JAK2 and p-STAT3 expression and induced cleaved-caspase-3 expression. These results indicate that the anticancer effect of LCD is due to the direct targeting of JAK2 kinase. Therefore, LCD can be used for therapeutic application against OSCC.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Chalconas/farmacología , Janus Quinasa 2/antagonistas & inhibidores , Inhibidores de las Cinasas Janus/farmacología , Neoplasias de la Boca/tratamiento farmacológico , Carcinoma de Células Escamosas de Cabeza y Cuello/tratamiento farmacológico , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Humanos , Janus Quinasa 2/metabolismo , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/patología , Terapia Molecular Dirigida , Neoplasias de la Boca/enzimología , Neoplasias de la Boca/patología , Transducción de Señal , Carcinoma de Células Escamosas de Cabeza y Cuello/enzimología , Carcinoma de Células Escamosas de Cabeza y Cuello/patología , Carga Tumoral/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
To explore the reactivity of copper-alkylperoxo species enabled by the heterolytic peroxide activation, room-temperature stable mononuclear nonheme copper(II)-alkylperoxo complexes bearing a N-(2-ethoxyethanol)-bis(2-picolyl)amine ligand (HN3O2), [CuII(OOR)(HN3O2)]+ (R = cumyl or tBu), were synthesized and spectroscopically characterized. A combined experimental and computational investigation on the reactivity and reaction mechanisms in the phosphorus oxidation, C-H bond activation, and aldehyde deformylation reactions by the copper(II)-alkylperoxo complexes has been conducted. DFT-optimized structures suggested that a hydrogen bonding interaction exists between the ethoxyethanol backbone of the HN3O2 ligand and either the proximal or distal oxygen atom of the alkylperoxide moiety, and this interaction consequently results in the enhanced stability of the copper(II)-alkylperoxo species. In the phosphorus oxidation reaction, both experimental and computational results indicated that a phosphine-triggered heterolytic O-O bond cleavage occurred to yield phosphine oxide and alcohol products. DFT calculations suggested that (i) the H-bonding between the ethoxyethanol backbone and distal oxygen of the alkylperoxide moiety and (ii) the phosphine binding to the proximal oxygen of the alkylperoxide moiety engendered the heterolytic peroxide activation. In the C-H bond activation reactions, temperature-dependent reactivity of the copper(II)-alkylperoxo complexes was observed, and a relatively strong activation energy of 95 kcal mol-1 was required to promote the homolytic peroxide activation. A rate-limiting hydrogen atom abstraction reaction of xanthene by the putative copper(II)-oxyl radical resulted in the formation of the dimeric copper product and the substrate radical that further underwent autocatalytic oxidation reactions to form an oxygen incorporated product. Finally, amphoteric reactivity of copper(II)-alkylperoxo complexes has been assessed by conducting kinetic studies and product analysis of the aldehyde deformylation reaction.
RESUMEN
Solubility is the prime criterion for determining the quality of recombinant proteins, yet it often fails to represent functional activity due to the involvement of non-functional, misfolded, soluble aggregates, which compromise the quality of recombinant proteins. However, guidelines for the quality assessment of soluble proteins have neither been proposed nor rigorously validated experimentally. Using the aggregation-prone enhanced green-fluorescent protein (EGFP) folding reporter system, we evaluated the folding status of recombinant proteins by employing the commonly used sonication and mild lysis of recombinant host cells. We showed that the differential screening of solubility and folding competence is crucial for improving the quality of recombinant proteins without sacrificing their yield. These results highlight the importance of screening out incorrectly folded soluble aggregates at the initial purification step to ensure the functional quality of recombinant proteins.
Asunto(s)
Agregado de Proteínas , Pliegue de Proteína , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Cromatografía en Gel , Tamaño de la Partícula , Proteínas Recombinantes de Fusión , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Solubilidad , SonicaciónRESUMEN
Esophageal squamous cell carcinoma (ESCC) is a poor prognostic cancer with a low five-year survival rate. Echinatin (Ech) is a retrochalone from licorice. It has been used as a herbal medicine due to its anti-inflammatory and anti-oxidative effects. However, its anticancer activity or underlying mechanism has not been elucidated yet. Thus, the objective of this study was to investigate the anti-tumor activity of Ech on ESCC by inducing ROS and ER stress dependent apoptosis. Ech inhibited ESCC cell growth in anchorage-dependent and independent analysis. Treatment with Ech induced G2/M phase of cell cycle and apoptosis of ESCC cells. It also regulated their related protein markers including p21, p27, cyclin B1, and cdc2. Ech also led to phosphorylation of JNK and p38. Regarding ROS and ER stress formation associated with apoptosis, we found that Ech increased ROS production, whereas its increase was diminished by NAC treatment. In addition, ER stress proteins were induced by treatment with Ech. Moreover, Ech enhanced MMP dysfunction and caspases activity. Furthermore, it regulated related biomarkers. Taken together, our results suggest that Ech can induce apoptosis in human ESCC cells via ROS/ER stress generation and p38 MAPK/JNK activation.
Asunto(s)
Apoptosis/genética , Chalconas/farmacología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Neoplasias Esofágicas/tratamiento farmacológico , Carcinoma de Células Escamosas de Esófago/tratamiento farmacológico , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacos , Biomarcadores de Tumor/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Neoplasias Esofágicas/metabolismo , Carcinoma de Células Escamosas de Esófago/metabolismo , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de los fármacos , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Fosforilación/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
In the present study, various extracts of C. tricuspidata fruit were prepared with varying ethanol contents and evaluated for their biomarker and biological properties. The 80% ethanolic extract showed the best tyrosinase inhibitory activity, while the 100% ethanolic extract showed the best total phenolics and flavonoids contents. The HPLC method was applied to analyze the chlorogenic acid in C. tricuspidata fruit extracts. The results suggest that the observed antioxidant and tyrosinase inhibitory activity of C. tricuspidata fruit extract could partially be attributed to the presence of marker compounds in the extract. In this study, we present an analytical method for standardization and optimization of C. tricuspidata fruit preparations. Further investigations are warranted to confirm the in vivo pharmacological activity of C. tricuspidata fruit extract and its active constituents and assess the safe use of the plant for the potential development of the extract as a skin depigmentation agent.
Asunto(s)
Antioxidantes/farmacología , Ácido Clorogénico/farmacología , Inhibidores Enzimáticos/farmacología , Monofenol Monooxigenasa/antagonistas & inhibidores , Moraceae/química , Antioxidantes/química , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Ácido Clorogénico/química , Cromatografía Líquida de Alta Presión , Inhibidores Enzimáticos/química , Flavonoides/aislamiento & purificación , Frutas/química , Humanos , Fenoles/aislamiento & purificación , Extractos Vegetales/aislamiento & purificaciónRESUMEN
Oral cancer is of an aggressive malignancy that arises on oral cavity and lip, 90% of cancers histologically originated in the squamous cells. Licochalcone (LC)C has been known as natural phenolic chalconoid substances, and its origin is the root of Glycyrrhiza glabra or Glycyrrhiza inflata. LCC inhibited oral squamous cell carcinoma (OSCC) cell viability, mitochondrial function, and anchorage-independent growth in a dose-dependent manner. To investigate the ability of LCC to target Janus kinase 2 (JAK2), we performed pull-down binding assay, kinase assay, and docking simulation. The molecular docking studies were performed between JAK2 and the potent inhibitor LCC. It was shown that LCC tightly interacted with ATP-binding site of JAK2. In addition, LCC inhibited the JAK2/signal transducer and activator of transcription 3 pathway, upregulated p21, and downregulated Bcl-2, Mcl-1, and Survivin, while it disrupted mitochondrial membrane potential and subsequently caused cytochrome c release with activation of multi-caspase, eventually leading to apoptosis in HN22 and HSC4 cells. LCC elevated the protein levels of Bax, cleaved Bid and PARP, and increased Apaf-1, and this effect was reversed by LCC treatment. Our results demonstrated that treatment of OSCC cells with LCC induced the death receptor (DR)4 and DR5 expression level with the generation of reactive oxygen species and the upregulation of CHOP protein expression. Taken together, these results could provide the basis for clinical application as a new therapeutic strategy in the treatment of oral cancer.
Asunto(s)
Carcinoma de Células Escamosas/tratamiento farmacológico , Chalconas/farmacología , Janus Quinasa 2/genética , Neoplasias de la Boca/tratamiento farmacológico , Factor de Transcripción STAT3/genética , Apoptosis/efectos de los fármacos , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/efectos de los fármacos , Simulación del Acoplamiento Molecular , Neoplasias de la Boca/genética , Neoplasias de la Boca/patología , Especies Reactivas de Oxígeno/metabolismo , Transducción de Señal/efectos de los fármacosRESUMEN
Licochalcone A (LCA), isolated from the root of Glycyrrhiza inflata, are known to have medicinal effect such as anti-oxidant, anti-bacterial, anti-viral, and anti-cancer. Though, as a pharmacological mechanism regulator, anti-cancer studies on LCA were not investigated in human breast cancer. We investigated the anti-proliferative and apoptotic effect of LCA in human breast cancer cells MCF-7 and MDA-MB-231 through MTS assay, PI staining, Annexin-V/7-AAD assay, mitochondrial membrane potential assay, multi-caspase assay, RT-PCR, Western blot analysis, and anchorage-independent cell transformation assay. Our results showed the little difference between two cells, as MCF-7 cell is both estrogen/progesterone receptor positive, there were only effect on Sp1 protein level, but not in mRNA level. Adversely, estrogen/progesterone/human epidermal growth factor receptor 2 triple negative, MDA-MB-231 showed decreased Sp1 mRNA, and protein levels. To confirm the participation of Sp1 in breast cancer cell viability, siRNA techniques were introduced. Both cells showed dysfunction of mitochondrial membrane potential and mitochondrial ROS production, which reflects it passed intracellular mitochondrial apoptosis pathway. Additionally, LCA showed the anti-proliferative and apoptotic effect in breast cancer cells through regulating Sp1 and apoptosis-related proteins in a dose- and a time-dependent manner. Consequently, LCA might be a potential anti-breast cancer drug substitute. J. Cell. Biochem. 118: 4652-4663, 2017. © 2017 Wiley Periodicals, Inc.
Asunto(s)
Neoplasias de la Mama/tratamiento farmacológico , Chalconas/farmacología , Proteínas de Neoplasias/metabolismo , Factor de Transcripción Sp1/metabolismo , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Supervivencia Celular/efectos de los fármacos , Femenino , Humanos , Células MCF-7 , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/patologíaRESUMEN
Licochalcone B (Lico B), which is normally isolated from the roots of Glycyrrhiza inflata (Chinese Licorice), generally classified into organic compounds including retrochalcones. Potential pharmacological properties of Lico B include anti-inflammatory, anti-bacterial, anti-oxidant, and anti-cancer activities. However, its biological effects on melanoma and squamous cell carcinoma (SCC) are unknown. Based on these known facts, this study investigated the role of Lico B in apoptosis, through the extrinsic and intrinsic pathways and additional regulation of specificity protein 1 in human skin cancer cell lines. Annexin V/7-aminoactinomycin D staining, western blot analysis, mitochondrial membrane potential assay, and an anchorage-independent cell transformation assay demonstrated that Lico B treatment of human melanoma and SCC cells significantly inhibited cell proliferation and induced apoptotic cell death. More specifically, Lico B induced apoptosis through the regulation of specificity protein 1 and apoptosis-related proteins including CCAAT/enhancer-binding protein homologous protein, death receptors, and poly (ADP-ribose) polymerase. These results indicate that Lico B has apoptotic effect on A375 and A431 skin cancer cells, suggesting the potential value of Lico B for the treatment of human melanoma and SCC. Copyright © 2017 John Wiley & Sons, Ltd.
Asunto(s)
Apoptosis/efectos de los fármacos , Productos Biológicos/química , Carcinoma de Células Escamosas/tratamiento farmacológico , Chalconas/química , Melanoma/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Carcinoma de Células Escamosas/patología , Humanos , Melanoma/patología , Neoplasias Cutáneas/patología , Melanoma Cutáneo MalignoRESUMEN
Experimental autoimmune uveoretinitis (EAU) is an autoimmune disease that models human uveitis. Caffeic acid phenethyl ester (CAPE), a phenolic compound isolated from propolis, possesses anti-inflammatory and immunomodulatory properties. CAPE demonstrates therapeutic potential in several animal disease models through its ability to inhibit NF-κB activity. To evaluate these therapeutic effects in EAU, we administered CAPE in a model of EAU that develops after immunization with interphotoreceptor retinal-binding protein (IRBP) in B10.RIII and C57BL/6 mice. Importantly, we found that CAPE lessened the severity of EAU symptoms in both mouse strains. Notably, treated mice exhibited a decrease in the ocular infiltration of immune cell populations into the retina; reduced TNF-α, IL-6, and IFN-γ serum levels: and inhibited TNF-α mRNA expression in retinal tissues. Although CAPE failed to inhibit IRBP-specific T cell proliferation, it was sufficient to suppress cytokine, chemokine, and IRBP-specific antibody production. In addition, retinal tissues isolated from CAPE-treated EAU mice revealed a decrease in NF-κB p65 and phospho-IκBα. The data identify CAPE as a potential therapeutic agent for autoimmune uveitis that acts by inhibiting cellular infiltration into the retina, reducing the levels of pro-inflammatory cytokines, chemokine, and IRBP-specific antibody and blocking NF-κB pathway activation.
Asunto(s)
Enfermedades Autoinmunes/tratamiento farmacológico , Ácidos Cafeicos/uso terapéutico , Modelos Animales de Enfermedad , FN-kappa B/antagonistas & inhibidores , Alcohol Feniletílico/análogos & derivados , Retinitis/tratamiento farmacológico , Uveítis/tratamiento farmacológico , Animales , Enfermedades Autoinmunes/metabolismo , Enfermedades Autoinmunes/patología , Western Blotting , Proteínas del Ojo/inmunología , Citometría de Flujo , Inmunoglobulina G/sangre , Interferón gamma/sangre , Interleucina-6/metabolismo , Activación de Linfocitos/inmunología , Ratones , Ratones Endogámicos C57BL , Alcohol Feniletílico/uso terapéutico , ARN Mensajero/metabolismo , Retinitis/metabolismo , Retinitis/patología , Proteínas de Unión al Retinol/inmunología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Linfocitos T/inmunología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Uveítis/metabolismo , Uveítis/patologíaRESUMEN
Dendropanax morbifera Leveille (Araliaceae) is well known in Korean traditional medicine for a variety of diseases. Rotenone is a commonly used neurotoxin to produce in vivo and in vitro Parkinson's disease models. This study was designed to elucidate the processes underlying neuroprotection of rutin, a bioflavonoid isolated from D. morbifera Leveille in cellular models of rotenone-induced toxicity. We found that rutin significantly decreased rotenone-induced generation of reactive oxygen species levels in SH-SY5Y cells. Rutin protected the increased level of intracellular Ca(2+) and depleted level of mitochondrial membrane potential (ΔΨm) induced by rotenone. Furthermore, it prevented the decreased ratio of Bax/Bcl-2 caused by rotenone treatment. Additionally, rutin protected SH-SY5Y cells from rotenone-induced caspase-9 and caspase-3 activation and apoptotic cell death. We also observed that rutin repressed rotenone-induced c-Jun N-terminal kinase and p38 mitogen-activated protein kinase phosphorylation. These results suggest that rutin may have therapeutic potential for the treatment of neurodegenerative diseases associated with oxidative stress.
Asunto(s)
Neuronas Dopaminérgicas/enzimología , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Rotenona/toxicidad , Rutina/farmacología , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidores , Araliaceae , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Neuronas Dopaminérgicas/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Fármacos Neuroprotectores/aislamiento & purificación , Fármacos Neuroprotectores/farmacología , Extractos Vegetales/aislamiento & purificación , Extractos Vegetales/farmacología , Inhibidores de Proteínas Quinasas/aislamiento & purificación , Rotenona/antagonistas & inhibidores , Rutina/aislamiento & purificación , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
In plants, timing of flowering is an essential factor that controls the survival rates of descendants. The circadian clock genes E1 and GIGANTEA (GI) play a central role in transmitting signals to flowering locus T (FT) in leguminous plants. Lotus japonicus is a wild Japanese species that ranges from northern Hokkaido to the southern Ryukyus and exhibits a wide range in terms of the time between seeding and first flowering. In this study, we first identified LjGI and analyzed polymorphisms of LjE1 and LjGI among wild populations covering the entire distribution range of this species in Japan. LjGI had a coding sequence (CDS) length of 3495 bp and included 14 exons. The homologies of DNA and amino acid sequences between LjGI and GmGI were 89 and 88% (positive rate was 92%), respectively. LjE1 harbored five nucleic acid changes in a 552 bp CDS, all of which were nonsynonymous; four of the changes were located in the core function area. LjE1 alleles exhibited partial north-south differentiation and non-neutrality. In contrast, the LjGI harbored one synonymous and one nonsynonymous change. Thus, our study suggests that LjE1 may be involved in the control of flowering times, whereas LjGI may be under strong purifying selection.
Asunto(s)
Flores/genética , Lotus/genética , Proteínas de Plantas/genética , Polimorfismo Genético , Selección Genética , Secuencia de Aminoácidos , Evolución Molecular , Flores/crecimiento & desarrollo , Japón , Lotus/metabolismo , Datos de Secuencia Molecular , Proteínas de Plantas/metabolismoRESUMEN
Poly(hexamethylenebicyanoguanide-hexamethylenediamine) hydrochloride (PHMB) is a biocide with a broad spectrum of antibacterial activity. Its use as a disinfectant and preservative in consumer products results in human exposure to PHMB. Toxicity studies on PHMB mainly focus on systemic toxicity or skin irritation; however, its effects on developmental neurotoxicity (DNT) and the underlying mechanisms are poorly understood. In this study, the DNT effects of PHMB were evaluated using IMR-32 and SH-SY5Y cell lines and zebrafish. In both cell lines, PHMB concentrations ≥ 10 µM reduced neurite outgrowth, and cytotoxicity was observed at concentrations up to 40 µM. PHMB regulated expression of neurodevelopmental genes and induced reactive oxygen species (ROS) production and mitochondrial dysfunction. Treatment with N-acetylcysteine reversed the toxic effects of PHMB. Toxicity tests on zebrafish embryos showed that PHMB reduced viability and heart rate and caused irregular hatching. PHMB concentrations of 1-4 µM reduced the width of the brain and spinal cord of transgenic zebrafish and attenuated myelination processes. Furthermore, PHMB modulated expression of neurodevelopmental genes in zebrafish and induced ROS accumulation. These results suggested that PHMB exerted DNT effects in vitro and in vivo through a ROS-dependent mechanism, highlighting the risk of PHMB exposure.
Asunto(s)
Diaminas , Desinfectantes , Neuroblastoma , Animales , Humanos , Especies Reactivas de Oxígeno/metabolismo , Pez Cebra/metabolismo , Neuroblastoma/metabolismo , Estrés Oxidativo , Desinfectantes/toxicidad , Embrión no MamíferoRESUMEN
α-mangostin (α-MG) demonstrates antibacterial activity against Staphylococcus species. Therefore, this study aimed to explore the antibacterial activity of α-MG-rich mangosteen pericarp extract (MPE)-loaded liposomes against Staphylococcus isolates from companion animal skin diseases in vitro and evaluated their therapeutic potential in a murine model of superficial skin infection caused by S. pseudintermedius. α-MG-rich extract was purified from mangosteen pericarp and then complexed with γ-cyclodextrin (γ-CD), forming the inclusion complexes. Nanoliposomes containing MPE and γ-CD complexes were prepared by adding lecithin and casein. Minimum inhibitory concentrations (MICs) and minimum bactericidal concentrations (MBCs) of MPE-loaded liposomes were determined using agar dilution and broth microdilution methods. The therapeutic potential of MPE-loaded liposomes was evaluated in vivo on tape-stripped skin lesions infected with S. pseudintermedius. Purified MPE and MPE-loaded liposomes contained 402.43 mg/g and 18.18 mg/g α-MG, respectively. MPE-loaded liposomes showed antibacterial activity against clinical Staphylococcus isolates in vitro but did not show antibacterial activity against Gram-negative bacterial isolates. MPE-loaded liposomes demonstrated consistent MICs and MBCs against Staphylococcus isolates. These liposomes significantly reduced bacterial numbers and lesional sizes in a superficial skin infection model. Moreover, they reconstructed the epidermal barrier in skin lesions. The therapeutic concentrations of MPE-loaded liposomes did not induce cytotoxicity in canine progenitor epidermal keratinocyte cells. In conclusion, MPE-loaded liposomes hold promise for the development of a prospective topical formulation to treat superficial pyoderma in companion animals.
RESUMEN
Previous studies on copper pyrithione (CPT) and zinc pyrithione (ZPT) as antifouling agents have mainly focused on marine organisms. Even though CPT and ZPT pose a risk of human exposure, their neurotoxic effects remain to be elucidated. Therefore, in this study, the cytotoxicity and neurotoxicity of CPT and ZPT were evaluated after the exposure of human SH-SY5Y/astrocytic co-cultured cells to them. The results showed that, in a co-culture model, CPT and ZPT induced cytotoxicity in a dose-dependent manner (~ 400 nM). Exposure to CPT and ZPT suppressed all parameters in the neurite outgrowth assays, including neurite length. In particular, exposure led to neurotoxicity at concentrations with low or no cytotoxicity (~ 200 nM). It also downregulated the expression of genes involved in neurodevelopment and maturation and upregulated astrocyte markers. Moreover, CPT and ZPT induced mitochondrial dysfunction and promoted the generation of reactive oxygen species. Notably, N-acetylcysteine treatment showed neuroprotective effects against CPT- and ZPT-mediated toxicity. We concluded that oxidative stress was the major mechanism underlying CPT- and ZPT-induced toxicity in the co-cultured cells.
Asunto(s)
Neuroblastoma , Compuestos Organometálicos , Humanos , Astrocitos/metabolismo , Técnicas de Cocultivo , Estrés Oxidativo , Compuestos Organometálicos/toxicidad , Compuestos Organometálicos/metabolismo , Células CultivadasRESUMEN
Biocidal disinfectants (BDs) that kill microorganisms or pathogens are widely used in hospitals and other healthcare fields. Recently, the use of BDs has rapidly increased as personal hygiene has become more apparent owing to the pandemic, namely the coronavirus outbreak. Despite frequent exposure to BDs, toxicity data of their potential neurotoxicity (NT) are lacking. In this study, a human-derived SH-SY5Y/astrocyte was used as a co-culture model to evaluate the chemical effects of BDs. Automated high-content screening was used to evaluate the potential NT of BDs through neurite growth analysis. A set of 12 BD substances classified from previous reports were tested. Our study confirms the potential NT of benzalkonium chloride (BKC) and provides the first evidence of the potential NT of poly(hexamethylenebicyanoguanide-hexamethylenediamine) hydrochloride (PHMB). BKC and PHMB showed significant NT at concentrations without cytotoxicity. This test system for analyzing the potential NT of BDs may be useful in early screening studies for NT prior to starting in vivo studies.