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(-)-Guaiol is a sesquiterpenoid found in many traditional Chinese medicines with potent antitumor activity. However, its therapeutic effect and mechanism in non-small cell lung cancer (NSCLC) have not been fully elucidated. In this study, (-)-Guaiol was found to induce immunogenic cell death (ICD) in NSCLC in vitro. Using (-)-Guaiol in vivo, we found that (-)-Guaiol could suppress tumor growth, increase dendritic cell activation, and enhance T-cell infiltration. Vaccination experiments suggest that cellular immunoprophylaxis after (-)-Guaiol intervention can suppress tumor growth. Previous studies have found that (-)-Guaiol induces apoptosis and autophagy in NSCLC. Apoptosis and autophagy are closely related to ICD. To explore whether autophagy and apoptosis are involved in (-)-Guaiol-induced ICD, we used inhibitors of apoptosis and autophagy. The results showed that the release of damage-associated molecular patterns (DAMPs) was partly reversed after inhibition of apoptosis and autophagy. In conclusion, these results suggested that the (-)-Guaiol triggers immunogenic cell death and inhibits tumor growth in NSCLC.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Humanos , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Neoplasias Pulmonares/patología , Muerte Celular Inmunogénica , Línea Celular Tumoral , Apoptosis , AutofagiaRESUMEN
Our study investigated the differences in pharmacokinetics of three major components of crude Cimicifuga foetida L. and its fried product and honey- and liquor-prepared products. A rapid and sensitive ultra-high performance liquid chromatography with tandem mass spectrometry approach was established for determing caffeic acid, isoferulic acid and ferulic acid in rat plasma. The approach has good linearity, precision, accuracy, recovery and stability. Phenolic acid was rapidly absorbed. The times to peak concentration were shorter in the processed group than those for the crude product, with their values of <30 min. The peak concentration values of caffeic acid and isoferulic acid were higher in the crude group than in the processed groups (p < 0.05). Area under the curve values of the three phenolics in the crude group were significantly higher than those of the processed groups (p < 0.05).
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Cimicifuga/química , Cinamatos/sangre , Cinamatos/farmacocinética , Medicamentos Herbarios Chinos , Animales , Cromatografía Líquida de Alta Presión/métodos , Cinamatos/química , Medicamentos Herbarios Chinos/administración & dosificación , Medicamentos Herbarios Chinos/farmacocinética , Modelos Lineales , Masculino , Ratas , Reproducibilidad de los Resultados , Sensibilidad y Especificidad , Espectrometría de Masas en Tándem/métodosRESUMEN
BACKGROUND: Hepatic Golgi protein-73 (GP73) expression is related to hepatocellular carcinoma (HCC) progression. The aim of this study was to investigate the dynamic expression of GP73 mRNA and protein during hepatocytes malignant transformation. METHODS: Human GP73 expressions in 88 HCC tissues and their self-control surrounding tissues were examined by immunohistochemistry, and survival time of HCC patients was evaluated by the Kaplan-Meier method. HCC model of Sprague-Dawley rats was made by diet containing 2-fluorenylacetamide. The rats were divided into the control, hepatocyte degeneration, precanceration, and HCC groups to observe GP73 protein and mRNA alterations during hepatocytes malignant transformation. RESULTS: The GP73 expression was significantly higher in the cancerous tissues than that in the surrounding tissues, with shorter survival time, and the positive rates of GP73 protein in human HCC tissues were 53.3% at stage I, 84.0% at stage II, 84.6% at stage III, and 60.0% at stage IV, respectively. The positive rates of hepatic GP73 protein and mRNA in the rat models were none in the control group, 66.7% and 44.4% in the hepatocytes degeneration group, 88.9% and 77.8% in the hepatocytes precanceration group, and 100% in the HCC group, respectively. There was a positive correlation (r = 0.91, P<0.01) between hepatic GP73 and serum GP73 during rat hepatocytes malignant transformation. CONCLUSIONS: Abnormal GP73 expression may be a sensitive and valuable biomarker in hepatocarcinogensis.
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Biomarcadores de Tumor/metabolismo , Carcinoma Hepatocelular/metabolismo , Transformación Celular Neoplásica/metabolismo , Hepatocitos/metabolismo , Neoplasias Hepáticas/metabolismo , Proteínas de la Membrana/metabolismo , Adulto , Anciano , Animales , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/mortalidad , Carcinoma Hepatocelular/patología , Transformación Celular Neoplásica/genética , Transformación Celular Neoplásica/patología , Femenino , Regulación Neoplásica de la Expresión Génica , Hepatocitos/patología , Humanos , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/mortalidad , Neoplasias Hepáticas/patología , Masculino , Proteínas de la Membrana/genética , Persona de Mediana Edad , Estadificación de Neoplasias , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
BACKGROUND: Non-small cell lung cancer (NSCLC) is one of the malignant tumors with the highest incidence and mortality. This meta-analysis aimed to explore the efficacy of the supplementing Qi and nourishing Yin method combined with chemotherapy for the treatment of NSCLC based on qualified randomized controlled trials. METHOD: PubMed, Cochrane, and Embase databases were searched by the index words to identify the eligible studies, and relevant literature sources were also searched. The latest research was performed in December 2017. Of the eligible studies, only those involving randomized controlled trials were included. Relative risks (RR) along with 95% confidence interval (95% CI) were used to analyze the main outcomes. RESULT: A total of 41 studies were involved in the meta-analysis with 1335 objects in the treatment group and 1272 objects in the control group. Compared with chemotherapy, supplementing Qi and nourishing Yin combined with chemotherapy significantly increased the effective rate (RR: 1.37, 95% CI: 1.24-1.51), increased the Karnofsky score (RR: 1.50, 95% CI: 1.39-1.61), and improved the TCM symptom (RR: 1.69, 95% CI: 1.55-1.85). CONCLUSION: These results demonstrated that supplementing Qi and nourishing Yin combined with chemotherapy would have better clinical efficacy in effective rate, the Karnofsky score, and TCM symptom. Most of the included studies had a low Jadad score; however, there is still a need for high-quality, larger-sample, multicentric, and long-term follow-up randomized controlled trials to confirm our conclusion.
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Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Quimioterapia/métodos , Medicamentos Herbarios Chinos/uso terapéutico , Neoplasias Pulmonares/tratamiento farmacológico , Terapia Combinada , Humanos , Estado de Ejecución de Karnofsky , Qi , Análisis de Supervivencia , Resultado del Tratamiento , Yin-YangRESUMEN
OBJECTIVES: To determine the feasibility of delivering inhaled treprostinil during mechanical ventilation and spontaneous unassisted ventilation using the Tyvaso Inhalation System and the vibrating mesh nebulizer. We sought to compare differences in fine particle fraction, and absolute inhaled treprostinil mass delivered to neonatal, pediatric, and adult models affixed with a face mask, conventional, and high-frequency ventilation between Tyvaso Inhalation System and with different nebulizer locations between Tyvaso Inhalation System and vibrating mesh nebulizer. DESIGN: Fine particle fraction was first determined via impaction with both the Tyvaso Inhalation System and vibrating mesh nebulizer. Next, a test lung configured with neonatal, pediatric, and adult mechanics and a filter to capture medication was attached to a realistic face model during spontaneous breathing or an endotracheal tube during conventional ventilation and high-frequency oscillator ventilator. Inhaled treprostinil was then nebulized with both the Tyvaso Inhalation System and vibrating mesh nebulizer, and the filter was analyzed via high-performance liquid chromatography. Testing was done in triplicate. Independent two-sample t tests were used to compare mean fine particle fraction and inhaled mass between devices. Analysis of variance with Tukey post hoc tests were used to compare within device differences. SETTING: Academic children's hospital aerosol research laboratory. MEASUREMENTS AND MAIN RESULTS: Fine particle fraction was not different between the Tyvaso Inhalation System and vibrating mesh nebulizer (0.78 ± 0.04 vs 0.77 ± 0.08, respectively; p = 0.79). The vibrating mesh nebulizer delivered the same or greater inhaled treprostinil than the Tyvaso Inhalation System in every simulated model and condition. When using the vibrating mesh nebulizer, delivery was highest when using high-frequency oscillator ventilator in the neonatal and pediatric models, and with the nebulizer in the distal position in the adult model. CONCLUSIONS: The vibrating mesh nebulizer is a suitable alternative to the Tyvaso Inhalation System for inhaled treprostinil delivery. Fine particle fraction is similar between devices, and vibrating mesh nebulizer delivery meets or exceeds delivery of the Tyvaso Inhalation System. Delivery for infants and children during high-frequency oscillator ventilator with the vibrating mesh nebulizer may result in higher than expected dosages.
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Antihipertensivos/administración & dosificación , Sistemas de Liberación de Medicamentos/instrumentación , Epoprostenol/análogos & derivados , Hipertensión Pulmonar/terapia , Nebulizadores y Vaporizadores , Respiración Artificial , Administración por Inhalación , Adulto , Aerosoles , Antihipertensivos/uso terapéutico , Niño , Preescolar , Terapia Combinada , Epoprostenol/administración & dosificación , Epoprostenol/uso terapéutico , Estudios de Factibilidad , Humanos , Recién Nacido , Modelos Anatómicos , Tamaño de la Partícula , VibraciónRESUMEN
Retronecine-, otonecine-, and heliotridine-type pyrrolizidine alkaloids (PAs) are all reported to be hepatotoxic. These PAs are suggested to be metabolized to the corresponding electrophilic dehydropyrrolizidine alkaloids (dehydro-PAs) and subsequently conjugated with macromolecules, such as glutathione (GSH). In the present study, a total of five glutathione conjugates, named M1-M5, were detected in rat and human liver microsomal incubations with three retrornecine-type PAs (isoline, retrorsine, or monocrotaline) in the presence of glutathione, and were chemically synthesized. M1 and M3 were unambiguously identified as a pair of epimers of 7-glutathionyl-6,7-dihydro-1-hydroxymethyl-5H-pyrrolizine (7-GSH-DHP), and M4 and M5 were epimers of 7,9-diglutathionyl-6,7-dihydro-1-hydroxymethyl-5H-pyrrolizine (7,9-diGSH-DHP). M2, an extremely unstable conjugate, was proposed to be 9-glutathionyl-6,7-dihydro-1-hydroxymethyl-5H-pyrrolizine (9-GSH-DHP). It was the most abundant among the five GSH conjugates, and the finding corrects the mistake that 7-GSH-DHP is the predominant GSH conjugate derived from dehydro-PAs. Similar patterns in glutathione conjugate profile were observed in the bile of rats treated with the PAs. This is the first study to describe 9-GSH-DHP as a major pyrrolic GSH conjugate of retronecine-type PAs, providing insight into the interactions of dehydro-PAs with biomolecules.
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Glutatión/análogos & derivados , Glutatión/química , Microsomas Hepáticos/metabolismo , Pirroles/análisis , Alcaloides de Pirrolicidina/química , Animales , Bilis/química , Bilis/metabolismo , Cromatografía Líquida de Alta Presión , Glutatión/análisis , Glutatión/metabolismo , Semivida , Humanos , Masculino , Pirroles/química , Alcaloides de Pirrolicidina/metabolismo , Ratas , Ratas Sprague-Dawley , Espectrometría de Masas en TándemRESUMEN
Tetrandrine is a diaryl ether-type bisbenzylisoquinoline alkaloid and has shown multiple pharmacological activities. Our early work demonstrated that tetrandrine produced acute pulmonary toxicity and that tetrandrine was biotransformed to a quinone methide-derived metabolite mediated by CYP3A enzymes. The formation of the reactive intermediate is suggested to be responsible for the pulmonary toxicity induced by tetrandrine. In the present study, a WI-38-based Cyp3a5 transgenic cell line (WI-38/Cyp3a5) was established to investigate the role of CYP3A5 in tetrandrine-induced cytotoxicity. The transgenic cells were found to be more susceptible to the cytotoxicity of tetrandrine than the wild-type cells (WI-38/Vector). WI-38/Cyp3a5 cells showed higher cellular ROS levels, higher LDH activities in culture media, but lower cellular GSH contents than those observed in WI-38/Vector cells after exposure to tetrandrine. And severer apoptosis were observed in WI-38/Cyp3a5 cells after treatment with tetrandrine: WI-38/Cyp3a5 cells had higher proportion of early and late apoptotic cells, higher expression levels of caspase-3, but lower level of Bcl-2 than WI-38/Vector cells. This study provided strong evidence that CYP3A5 participated in tetrandrine-induced cytotoxicity.
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Apoptosis/efectos de los fármacos , Bencilisoquinolinas/toxicidad , Citocromo P-450 CYP3A/metabolismo , Pulmón/efectos de los fármacos , Activación Metabólica/efectos de los fármacos , Bencilisoquinolinas/metabolismo , Western Blotting , Técnicas de Cultivo de Célula , Línea Celular , Supervivencia Celular/efectos de los fármacos , Citocromo P-450 CYP3A/genética , Citometría de Flujo , Glutatión/metabolismo , Humanos , L-Lactato Deshidrogenasa/metabolismo , Pulmón/enzimología , Pulmón/patología , Plásmidos , Especies Reactivas de Oxígeno/metabolismo , TransgenesRESUMEN
RATIONALE: Glutathione (GSH) is a very important molecule that participates in various physiologically important events. Depletion of cellular GSH has been suggested as a biomarker of early stage of cell injury. Assessment of GSH with high selectivity and sensitivity is in demand in many fields of life sciences. METHODS: Cell lysates were deproteinated by 5-sulfosalicylic acid. Glutathione was derivatized with monobromobimane to form a GSH conjugate. S-Hexylglutathione was employed as internal standard. The resulting samples were analyzed using a high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS) system. RESULTS: The limit of detection was as low as 500 amol with high selectivity. The approach was sensitive enough to detect GSH content in a single human erythrocyte. The technique is the most sensitive approach among the reported MS-based GSH assays. CONCLUSIONS: A monobromobimane derivatization and mass spectrometry based method was developed for the assessment of GSH. The derivatized GSH was chemically stable against autooxidation. With the great selectivity and sensitivity, the approach is anticipated to serve as a routine method for assessment of GSH and possibly other important endogenous low molecular weight thiols.
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Cromatografía Líquida de Alta Presión/métodos , Glutatión/análisis , Análisis de la Célula Individual/métodos , Espectrometría de Masa por Ionización de Electrospray/métodos , Espectrometría de Masas en Tándem/métodos , Animales , Células Cultivadas , Humanos , Límite de Detección , Ratas , Reproducibilidad de los Resultados , Sensibilidad y EspecificidadRESUMEN
Pulmonary toxicity of styrene is initiated by cytochromes P450-dependent metabolic activation. P450 2E1 and P450 2F2 are considered to be two main cytochrome P450 enzymes responsible for styrene metabolism in mice. The objective of the current study was to determine the correlation between the formation of styrene metabolites (i.e., styrene oxide and 4-vinylphenol) and pulmonary toxicity of styrene, using Cyp2e1- and Cyp2f2-null mouse models. A dramatic decrease in the formation of styrene glycol and 4-vinylphenol was found in Cyp2f2-null mouse lung microsomes relative to that in the wild-type mouse lung microsomes; however, no significant difference in the production of the styrene metabolites was observed between lung microsomes obtained from Cyp2e1-null and the wild-type mice. The knockout and wild-type mice were treated with styrene (6.0 mmol/kg, ip), and cell counts and LDH activity in bronchoalveolar lavage fluids were monitored to evaluate the pulmonary toxicity induced by styrene. Cyp2e1-null mice displayed a susceptibility to lung toxicity of styrene similar to that of the wild-type animals; however, Cyp2f2-null mice were resistant to styrene-induced pulmonary toxicity. In conclusion, both P450 2E1 and P450 2F2 are responsible for the metabolic activation of styrene. The latter enzyme plays an important role in styrene-induced pulmonary toxicity. Both styrene oxide and 4-vinylphenol are suggested to participate in the development of lung injury induced by styrene.
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Sistema Enzimático del Citocromo P-450/metabolismo , Compuestos Epoxi/metabolismo , Microsomas/metabolismo , Fenoles/metabolismo , Estireno/metabolismo , Animales , Sistema Enzimático del Citocromo P-450/deficiencia , Compuestos Epoxi/química , Pulmón/citología , Pulmón/metabolismo , Ratones , Ratones Noqueados , Microsomas Hepáticos/metabolismo , Estructura Molecular , Fenoles/química , Estireno/químicaRESUMEN
Lung cancer still is one of the most common malignancy tumors in the world. However, the mechanisms of its occurrence and development have not been fully elucidated. Zinc finger protein family (ZNFs) is the largest transcription factor family in human genome. Recently, the more and more basic and clinical evidences have confirmed that ZNFs/Krüppel-like factors (KLFs) refer to a group of conserved zinc finger-containing transcription factors that are involved in lung cancer progression, with the functions of promotion, inhibition, dual roles and unknown classifications. Based on the recent literature, some of the oncogenic KLFs are promising molecular biomarkers for diagnosis, prognosis or therapeutic targets of lung cancer. Interestingly, a novel computational approach has been proposed by using machine learning on features calculated from primary sequences, the XGBoost-based model with accuracy of 96.4 % is efficient in identifying KLF proteins. This paper reviews the recent some progresses of the oncogenic KLFs with their potential values for diagnosis, prognosis and molecular target in lung cancer.
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RATIONALE: Immune checkpoint inhibitors have shown high efficacies as the first-line treatment of various advanced malignancies. Yet, the effect and practice patterns of immune checkpoint inhibitors on the second primary tumors are still unclear. Second primary malignancy post immunotherapy, there is paucity in such cases being reported. PATIENT CONCERNS: We report 2 cases of a 57-year-old woman with nonsmall cell lung cancer and a 69-year-old man with metastatic clear cell renal carcinoma treated with immunotherapy who developed second primary malignancies during the therapy. DIAGNOSIS: Second primary malignancy during the therapy. INTERVENTIONS: In addition to the treatments of the second primary malignancies, maintenance immunotherapy was continued for the patients. OUTCOMES: Overall survival in both patients was longer than 12 months, and the treatments were well tolerated. The adverse reactions mainly included depigmentation of hair and facial and limb skin in patient 1 and diarrhea in patient 2. LESSONS: It is necessary to recognize that the second primary malignancy may occur during the immunotherapy, and more clinical studies and practices are still needed for the adjustment of the regimens of immunotherapy. Full diagnosis, timely treatment, and long-term regular follow-up have important significance for patients with malignancies.
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Carcinoma de Pulmón de Células no Pequeñas , Neoplasias Pulmonares , Neoplasias Primarias Secundarias , Masculino , Femenino , Humanos , Persona de Mediana Edad , Anciano , Carcinoma de Pulmón de Células no Pequeñas/tratamiento farmacológico , Neoplasias Primarias Secundarias/tratamiento farmacológico , Neoplasias Pulmonares/patología , Inhibidores de Puntos de Control Inmunológico/uso terapéutico , Inmunoterapia/efectos adversosRESUMEN
Dauricine is the major bioactive component isolated from the root of Menispermum dauricum DC and has shown promising pharmacologic activities with a great potential for clinical use. Recently, we found that intraperitoneal exposure of dauricine produced selective pulmonary injury in mice. A quinone methide metabolite of dauricine was identified and is suggested to be associated with the pulmonary toxicity of dauricine. The present study evaluated the apoptotic effect of dauricine in cultured cells and mice, determined the change in cellular glutathione (GSH) contents after exposure to dauricine, investigated the role of GSH depletion in dauricine-induced cytotoxicity and apoptosis, and examined the role of CYP3A in dauricine-induced GSH depletion and apoptosis. Dauricine was found to induce apoptosis in NL-20 cells. Additionally, intraperitoneal administration of dauricine caused GSH depletion and apoptosis in lungs of mice. Treatment with ketoconazole, an inhibitor of CYP3A, reversed cellular GSH depletion in lungs of mice given dauricine and showed protective effect on dauricine-induced apoptosis in lungs of mice. This indicates that metabolic activation is involved in dauricine-induced GSH-depletion, cytotoxicity and apoptosis. The glutathione depletor L-buthionine sulfoximine showed potentiating effect on cytotoxicity and apoptosis induced by dauricine. We propose that dauricine is metabolized to a quinone methide intermediate which depletes cellular GSH, and the depletion of GSH may trigger and/or intensify the cytotoxicity and apoptosis induced by dauricine.
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Apoptosis/efectos de los fármacos , Bencilisoquinolinas/toxicidad , Bronquios/citología , Citocromo P-450 CYP3A/fisiología , Células Epiteliales/efectos de los fármacos , Pulmón/citología , Tetrahidroisoquinolinas/toxicidad , Animales , Western Blotting , Bronquios/efectos de los fármacos , Butionina Sulfoximina/farmacología , Caspasa 3/metabolismo , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Inhibidores del Citocromo P-450 CYP3A , Citocromos c/metabolismo , Inhibidores Enzimáticos/farmacología , Citometría de Flujo , Glutatión/metabolismo , Humanos , Etiquetado Corte-Fin in Situ , Cetoconazol/farmacología , Pulmón/efectos de los fármacos , Masculino , Ratones , Proteína Letal Asociada a bcl/metabolismoRESUMEN
Metabolic activation is considered to be a critical step for styrene-induced pulmonary toxicity. Styrene-7,8-oxide is a primary oxidative metabolite generated by vinyl epoxidation of styrene. In addition, urinary 4-vinylphenol (4-VP), a phenolic metabolite formed by aromatic hydroxylation, has been detected in workers and experimental animals after exposure to styrene. In the present study, new oxidative metabolites of styrene, including 2-vinylphenol (2-VP), 3-vinylphenol (3-VP), vinyl-1,4-hydroquinone, and 2-hydroxystyrene glycol were detected in mouse liver microsomal incubations. The production rates of 2-VP, 3-VP, 4-VP, and styrene glycol were 0.0527 ± 0.0045, 0.0019 ± 0.0006, 0.0053 ± 0.0002, and 4.42 ± 0.33 nmol/(min · mg protein) in mouse liver microsomes, respectively. Both disulfiram (100 µM) and 5-phenyl-1-pentyne (5 µM) significantly inhibited the formation of the VPs and styrene glycol. 2-VP, 3-VP, and 4-VP were metabolized in mouse liver microsomes at rates of 2.50 ± 0.30, 2.63 ± 0.13, and 3.45 ± 0.11 nmol/(min · mg protein), respectively. The three VPs were further metabolized to vinylcatechols and/or vinyl-1,4-hydroquinone and the corresponding glycols. Pulmonary toxicity of 2-VP, 3-VP, and 4-VP was evaluated in CD-1 mice, and 4-VP was found to be more toxic than 2-VP and 3-VP.
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Contaminantes Ambientales/metabolismo , Hígado/metabolismo , Pulmón/metabolismo , Microsomas/metabolismo , Fenoles/análisis , Estireno/metabolismo , Animales , Biotransformación , Contaminantes Ambientales/farmacocinética , Contaminantes Ambientales/toxicidad , Cromatografía de Gases y Espectrometría de Masas , Hidroxilación , Técnicas In Vitro , Hígado/efectos de los fármacos , Hígado/enzimología , Pulmón/efectos de los fármacos , Pulmón/enzimología , Masculino , Ratones , Ratones Endogámicos , Microsomas Hepáticos/metabolismo , Fenoles/síntesis química , Fenoles/metabolismo , Fenoles/toxicidad , Estireno/farmacocinética , Estireno/toxicidadRESUMEN
Styrene has been reported to be pneumotoxic and hepatotoxic in humans and animals. Styrene oxide, a major reactive metabolite of styrene, has been found to form covalent binding with proteins, such as albumin and hemoglobin. Styrene oxide has two optical isomers and it was reported that the (R)-enantiomer was more toxic than the (S)-enantiomer. The purpose of this study was to develop polyclonal antibodies that can stereoselectively recognize proteins modified by styrene oxide enantiomers at cysteine residues. Immunogens were prepared by alkylation of thiolated keyhole limpet hemocyanin (KLH) with styrene oxide enantiomers. Polyclonal antibodies were raised by immunization of rabbits with the chiral immunogens. Titration tests showed all six rabbits generated high titers of antisera that recognize (R)- or (S)-coating antigens accordingly. No cross-reaction was observed toward the carrier protein (BSA). All three rabbits immunized with (R)-immunogen produced antibodies that show enantioselectivity to the corresponding antigen, while only one among the three rabbits immunized with (S)-immunogen generated antibodies with enantioselectivity of the recognition. The enantioselectivity was also observed in competitive ELISA and immunoblot analysis. Additionally, competitive ELISA tests showed that the immunorecognition required the hydroxyl group of the haptens. Immunoblot analysis demonstrated that the immunorecognition depended on the amount of protein adducts blotted and hapten loading in protein adducts. In summary, we successfully developed polyclonal antibodies to stereoselectively detect protein adducts modified by styrene oxide enantiomers.
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Anticuerpos/inmunología , Especificidad de Anticuerpos , Compuestos Epoxi/química , Inmunoensayo/métodos , Proteínas/análisis , Proteínas/química , Animales , Western Blotting , Ensayo de Inmunoadsorción Enzimática , Sueros Inmunes/inmunología , Proteínas/inmunología , Estereoisomerismo , Estirenos/toxicidadRESUMEN
Styrene and 1,3-butadiene are important intermediates used extensively in the plastics industry. They are metabolized mainly through cytochrome P450-mediated oxidation to the corresponding epoxides, which are subsequently converted to diols by epoxide hydrolase or through spontaneous hydration. The resulting styrene glycol and 3-butene-1,2-diol have been suggested as biomarkers of exposure to styrene and 1,3-butadiene, respectively. Unfortunately, poor ionization of the diols within electrospray mass spectrometers becomes an obstacle to the detection of the two diols by liquid chromatography/electrospray ionization-mass spectrometry (LC/ESI-MS). We developed an LC/ESI-MS approach to analyze styrene glycol and 3-butene-1,2-diol by means of derivatization with 2-bromopyridine-5-boronic acid (BPBA), which not only dramatically increases the sensitivity of diol detection but also facilitates the identification of the diols. The analytical approach developed was simple, quick, and convincing without the need for complicated chemical derivatization. To evaluate the feasibility of BPBA as a derivatizing reagent of diols, we investigated the impact of diol configuration on the affinity of a selection of diols to BPBA using the established LC/ESI-MS approach. We found that both cis and trans diols can be derivatized by BPBA. In conclusion, BPBA may be used as a general derivatizing reagent for the detection of vicinal diols by LC/MS.
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Butadienos/química , Carcinógenos/análisis , Cromatografía Liquida/métodos , Glicoles de Etileno/análisis , Glicoles/análisis , Espectrometría de Masa por Ionización de Electrospray/métodos , Estireno/química , Biomarcadores/análisis , Ácidos Borónicos/química , Humanos , Piridinas/químicaRESUMEN
Benzene, toluene, xylene, and formaldehyde are well-known indoor air pollutants, especially after house decoration. They are also common pollutants in the working places of the plastic industry, chemical industry, and leather industry. It has been reported that these pollutants cause people to be irritated, sick, experience a headache, and be dizzy. They also have the potential to induce asthma, aplastic anemia, and leukemia, even cause abortion or fetus malformation in humans. In this study, the airborne toxicity of benzene, toluene, xylene, and formaldehyde to murine embryonic stem cells (mES cells) were tested using airborne exposure technique to evaluate the mES cell airborne exposure model on embryotoxicity prediction. Briefly, mES cells were cultured on Transwell inserts and were exposed to an airborne surrounding of test chemicals in a chamber for 1 h at 37 degrees C. Cytotoxicity was determined using the MTT assay after further culture for 18 h at 37 degrees C in normal medium. The airborne IC(50) (50% inhibition concentration) of benzene, toluene, xylene, and formaldehyde derived from the fitted dose-response curves were 17,400 +/- 1290, 16,000 +/- 250, 4680 +/- 500, and 620 +/- 310 ppm, respectively. Formaldehyde was found to be the compound most toxic to mES cells compared to benzene homologues. The toxicity data had good correlation with the in vivo data. The results showed that the mES airborne exposure model may be used to predict embryotoxicity of volatile organic compounds.
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Contaminantes Ocupacionales del Aire/toxicidad , Contaminación del Aire Interior/efectos adversos , Células Madre Embrionarias/efectos de los fármacos , Solventes/toxicidad , Algoritmos , Animales , Cámaras de Exposición Atmosférica , Células Cultivadas , Colorantes , Interpretación Estadística de Datos , Relación Dosis-Respuesta a Droga , Ratones , Sales de Tetrazolio , TiazolesRESUMEN
Many traditional Chinese medicine tonifying prescriptions for kidney and spleen have been proved to play various roles in osteoporosis. This study aimed to explore whether Anti-Osteoporosis Decoction (AOD) and Yougui Pill (YGP) have potential therapeutic effects on osteoporosis in ovariectomy-induced rat model. The osteoporosis rat model was established with female Wistar rats by the way of ovariectomy. The chosen rats were randomly divided into five groups (control group, model group, sham group, model + AOD group, and model + YGP group). H&E staining was used to detect the bone histological pathology changes. The bone mineral density (BMD) was assessed with dual-energy X-ray absorptiometry. In addition, western blotting assay was applied to explore the expressions of BMP2, Runx2, Collagen I and Opn. Next, we examined the expression of collagen I by immunohistochemistry staining. Finally, the levels of Alkaline phosphatase (ALP), procollagen type I N propeptide (PINP) and ß-C-terminal telopeptide of type I collagen (ß-CTX) were detected. The results revealed that the OVX osteoporosis model was successfully established. AOD and YGP treatment effectively inhibited osteoporosis and reduced the broken trabecular bones. BMD was increased in AOD and YGP treatment. In addition, AOD and YGP treatment groups significantly increased the ALP levels. Furthermore, AOD and YGP significantly increased the expressions of BMP2, Runx2, Collagen I and Opn, while reduced the levels of PINP and ß-CTX in serum compared to OVX model group. In conclusion, AOD and YGP exert regulatory effects on osteoporosis in ovariectomized rats. They may be potential candidates for the therapy and cure of human osteoporosis.
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Intervertebral disc degeneration (IDD) has become the most common cause of lowback pain, and it imposes a heavy burden on patients with IDD and society. The effects of long noncoding RNAs on the proliferation and development of IDD have attracted increasing attention. The present study aimed to investigate the role and molecular mechanism of Fasassociated protein factor1 (FAF1) in IDD. The expression of FAF1 was detected by reverse transcriptionquantitative polymerase chain reaction. CCK8 and immunofluorescence staining were used to determine cell proliferation. Flow cytometry was performed to measure the cell cycle and apoptosis. Western blotting was used to test pErk expression. The results of the present study demonstrated that the expression of FAF1 was upregulated in patients with disc bulging, herniation and IDD, and the expression of FAF1 was positively correlated with the grade of disc degeneration according to the patients' Pfirrmann score. The overexpression of FAF1 in nucleus pulposus (NP) cells promoted cell proliferation by increasing the percentage of cells in the Sphase of the cell cycle. The expression of phosphorylated extracellular signalregulated kinase (Erk), a possible target of FAF1, was consistent with the expression of FAF1. In addition, it was elucidated that inactivation of the Erk signaling pathway by PD98059 reversed the effect of FAF1 on NP cell proliferation. Taken together, these results demonstrated that FAF1 was vital in the proliferation of NP cells by modulating the Erk signaling pathway, which suggests that FAF1 may be a novel marker in the early diagnosis of IDD and a therapeutic target for patients.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/genética , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Degeneración del Disco Intervertebral/patología , ARN Largo no Codificante/metabolismo , Proteínas Adaptadoras Transductoras de Señales/antagonistas & inhibidores , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Adulto , Anciano , Proteínas Reguladoras de la Apoptosis , Proliferación Celular , Femenino , Flavonoides/farmacología , Humanos , Degeneración del Disco Intervertebral/diagnóstico , Degeneración del Disco Intervertebral/genética , Antígeno Ki-67/metabolismo , Masculino , Persona de Mediana Edad , Núcleo Pulposo/citología , Núcleo Pulposo/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Puntos de Control de la Fase S del Ciclo Celular , Transducción de Señal , Regulación hacia ArribaRESUMEN
OBJECTIVE: To investigate the tumor inhibition effect of Yangfei Kongliu Formula (YKF), a compound Chinese herbal medicine, combined with cisplatin (DDP) and its action mechanisms. METHODS: C57BL/6 mice with Lewis lung carcinoma were divided into six groups: control group (C), DDP group (2 mg/kg, DDP), low-dose YKF group (2.43 g/kg, L), high-dose YKF group (24.3 g/kg, H), low-dose YKF combined with DDP group (L + DDP) and high-dose YKF combined with DDP group (H + DDP). Transforming growth factor-ß1 (TGF-ß1), mothers against decapentaplegic homolog 3 (Smad3) and Smad7 levels were measured with quantitative real-time polymerase chain reaction (qPCR), Western blotting and immunohistochemistry. An enzyme-linked immunosorbent assay was used to analyze the expressions of interleukin-2 (IL-2) and tumor necrosis factor-α (TNF-α). RESULTS: YKF combined with DDP significantly inhibited the growth and metastasis of tumors relative to the control group, and YKF groups (P < 0.05). There was no significant difference between high-dose YKF group and low-dose YKF group (P > 0.05). We also found that the expression levels of TGF-ß1 and Smad3 were both significantly decreased by YKF relative to the control group (P < 0.05). Furthermore, after treatment with YKF combined with DDP, the expression levels of TGF-ß1 and Smad3 were decreased but the expression level of Smad7 was increased relative to the DDP group (P < 0.05). Compared to the DDP group, the combination of YKF and DDP enhanced the effect of tumor inhibition (P < 0.05), showing obvious synergy between YKF and DDP. Treatment with DDP or YKF decreased serum levels of IL-2 and TNF-α relative to the control group (P < 0.05). Furthermore, the expression levels of IL-2 and TNF-α were significantly decreased when treated with YKF in combination with DDP. Co-treatment with YKF and DDP significantly inhibited tumor growth, decreased the expressions of TGF-ß1, Smad3, IL-2 and TNF-α and increased the expression of Smad7; these differences were significant relative to both YKF groups and the control group (P < 0.05). CONCLUSION: YKF can inhibit tumor growth synergistically with DDP, mainly through the TGF-ß1 signaling pathway.
Asunto(s)
Antineoplásicos Fitogénicos/uso terapéutico , Carcinoma Pulmonar de Lewis/tratamiento farmacológico , Cisplatino/uso terapéutico , Medicamentos Herbarios Chinos/uso terapéutico , Neoplasias Pulmonares/tratamiento farmacológico , Fitoterapia , Factor de Crecimiento Transformador beta1/metabolismo , Animales , Antineoplásicos/farmacología , Antineoplásicos/uso terapéutico , Antineoplásicos Fitogénicos/farmacología , Carcinoma Pulmonar de Lewis/metabolismo , Quimioterapia Combinada , Medicamentos Herbarios Chinos/farmacología , Interleucina-2/metabolismo , Neoplasias Pulmonares/metabolismo , Masculino , Ratones Endogámicos C57BL , Transducción de Señal , Proteína smad3/metabolismo , Proteína smad7/metabolismo , Factores de Crecimiento Transformadores , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
A reversed-phase HPLC method for the simultaneous quantitative determination of mandelic acid enantiomers (MA) and phenylglyoxylic acid (PGA) in urine is described. MA and PGA were extracted with ethyl acetate from urine at acidic pH and derivatized with S-(-)-1-(1-naphthyl) ethylamine. A ZORBAX SB-C(18) column (250 mm x 4.6mm i.d., 5 microm, Agilent, USA) was used with a mobile phase composed of methanol-10 mmol/L phosphate buffer [pH 2.5 (65:35, v/v)] at a flow-rate of 0.8 ml/min. Detection was set at UV wavelength of 254 nm. The mean absolute recoveries were 94.2%, 91.9%, 92.5% and 86.3% for S-MA, R-MA, PGA and salicylic acid (I.S.), respectively. The intra- and inter-day precisions determined at three different concentrations ranged from 2.8% to 4.8%, 0.7% to 7.7% and 1.3% to 6.8%, respectively. The lower limits of detection for MA enantiomers and PGA in urine were 1 microg/ml and the lower limits of quantification were 5 microg/ml (R.S.D.<10%, n=5). The method has been applied to determine the urinary excretion of MA enantiomers and PGA from Sprague-Dawley rats after orally administered with styrene.