RESUMEN
BACKGROUND: Clinical evidence gathered in Chinese communities suggested that acupoint sticking therapy could be an alternative treatment for asthma-related diseases. However, its underlying mechanism is still poorly understood. AIM/HYPOTHESIS: In this study, we aimed to investigate the mechanism of the anti-inflammatory effect of acupoint sticking application with 'Treatment of Winter Disease in Summer' (TWDS) prescription by using metabolomics. METHODS: Allergic asthma in guinea pig was sensitized and challenged by ovalbumin (OVA). Histopathological evaluation of the lung tissue was performed by hematoxylin and eosin (H&E) staining and Masson's trichrome staining. The levels of Th2 cytokine and IgE level in serum were measured using enzyme-linked immunoassay (ELISA). The mRNA expression levels of IL-4, IL-5, IL-13 and orosomucoid-like 3 (ORMDL3) were measured using quantitative reverse transcription polymerase chain reaction (RT-qPCR). Proteins of NF-κB signaling pathway were measured using western blot. The serum metabolomics profiles were obtained by using ultra-performance liquid chromatography combined with electrospray ionization quadrupole time-of-flight mass spectrometry (UPLC-ESI-QTOF-MS). RESULTS: The overall results confirmed that AST with TWDS prescription had a significant protective effect against OVA-induced allergic asthma in guinea pig. This treatment not only attenuated airway inflammation and collagen deposition in the airway, but also decreased the levels of IL-4, IL-5, IL-13 and IgE in serum. In addition, metabolomics results indicated that metabolisms of phospholipid, sphingolipid, purine, amino acid and level of epinephrine were restored back to the normal control level. Moreover, results of the gene expression of ORMDL3 in lung tissues indicated that AST using TWDS could alter the sphingolipid metabolism. Further western blotting analysis also showed that its anti-inflammatory mechanism was by decreasing the phosphorylation of p65 and IκB. CONCLUSION: The study demonstrated that metabolomics provides a better understanding of the actions of TWDS acupoint sticking therapy on OVA-induced allergic asthma.
Asunto(s)
Terapia por Acupuntura/métodos , Antiasmáticos/farmacología , Asma/terapia , Medicamentos Herbarios Chinos/farmacología , Hipersensibilidad/terapia , Animales , Asma/metabolismo , Citocinas/sangre , Citocinas/genética , Citocinas/metabolismo , Cobayas , Hipersensibilidad/metabolismo , Inmunoglobulina E/sangre , Pulmón/metabolismo , Pulmón/patología , Masculino , Proteínas de la Membrana/genética , Metabolómica , FN-kappa B/metabolismo , Ovalbúmina/efectos adversos , Transducción de Señal/efectos de los fármacosRESUMEN
INTRODUCTION: Melanin is synthesized by melanocytes, which are located in the basal layer of the skin. After synthesis, melanin is further deposited on the surface of the skin to form black spots or chloasma. Tyrosinase is a rate-limiting enzyme that plays an important role in melanogenesis. Currently, there are many drugs that inhibit tyrosinase expression to further reduce melanogenesis. Nevertheless, some of these could reverse the pharmacological effect of other drugs, when used simultaneously. MATERIALS AND METHODS: B16 mouse melanoma cells were treated with the tyrosinase inhibitors licochalcone A and ß-arbutin, alone or in combination with capsaicin, an alkaloid found in peppers. Cytotoxicity, melanin content, and tyrosinase activity and expression were determined. RESULTS: Licochalcone A/ß-arbutin inhibited tyrosinase expression and further hindered melanin synthesis when applied individually to B16 mouse melanoma cells. However, licochalcone A/ß-arbutin combined with 50 µmol/L capsaicin enhanced the expression of tyrosinase in these cells and further increased melanin content. CONCLUSION: Our data implied that capsaicin could reverse the inhibitory effect of licochalcone A/ß-arbutin on tyrosinase expression in B16 mouse melanoma cells. SUMMARY: B16 mouse melanoma cells were treated with the tyrosinase inhibitors licochalcone A and ß-arbutin, alone or in combination with capsaicin, an alkaloid found in peppers. Cytotoxicity, melanin content, and tyrosinase activity and expression were determined. Licochalcone A/ß-arbutin inhibited tyrosinase expression and further hindered melanin synthesis when applied individually to B16 mouse melanoma cells. However, licochalcone A/ß-arbutin combined with 50 µmol/L capsaicin enhanced the expression of tyrosinase in these cells and further increased melanin content. Our research implied that capsaicin could reverse the inhibitory effect of licochalcone A/ß-arbutin on tyrosinase expression in B16 mouse melanoma cells. Abbreviations used: B16: B16 mouse melanoma cells; L-DOPA: 3, 4-L-dihydroxyphenylalanine; TYR: Tyrosinase; USP: United States Pharmacopeia; FBS: Fetal bovine serum; EDTA: Ethylenediaminetetraacetic acid; DMSO: Dimethyl sulfoxide; RPMI: Roswell Park Memorial Institute; MTT3: 4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide, NaOH: Sodium hydroxide; PBS: Phosphate-buffered saline; RIPA: Radio-immunoprecipitation assay; PMSF: Phenylmethanesulfonyl fluoride or phenylmethylsulfonyl fluoride; SDS: Sodium dodecyl sulfate, sodium salt; PVDF: Polyvinylidene fluoride; ECL: Enhanced chemiluminescence.