RESUMEN
Idiopathic pulmonary fibrosis (IPF) is a chronic and refractory interstitial lung disease. Although there is no cure for IPF, the development of drugs with improved efficacy in the treatment of IPF is required. Daphnetin, a natural coumarin derivative, has immunosuppressive, anti-inflammatory, and antioxidant activities. However, its antifibrotic effects have not yet been elucidated. In this study, we investigated the antifibrotic effects of daphnetin on pulmonary fibrosis and the associated molecular mechanism. We examined the effects of daphnetin on splenocytes cultured in Th17 conditions, lung epithelial cells, and a mouse model of bleomycin (BLM)-induced pulmonary fibrosis. We identified that daphnetin inhibited IL-17A production in developing Th17 cells. We also found that daphnetin suppressed epithelial-to-mesenchymal transition (EMT) in TGF-ß-treated BEAS2B cells through the regulation of AKT phosphorylation. In BLM-treated mice, the oral administration of daphnetin attenuated lung histopathology and improved lung mechanical functions. Our findings clearly demonstrated that daphnetin inhibited IL-17A and EMT both in vitro and in vivo, thereby protecting against BLM-induced pulmonary fibrosis. Taken together, these results suggest that daphnetin has potent therapeutic effects on lung fibrosis by modulating both Th17 differentiation and the TGF-ß signaling pathway, and we thus expect daphnetin to be a drug candidate for the treatment of IPF.
Asunto(s)
Bleomicina , Fibrosis Pulmonar Idiopática , Ratones , Animales , Bleomicina/efectos adversos , Interleucina-17/metabolismo , Fibrosis Pulmonar Idiopática/inducido químicamente , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Factor de Crecimiento Transformador beta/metabolismoRESUMEN
Idiopathic pulmonary fibrosis (IPF) is a chronic and refractory interstitial lung disease. Although there are two approved drugs for IPF, they were not able to completely cure the disease. Therefore, the development of new drugs is required for the effective treatment of IPF. In this study, we investigated the effect of theophylline, which has long been used for the treatment of asthma, on pulmonary fibrosis. The administration of theophylline attenuated the fibrotic changes of lung tissues and improved mechanical pulmonary functions in bleomycin (BLM)-induced pulmonary fibrosis. Theophylline treatment suppressed IL-17 production through inhibiting cytokines controlling Th17 differentiation; TGF-ß, IL-6, IL-1ß, and IL-23. The inhibition of IL-6 and IL-1ß by theophylline is mediated by suppressing BLM-induced ROS production and NF-κB activation in epithelial cells. We further demonstrated that theophylline inhibited TGF-ß-induced epithelial-to-mesenchymal transition in epithelial cells through suppressing the phosphorylation of Smad2/3 and AKT. The inhibitory effects of theophylline on the phosphorylation of Smad2/3 and AKT were recapitulated in BLM-treated lung tissues. Taken together, these results demonstrated that theophylline prevents pulmonary fibrosis by inhibiting Th17 differentiation and TGF-ß signaling.
Asunto(s)
Bleomicina , Fibrosis Pulmonar Idiopática , Animales , Ratones , Bleomicina/toxicidad , Teofilina/farmacología , Interleucina-6/farmacología , Proteínas Proto-Oncogénicas c-akt , Pulmón , Diferenciación Celular , Factor de Crecimiento Transformador beta/farmacología , Fibrosis Pulmonar Idiopática/inducido químicamente , Fibrosis Pulmonar Idiopática/tratamiento farmacológico , Ratones Endogámicos C57BLRESUMEN
We synthesized biodegradable polycationic PAMAM (polyamidoamine) esters (e-PAM-R, e-PAM-K) that contain arginines or lysines at the peripheral ends of PAMAM-OH dendrimer through ester bond linkages. The PAMAM esters were readily degradable under physiological conditions (pH 7.4, 37 degrees C), with more than 50% of the grafted amino acids hydrolyzed within 5h. However, polyplexes were very stable and were hardly degraded in the endosomal pH range. Moreover, these amino-acid-modified polymers showed excellent buffering capacities between pH 5.1 and 7.4, facilitating endosomal escape of polyplexes. While the lysine-grafted PAMAM ester did not display significant improvement in transfection efficiency, the arginine-conjugated PAMAM ester-mediated transfection of a luciferase gene showed better transfection efficiency than the branched 25 kDa PEI (polyethylenimine) and PAM-R (peptide bond), and lower cytotoxicity, especially with primary cells such as HUVECs (human umbilical vein endothelial cells) and SMCs (primary rat aorta vascular smooth muscle cells). Furthermore, after DNA release, free e-PAM-R degraded completely into nontoxic PAMAM-OH and arginines by hydrolysis, which resulted in lower cytotoxicity in contrast to the poorly degradable arginine-modified PAMAM with amide bonds. These findings demonstrated that the arginine-grafted biodegradable PAMAM dendrimer, e-PAM-R, is a potential candidate as a safe and efficient gene delivery carrier for gene therapy.
Asunto(s)
Ésteres/síntesis química , Poliaminas/síntesis química , Transfección , Animales , Arginina , Tampones (Química) , Muerte Celular/efectos de los fármacos , Línea Celular , Dendrímeros , Electroforesis en Gel de Agar , Ésteres/química , Ésteres/farmacología , Humanos , Lisina , Microscopía Confocal , Plásmidos/metabolismo , Poliaminas/química , Poliaminas/farmacología , Polímeros , Ratas , VolumetríaRESUMEN
It is a matter of concern to develop and design synthetic non-viral gene carriers with high transfection efficiency and low cytotoxicity in gene therapy. Recently, various arginine conjugated dendrimers showed better performance in transfection and greater viability than polyethyleneimine (PEI). In this study, we synthesized and investigated e-PAM-R G2, 3 and 4 which are biodegradable polyamidoamine (PAMAM) dendrimers modified with arginine and compared that with PAMAM-R series containing amide bonds for gene carriers. For plasmid DNA delivery, the transfection efficiency of e-PAM-R G4 was higher than G3 and G2 and similar to PAMAM-R G4 with favorable cell viability. Moreover, they indicated significantly higher suppression of TEL/AML1 protein, maybe due to rapid olidonucleotide (ODNs) release through biodegradability of e-PAM-R. These results suggest that biodegradable and non-toxic e-PAM-R may be useful carriers for the gene including plasmid DNA, antisense ODNs and si-RNA.