Fehrholz Markus, Glaser Kirsten, Speer Christian P, Seidenspinner Silvia, Ottensmeier Barbara, Kunzmann Steffen
University Children's Hospital, University of Wuerzburg, Josef-Schneider-Str. 2, 97080, Wuerzburg, Germany.
Clinic of Neonatology, Buergerhospital Frankfurt am Main, Nibelungenallee 37-41, 60318, Frankfurt am Main, Germany.
Respir Res. 2017 Mar 23;18(1):51. doi: 10.1186/s12931-017-0535-8.
Although caffeine and glucocorticoids are frequently used to treat chronic lung disease in preterm neonates, potential interactions are largely unknown. While anti-inflammatory effects of glucocorticoids are well defined, their impact on airway remodeling is less characterized. Caffeine has been ascribed to positive effects on airway inflammation as well as remodeling. Connective tissue growth factor (CTGF, CCN2) plays a key role in airway remodeling and has been implicated in the pathogenesis of chronic lung diseases such as bronchopulmonary dysplasia (BPD) in preterm infants. The current study addressed the impact of glucocorticoids on the regulation of CTGF in the presence of caffeine using human lung epithelial and fibroblast cells.
The human airway epithelial cell line H441 and the fetal lung fibroblast strain IMR-90 were exposed to different glucocorticoids (dexamethasone, budesonide, betamethasone, prednisolone, hydrocortisone) and caffeine. mRNA and protein expression of CTGF, TGF-β1-3, and TNF-α were determined by means of quantitative real-time PCR and immunoblotting. H441 cells were additionally treated with cAMP, the adenylyl cyclase activator forskolin, and the selective phosphodiesterase (PDE)-4 inhibitor cilomilast to mimic caffeine-mediated PDE inhibition.
Treatment with different glucocorticoids (1 μM) significantly increased CTGF mRNA levels in H441 (p < 0.0001) and IMR-90 cells (p < 0.01). Upon simultaneous exposure to caffeine (10 mM), both glucocorticoid-induced mRNA and protein expression were significantly reduced in IMR-90 cells (p < 0.0001). Of note, 24 h exposure to caffeine alone significantly suppressed basal expression of CTGF mRNA and protein in IMR-90 cells. Caffeine-induced reduction of CTGF mRNA expression seemed to be independent of cAMP levels, adenylyl cyclase activation, or PDE-4 inhibition. While dexamethasone or caffeine treatment did not affect TGF-β1 mRNA in H441 cells, increased expression of TGF-β2 and TGF-β3 mRNA was detected upon exposure to dexamethasone or dexamethasone and caffeine, respectively. Moreover, caffeine increased TNF-α mRNA in H441 cells (6.5 ± 2.2-fold, p < 0.05) which has been described as potent inhibitor of CTGF expression.
In addition to well-known anti-inflammatory features, glucocorticoids may have adverse effects on long-term remodeling by TGF-β1-independent induction of CTGF in lung cells. Simultaneous treatment with caffeine may attenuate glucocorticoid-induced expression of CTGF, thereby promoting restoration of lung homeostasis.
尽管咖啡因和糖皮质激素常用于治疗早产儿慢性肺部疾病，但它们之间潜在的相互作用大多未知。糖皮质激素的抗炎作用已明确，但其对气道重塑的影响尚不清楚。咖啡因已被证实对气道炎症和重塑有积极作用。结缔组织生长因子（CTGF，CCN2）在气道重塑中起关键作用，并与早产儿慢性肺部疾病如支气管肺发育不良（BPD）的发病机制有关。本研究利用人肺上皮细胞和成纤维细胞探讨了在咖啡因存在的情况下糖皮质激素对CTGF调控的影响。
将人气道上皮细胞系H441和胎儿肺成纤维细胞系IMR-90暴露于不同的糖皮质激素（地塞米松、布地奈德、倍他米松、泼尼松龙、氢化可的松）和咖啡因中。通过定量实时PCR和免疫印迹法测定CTGF、TGF-β1-3和TNF-α的mRNA和蛋白表达。另外，用cAMP、腺苷酸环化酶激活剂福斯高林和选择性磷酸二酯酶（PDE）-4抑制剂西洛司特处理H441细胞，以模拟咖啡因介导的PDE抑制作用。
用不同的糖皮质激素（1 μM）处理显著增加了H441细胞（p < 0.0001）和IMR-90细胞（p < 0.01）中CTGF的mRNA水平。同时暴露于咖啡因（10 mM）时，IMR-90细胞中糖皮质激素诱导的mRNA和蛋白表达均显著降低（p < 0.0001）。值得注意的是，单独暴露于咖啡因24小时显著抑制了IMR-90细胞中CTGF mRNA和蛋白的基础表达。咖啡因诱导的CTGF mRNA表达降低似乎与cAMP水平、腺苷酸环化酶激活或PDE-4抑制无关。虽然地塞米松或咖啡因处理不影响H441细胞中TGF-β1 mRNA，但分别暴露于地塞米松或地塞米松和咖啡因后，检测到TGF-β2和TGF-β3 mRNA表达增加。此外，咖啡因增加了H441细胞中TNF-α mRNA（6.5±2.2倍，p < 0.05），TNF-α被认为是CTGF表达的有效抑制剂。
除了众所周知的抗炎特性外，糖皮质激素可能通过非TGF-β1依赖性诱导肺细胞中的CTGF对长期重塑产生不利影响。同时用咖啡因治疗可能会减弱糖皮质激素诱导的CTGF表达，从而促进肺内环境稳态的恢复。
