RESUMO
Aging is a high-risk factor for obstructive and fibrotic lung diseases. Fibrotic lung disease leading to decreased lung function is characterized by interstitial remodeling and tissue scarring (sclerosis), with destruction of alveoli and excess deposition of type I collagen, an extracellular matrix component secreted by fibroblasts. Therefore, regulating transforming growth factor-ß (TGF-ß) as a profibrotic signal is essential to suppress pulmonary fibrosis. In pulmonary fibrosis, TGF-ß signaling is mediated by Smad and YAP/TAZ, and TAZ linked to the pathology of pulmonary function is observed in lung fibroblasts from patients with idiopathic pulmonary fibrosis. Although fibrosis is thought to be irreversible, it is an interventional condition. Decorin (DCN) blocks TGF-ß signaling in pulmonary fibrosis, although there are no cellular pharmacological methods to stimulate DCN secretion. We previously showed that chicken eggshell membrane (ESM, a well-known wound-healing material) promotes dcn gene expression in fibroblasts. In this study, we investigated whether ESM stimulates DCN secretion as an endogenous mediator and ameliorates pulmonary fibrosis. Decorin secretion was significantly enhanced in the WI-38 lung fibroblast culture supernatants supplemented with ESM. This effect was increased with major component lysozyme and maximally promoted in experiments with lysozyme and ovotransferrin (the two main proteins in soluble ESM) at a 16:1 concentration ratio, the ratio in the ESM extract. Decorin secretion by ESM modulates TGF-ß signaling in lung fibroblasts by reducing TAZ and pSmad2 nuclear localization. Decorin siRNA experiments confirmed that nuclear localization of TAZ is DCN-dependent. In a mouse model of bleomycin-induced pulmonary fibrosis, all fibrotic markers of ESM treatment group such as hydroxyproline (a collagen deposition marker), and both evaluation of fibrosis density by automated thresholding of picrosirius red-stained lung tissue scan images and Ashcroft fibrosis scores, and also the nuclear localization of TAZ were reduced after 2 weeks compared with control group. Furthermore, long-term (22 week) ESM consumption by healthy individuals significantly improved vital capacity and the forced expiratory volume in 1 s to forced vital capacity ratio (FEV1/FVC). This study reveals that ESM, a well-established wound-healing material, may be a potential preventive medicine for pulmonary fibrosis.
RESUMO
αB-crystallin is highly expressed in the heart and slow skeletal muscle; however, the roles of αB-crystallin in the muscle are obscure. Previously, we showed that αB-crystallin localizes at the sarcomere Z-bands, corresponding to the focal adhesions of cultured cells. In myoblast cells, αB-crystallin completely colocalizes with microtubules and maintains cell shape and adhesion. In this study, we show that in beating cardiomyocytes α-tubulin and αB-crystallin colocalize at the I- and Z-bands of the myocardium, where it may function as a molecular chaperone for tubulin/microtubules. Fluorescence recovery after photobleaching (FRAP) analysis revealed that the striated patterns of GFP-αB-crystallin fluorescence recovered quickly at 37°C. FRAP mobility assay also showed αB-crystallin to be associated with nocodazole-treated free tubulin dimers but not with taxol-treated microtubules. The interaction of αB-crystallin and free tubulin was further confirmed by immunoprecipitation and microtubule sedimentation assay in the presence of 1-100 µM calcium, which destabilizes microtubules. Förster resonance energy transfer analysis showed that αB-crystallin and tubulin were at 1-10 nm apart from each other in the presence of colchicine. These results suggested that αB-crystallin may play an essential role in microtubule dynamics by maintaining free tubulin in striated muscles, such as the soleus or cardiac muscles.