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The endothelins are a family of endothelium-derived peptides that possess a variety of functions, including vasoconstriction. Endothelin-1 (ET-1) is up-regulated during tissue repair and promotes myofibroblast contraction and migration, hence contributing to matrix remodeling during tissue repair. Here, we show that addition of ET-1 to normal lung fibroblasts induces expression of proteins that contribute to a contractile phenotype, including ␣-smooth muscle actin (␣-SMA), ezrin, moesin, and paxillin. We confirm that ET-1 enhances the ability of lung fibroblasts to contract extracellular matrix, a function essential for tissue repair, through induction of de novo protein synthesis. Blockade of the Akt/ phosphoinositide 3-kinase (PI3-kinase) pathway with LY294002 and wortmannin prevents the ability of ET-1 to induce ␣-SMA, ezrin, paxillin, and moesin and to promote matrix contraction. Dominant negative rac and Akt blocked the ability of ET-1 to promote formation of ␣-SMA stress fibers. Using specific ET-1 receptor inhibitors, we show that ET-1 induces collagen matrix contraction through the ETA, but not the ETB, receptor. Relative to normal pulmonary fibroblasts, fibroblasts cultured from scars of patients with the fibrotic disease systemic sclerosis (scleroderma) show enhanced ET-1 expression and binding. Systemic sclerosis lung fibroblasts show increased ability to contract a collagen matrix and elevated expression of the procontractile proteins ␣-SMA, ezrin, paxillin, and moesin, which are greatly reduced by antagonizing endogenous ET-1 signaling. Thus, blocking ET-1 or the PI3-kinase/Akt cascades might be beneficial in reducing scar formation in pulmonary fibrosis. INTRODUCTIONA complex histological and architectural structure is a prerequisite for effective lung function. In the lung, specialized structures, the alveoli, increase the surface area of the lung, allowing for efficient gas exchange. The maintenance of these specialized structures is in turn dependent on the underlying connective tissue, comprised principally of fibroblasts and extracellular matrix (ECM; for review, see Gadek et al., 1984), which is essential for the mechanical and structural integrity of the lung. As a response to environmental insults, or as a consequence of local inflammatory processes, structural damage to the lung can occur, resulting in a wound healing response. This response consists of an integrated series of biochemical, immunological, and structural changes that result in the de novo synthesis of a new epithelium, blood vessels, and connective tissue (Razzaque and Taguchi, 2003). The proper repair of connective tissue requires synthesis of new ECM components, such as collagen and fibronectin (Badylak, 2002). In addition, repair of connective tissue requires the proper reconstitution of its support function; that is, an appropriate tensile strength must be recreated. This tensile strength results from the remodeling of the newly formed ECM through a combination of cell locomotion and translocation of the flexible collage...

Antisense oligonucleotide-mediated alternative splicing has great potential for treatment of Duchenne muscular dystrophy (DMD) caused by mutations within nonessential regions of the dystrophin gene. We have recently shown in the dystrophic mdx mouse that exon 23, bearing a nonsense mutation, can be skipped after intramuscular injection of a specific 2 -O-methyl phosphorothioate antisense oligoribonucleotide (2OMeAO). This skipping created a shortened, but in-frame, transcript that is translated to produce near-normal levels of dystrophin expression. This expression, in turn, led to improved muscle function. However, because DMD affects muscles body-wide, effective treatment requires dystrophin induction ideally in all muscles. Here, we show that systemic delivery of specific 2OMeAOs, together with the triblock copolymer F127, induced dystrophin expression in all skeletal muscles but not in cardiac muscle of the mdx dystrophic mice. The highest dystrophin expression was detected in diaphragm, gastrocnemius, and intercostal muscles. Large numbers of fibers with near-normal level of dystrophin were observed in focal areas. Three injections of 2OMeAOs at weekly intervals enhanced the levels of dystrophin. Dystrophin mRNA lacking the targeted exon 23 remained detectable 2 weeks after injection. No evidence of tissue damage was detected after 2OMeAO and F127 treatment either by serum analysis or histological examination of liver, kidney, lung, and muscles. The simplicity and safety of the antisense protocol provide a realistic prospect for treatment of the majority of DMD mutations. We conclude that a significant therapeutic effect may be achieved by further optimization in dose and regime of administration of antisense oligonucleotide.exon skipping ͉ muscular dystrophy