Titration of non-replicating adenovirus as a vector for transducing active TGF-β1 gene expression causing inflammation and fibrogenesis in the lungs of C57BL/6 mice

Transforming growth factor beta 1 (TGF-β1) is a potent inducer of inflammation and extracellular matrix production in the lung as well as in other organ systems (Border & Noble 1994). TGF-β1 has been overexpressed in the lungs of rats by means of an adenovirus vector, resulting in severe inflammation and persistent fibrosis (Sime et al. 1997). Similar techniques also were used to transduce tumour necrosis factor alpha (TNF-α) in rat lungs (Sime et al. 1998). These and a number of other studies have demonstrated the utility of replication-deficient adenovirus vector for introducing biologically active peptide growth factors (GF) (Xing et al. 1996; Zhao et al. 1999; Morikawa et al. 2000), cytokines (Kolb et al. 2001) or their receptors (Kolls et al. 1994) into anatomical sites where their roles in disease development can be studied. We have focused on four GF in the lungs of rats and mice that develop interstitial pulmonary fibrosis (IPF) as a result of inhaling aerosolized asbestos fibres (Perdue & Brody 1994; Liu et al. 1996; Liu et al. 1997; Liu et al. 1998; Liu et al. 2001). Obviously, there are numerous factors that are synthesized and secreted as IPF develops (Adler et al. 1994; Broekelmann et al. 1991; Zhang & Phan 1996; Baughman et al. 1999), but it is possible to consider effectively only a very few and ask what roles they might play in mediating the disease process. The factors we have identified during the initial phases of developing fibrotic lesions are transforming growth factor alpha (TGF-α) (Liu et al. 1996), TGF-β1 (Perdue & Brody 1994; Brass et al. 1999; Liu & Brody 2001), the platelet-derived GF isoforms A and B (PDGF-A and -B) (Liu et al. 1997) and TNF-α (Liu et al. 1998). TGF-α is a strong mitogenic factor for epithelial cells (Korfhagen et al. 1994), whereas the PDGF isoforms exert a potent proliferative influence on mesenchymal cells (Ross et al. 1986). TNF-α is a pleiotropic cytokine, with its action dependent upon several conditions, including cell type, stage of cell maturation and position in the cell cycle (Sherry & Cerami 1988; Phan et al. 1992; Piguet 1993). TGF-β1 has been studied extensively and apparently is the most potent inducer of extracellular matrix by mesenchymal cells (Ignotz & Massague 1986; Rossi et al. 1988; Khalil & Greenberg 1991). It also is a strong inhibitor of epithelial and mesenchymal cell growth (Roberts et al. 1985; Fine & Goldstein 1987; Cazals et al. 1994). TGF-β1 is such a dependable blocker of epithelial proliferation that it is used to inhibit growth of mink lung epithelial cells in a popular assay to quantify the GF protein (Garrigue-Antar et al. 1995). In its primary role as a stimulator of matrix production, TGF-β1 could operate as a central mediator of fibroproliferative lung disease (Leof et al. 1986; Soma & Grotendorst 1989; Kelley et al. 1991; Bonner et al. 1993; Perdue & Brody 1994; Lasky et al. 1995). In the studies reported here, we have used a range of adenoviral vector concentrations to transduce the expression of biologically active TGF-β1 in the lungs of normal mice. It is necessary to know the dose of viral vector that modulates the development of interstitial inflammation and fibrogenesis through transduction of TGF-β1 expression. The concept is to sort out the complex effects of this growth factor because it acts as a chemotactic factor, growth factor and inducer of extracellular matrix production in the lung. We have carried out a series of dose–response experiments in which a recombinant adenovirus transduces TGF-β1 expression at a no-effect level, a minimal-effect level and through severe disease. We demonstrate progression and resolution of disease, inflammation, fibrosis, quantification of TGF-β1 protein and apparent suppression of epithelial proliferation.