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Inflammation is a prominent feature of many disorders that may lead to lung fibrosis, such as desquamative interstitial pneumonia, non-specific interstitial pneumonia, radiation pneumonitis, drug-related lung injury and others (Katzenstein & Myers 1998). Bleomycin-induced lung fibrosis in rodents is a commonly employed animal model of fibrotic lung disease (Snider et al. 1978; Hesterberg et al. 1981; Chandler et al. 1983; Lazo et al. 1990). Bleomycin-induced lung injury involves a complex interaction of many cell types, some resident in the lung, others migratory to the lung from the circulation. The role of lymphocytes in the pathogenesis of the pulmonary fibrotic response to bleomycin is somewhat obscure. T lymphocytes are capable of generating fibroblast chemotactic factors and factors that influence both fibroblast proliferation and collagen synthesis (Johnson & Ziff 1976; Hibbs et al. 1983; Postlethwaite et al. 1984). The inflammatory response to bleomycin instillation is demonstrated by means of analysis of BAL cells. An early increase in the percentage of neutrophils is followed by a more sustained BAL lymphocytosis (Izbicki et al. 2002b). In a study of parenchymal lung lymphocytes in bleomycin-treated rats, the normal CD4 : CD8 ratio of 1 : 1 was increased 14 days post-bleomycin to 2 : 1, reversing to 1 : 2 in the later stages of the disease process (Thrall & Barton 1984). Intriguingly, the increase in helper T cells occurs when collagen synthesis is most active, and the increase in suppressor T cells occurs when collagen synthesis has decreased, suggesting that helper T cells might enhance, and suppressor T cells suppress, collagen deposition in bleomycin lung injury. A number of studies have attempted to define the role of lymphocytes in the pathogenesis of bleomycin lung fibrosis. Treatment of animals with anti-CD3 monoclonal antibody (Sharma et al. 1996), antilymphocyte globulin (Thrall et al. 1979) or with thymectomy and irradiation (Thrall et al. 1980) resulted in a decreased collagen deposition in bleomycin-exposed animals. However, SCID mice appear to develop lung injury of similar severity to wild-type animals (Zhu et al. 1996; Helene et al. 1999). Moreover, while one study of the response to bleomycin in athymic nude mice showed no difference in comparison with euthymic mice (Szapiel et al. 1979), two further studies demonstrated an attenuated response to bleomycin (Schrier et al. 1983) and peplomycin (Ekimoto et al. 1985), a bleomycin analogue, in nude mice. Thus, the pathogenetic role of lymphocytes in lung fibrosis is somewhat controversial, but there appears to be considerable evidence that supports a role for lymphocytes in the development of fibrosis. We, therefore, hypothesized that elimination of activated lymphocytes by interleukin-2 receptor (IL-2R)-targeted chimeric proteins might ameliorate lung fibrosis. IL-2 and its receptor (IL-2R) regulate the magnitude and duration of the T-cell immune response. The IL-2R α-subunit, which confers high affinity for IL-2, is expressed on activated T cells, but not on normal resting cells. This selective expression of the high-affinity IL-2R provides the scientific basis for therapeutic strategies that target IL-2R-expressing cells (Waldmann 1993). One such strategy uses chimeric proteins, such as IL-2-based chimeras, in which IL-2 is responsible for the binding of the protein to activated lymphocytes. IL-2 is fused at the cDNA level to a cytotoxic protein, which comprises the cytotoxic component of the chimeric protein. Bacterial toxins, such as Pseudomonas exotoxin (PE) or diphtheria toxin have been used as cytotoxic components of targeted chimeric proteins (Brinkmann & Pastan 1994). One such molecule is IL-2-PE40, a chimeric protein linking human IL-2 to a modified (truncated) form of PE, so that the toxin will no longer indiscriminately bind and kill normal cells but will, instead, selectively target and kill cells expressing the ligand identified by the binding component (Ogata et al. 1988; Lorberboum-Galski et al. 1988a; Lorberboum-Galski et al. 1988b). IL-2-PE40 effectively attenuates several experimental autoimmune conditions, such as adjuvant-induced arthritis (Case et al. 1989; Lorberboum-Galski et al. 1991), experimental autoimmune encephalomyelitis (Rose et al. 1991) and autoimmune uveoretinitis (Roberge et al. 1989), and prevents the rejection of heart and corneal allografts in rodents (Lorberboum-Galski et al. 1989; Herbort et al. 1991). IL-2-PE40 has also been showed to suppress the growth of a T-cell lymphoma in mice (Kozak et al. 1990). The effect of IL-2-PE40 has been showed to be specific in that IL-2-PE40Asp553, a mutant chimeric protein in which the enzymatic domain of PE is inactive, has no effect (Case et al. 1989; Lorberboum-Galski et al. 1989). A disadvantage of this class of targeted chimeric toxins is that the bacterial toxins are recognized as foreign by the immune system of the recipient organism. In order to overcome this problem, a new generation of chimeric proteins exploiting human apoptosis-inducing proteins was developed. IL-2-Bax is the first prototype of this class. Bax is a pro-apoptotic member of the Bcl-2 protein family. IL-2-Bax specifically targets IL-2R-expressing cells and induces cell-specific, dose-dependent apoptosis (Aqeilan et al. 1999; Aqeilan et al. 2002). IL-2-BaxS184D, a mutant in which Bax is inactive, is taken up by IL-2R-expressing cells, such as IL-2-Bax, but does not activate the apoptotic pathway (Aqeilan et al. 2002). In this study, we investigated the effects of the novel apoptosis-inducing IL-2R-targeted chimeric protein IL-2-Bax on bleomycin-induced lung injury in mice. |
