Partial inhibition of TGF-beta using alpha(v)beta6 integrin antibodies is effective in blocking murine pulmonary fibrosis without exacerbating inflammation. In addition, the elevated expression of alpha(v)beta6, an activator of the fibrogenic cytokine, TGF-beta, in human pulmonary fibrosis suggests that alpha(v)beta6 monoclonal antibodies could represent a promising new therapeutic strategy for treating pulmonary fibrosis.

Rationale: In experimental models, lung fibrosis is dependent on transforming growth factor (TGF)-b signaling. TGF-b is secreted in a latent complex with its propeptide, and TGF-b activators release TGF-b from this complex. Because the integrin avb6 is a major TGF-b activator in the lung, inhibition of avb6-mediated TGF-b activation is a logical strategy to treat lung fibrosis. Objectives: To determine, by genetic and pharmacologic approaches, whether murine radiation-induced lung fibrosis is dependent on avb6. Methods: Wild-type mice, avb6-deficient (Itgb6 2/2 ) mice, and mice heterozygous for a Tgfb1 mutation that eliminates integrin-mediated activation (Tgfb1 1/RGE ) were exposed to 14 Gy thoracic radiation. Some mice were treated with an anti-avb6 monoclonal antibody or a soluble TGF-b receptor fusion protein. avb6 expression was determined by immunohistochemistry. Fibrosis, inflammation, and gene expression patterns were assessed 20-32 weeks postirradiation. Measurements and Main Results: b6 Integrin expression increased within the alveolar epithelium 18 weeks postirradiation, just before onset of fibrosis. Itgb6 2/2 mice were completely protected from fibrosis, but not from late radiation-induced mortality. Anti-avb6 therapy (1-10 mg/kg/wk) prevented fibrosis, but only higher doses (6-10 mg/kg/wk) caused lung inflammation similar to that in Itgb6 2/2 mice. Tgfb1-haploinsufficient mice were also protected from fibrosis. Conclusions: avb6-Mediated TGF-b activation is required for radiationinduced lung fibrosis. Together with previous data, our results demonstrate a robust requirement for avb6 in distinct fibrosis models. Inhibition of avb6-mediated TGF-b activation is a promising new approach for antifibrosis therapy.

SummaryMice that lack activity of αvβ6-and αvβ8-integrins reproduce the abnormalities of Tgfb1-and Tgfb3-null mice

The transforming growth factor (TGF)-␤-inducible integrin ␣v␤6 is preferentially expressed at sites of epithelial remodeling and has been shown to bind and activate latent precursor TGF-␤. Herein , we show that ␣v␤6 is overexpressed in human kidney epithelium in membranous glomerulonephritis , diabetes mellitus , IgA nephropathy , Goodpasture's syndrome , and Alport syndrome renal epithelium. To assess the potential regulatory role of ␣v␤6 in renal disease , we studied the effects of functionblocking ␣v␤6 monoclonal antibodies (mAbs) and genetic ablation of the ␤6 subunit on kidney fibrosis in Col4A3 ؊/؊ mice , a mouse model of Alport syndrome. Expression of ␣v␤6 in Alport mouse kidneys was observed primarily in cortical tubular epithelial cells and in correlation with the progression of fibrosis. Treatment with ␣v␤6-blocking mAbs inhibited accumulation of activated fibroblasts and deposition of interstitial collagen matrix. Similar inhibition of renal fibrosis was observed in ␤6-deficient Alport mice. Transcript profiling of kidney tissues showed that ␣v␤6-blocking mAbs significantly inhibited disease-associated changes in expression of fibrotic and inflammatory mediators. Similar patterns of transcript modulation were produced with recombinant soluble TGF-␤ RII treatment , suggesting shared regulatory functions of ␣v␤6 and TGF-␤. These findings demonstrate that ␣v␤6 can contribute to the regulation of renal fibrosis and suggest this integrin as a potential therapeutic target.

Abstract-Interleukin (IL)-1␤ has previously been shown to be among the most biologically active cytokines in the lungs of patients with acute lung injury (ALI). Furthermore, there is experimental evidence that lung vascular permeability increases after short-term exposure to IL-1 protein, although the exact mechanism is unknown. Therefore, the objective of this study was to determine the mechanisms of IL-1␤-mediated increase in lung vascular permeability and pulmonary edema following transient overexpression of this cytokine in the lungs by adenoviral gene transfer. Lung vascular permeability increased with intrapulmonary IL-1␤ production with a maximal effect 7 days after instillation of the adenovirus. Furthermore, inhibition of the ␣v␤6 integrin and/or transforming growth factor-␤ attenuated the IL-1␤-induced ALI. The results of in vitro studies indicated that IL-1␤ caused the activation of transforming growth factor-␤ via RhoA/␣v␤6 integrin-dependent mechanisms and the inhibition of the ␣v␤6 integrin and/or transforming growth factor-␤ signaling completely blocked the IL-1␤-mediated protein permeability across alveolar epithelial cell monolayers. In addition, IL-1␤ increased protein permeability across lung endothelial cell monolayers via RhoA-and ␣v␤5 integrin-dependent mechanisms. The final series of in vivo experiments demonstrated that pretreatment with blocking antibodies to both the ␣v␤5 and ␣v␤6 integrins had an additive protective effect against IL-1␤-induced ALI. In summary, these results demonstrate a critical role for the ␣v␤5/␤6 integrins in mediating the IL-1␤-induced ALI and indicate that these integrins could be a potentially attractive therapeutic target in ALI. Key Words: lung Ⅲ cytokines Ⅲ inflammation Ⅲ endothelial cells Ⅲ epithelial cells Ⅲ rodents A cute lung injury (ALI) is a devastating clinical syndrome in critically ill patients with an overall mortality rate of 30% to 40%. 1 The syndrome is characterized by alveolar epithelial and lung endothelial injury leading to increased permeability across the alveolar-capillary barrier, pulmonary edema, and acute respiratory failure. 2 Despite an improved understanding of the pathogenesis of ALI in recent years, the molecular steps regulating the development of increased lung endothelial and epithelial permeability remain poorly understood, and no specific pharmacological therapies are currently available.During the early phase of ALI, a variety of inflammatory mediators are released into the distal air spaces. 2 Among those, interleukin (IL)-1␤ has been shown to be among the most biologically active cytokines in the lungs early after the onset of ALI. 3-5 Furthermore, IL-1␤ stimulates the production of a variety of chemokines (eg, IL-8, monocyte chemotactic protein [MCP]-1, and macrophage inflammatory protein [MIP]-1␣) 6 involved in epithelial wound repair 7,8 and is a potent inducer of lung fibrosis. 9,10 It has been previously shown in rats that lung vascular permeability increases after short-term exposure of IL-1␣ and IL-1␤ protein when ...