Research Article: The Role of the Receptor for Advanced Glycation End-Products in a Murine Model of Silicosis

Date Published: March 19, 2010

Publisher: Public Library of Science

Author(s): Lasse Ramsgaard, Judson M. Englert, Jacob Tobolewski, Lauren Tomai, Cheryl L. Fattman, Adriana S. Leme, A. Murat Kaynar, Steven D. Shapiro, Jan J. Enghild, Tim D. Oury, Rory Edward Morty.

Abstract: The role of the receptor for advanced glycation end-products (RAGE) has been shown to differ in two different mouse models of asbestos and bleomycin induced pulmonary fibrosis. RAGE knockout (KO) mice get worse fibrosis when challenged with asbestos, whereas in the bleomycin model they are largely protected against fibrosis. In the current study the role of RAGE in a mouse model of silica induced pulmonary fibrosis was investigated.

Wild type (WT) and RAGE KO mice received a single intratracheal (i.t.) instillation of silica in saline or saline alone as vehicle control. Fourteen days after treatment mice were subjected to a lung mechanistic study and the lungs were lavaged and inflammatory cells, protein and TGF-β levels in lavage fluid determined. Lungs were subsequently either fixed for histology or excised for biochemical assessment of fibrosis and determination of RAGE protein- and mRNA levels. There was no difference in the inflammatory response or degree of fibrosis (hydroxyproline levels) in the lungs between WT and RAGE KO mice after silica injury. However, histologically the fibrotic lesions in the RAGE KO mice had a more diffuse alveolar septal fibrosis compared to the nodular fibrosis in WT mice. Furthermore, RAGE KO mice had a significantly higher histologic score, a measure of affected areas of the lung, compared to WT silica treated mice. A lung mechanistic study revealed a significant decrease in lung function after silica compared to control, but no difference between WT and RAGE KO. While a dose response study showed similar degrees of fibrosis after silica treatment in the two strains, the RAGE KO mice had some differences in the inflammatory response compared to WT mice.

Aside from the difference in the fibrotic pattern, these studies showed no indicators of RAGE having an effect on the severity of pulmonary fibrosis following silica injury.

Partial Text: Pulmonary fibrosis can have various causes and can be a debilitating progressive condition with a poor prognosis. Fibrosis can develop as a response to inhaled fibrogenic fibers or particles, such as asbestos fibers or silica particles [1], [2]. It can also develop as a side effect of treatment with chemotherapeutic agents or be idiopathic (IPF) with no identifiable etiology [3], [4]. Although silicosis is 100% preventable by proper personal protection, it is still a widespread disease. In the United States the annual number of deaths with silicosis as the underlying or contributing cause has decreased from over 1,100 back in 1968 to under 200 in 2004 [5]. In contrast, China recorded 500,000 cases of silicosis in the period 1991–1995 with an incidence of around 6,000 per year with more than 24,000 deaths annually [6].

The role of RAGE has previously been studied in an asbestos and a bleomycin model of pulmonary fibrosis [21], [22]. Notably, the absence of RAGE was found to lead to spontaneous fibrosis with aging and increased fibrosis in response to asbestos injury [21]. In contrast, RAGE KO mice were found to be largely protected against bleomycin induced fibrosis [22]. Here, the role of RAGE in a silica model of pulmonary fibrosis was studied. Injury led to a decrease in RAGE mRNA levels and a decrease in total RAGE protein in total lung homogenates. This is consistent with findings in both the bleomycin model and asbestos model as well as in lungs from human IPF patients [20]–[22]. The decrease in membrane RAGE protein is likely to be a direct effect of loss of type I epithelial cells and may also be due to cleavage of membrane RAGE by proteases to form soluble RAGE [35], [36]. The loss of soluble RAGE is more likely to be an indicator of tissue damage and damage to the extracellular matrix, but may be due to decreased production from injured epithelial cells. Previous studies have shown that shedding of syndecan-1, a heparin sulfate proteoglycan highly abundant in the extracellular matrix, occurs in models of pulmonary fibrosis [37]. Since soluble RAGE has high affinity to heparin sulfate, collagen and other extracellular matrix proteins [29], [36], [38], it is likely that damage to those structures will result in release of bound soluble RAGE. In fact, soluble RAGE has been shown to accumulate in the BALF after acute lung injury [33]. However, barely any soluble RAGE was detected in the BALF of WT mice both before and after silica injury. This may be due to clearance of soluble RAGE from the BALF after the initial injury.