The definition of fibrogenic processes in fibroblastic foci of idiopathic pulmonary fibrosis based on morphometric quantification of extracellular matrices

Summary

There is limited information regarding the process of tissue remodeling in fibroblastic foci associated with idiopathic pulmonary fibrosis. The aim of this study was to identify the different pathologic stages of tissue remodeling in fibroblastic foci based on the histopathologic differences in the glycosaminoglycan distribution and collagen deposition. In addition, we also aimed at clarifying the stage-specific characteristics by taking into consideration the expression pattern of matrix metalloproteinase and angiogenesis. Lung biopsies of 16 patients with idiopathic pulmonary fibrosis were used. The presence of glycosaminoglycans was detected by Alcian blue staining, and type I collagen was detected by immunohistochemical analysis with a primary antibody specific to the cross-linked carboxyterminal telopeptide of type I collagen. The fibroblastic foci characterized by the expression intensity of Alcian blue and telopeptide of type I collagen were divided into 3 groups, namely, Alcian blue⁺telopeptide of type I collagen^weak, Alcian blue⁺telopeptide of type I collagen⁺, and Alcian blue^weaktelopeptide of type I collagen⁺; consequently, 3 new stages were defined—stages I, II, and III, respectively. A significant inverse correlation was observed between the area densities of Alcian blue⁺ and telopeptide of type I collagen⁺ in fibroblastic foci. Stage I was characterized by the expression of matrix metalloproteinase-2 and tissue inhibitor of matrix metalloprotease-2 in fibroblasts and the overlying epithelium of fibroblastic foci, and also the absence of capillary angiogenesis. In contrast, the expression of these proteins was attenuated in stage III, except for that of matrix metalloproteinase-2 in fibroblasts. In stages II and III, capillary angiogenesis was observed. Lymphangiogenesis was undetected in all the 3 stages. Thus, pathologic staging helps understand the roles of the factors involved in tissue remodeling in idiopathic pulmonary fibrosis.

Introduction

Idiopathic pulmonary fibrosis (IPF) is a chronic and progressive fibrogenic disease of unknown etiology, and it is believed to be a consequence of aberrant healing of a lung injury [1]. IPF is histologically characterized by chronic interstitial pneumonia, small aggregates of myofibroblasts and fibroblasts (termed as fibroblastic foci [FF]), and dense collagen and honeycomb change [2]. FF consist of an extracellular matrix (ECM), overlying epithelium, and a small aggregate of fibroblasts expressing procollagen proteins [2]. It has been considered that FF represent a leading edge of fibrogenesis during the development of IPF [3], [4], [5], [6], [7], although the prognostic significance of FF in IPF patients is still controversial [8], [9], [10].

A variety of factors that are involved in tissue remodeling of IPF have been examined. The synthesis/degradation of the ECM in FF, in which collagen, cell adhesion protein, and proteoglycans (PGs) are present [11], [12], [13], [14], [15], has been of particular interest in this pathologic condition. Matrix metalloproteinase-2 (MMP-2) is a regulator of matrix degradation, and tissue inhibitor of matrix metalloproteinase (TIMP-2) inhibits the activity of MMP-2 by binding to pro-MMP-2. The expression of MMP-2 and TIMP-2 has been observed in fibroblasts and in the overlying epithelium of FF. It has been suggested that these factors play a role in the excessive collagen deposition in the ECM [16], [17], [18], [19], [20], [21], [22], [23]. It has also been shown that capillary angiogenesis is involved in tissue remodeling in pulmonary fibrosis [24], [25], [26], [27], [28], [29]. In particular, in the case of IPF, CD34⁺ capillaries are barely detectable in FF [28], [29]. It has been speculated that the absence of capillary angiogenesis in FF leads to the poor prognosis of IPF.

There is limited information regarding the fibrogenic processes occurring in FF during the development of IPF. It has been reported that versican—a type of PG that consists of a core protein linked to glycosaminoglycans (GAGs)—is observed in FF and that mature collagen is barely detected in versican-rich lesions [30]. This report suggests that multiple temporal phases may be recognizable in the matrix turnover of FF. In the present study, we performed the pathologic staging of FF by characterizing GAG distribution and collagen deposition in FF. Based on the staging, we investigated the expression patterns of MMP-2 and TIMP-2 and the occurrence of capillary angiogenesis and lymphangiogenesis in FF. In a recent study, lymphangiogenesis has been shown to occur in fibrogenic conditions [31], [32], [33]. The pathologic staging of FF provides a deeper understanding of tissue remodeling that occurs during IPF.