Elsevier

Experimental Gerontology

Volume 42, Issue 10, October 2007, Pages 1003-1011
Experimental Gerontology

Natural aging, expression of fibrosis-related genes and collagen deposition in rat lung

https://doi.org/10.1016/j.exger.2007.06.016Get rights and content

Abstract

Aging lung is characterized by morpho-structural modifications, including progressive fibrosis, that lead to an altered function. Here we provide a comprehensive description of lung collagen expression and metabolism during natural aging of rats. Peribronchial collagen increased significantly in the oldest animals (p = 0.05 2- vs. 6- and 19-month-old rats), as a consequence of Collagen-I and Collagen-III (COL-I, COL-III) protein accumulation (p < 0.05 in 6-, 12- and 19-month-old rats versus the youngest). No changes in fibronectin (FN) protein expression and in COL-III and transforming grow factorβ-1 (TGFβ-1) mRNA expression were observed. Conversely the transcription activity of the COL-I gene was overexpressed in the oldest animals (p < 0.05).

In the aged rats, the activity of lung matrix metalloproteinases (MMP), MMP-1 and MMP-2, dropped significantly (p < 0.05), whilst MMP-9 levels were slightly decreased. These changes were associated with a concomitant increase of tissue inhibitors of MMP (TIMP-1 and TIMP-2).

All together, these results suggest that, during natural aging, collagen accumulation in the lung and its progressive fibrosis are mainly due to a reduced proteolytic activity of MMP, in which TIMP-1 and -2 seem to be the major regulating factors.

Introduction

Aging affects organs, tissues and cell types of the same organism in different ways, resulting in differential rates of functional decline, but all together leading to the diminished ability to meet increased demand (Evers et al., 1994).

In the lung, this phenomenon is characterized by morphological and structural modifications associated with several functional changes (D’Errico et al., 1989). While the total capacity remains almost unchanged, the forced expiratory volume and the forced vital capacity decrease as a result of the diminished elastic forces (Enright et al., 1994, Tack et al., 1982, Peterson et al., 1981).

The pulmonary plastic deformation during respiration is carried out by the interstitial structures that comprise collagen, elastin, fibronectin, proteoglycans and other molecules, including the basal membranes of the endothelium (Meyer, 2005).

Collagen, the main component of the extracellular matrix (ECM), represents a family of proteins that, together with other ECM constituents, forms a complex network in the pulmonary tissue. There are 11 different types of collagenic proteins in the lung, that account for 15–20% of the total dry weight of the pulmonary tissue; type I and III (COL I - COL III) are the most representative of these and they comprise approximately 90% of the total collagen (Chambered and Laurent, 1997).

Most of the newly synthesized collagens are immediately degraded and the extent of this process is primarily regulated by the metalloproteinases (MMPs). The activity of the MMPs under physiological conditions is carefully regulated at the level of expression and activation or at the step of inhibition by MMP tissue inhibitors (TIMPs) (Angel et al., 1987, Brinckerhoff et al., 1986, Herron et al., 1986). Lasting perturbations of any of these steps can lead to collagen accumulation in the parenchyma (Ebihara et al., 2000, Iyer et al., 1999).

Fibronectin (FN) forms fibrils associated with matrix components and mesenchymal cells in the interstitium and, through specific binding sites, promotes cell adhesion and migration, cyto-differentiation, phagocytosis and cell growth. In the adult lung, FN is specifically localized at the basal membrane, around smooth muscle cells and in the alveolar lining fluid (Limper and Roman, 1992). During the early phase of active fibroplasia, FN production increases dramatically, and this augmentation is associated with the fibroblast proliferation thereafter responsible for excessive synthesis and deposition of the collagenic protein.

The overall balance of ECM deposition can also be affected by a family of growth factors, namely transforming growth factor β (TGF-β1), through a variety of biological activities.

TGF-β1 directly increases both the abundance and stability of COL-I and COL-III transcripts, but it also regulates the expression of several MMPs (Sporn et al., 1987, Ignotz and Massaguè, 1986). In fact, elevated circulating levels of TGF-β1 are associated with collagen accumulation and increased amounts of MMP-2 protein and TIMPs mRNA (Mozes et al., 1999).

Since fibrosis is a hallmark of aging in various organs (Abrass et al., 1995), the primary goal of this study was to characterize the gene and protein expression of several lung ECM components (Collagen type I, type III and FN).

Therefore, as an index of the ECM turnover, we measured both levels and activities of some metalloproteinases (MMP1, MMP2 and MMP9) and their inhibitors (TIMP-1 and TIMP-2), and, finally, we investigated the age-dependent change in the mRNA expression of TGF-β1.

Section snippets

Animals

Twenty-four male Sprague–Dawely rats (Iffa Credo S.A., a Charles River Company, Calco, Italy) were studied at 2, 6, 12 and 19 months of age (six animals for each group). On arrival, rats were individually housed for 15 days, with controlled temperature (25 °C) and 12 h alternate light–dark cycle, then weighed and killed under fentanyl anaesthesia. Procedures involving animals and their care were conducted in accordance with the institutional guidelines, in compliance with the international

Lung expression of fibrosis-related gene

COL-I gene expression remained essentially similar in rats aged between 2 and 12 months (1.81 ± 0.46, 1.22 ± 0.24, 1.90 ± 0.26), but significantly peaked in the 19-month-old rats (3.34 ± 0.71, +84% vs. 6 months p = 0.037). By contrast no changes in COL-III mRNA were evident at any age (2.06 ± 0.78, 2.18 ± 1.09, 1.62 ± 0.78, 2.12 ± 0.55; p = 0.635 at 2, 6, 12 and 19 months, respectively) (Fig. 1).

TGF-β1 gene transcription remained weakly detectable and unchanged throughout the observation time (0.63 ± 0.09, 0.45 ± 

Discussion

Clinical observations suggest that many respiratory disorders appear mostly in old age and that the prevailing pathologies are certainly those involving remodelling of airways and distal lung parenchyma (Malik et al., 2004, Lundback et al., 2003, Janssens et al., 2001).

Since fibrosis interferes with respiratory function, a better knowledge of the basic mechanisms that control this process in the lung would be useful (Green and Pinkerton, 2004).

Up to now, most published data relate to specific

Acknowledgement

We thank Dr. Luigi Flaminio Ghilardini (Milan University, Italy) for graphic assistance.

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