Skip to main page content
U.S. flag

An official website of the United States government

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2020 Feb 11;9(2):411.
doi: 10.3390/cells9020411.

Matrix Metalloproteinases Retain Soluble FasL-mediated Resistance to Cell Death in Fibrotic-Lung Myofibroblasts

Affiliations

Matrix Metalloproteinases Retain Soluble FasL-mediated Resistance to Cell Death in Fibrotic-Lung Myofibroblasts

David Nareznoi et al. Cells. .

Abstract

A prominent feature of obstructed tissue regeneration following injury in general, and fibrotic lung tissue in particular, is fibroblast proliferation and accumulation. The Fas/FasL apoptotic pathway has been shown to be involved in human idiopathic pulmonary fibrosis (IPF) and bleomycin-induced lung fibrosis in rodents. We previously showed that in normal injury repair, myofibroblasts' accumulation is followed by their decline by FasL+ T cell-induced cell death. In pathological lung fibrosis, myofibroblasts resist cell death and accumulate. Like other members of the tumor necrosis factor (TNF) family, membrane-bound FasL can be cleaved from the cell surface to generate a soluble form (sFasL). Metalloproteinases (MMPs) are known to convert the membrane-bound form of FasL to sFasL. MMP-7 knockout (KO) mice were shown to be protected from bleomycin (BLM)-induced lung fibrosis. In this study, we detected increased levels of sFasL in their blood serum, as in the lungs of patients with IPF, and IPF-lung myofibroblast culture medium. In this study, using an MMP-inhibitor, we showed that sFasL is decreased in cultures of IPF-lung myofibroblasts and BLM-treated lung myofibroblasts, and in the blood serum of MMP-7KO mice. Moreover, resistant fibrotic-lung myofibroblasts, from the lungs of humans with IPF and of BLM-treated mice, became susceptible to T-cell induced cell death in a co-culture following MMP-inhibition- vs. control-treatment or BLM-treated MMP-7KO vs. wild-type mice, respectively. sFasL may be an unrecognized mechanism for MMP-7-mediated decreased tissue regeneration following injury and the evolution of lung fibrosis.

Keywords: cell death; lung myofibroblasts; matrix metalloproteinase (MMP); pulmonary fibrosis; soluble FasL (sFasL).

PubMed Disclaimer

Conflict of interest statement

Acknowledgments: We thank Shifra Fraifeld for her editorial assistance in preparing this manuscript.

Figures

Figure 1
Figure 1
The decreased soluble and increased membrane FasL levels in fibrotic-(IPF) and normal (NL)-lung myofibroblasts, following exposure to the batimastat MMP inhibitor. Western blot of: (A) sFasL in culture medium and (B) mFasL, of fibroblast cell lines (3 × 105) of fibrotic-lung/ATCC191 (IPF)- or normal/ATCC151 (NL)-lungs; Graphical presentation and blots (insert) with optical density ratios normalized to fibroblasts GAPDH after treatment with control-vehicle (0.1% DMSO) or batimastat (24 h, 10 µM). Mean ± standard deviation; n = 4; * p < 0.05, NS—p > 0.05.
Figure 2
Figure 2
The MMP inhibitor (batimastat) decreases IPF-lung myofibroblast’ resistance to T cell-induced cell death. (A) Light microscope images with trypan blue exclusion (inserted numbers); and (B) Graphical presentation of the cell death percentage defined by trypan blue exclusion. Comparisons were made between IPF-lung myofibroblasts (ATCC-191 cell-line) cultured alone (inserts-Ctrl) and IPF-lung myofibroblasts co-cultured with T cells (1 × 106, 48 h), following being treated with vehicle (0.1% DMSO) or MMP-inhibitor (batimastat 10 µM, 24 h), (vehicle or MMP inhibitor, +T cells). The percentage of dead cells to total cell count in each sample was analyzed as the mean ± standard deviation; n = 3; * p < 0.05, NS p > 0.05.
Figure 3
Figure 3
sFasL is increased in the blood serum of wild type (WT), but not of MMP-7 knock-out (KO) bleomycin-treated, mice. The graphical presentation and inserts of WB with the OD analysis of sFasL assessed from equal volumes of blood serum samples (10 μL) of day 14 of (A) bleomycin (Bleo) vs. saline (NL)-treated WT mice, and (B) bleomycin-treated WT vs. MMP-7 KO mice. Mean ± standard deviation; n = 4; *p < 0.05.
Figure 4
Figure 4
The decreased levels of sFasL in the culture medium of MMP-7 KO lung myofibroblasts increase their susceptibility to cell death when further co-cultured with T cells. (A) Western blot of sFasL in culture medium, of lung fibroblasts (3 × 105), isolated from bleomycin-treated lungs (day 14) of WT vs. MMP-7 KO mice; Graphical presentation and blots (insert) with optical density ratios normalized to fibroblasts’ GAPDH. Mean ± standard deviation; n = 3; * p < 0.05. (B) Light microscope images with trypan blue exclusion (inserted numbers); and (C) Graphical presentation of the cell death percentage defined by trypan blue exclusion. Comparisons were made between myofibroblasts from lungs of bleomycin-treated wild type (WT) vs. MMP7 knockout (MMP-7 KO) cultured alone (inserts-Ctrl) and co-cultured with T cells (1 × 106, 48 h), (WT or MMP7-KO, +T cells). Mean ± standard deviation; n = 3; *p < 0.05.
Figure 5
Figure 5
Simplified scheme of the proposed model of sFasL, MMP-mediated, immune cell-induced myofibroblast death regulation during lung fibrosis. Lymphocytes are known to express mFasL that enables T cell-induced myofibroblast cell death with resolution of fibrosis. In fibrotic-lung myofibroblasts, in the presence of MMPs, high levels of sFasL are detected due to mFasL cleavage. High sFasL in the milieu competes with mFasL and limits T cell-induced cell death. sFasL secreted by fibrotic lung fibroblasts is an additional mechanism of their resistance to cell death and their uninterrupted accumulation in lung fibrosis.

Similar articles

Cited by

References

    1. King T.E. Diagnostic Advances in Idiopathic Pulmonary Fibrosis. Chest. 1991;100:238–241. doi: 10.1378/chest.100.1.238. - DOI - PubMed
    1. Marinelli W.A. Idiopathic Pulmonary Fibrosis: Progress and Challenge. Chest. 1995;108:297–298. doi: 10.1378/chest.108.2.297. - DOI - PubMed
    1. Daniil Z.D., Gilchrist F.C., Nicholson A.G., Hansell D.M., Harris J., Colby T.V., du Bois R.M. A histologic pattern of nonspecific interstitial pneumonia is associated with a better prognosis than usual interstitial pneumonia in patients with cryptogenic fibrosing alveolitis. Am. J. Respir. Crit. Care Med. 1999;160:899–905. doi: 10.1164/ajrccm.160.3.9903021. - DOI - PubMed
    1. Kuhn C., McDonald J.A. The roles of the myofibroblast in idiopathic pulmonary fibrosis. Ultrastructural and immunohistochemical features of sites of active extracellular matrix synthesis. Am. J. Pathol. 1991;138:1257–1265. - PMC - PubMed
    1. Zhang K., Rekhter M.D., Gordon D., Phan S.H. Myofibroblasts and their role in lung collagen gene expression during pulmonary fibrosis. A combined immunohistochemical and in situ hybridization study. Am. J. Pathol. 1994;145:114–125. - PMC - PubMed

Publication types

MeSH terms

LinkOut - more resources