Cellular Senescence as Fibrogenic Mechanism in Smoking Lung

Abstract

Age is a risk factor for many diseases including idiopathic pulmonary fibrosis (IPF) by affecting wound healing through abnormal fibrogenesis. Common cellular and molecular traits associated with aging can be found in senescent cells, which may occur throughout the lifespan playing physiological roles during normal development or limiting the proliferation of damaged and aged cells. In this study we investigated in smoking mice whether the senescence of lung cells, triggered by the oxidative stress response, is associated with molecular pathways that can influence lung fibrogenesis and contribute to the remodeling of the fibrotic tissue characterizing some phenotypes of COPD in humans. Among the complex mixture of hundreds senescence secreted factors that include proinflammatory cytokines, chemokines, growth factors, and proteases, we selected important factors involved in differentiation of myofibroblasts, their dedifferentiation and apoptosis, as well as in their metabolic function.The expression of these factors and the amount of matrix deposition was analyzed after chronic cigarette smoke (CS) exposure of C57 Bl/6 and DBA/2 mice sensitive to oxidative, in different lung compartmentscharacterized by a fibrotic remodeling using histochemistry or immunohistochemistry.Unlike C57 Bl/6, DBA/2 mice at 4 months of exposure show marked positivity for 8‐OHdG, a marker of oxidative damage, in subpleural areas, in central parts of the lung parenchyma, on airways and on cells of the fibro‐muscular layer.In these areas, a marked positivity is observed for fibrogenic cytokines such as TGF‐β, PDGF‐b and CTGF, and for other factors such as the p16ink4A senescence marker, and proliferation markers PCNA and Ki67. In DBA/2 mice, these compartments are characterized by a progressive fibrous remodeling at various experimental time points from 5 months of CS exposure onwards as demonstrated by Masson’s trichrome staining.Fibroproliferative foci are present in sub‐pleural areas, in peripheral areas of airways, and in central parts of the lung parenchyma. They are characterized by increasing number of α‐SMA positive myofibroblasts, which express the senescence marker p16ink4A and the transcription factor MyoD that plays a crucial role in myoblast proliferation and differentiation. MyoD induces cell cycle arrest, promotes cellular terminal differentiation of fibroblasts into myofibroblasts, and in senescent myofibroblasts it has been reported to determine resistance to apoptosis and opposes their dedifferentiation.All these factors contribute to the development in smoking DBA/2 mice of lung lesions, which are like those seen in patients with “Combined Fibrosis/Emphysema Syndrome.”Despite consistent levels of TGF‐β, PDGF‐b and CTGF, C57 Bl/6 mice develop after chronic CS exposure fibrotic remodeling at lower extent. Of interest, a very low expression of factors indicative of senescence (p16ink4A and MyoD) and cell proliferation (PCNA and Ki67) is observed.Our results suggest that apoptosis, senescence, and proliferation induced at different rate by fibrotic cytokines and senescence associated secretory factors play important role in early or late appearance of fibrotic remodeling of lung and airways exposed to CS.