Müller, K. C.; Welker, L.; Paasch, K.; Feindt, B.; Erpenbeck, V.; Hohlfeld, J.; Krug, N.; Nakashima, M.; Branscheid, D.; Magnussen, H.; Jorres, R. A.; Holz, O.:
Lung fibroblasts from patients with emphysema show markers of senescence in vitro.
In: Respiratory Research
Background: The loss of alveolar walls is a hallmark of emphysema. As fibroblasts play an important role in the maintenance of alveolar structure, a change in fibroblast phenotype could be involved in the pathogenesis of this disease. In a previous study we found a reduced in vitro proliferation rate and number of population doublings of parenchymal lung fibroblasts from patients with emphysema and we hypothesized that these findings could be related to a premature cellular aging of these cells. In this study, we therefore compared cellular senescence markers and expression of respective genes between lung fibroblasts from patients with emphysema and control patients without COPD. Methods: Primary lung fibroblasts were obtained from 13 patients with moderate to severe lung emphysema ( E) and 15 controls ( C) undergoing surgery for lung tumor resection or volume reduction ( n = 2). Fibroblasts (8E/9C) were stained for senescence- associated beta-galactosidase (SA-beta-Gal). In independent cultures, DNA from lung fibroblasts (7E/8C) was assessed for mean telomere length. Two exploratory 12 k cDNA microarrays were used to assess gene expression in pooled fibroblasts (3E/ 3C). Subsequently, expression of selected genes was evaluated by quantitative PCR (qPCR) in fibroblasts of individual patients (10E/9C) and protein concentration was analyzed in the cell culture supernatant. Results: The median ( quartiles) percentage of fibroblasts positive for SA-beta-Gal was 4.4 (3.2; 4.7) % in controls and 16.0 (10.0; 24.8) % in emphysema ( p = 0.001), while telomere length was not different. Among the candidates for differentially expressed genes in the array ( factor = 3), 15 were upregulated and 121 downregulated in emphysema. qPCR confirmed the upregulation of insulin-like growth factor-binding protein ( IGFBP)-3 and IGFBP-rP1 ( p = 0.029, p = 0.0002), while expression of IGFBP-5, - rP2 ( CTGF), - rP4 (Cyr61), FOSL1, LOXL2, OAZ1 and CDK4 was not different between groups. In line with the gene expression we found increased cell culture supernatant concentrations of IGFBP-3 ( p = 0.006) in emphysema. Conclusion: These data support the hypothesis that premature aging of lung fibroblasts occurs in emphysema, via a telomere-independent mechanism. The upregulation of the senescence- associated IGFBP-3 and - rP1 in emphysema suggests that inhibition of the action of insulin and insulin-like growth factors could be involved in the reduced in vitro-proliferation rate.