Lung fibroblasts from patients with emphysema show markers of senescence in vitro

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.