Aging of stromal-derived human breast fibroblasts might contribute to breast cancer progression.

Age is an important factor in the development and spread of breast cancer. Stromal cells also contribute to breast cancer growth and metastasis through the production of extracellular matrix (ECM) modifiers such as urokinase type plasminogen activator (uPA), its receptor (uPAR), its inhibitors (PAI-1 and PAI-2), matrix metalloproteinases (MMPs), and growth factors, including the fibroblast and insulin-like growth factors (FGF's and IGF's). In the present study we have investigated whether breast fibroblasts aged in vitro through passage in culture display altered levels of the plasminogen activator system and growth factors that are known to modulate that system. With real-time RT-PCR we found that during passage human breast fibroblasts, whether derived from the tumour burden or from matched adjacent normal breast tissue, exhibited a consistent increase in PAI-1 and FGF-1 and a decrease in MMP-2 mRNA expression. In addition, in 5 out of 7 fibroblast strains we observed an induction of uPA expression in combination with a reduced IGF-1 expression. Interestingly, while during aging MMP-2 protein increased in all tumour-derived fibroblast strains, these protein levels were reduced in all normal tissue- derived fibroblasts. No other clear-cut age-dependent alterations were found in the all-together 25 factors investigated. We furthermore demonstrate in one tumour-derived fibroblast strain that the increases in uPA and PAI-1 mRNA and MMP-2 protein production are inversely related to the telomere length. Artificially increasing telomere length in this fibroblast strain by expressing human telomerase reverse transcriptase (hTERT) prevented senescence and resulted in late passage cultures with early passage uPA, PAI-1 and MMP-2 levels. Our results show that aging accompanied by telomere loss induces PAI-1 and FGF-1 mRNA expression in all breast fibroblast strains, increases uPA and decreases IGF-1 mRNA expression in a subset, and increases MMP-2 protein expression only in tumour-derived breast fibroblasts. These age-induced levels of PAI-1, FGF-1, uPA and MMP-2 in stromal breast fibroblast could contribute to breast cancer progression.