Abstract
The incidence of pancreatic cancer increases with age, suggesting that chronological aging is a significant risk factor for this disease. Fibroblasts are the major nonmalignant cell type in the stroma of human pancreatic ductal adenocarcinoma (PDAC). In this study, we investigated whether the chronological aging of normal human fibroblasts (NHFs), a previously underappreciated area in pancreatic cancer research, influences the progression and therapeutic outcomes of PDAC. Results from experiments with murine xenografts and 2D and 3D co-cultures of NHFs and PDAC cells revealed that older NHFs stimulate proliferation of and confer resistance to radiation therapy of PDAC. MS-based metabolite analysis indicated that older NHFs have significantly increased arachidonic acid 12-lipoxygenase (ALOX12) expression and elevated levels of its mitogenic metabolite, 12-(S)-hydroxy-5,8,10,14-eicosatetraenoic acid (12-(S)-HETE) compared with their younger counterparts. In co-cultures with older rather than with younger NHFs, PDAC cells exhibited increases in mitogen-activated protein kinase signaling and cellular metabolism, as well as a lower oxidation state that correlated with their enhanced proliferation and resistance to radiation therapy. Expression of ALOX12 was found to be significantly lower in PDAC cell lines and tumor biopsies, suggesting that PDAC cells rely on a stromal supply of mitogens for their proliferative needs. Pharmacological (hydroxytyrosol) and molecular (siRNA) interventions of ALOX12 in older NHFs suppressed their ability to stimulate proliferation of PDAC cells. We conclude that chronological aging of NHFs contributes to PDAC progression and that ALOX12 and 12-(S)-HETE may be potential stromal targets for interventions that seek to halt progression and improve therapy outcomes.
Highlights
The incidence of pancreatic cancer increases with age; median age at diagnosis is 71 y and the median age at death is 73 y, suggesting that chronological aging is a significant risk factor for pancreatic cancer
Results showed a significant increase in the percentage of MIA PaCa-2 cells in un-irradiated control co-cultures of 12-y and 61-y compared to 3-d normal human fibroblasts (NHFs); approximately, 15 and 30% MIA PaCa-2 cells in co-cultures of 12-y and 61-y NHFs compared to 5% MIA PaCa-2 cells in co-cultures of 3-d NHFs (Figure 4A)
Flow cytometry analysis of radiation induced toxicity of MIA PaCa-2 cells in co-cultures of NHFs showed maximal cell death in co-cultures of 3-d compared to 12-y and 61-y NHFs (Figure 4B). These results showed that quiescent cultures of NHFs from donors of older compared to younger healthy individuals stimulate proliferation and induce radiation therapy resistance of pancreatic ductal adenocarcinoma (PDAC) cells (Figures 1-4)
Summary
The incidence of pancreatic cancer increases with age; median age at diagnosis is 71 y and the median age at death is 73 y, suggesting that chronological aging is a significant risk factor for pancreatic cancer. The stroma of PDAC includes fibroblasts, myofibroblasts, stellate cells, immune cells, and endothelial cells [8]. Fibroblasts are the major non-cancerous cell type contributing to the cellularity of the stroma that include synthesis and remodeling of the extracellular matrix as well as the production of secretory and non-secretory factors that influence cancer cell proliferation and progression to end stage disease. Because age is a significant risk factor for PDAC, it is important to know whether the age of the stroma influences progression of PDAC and the mechanisms involved in this process
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