Abstract B35: Modeling interstitial fluid movement in the tumor microenvironment: A role in fibroblast activation?

Abstract

Abstract Introduction: The tissue dynamics of a growing tumor differ from normal tissue, with increased matrix stiffness and mechanical pressure causing alterations in interstitial fluid movement in the tumor microenvironment (TME). This change in fluid flow is known to affect the ability of chemotherapeutic drugs to penetrate tumors, but the effect of flow on the phenotype of cells of the tumor microenvironment has not been studied. The predominant cell type of the TME, the fibroblast, is known to become activated to a pro-tumorigenic phenotype upon exposure to factors such as TGF-β1 released by cancer cells, immune cells and also by mechanical forces. The effect of fluid flow on the fibroblast phenotype, however, remains undetermined. Objectives: We hypothesize that stromal fibroblasts are activated by interstitial fluid flow and thereby promote tumor growth. We aim to investigate this by studying the markers of fibroblast activation under mechanically relevant interstitial fluid flow conditions. Methods: Normal oral (NOF) and dermal (NHDF) fibroblasts were plated on collagen coated Thermonox cover slips, transferred into Quasi-vivo bioreactors and subjected to flow (150 μl/min) for 24 h. Expression of fibroblast activation markers, α-SMA and collagen IA1, were assessed by qPCR and microarray gene expression analysis was performed using RNA harvested from the cells. α-SMA and phospho-SMAD3 protein expression was studied using immunofluorescence and western blot. Results: Media flow altered gene expression in oral and skin fibroblasts, increasing expression of α-SMA by 5.2 fold (NHDF) and 2.6 fold (NOF) and collagen IA by 2.9 fold (NHDF) and 3.2 fold (NOF) when compared to cells in static culture. TGF-β1 stimulation in the presence of flow decreased expression of these markers, but increased them in static culture. In dermal fibroblasts, flow stimulated α-SMA protein expression and translocation of phosphorylated SMAD3 to the cell nucleus and enhanced the expression of caveolin-1 alpha, which regulates the TGF-β1 pathway. Dermal fibroblasts exposed to flow revealed significant variations in expression of 16348 genes that may regulate the TGF-β1 pathway. Conclusion: Flow activates fibroblasts to a similar extent as TGF-β1 stimulated static cultures. Concurrent TGF-β1 treatment with flow, however, does not cause fibroblast activation. It is possible that TGF-β1 stimulation of fibroblasts under flow leads to caveolin-mediated endocytosis of the receptor or negative regulation of SMAD intracellular signaling pathways resulting in decreased expression of activation markers. Regardless of their origin (oral or dermal), fibroblasts respond to even modest flow conditions. Our findings suggest caution should be used when extrapolating findings from static models to in vivo conditions. It further prompts exploration of novel anti-tumor strategies that alter the physical properties of the tumor microenvironment. Citation Format: Sadhvi Nithiananthan, Aileen Crawford, Daniel W. Lambert, Simon Whawell. Modeling interstitial fluid movement in the tumor microenvironment: A role in fibroblast activation? [abstract]. In: Proceedings of the AACR Special Conference: Function of Tumor Microenvironment in Cancer Progression; 2016 Jan 7–10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2016;76(15 Suppl):Abstract nr B35.