Induction of Reactive Bone Stromal Fibroblasts in 3D Models of Prostate Cancer Bone Metastases

Abstract

A dynamic interplay between prostate cancer (PCa) cells and reactive bone stroma modulates the growth of metastases within the bone microenvironment. Of the stromal cells, metastasis-associated fibroblasts (MAFs) are known to contribute but are the least studied cell type in PCa tumour progression. It is the aim of the current study to establish a biologically relevant 3D in vitro model that mimics the cellular and molecular profiles of MAFs found in vivo. Using 3D in vitro cell culture models, the bone-derived fibroblast cell line, HS-5, was treated with conditioned media from metastatic-derived PCa cell lines, PC3 and MDA-PCa 2b, or mouse-derived fibroblasts 3T3. Two corresponding reactive cell lines were propagated: HS5-PC3 and HS5-MDA, and evaluated for alterations in morphology, phenotype, cellular behaviour, plus protein and genomic profiles. HS5-PC3 and HS5-MDA displayed distinct alterations in expression levels of N-Cadherin, non-functional E-Cadherin, alpha-smooth muscle actin (α-SMA), Tenascin C, and vimentin, along with transforming growth factor receptor expression (TGF β R1 and R2), consistent with subpopulations of MAFs reported in vivo. Transcriptomic analysis revealed a reversion of HS5-PC3 towards a metastatic phenotype with an upregulation in pathways known to regulate cancer invasion, proliferation, and angiogenesis. The exploitation of these engineered 3D models could help further unravel the novel biology regulating metastatic growth and the role fibroblasts play in the colonisation process.

Extracellular Mechanical Stimuli Alters the Metastatic Progression of Prostate Cancer Cells within 3D Tissue Matrix

“…The interaction between metastatic cancer cells and the bone microenvironment plays a crucial role in driving cellular and molecular changes in tumor cells to promote their migration within the bone [6,7]. Efforts have been put into investigating interactions of metastatic cancer cells with the bone microenvironment [8][9][10].…”

“…Their results showed that smaller, poorly crystalline HA particles enhanced cancer cell adhesion and growth relative to larger, more crystalline HA particles, confirming HA particles' role in bone metastasis. Sieh et al [6] studied the interactions of osteoblast cells and CaP cells with polycaprolactonetricalcium phosphate (mPCL-TCP) scaffolds by wrapping mPCL-TCP scaffold within the osteoblast sheets to create a 3D matrix for CaP cell culture. CaP cells were then seeded on this 3D matrix to study CaP metastasis in a bone-like microenvironment, showing that CaP cell-bone matrix interactions lead to elevated levels of matrix metalloproteinase (MMPs) and prostate-specific antigens (PSAs), which are all associated with CaP metastasis.…”

“…However, without any exceptions, these studies and many others in the fields ignore the mechanical loading exerted on bone tissue due to the diurnal activities, and [22][23][24] increased constant pressure as a result of growing tumor [3,4]. For instance, metastasized cancer cells are exposed to a myriad of extracellular factors, including increased mechanical stimuli due to tumor growth within a confined space, matrix stiffness via stromal activation [5][6][7], interstitial fluid pressure due to ineffective vascularization, and production of soluble factors from cell-cell and cell-matrix interactions [1,[25][26][27][28]. While the effects of matrix stiffness, interstitial fluid pressure, and soluble factors on cancer metastasis have been extensively studied [29][30][31], the influence of extracellular mechanical stimuli upon cancer cell migration within 3D matrix and establishment of new metastases remains relatively unexplored.…”