Cancer-associated mesenchymal stroma fosters the stemness of osteosarcoma cells in response to intratumoral acidosis via NF-κB activation

Abstract

The role of mesenchymal stem cells (MSC) in osteosarcoma (OS), the most common primary tumor of bone, has not been extensively elucidated. We have recently shown that OS is characterized by interstitial acidosis, a microenvironmental condition that is similar to a wound setting, in which mesenchymal reactive cells are activated to release mitogenic and chemotactic factors. We therefore intended to test the hypothesis that, in OS, acid-activated MSC influence tumor cell behavior. Conditioned media or co-culture with normal MSC previously incubated with short-term acidosis (pH 6.8 for 10 hours, H+-MSC) enhanced OS clonogenicity and invasion. This effect was mediated by NF-κB pathway activation. In fact, deep-sequencing analysis, confirmed by Real-Time PCR and ELISA, demonstrated that H+-MSC differentially induced a tissue remodeling phenotype with increased expression of RelA, RelB, and NF-κB1, and downstream, of CSF2/GM-CSF, CSF3/G-CSF, and BMP2 colony-promoting factors, and of chemokines (CCL5, CXCL5, and CXCL1), and cytokines (IL6 and IL8), with an increased expression of CXCR4. An increased expression of IL6 and IL8 were found only in normal stromal cells, but not in OS cells, and this was confirmed in tumor-associated stromal cells isolated from OS tissue. Finally, H+-MSC conditioned medium differentially promoted OS stemness (sarcosphere number, stem-associated gene expression), and chemoresistance also via IL6 secretion. Our data support the hypothesis that the acidic OS microenvironment is a key factor for MSC activation, in turn promoting the secretion of paracrine factors that influence tumor behavior, a mechanism that holds the potential for future therapeutic interventions aimed to target OS.

RhoA knockout fibroblasts lose tumor-inhibitory capacity in vitro and promote tumor growth in vivo

“…An aspect here that remains elusive to the scope of the present study is how the loss of RhoA (which alters the mechanical and biochemical properties of normal stromal fibroblasts) can trigger a stem-like phenotype in these PC3 prostate cancer cells. Probably, increased level of proinflammatory genes plays role in inducing the expression of stemness-related properties of tumor cells (38,39). We have demonstrated that loss of RhoA changed the cytoskeleton, the contractile forces and cell stiffness of the cells, induced a proinflammatory state, and interfered with Rho signaling cascades.…”

The contribution of immune infiltrates and the local microenvironment in the pathogenesis of osteosarcoma

“…Mesenchymal stem cells, the low differentiated precursor of various cell-types including osteoblast [16], in contrast to differentiated osteoblasts appear unable to communicate with osteosarcoma cells through the establishment of functional gap junctions [50]. Whatever, several groups have demonstrated that extracellular vesicles represent an important way of communication between mesenchymal stem cells and osteosarcoma cells [51,52]. Under stress environment such as local acidosis associated to tumor growth and peritumoral osteolysis, mesenchymal stem cells could secrete extracellular vesicles carrying proteins, mRNA and microRNA modulating osteosarcoma cell proliferation and stemness [14,51,52].…”

“…Whatever, several groups have demonstrated that extracellular vesicles represent an important way of communication between mesenchymal stem cells and osteosarcoma cells [51,52]. Under stress environment such as local acidosis associated to tumor growth and peritumoral osteolysis, mesenchymal stem cells could secrete extracellular vesicles carrying proteins, mRNA and microRNA modulating osteosarcoma cell proliferation and stemness [14,51,52]. Acquired stemness properties (e.g.…”

“…Acquired stemness properties (e.g. ability to form sarcospheres, expression of stem-associated genes such as Nanog and Oct4) by osteosarcoma cells appeared mediated by the secretion of IL-6 by mesenchymal stem cells [52] [53]. The dialog between osteosarcoma cells and mesenchymal stem cells is bilateral, and mesenchymal stem cells can be educated by tumor-secreted extracellular vesicles [54].…”

Priming of MSCs with inflammation-relevant signals affects extracellular vesicle biogenesis, surface markers, and modulation of T cell subsets

“…MSC-EV have been shown to associate with TGF-β, and EVs may bind TGF-β on their surface (39,40). Acidosis primed MSCs upregulated osteosarcoma expression of CXCL5 and CCL5 (41). These chemokines have also been implicated in the formation and recruitment of Tregs, respectively (42)(43)(44).…”