No abstract

Simple stress or necrotic cell death with subsequent release of damage-associated molecular patterns (DAMPs) is a characteristic feature of most advanced tumors. DAMPs within the tumor microenvironment stimulate tumor-associated cells, including dendritic cells and mesenchymal stromal cells (MSCs). The presence of tumor-infiltrating MSCs is associated with tumor progression and metastasis. Oxidized necrotic material loses its stimulatory capacity for MSCs. As a DAMP, S100A4 is sensitive to oxidation whereas uric acid (UA) acts primarily as an antioxidant. We tested these two biologic moieties separately and in combination for their activity on MSCs. Similar to necrotic tumor material, S100A4 and UA both dose-dependently induced chemotaxis of MSCs with synergistic effects when combined. Substituting for UA, alternative antioxidants (vitamin C, DTT, and N-acetylcysteine) also enhanced the chemotactic activity of S100A4 in a synergistic manner. This emphasizes the reducing potential of UA being, at least in part, responsible for the observed synergy. With regard to MSC proliferation, both S100A4 and UA inhibited MSCs without altering survival or inducing differentiation toward adipo-, osteo-, or chondrocytes. In the presence of S100A4 or UA, MSCs gained an immunosuppressive capability and stably induced IL-10– and IDO-expressing lymphocytes that maintained their phenotype following proliferation. We have thus demonstrated that both S100A4 and UA act as DAMPs and, as such, may play a critical role in promoting some aspects of MSC-associated immunoregulation. Our findings have implications for therapeutic approaches targeting the tumor microenvironment and addressing the immunosuppressive nature of unscheduled cell death within the tumor microenvironment.

Chronic inflammatory response is found in most advanced solid tumors with subsequent necrosis associated with release of various damage-associated molecular patterns (DAMPs) including S100A4. Mesenchymal stromal cells (MSCs) seem to interfere with the anti-tumor-immune response resulting in a poor prognosis for cancer patients, given the immunosuppressive capacity of these cells. In previous experiments we already showed that S100A4 induces MSC proliferation with an optimum at 100ng/ml. We now tested higher S100A4 concentrations and observed a bimodal effect with a second optimum achieved when using more than 2000ng S100A4/ml. Furthermore, stimulation of MSCs with graded concentrations of S100A4 inhibited dose-dependently the proliferation of lymphocytes. Based on our previous observations that S100A4 activity is enhanced in the presence of uric acid, we now tested further reducing agents like ascorbic acid, dithiothreitole, and N-acetylcysteine and could demonstrate increased chemotactic activity of S100A4 on MSCs when mentioned antioxidants were present, while they had minimal - if any - effect by themselves. Considering that MSCs respond to necrosis-associated factors, we conclude that creating an oxidative environment could be one possible strategy to inactivate not only S100A4 but also further oxidation-sensitive DAMPs like HMGB1, which contribute to the immunosuppressive nature of tumor microenvironment.

Necrosis with subsequent release of damage-associated molecular patterns (DAMPs) is a characteristic feature of advanced solid tumors. DAMPs impact the tumor-microenvironment stimulating tumor-associated cells like mesenchymal stromal cells (MSCs). Tumor-infiltrating-MSCs are associated with tumor progression and metastasis. Oxidized DAMPs lose their stimulatory capacity on MSCs. As a DAMP-member S100A4 is sensitive to oxidation while uric acid (UA) acts as an antioxidant. We tested these two DAMP-members separately and in combination for their biologic activity on MSCs. S100A4 and UA showed a dose-dependent chemotactic activity on MSCs with synergistic effects when both DAMPs were combined. Substituting for UA, alternative antioxidants (Vitamin-C, dithiothreitol, and acetylcystein) had also a comparable synergistic effect on the chemotactic activity of S100A4, emphasizing the reducing potential of UA being responsible for the observed synergy. Yet, S100A4 and UA inhibited MSC proliferation without impacting their viability. Nevertheless, in the presence of S100A4 or UA, MSCs gained immunosuppressive capacities by inducing IL-10 and indoleamine dioxygenase expressing lymphocytes. We characterized S100A4 and UA as necrosis-associated factors playing a crucial role within MSC biology and thus immunoregulation. Our results have implications for therapeutic approaches targeting the tumor microenvironment and addressing the immunosuppressive nature of necrotic tumor.