Shared and Divergent Stimulatory Effects of S100A4 and Uric Acid on Proliferation and Chemotaxis of Mesenchymal Stem Cells (MSCs). Interplay Between Necrotic Tumor Material and Immunosuppressive MSCs within Tumor Microenvironment,

Necrotic cell death is a characteristic feature of advanced solid tumor. S100 proteins and uric acid (UA) are released from necrotic (tumor) tissue regardless of tumor's origin. Both of these factors are known to influence immune response. Mesenchymal stem cells (MSCs) are often found within tumor microenvironment and are associated with poor prognosis of cancer patients in terms of metastasis and survival. MSCs seem to play a crucial role within tumor microenvironment probably due to their immunosuppressive capacity interfering with the specific anti-tumor immune response. Underlying mechanisms for MSC accumulation and stimulation in tumor tissue are not well characterized yet. S100A4 is known to promote metastasis in certain tumors such as colorectal cancer, but the exact mechanism of this effect still remains unclear. UA not only induces an acute inflammatory response but is also known to maintain chronic inflammation. In our previous experiments, we could already show that necrotic tumor material is capable of inducing chemotaxis and proliferation of MSCs. Focusing on S100A4 and UA, we now sought to characterize individual known factors within necrotic material which may be responsible for described effects.

Proliferation assays were performed using CYQUANT Assay (Invitrogen) based on fluorescent staining of nucleic acid. Migration of fluorescent-labeled MSCs was assessed using FluoroBlok (BD) two-chamber chemotaxis plates, fluorescence within the bottom chamber of migration plates was measured with PolarStar plate reader (BMG).

We tested the influence of recombinant human S100A4 at concentrations between 50 and 1000 ng/ml on MSC proliferation and could demonstrate divergent effects depending on the chosen S100A4 concentration: S100A4 at concentrations not higher than 100 ng/ml enhanced the proliferation of MSCs in a dose dependent manner up to 50%, whereas concentrations above 100 ng/ml inhibited MSC proliferation down to 50% compared to medium containing 5% human serum without S100A4. We tested the chemotactic activity of recombinant human S100A4 at concentrations between 0.01 and 1000 ng/ml on MSCs and could demonstrate a dose dependent chemotactic activity up to 80% of the response which was achieved by positive control (50% FBS). By adding UA to suboptimal concentration of S100A4 (100ng/ml), we could enhance the chemotactic activity of S100A4 on MSCs. In contrast to its enhancing effect on MSC migration, UA inhibited dose dependently the proliferation inducing effect of S100A4 with strongest effect at 300μg UA/ml resulting in about 50% inhibition of proliferation compared to medium containing 100ng/ml S100A4 without additional UA. ELISA assays confirmed the presence of S100A4 in the necrotic tumor material which we had used in our previous experiments with similar stimulatory effects on MSCs in terms of proliferation and chemotaxis.

MSCs are often found within tumor tissue and may influence the biologic behavior of tumor and host's immune response to tumor. We could characterize UA and S100A4 as bioactive factors within necrotic (tumor) material. Whereas S100A4 stimulated both proliferation as well as chemotaxis of MSCs, UA enhanced the chemotactic acitivity of S100A4 while inhibiting its proliferation inducing effect. Comparing our new data with our previous observations using tumor lysates to induce chemotaxis and proliferation of MSCs the overall effect of necrotic material seems to be rather proliferative and chemotactic even though some factors like UA may interfere with mentioned overall effects. Keeping in mind that MSCs can act as potent immunosuppressive cells, we conclude that tumor necrosis rather inhibits an effective anti-tumor immune response by favoring accumulation and proliferation of MSCs within tumor microenvironment. Our observations shed some light into the biology of MSCs within tumor microenvironment and open new questions concerning the interplay between MSCs, necrotic tumor cells and tumor progression. Possible strategies to break the described vicious circle by rather inducing an apoptotic tumor cell death or by reducing bioactive factors released from necrotic tissue within tumor microenvironment may be considered. Additionally, clinical investigations using specific antibodies to S100A4 or influencing serum concentrations of UA in tumor patients may be performed.

No relevant conflicts of interest to declare.