Obesity Triggers Enhanced MDSC Accumulation in Murine Renal Tumors via Elevated Local Production of CCL2

<div><p>Obesity is one of the leading risk factors for developing renal cell carcinoma, an immunogenic tumor that is treated clinically with immunostimulatory therapies. Currently, however, the mechanisms linking obesity with renal cancer incidence are unclear. Using a model of diet-induced obesity, we found that obese BALB/c mice with orthotopic renal tumors had increased total frequencies of myeloid-derived suppressor cells (MDSC) in renal tumors and spleens by d14 post-tumor challenge, relative to lean counterparts. Renal tumors from obese mice had elevated concentrations of the known myeloid cell chemoattractant CCL2, which was produced locally by increased percentages of dendritic cells, macrophages, B cells, and CD45^- cells in tumors. MDSC expression of the CCL2 receptor, CCR2, was unaltered by obesity but greater percentages of CCR2⁺ MDSCs were present in renal tumors from obese mice. Of note, the intracellular arginase levels and per-cell suppressive capacities of tumor-infiltrating and splenic MDSCs were unchanged in obese mice relative to lean controls. Thus, our findings suggest that obesity promotes renal tumor progression via development of a robust immunosuppressive environment that is characterized by heightened local and systemic MDSC prevalence. Targeted intervention of the CCL2/CCR2 pathway may facilitate immune-mediated renal tumor clearance in the obese.</p></div

Obese mice have elevated concentrations of the MDSC chemoattractant CCL2 in renal tumors.

<p>(A) DIO and NW mice were challenged as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118784#pone.0118784.g001" target="_blank">Fig. 1</a>. Tumor-bearing kidneys (TK) or tumor-free contralateral kidneys (CK) were harvested at the times indicated. Homogenates were tested via Multiplex array for CCL2. n = 4–8 mice per group, combined from three experiments. Bars indicate mean +/- sd. * = p < 0.05. (B) Histograms showing intracellular CCL2 for the indicated cell populations. (C) Intracellular CCL2 from pooled DIO versus NW mice for the indicated cell populations, with n = 3–5 mice per group. Bars indicate mean +/- sd. * = p < 0.05.</p

Obesity does not alter the suppressive capacity of MDSC.

<p>Tumor-bearing kidneys and spleens from the same mice were harvested between days 21–23. (A) Monocytic (Ly6C⁺) and granulocytic MDSC (Ly6G⁺) were sort-purified from each organ, then placed into culture with naive DUC18 T cells and stimulatory splenic DC from tumor-free NW mice. MDSC and stimulatory DC were pulsed with the DUC18 cognate peptide tERK. n = 6–8 mice per group, combined from three experiments. Bars indicate mean +/- sd. * = p < 0.05. TI = tumor-infiltrating: sp = splenic. (B) Bulk tumor and spleen homogenates were surface stained as described in Methods to distinguish monocytic versus granulocytic MDSC subpopulations, then stained intracellularly for arginase-1 expression. Dots indicate results from individual mice. No statistical differences were present in the percentages of arginase⁺ MDSC subpopulations from like organs in obese versus NW mice.</p

Equivalent surface expression of CCR2 on MDSCs from DIO and NW mice.

<p>(A) DIO and NW mice were challenged as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0118784#pone.0118784.g001" target="_blank">Fig. 1</a>. Tumor-bearing kidneys (TK) or tumor-free contralateral kidneys (CK) were harvested at the times indicated. Histograms show representative surface expression of CCR2 on Ly6C⁺ or Ly6G⁺ MDSC from DIO or NW mice. Open histograms = isotype control; shaded histograms = CCR2 staining. (B) Pooled data showing the % of MDSCs that express CCR2 in tumor-bearing kidneys (TK) and contralateral kidneys (CK). Gating on Ly6C⁺ or Ly6G⁺ MDSC shows strong and equivalent expression of CCR2 on MDSC subpopulations from DIO and NW mice. (C) Increased overall frequencies of CCR2⁺ MDSC from both Ly6C⁺ and Ly6G⁺ subsets when calculated as a percentage of total live cells within tumor-bearing kidneys. For B and C, n = 4–6 mice per group from two experiments. Bars indicate mean +/- sd. * = p < 0.05.</p