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TNF Neutralization Results in the Delay of Transplantable Tumor Growth and Reduced MDSC Accumulation.

Atretkhany KS, Nosenko MA, Gogoleva VS, Zvartsev RV, Qin Z, Nedospasov SA, Drutskaya MS - Front Immunol (2016)

Bottom Line: Under the influence of inflammatory cytokines, these cells become MDSCs, acquire immunosuppressive phenotype, and accumulate in the affected tissue, as well as in the periphery.TNF is a critical factor for the induction, expansion, and suppressive activity of MDSCs.Both etanercept and infliximab treatments resulted in a delayed growth of MCA 205 fibrosarcoma in hTNF KI mice, significantly reduced tumor volume, and also resulted in less accumulated MDSCs in the blood 3 weeks after tumor cell inoculation.

View Article: PubMed Central - PubMed

Affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia; Immunology Department, Faculty of Biology, Beloszersky Institue of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, Russia.

ABSTRACT
Myeloid-derived suppressor cells (MDSCs) represent a heterogeneous population of immature myeloid cells (IMCs) that, under normal conditions, may differentiate into mature macrophages, granulocytes, and dendritic cells. However, under pathological conditions associated with inflammation, cancer, or infection, such differentiation is inhibited leading to IMC expansion. Under the influence of inflammatory cytokines, these cells become MDSCs, acquire immunosuppressive phenotype, and accumulate in the affected tissue, as well as in the periphery. Immune suppressive activity of MDSCs is partly due to upregulation of arginase 1, inducible nitric oxide synthase, and anti-inflammatory cytokines, such as IL-10 and TGF-β. These suppressive factors can enhance tumor growth by repressing T-cell-mediated anti-tumor responses. TNF is a critical factor for the induction, expansion, and suppressive activity of MDSCs. In this study, we evaluated the effects of systemic TNF ablation on tumor-induced expansion of MDSCs in vivo using TNF humanized (hTNF KI) mice. Both etanercept and infliximab treatments resulted in a delayed growth of MCA 205 fibrosarcoma in hTNF KI mice, significantly reduced tumor volume, and also resulted in less accumulated MDSCs in the blood 3 weeks after tumor cell inoculation. Thus, our study uncovers anti-tumor effects of systemic TNF ablation in vivo.

No MeSH data available.


Related in: MedlinePlus

Infliximab or etanercept efficiently inhibit suppressive functions of MDSCs. (A) Blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with etanercept (red) or infliximab (blue) have reduced levels of NO compared to PBS-treated mice (black). Data are representative of two independent experiments. (B) Purified splenic MDSCs from tumor-bearing hTNF KI mice undergoing systemic TNF ablation with etanercept (red) or infliximab (blue) fail to suppress T-cell proliferation. MDSCs and T-cells were co-cultured at ratio 5:1, 2 × 106 and 4 × 105 cells, respectively. (C)In vitro proliferation of purified CFSE-labeled T-cells from the spleens of naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies or (D) unstimulated T-cells. (E) CFSE-labeled T-cells from naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies in the presence of purified splenic MDSCs from tumor-bearing mice treated with PBS (left), etanercept (middle), or infliximab (right). Pink and red peaks were defined as non-proliferating and proliferating cells, respectively. Cells were gated as VD−CD4+. Data are representative of three independent experiments with each group represented by six mice, and MDSCs were isolated from pooled splenocytes from two to three mice. *p < 0.5 and **p < 0.01.
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Figure 4: Infliximab or etanercept efficiently inhibit suppressive functions of MDSCs. (A) Blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with etanercept (red) or infliximab (blue) have reduced levels of NO compared to PBS-treated mice (black). Data are representative of two independent experiments. (B) Purified splenic MDSCs from tumor-bearing hTNF KI mice undergoing systemic TNF ablation with etanercept (red) or infliximab (blue) fail to suppress T-cell proliferation. MDSCs and T-cells were co-cultured at ratio 5:1, 2 × 106 and 4 × 105 cells, respectively. (C)In vitro proliferation of purified CFSE-labeled T-cells from the spleens of naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies or (D) unstimulated T-cells. (E) CFSE-labeled T-cells from naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies in the presence of purified splenic MDSCs from tumor-bearing mice treated with PBS (left), etanercept (middle), or infliximab (right). Pink and red peaks were defined as non-proliferating and proliferating cells, respectively. Cells were gated as VD−CD4+. Data are representative of three independent experiments with each group represented by six mice, and MDSCs were isolated from pooled splenocytes from two to three mice. *p < 0.5 and **p < 0.01.

Mentions: To study the impact of systemic TNF ablation on the functional properties of MDSCs, we examined ROS and NO production in vivo by blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with TNF inhibitors. We detected a significant reduction in NO in CD11b+Ly6C+ myeloid cells in the blood of both etanercept- and infliximab-treated mice as compared to PBS-treated hTNF KI tumor-bearing mice (Figure 4A), whereas significant difference in ROS production was only detected in etanercept-treated tumor-bearing hTNF KI mice (Figure S2B in Supplementary Material). Furthermore, to address possible impact of anti-TNF treatment on MDSC functions, we evaluated suppressive activity of myeloid cells on T-cell proliferation using a co-culture of CFSE-labeled T-cells and purified MDSCs in the presence of agonistic anti-CD3 and anti-CD28 antibodies. MDSCs were isolated from the spleens of hTNF KI tumor-bearing mice undergoing treatment with anti-TNF blockers or with PBS, as control. After 3 days of co-culture, we determined the label distribution in the population of CD4+ T-cells (Figure 4B). As expected, purified MDSCs isolated from PBS-treated tumor-bearing mice completely suppressed T-cell proliferation (Figures 4B,E, left). Strikingly, MDSCs isolated from tumor-bearing mice under infliximab or etanercept treatment were not able to prevent T-cell proliferation indicating that their suppressive function was compromised (Figures 4B,E, middle and right). Unstimulated and stimulated T-cells in the absence of MDSCs are shown as control stainings in Figures 4C,D). We also analyzed gene expression and evaluated their suppressive activity ex vivo. For this, we isolated MDSCs from the spleens of wild-type tumor-bearing mice after etanercept administration using magnetic bead separation (Figure S4A in Supplementary Material). We then performed gene expression analysis to see how anti-TNF treatment may affect the transcriptome of MDSCs (Figure S4B in Supplementary Material). We specifically looked for genes that encoded factors known to play an important role in MDSC function and found that at least some of them were indeed affected by systemic anti-TNF treatment. In particular, we observed significant reduction in the expression of il10, adam17, and tgfb, while the expression of arginase-1 gene was increased (Figure S4B in Supplementary Material). Taken together, our data demonstrated that systemic TNF inhibitors may not only reduce MDSC numbers but also affect their suppressive function.


TNF Neutralization Results in the Delay of Transplantable Tumor Growth and Reduced MDSC Accumulation.

Atretkhany KS, Nosenko MA, Gogoleva VS, Zvartsev RV, Qin Z, Nedospasov SA, Drutskaya MS - Front Immunol (2016)

Infliximab or etanercept efficiently inhibit suppressive functions of MDSCs. (A) Blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with etanercept (red) or infliximab (blue) have reduced levels of NO compared to PBS-treated mice (black). Data are representative of two independent experiments. (B) Purified splenic MDSCs from tumor-bearing hTNF KI mice undergoing systemic TNF ablation with etanercept (red) or infliximab (blue) fail to suppress T-cell proliferation. MDSCs and T-cells were co-cultured at ratio 5:1, 2 × 106 and 4 × 105 cells, respectively. (C)In vitro proliferation of purified CFSE-labeled T-cells from the spleens of naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies or (D) unstimulated T-cells. (E) CFSE-labeled T-cells from naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies in the presence of purified splenic MDSCs from tumor-bearing mice treated with PBS (left), etanercept (middle), or infliximab (right). Pink and red peaks were defined as non-proliferating and proliferating cells, respectively. Cells were gated as VD−CD4+. Data are representative of three independent experiments with each group represented by six mice, and MDSCs were isolated from pooled splenocytes from two to three mice. *p < 0.5 and **p < 0.01.
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Figure 4: Infliximab or etanercept efficiently inhibit suppressive functions of MDSCs. (A) Blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with etanercept (red) or infliximab (blue) have reduced levels of NO compared to PBS-treated mice (black). Data are representative of two independent experiments. (B) Purified splenic MDSCs from tumor-bearing hTNF KI mice undergoing systemic TNF ablation with etanercept (red) or infliximab (blue) fail to suppress T-cell proliferation. MDSCs and T-cells were co-cultured at ratio 5:1, 2 × 106 and 4 × 105 cells, respectively. (C)In vitro proliferation of purified CFSE-labeled T-cells from the spleens of naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies or (D) unstimulated T-cells. (E) CFSE-labeled T-cells from naive mice after three days of stimulation with anti-CD3 and anti-CD28 antibodies in the presence of purified splenic MDSCs from tumor-bearing mice treated with PBS (left), etanercept (middle), or infliximab (right). Pink and red peaks were defined as non-proliferating and proliferating cells, respectively. Cells were gated as VD−CD4+. Data are representative of three independent experiments with each group represented by six mice, and MDSCs were isolated from pooled splenocytes from two to three mice. *p < 0.5 and **p < 0.01.
Mentions: To study the impact of systemic TNF ablation on the functional properties of MDSCs, we examined ROS and NO production in vivo by blood MDSCs of tumor-bearing hTNF KI mice undergoing treatment with TNF inhibitors. We detected a significant reduction in NO in CD11b+Ly6C+ myeloid cells in the blood of both etanercept- and infliximab-treated mice as compared to PBS-treated hTNF KI tumor-bearing mice (Figure 4A), whereas significant difference in ROS production was only detected in etanercept-treated tumor-bearing hTNF KI mice (Figure S2B in Supplementary Material). Furthermore, to address possible impact of anti-TNF treatment on MDSC functions, we evaluated suppressive activity of myeloid cells on T-cell proliferation using a co-culture of CFSE-labeled T-cells and purified MDSCs in the presence of agonistic anti-CD3 and anti-CD28 antibodies. MDSCs were isolated from the spleens of hTNF KI tumor-bearing mice undergoing treatment with anti-TNF blockers or with PBS, as control. After 3 days of co-culture, we determined the label distribution in the population of CD4+ T-cells (Figure 4B). As expected, purified MDSCs isolated from PBS-treated tumor-bearing mice completely suppressed T-cell proliferation (Figures 4B,E, left). Strikingly, MDSCs isolated from tumor-bearing mice under infliximab or etanercept treatment were not able to prevent T-cell proliferation indicating that their suppressive function was compromised (Figures 4B,E, middle and right). Unstimulated and stimulated T-cells in the absence of MDSCs are shown as control stainings in Figures 4C,D). We also analyzed gene expression and evaluated their suppressive activity ex vivo. For this, we isolated MDSCs from the spleens of wild-type tumor-bearing mice after etanercept administration using magnetic bead separation (Figure S4A in Supplementary Material). We then performed gene expression analysis to see how anti-TNF treatment may affect the transcriptome of MDSCs (Figure S4B in Supplementary Material). We specifically looked for genes that encoded factors known to play an important role in MDSC function and found that at least some of them were indeed affected by systemic anti-TNF treatment. In particular, we observed significant reduction in the expression of il10, adam17, and tgfb, while the expression of arginase-1 gene was increased (Figure S4B in Supplementary Material). Taken together, our data demonstrated that systemic TNF inhibitors may not only reduce MDSC numbers but also affect their suppressive function.

Bottom Line: Under the influence of inflammatory cytokines, these cells become MDSCs, acquire immunosuppressive phenotype, and accumulate in the affected tissue, as well as in the periphery.TNF is a critical factor for the induction, expansion, and suppressive activity of MDSCs.Both etanercept and infliximab treatments resulted in a delayed growth of MCA 205 fibrosarcoma in hTNF KI mice, significantly reduced tumor volume, and also resulted in less accumulated MDSCs in the blood 3 weeks after tumor cell inoculation.

View Article: PubMed Central - PubMed

Affiliation: Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow, Russia; Immunology Department, Faculty of Biology, Beloszersky Institue of Physico-Chemical Biology, Lomonosov Moscow State University, Moscow, Russia.

ABSTRACT
Myeloid-derived suppressor cells (MDSCs) represent a heterogeneous population of immature myeloid cells (IMCs) that, under normal conditions, may differentiate into mature macrophages, granulocytes, and dendritic cells. However, under pathological conditions associated with inflammation, cancer, or infection, such differentiation is inhibited leading to IMC expansion. Under the influence of inflammatory cytokines, these cells become MDSCs, acquire immunosuppressive phenotype, and accumulate in the affected tissue, as well as in the periphery. Immune suppressive activity of MDSCs is partly due to upregulation of arginase 1, inducible nitric oxide synthase, and anti-inflammatory cytokines, such as IL-10 and TGF-β. These suppressive factors can enhance tumor growth by repressing T-cell-mediated anti-tumor responses. TNF is a critical factor for the induction, expansion, and suppressive activity of MDSCs. In this study, we evaluated the effects of systemic TNF ablation on tumor-induced expansion of MDSCs in vivo using TNF humanized (hTNF KI) mice. Both etanercept and infliximab treatments resulted in a delayed growth of MCA 205 fibrosarcoma in hTNF KI mice, significantly reduced tumor volume, and also resulted in less accumulated MDSCs in the blood 3 weeks after tumor cell inoculation. Thus, our study uncovers anti-tumor effects of systemic TNF ablation in vivo.

No MeSH data available.


Related in: MedlinePlus