Myeloid-derived suppressor cells enhance IgE-mediated mast cell responses

Abstract

Mast cells and MDSCs are increased by parasitic infection and tumor growth. We previously demonstrated that enhanced MDSC development in ADAM10 transgenic mice yielded resistance to Nb infection and that coculturing MDSCs and mast cells enhanced cytokine production. In the current work, we show that MDSC-mast cell coculture selectively enhances IgE-mediated cytokine secretion among mast cells, without increasing MDSC cytokine production. This effect was independent of cell contact and elicited by Ly6C(+) and Ly6C/G+ MDSC subsets. These interactions were functionally important. MDSC depletion with the FDA-approved drug gemcitabine exacerbated Nb or Trichinella spiralis infection and reduced mast cell-dependent AHR and lung inflammation. Adoptive transfer of MDSC worsened AHR in WT but not mast cell-deficient Wsh/Wsh mice. These data support the hypothesis that MDSCs enhance mast cell inflammatory responses and demonstrate that this interaction can be altered by an existing chemotherapeutic.

Keywords: Nippostrongylus; Trichinella; allergy; asthma; inflammation.

Figure 1.. Mast cell-MDSC coculture enhances IgE-induced cytokine production.

MDSCs isolated from the spleens of 4T1 tumor-bearing BALB/c mice were cultured at a 1:1 ratio with BALB/c BMMCs that were presensitized with IgE. After overnight culture, cells were activated by antigen-induced IgE crosslinkage (XL). Supernatants were collected after 6 hours (TNF) or 18 h and analyzed by ELISA. Data are mean and sem from three to 34 measurements, obtained from two to four independent experiments. In the indicated samples, MDSCs and BMMCs were separated by 0.4 μm membranes (Transwells). *P < 0.05; **P < 0.01; ***P < 0.0001, when comparing the indicated samples with BMMCs or MDSCs alone.

Figure 2.. Enhanced cytokine production in mast cell-MDSC cocultures is restricted to mast cells.

MDSCs derived from the spleens of 4T1 tumor-bearing BALB/c mice were cocultured at a 1:1 ratio with IgE-presensitized BALB/c BMMCs. After overnight coculture, BMMCs were activated by IgE crosslinkage, and cells were assessed for intracellular IL-6 or TNF content by flow cytometry, as described in Materials and Methods. (A) Representative flow cytometry analyses after gating on Gr1+ MDSCs or Gr1− mast cells (MC). (B) Summary of percent-positive cells for each population. Data are averages of five to six samples/point. FL4/2-H, Fluorescence 4/2-height.

Figure 3.. The ability to enhance mast cell cytokine production is not restricted by Ly6C/G subsets.

Splenocytes from 4T1 tumor-bearing BALB/c mice were sorted based on Ly6C+ or Ly6C/G+ expression patterns. These cells were cocultured as described in Fig. 1. Data shown are mean and sem of six samples from one of two experiments that gave similar results. ND, Not detectable within the limits of this assay. *P < 0.05; **P < 0.01; ***P < 0.0001.

Figure 4.. MDSC depletion exacerbates Nb infection.

(A) BALB/c mice were inoculated with 650 infective Nb larvae and were subsequently treated with PBS or gemcitabine (1.2 mg i.p. injection; 200 μl vol) on Days 4 and 8 (indicated by arrowheads). Feces were collected daily and weighed and eggs counted and normalized/gram of feces. (B) Number of adult worms in the intestine on Day 11 postinoculation. (C) Flow cytometry of splenocytes on Day 11 for indicated surface markers. All data are mean and sem for five to six mice/group. *P < 0.05; **P < 0.01; ***P < 0.0001.

Figure 5.. MDSC depletion exacerbates T. spiralis infection.

(A) BALB/c mice were inoculated with T. spiralis muscle stage-infective larvae and then treated with gemcitabine (1.2 mg i.p. injection; 200 μl vol) or PBS on Days 1, 4, and 7 postinoculation before death on Day 9. Adult worms were counted in the intestine. (B) A second set of mice was treated as described in A, with gemcitabine injections continuing every 7 days, from Days 10 through 38 until T. spiralis larvae in muscle tissues were assessed on Day 41. Data shown represent the mean and sem from five mice. *P < 0.05; ***P < 0.0001.

Figure 6.. MDSC depletion reduces AHR.

BALB/c mice were sensitized and challenged with OVA, as described in Materials and Methods. After establishing inflammation with OVA challenge, i.n. gemcitabine treatment was given every 3 days from Days 47 through 75. OVA restimulation was performed on Days 66 through 75, with AHR and inflammation assessed on Day 76. (A) Airway resistance to methacholine challenge, as measured by Flexivent. (B) Trypan blue cell counts of cellular infiltrate recovered by BAL fluid (BALF). (C) Quantification of IgE levels in the serum by ELISA. Data shown represent the mean and sem from four to six mice/group. *P < 0.05; ***P < 0.0001.

Figure 7.. MDSC transfer exacerbates AHR and requires mast cells.

C57BL/6 or Wsh/Wsh mice were sensitized and challenged with OVA, as described in Materials and Methods. MDSCs (1×10⁷/mouse) were injected i.p. every 3 days from Days 18 through 27. Mice were challenged with OVA on Days 22–27. AHR was assessed on Day 28. Data shown represent the mean and sem from three to five mice/group. ΔR_L, change in lung resistance.