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Inhibition of breast cancer resistance protein (ABCG2) in human myeloid dendritic cells induces potent tolerogenic functions during LPS stimulation.

Jin JO, Zhang W, Wong KW, Kwak M, van Driel IR, Yu Q - PLoS ONE (2014)

Bottom Line: Moreover, inhibition of ABCG2 in monocyte-derived DCs (MDDCs) abrogated the up-regulation of co-stimulatory molecules and production of pro-inflammatory cytokines in these cells in response to LPS.Furthermore, CD1c+ mDCs stimulated with LPS plus Ko143 inhibited the proliferation of allogeneic and superantigen-specific syngenic CD4+ T cells and promoted expansion of CD25+FOXP3+ regulatory T (Treg) cells in an IL-10-dependent fashion.These tolerogenic effects of ABCG2 inhibition could be abolished by ERK inhibition.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Public Health Clinical Center, Shanghai Medical College, Fudan University, Shanghai, China.

ABSTRACT
Breast cancer resistance protein (ABCG2), a member of the ATP-binding cassette transporters has been identified as a major determinant of multidrug resistance (MDR) in cancer cells, but ABC transporter inhibition has limited therapeutic value in vivo. In this research, we demonstrated that inhibition of efflux transporters ABCG2 induced the generation of tolerogenic DCs from human peripheral blood myeloid DCs (mDCs). ABCG2 expression was present in mDCs and was further increased by LPS stimulation. Treatment of CD1c+ mDCs with an ABCG2 inhibitor, Ko143, during LPS stimulation caused increased production of IL-10 and decreased production of pro-inflammatory cytokines and decreased expression of CD83 and CD86. Moreover, inhibition of ABCG2 in monocyte-derived DCs (MDDCs) abrogated the up-regulation of co-stimulatory molecules and production of pro-inflammatory cytokines in these cells in response to LPS. Furthermore, CD1c+ mDCs stimulated with LPS plus Ko143 inhibited the proliferation of allogeneic and superantigen-specific syngenic CD4+ T cells and promoted expansion of CD25+FOXP3+ regulatory T (Treg) cells in an IL-10-dependent fashion. These tolerogenic effects of ABCG2 inhibition could be abolished by ERK inhibition. Thus, we demonstrated that inhibition of ABCG2 in LPS-stimulated mDCs can potently induce tolerogenic potentials in these cells, providing crucial new information that could lead to development of better strategies to combat MDR cancer.

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LPS plus Ko143-treated mDCs promote expansion of Treg cells.Ko143-, LPS- or LPS plus Ko143-treated CD1c+ mDCs were co-cultured with CFSE-labeled allogeneic CD4+ T cells (1×105) in a 1∶10 ratio for 4 days. (A) Surface expression of CD25 and CFSE dilution was analyzed in CD4+ T cells by flow cytometry. (B) Flow cytometry of CD4, CD25 and FOXP3 expression was shown. Data are representative of analyses of 3 samples from 3 donors. (C) Flow cytometry of intracellular IL-10 and IFN-γ production in gated CD4+ T cells was shown. (D) Intracellular IL-10 expression in CD4+FOXP3+ T cells (left panel) and mean percentage of IL-10+FOXP3+ or IL-10+FOXP3− cells (right panel) were shown. Data are representative or the average of analyses of 3 samples from 3 donors. (E) CFSE-labeled PBMCs were incubated with soluble anti-CD3 and CD28 Abs for 4 days in the presence or absence of CD25+ Treg cells. CFSE dilution was analyzed in CD4+CD25− and CD8+ T cells (left panel) and mean percentage of proliferating cells (right panel) was shown. Data are representative or the average of analyses of 4 samples from 4 donors. (F) CD1c+ mDCs were cultured with LPS and CFSE-labeled syngenic CD4 T cells, in the presence or absence of Staphylococcal Enterotoxin B (SEB) and Ko143 for 4 days. CFSE dilution was analyzed in CD4+TCRVβ3+ T cells by flow cytometry. Data are representative of analyses of 3 samples from 3 donors.
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pone-0104753-g004: LPS plus Ko143-treated mDCs promote expansion of Treg cells.Ko143-, LPS- or LPS plus Ko143-treated CD1c+ mDCs were co-cultured with CFSE-labeled allogeneic CD4+ T cells (1×105) in a 1∶10 ratio for 4 days. (A) Surface expression of CD25 and CFSE dilution was analyzed in CD4+ T cells by flow cytometry. (B) Flow cytometry of CD4, CD25 and FOXP3 expression was shown. Data are representative of analyses of 3 samples from 3 donors. (C) Flow cytometry of intracellular IL-10 and IFN-γ production in gated CD4+ T cells was shown. (D) Intracellular IL-10 expression in CD4+FOXP3+ T cells (left panel) and mean percentage of IL-10+FOXP3+ or IL-10+FOXP3− cells (right panel) were shown. Data are representative or the average of analyses of 3 samples from 3 donors. (E) CFSE-labeled PBMCs were incubated with soluble anti-CD3 and CD28 Abs for 4 days in the presence or absence of CD25+ Treg cells. CFSE dilution was analyzed in CD4+CD25− and CD8+ T cells (left panel) and mean percentage of proliferating cells (right panel) was shown. Data are representative or the average of analyses of 4 samples from 4 donors. (F) CD1c+ mDCs were cultured with LPS and CFSE-labeled syngenic CD4 T cells, in the presence or absence of Staphylococcal Enterotoxin B (SEB) and Ko143 for 4 days. CFSE dilution was analyzed in CD4+TCRVβ3+ T cells by flow cytometry. Data are representative of analyses of 3 samples from 3 donors.

Mentions: Recent reports have demonstrated that IL-10-producing DCs induce Treg cell differentiation [23]. To determine whether CD1c+ mDCs treated with Ko143 and LPS can also drive Treg cell differentiation, we co-cultured LPS and Ko143-stimulated CD1c+ mDCs with allogeneic CD4 T cells. CD1c+ mDCs treated with the combination of LPS and Ko143 efficiently promoted the expansion of CD4+CD25+ T cells, whereas those treated with LPS alone did not (Figure 4A). Moreover, proliferation of CD4 T cells was reduced when cultured with LPS plus Ko143-treated mDCs compared to those cultured with LPS-stimulated mDCs, as measured by CFSE-labeling assay (Figure 4A). We next determined whether the expanded CD4+CD25+ T cells express FOXP3, the key transcription factor controlling Treg cell development and function. As shown in Figure 4B, a large proportion of CD25+FOXP3+ T cells were induced only by CD1c+ mDCs treated with LPS plus Ko143. In addition, CD1c+ mDCs treated with LPS plus Ko143 induced significant higher frequency of IL-10-producing CD4 T cells and lower frequency of IFN-γ-producing CD4 T cells compared to those treated with LPS alone (Figure 4C). Moreover, the majority of IL-10-producing T cells induced by LPS plus Ko143-treated CD1c+ mDCs were FOXP3+ cells, thus possessing the characteristics of Treg cells (Figure 4D).


Inhibition of breast cancer resistance protein (ABCG2) in human myeloid dendritic cells induces potent tolerogenic functions during LPS stimulation.

Jin JO, Zhang W, Wong KW, Kwak M, van Driel IR, Yu Q - PLoS ONE (2014)

LPS plus Ko143-treated mDCs promote expansion of Treg cells.Ko143-, LPS- or LPS plus Ko143-treated CD1c+ mDCs were co-cultured with CFSE-labeled allogeneic CD4+ T cells (1×105) in a 1∶10 ratio for 4 days. (A) Surface expression of CD25 and CFSE dilution was analyzed in CD4+ T cells by flow cytometry. (B) Flow cytometry of CD4, CD25 and FOXP3 expression was shown. Data are representative of analyses of 3 samples from 3 donors. (C) Flow cytometry of intracellular IL-10 and IFN-γ production in gated CD4+ T cells was shown. (D) Intracellular IL-10 expression in CD4+FOXP3+ T cells (left panel) and mean percentage of IL-10+FOXP3+ or IL-10+FOXP3− cells (right panel) were shown. Data are representative or the average of analyses of 3 samples from 3 donors. (E) CFSE-labeled PBMCs were incubated with soluble anti-CD3 and CD28 Abs for 4 days in the presence or absence of CD25+ Treg cells. CFSE dilution was analyzed in CD4+CD25− and CD8+ T cells (left panel) and mean percentage of proliferating cells (right panel) was shown. Data are representative or the average of analyses of 4 samples from 4 donors. (F) CD1c+ mDCs were cultured with LPS and CFSE-labeled syngenic CD4 T cells, in the presence or absence of Staphylococcal Enterotoxin B (SEB) and Ko143 for 4 days. CFSE dilution was analyzed in CD4+TCRVβ3+ T cells by flow cytometry. Data are representative of analyses of 3 samples from 3 donors.
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pone-0104753-g004: LPS plus Ko143-treated mDCs promote expansion of Treg cells.Ko143-, LPS- or LPS plus Ko143-treated CD1c+ mDCs were co-cultured with CFSE-labeled allogeneic CD4+ T cells (1×105) in a 1∶10 ratio for 4 days. (A) Surface expression of CD25 and CFSE dilution was analyzed in CD4+ T cells by flow cytometry. (B) Flow cytometry of CD4, CD25 and FOXP3 expression was shown. Data are representative of analyses of 3 samples from 3 donors. (C) Flow cytometry of intracellular IL-10 and IFN-γ production in gated CD4+ T cells was shown. (D) Intracellular IL-10 expression in CD4+FOXP3+ T cells (left panel) and mean percentage of IL-10+FOXP3+ or IL-10+FOXP3− cells (right panel) were shown. Data are representative or the average of analyses of 3 samples from 3 donors. (E) CFSE-labeled PBMCs were incubated with soluble anti-CD3 and CD28 Abs for 4 days in the presence or absence of CD25+ Treg cells. CFSE dilution was analyzed in CD4+CD25− and CD8+ T cells (left panel) and mean percentage of proliferating cells (right panel) was shown. Data are representative or the average of analyses of 4 samples from 4 donors. (F) CD1c+ mDCs were cultured with LPS and CFSE-labeled syngenic CD4 T cells, in the presence or absence of Staphylococcal Enterotoxin B (SEB) and Ko143 for 4 days. CFSE dilution was analyzed in CD4+TCRVβ3+ T cells by flow cytometry. Data are representative of analyses of 3 samples from 3 donors.
Mentions: Recent reports have demonstrated that IL-10-producing DCs induce Treg cell differentiation [23]. To determine whether CD1c+ mDCs treated with Ko143 and LPS can also drive Treg cell differentiation, we co-cultured LPS and Ko143-stimulated CD1c+ mDCs with allogeneic CD4 T cells. CD1c+ mDCs treated with the combination of LPS and Ko143 efficiently promoted the expansion of CD4+CD25+ T cells, whereas those treated with LPS alone did not (Figure 4A). Moreover, proliferation of CD4 T cells was reduced when cultured with LPS plus Ko143-treated mDCs compared to those cultured with LPS-stimulated mDCs, as measured by CFSE-labeling assay (Figure 4A). We next determined whether the expanded CD4+CD25+ T cells express FOXP3, the key transcription factor controlling Treg cell development and function. As shown in Figure 4B, a large proportion of CD25+FOXP3+ T cells were induced only by CD1c+ mDCs treated with LPS plus Ko143. In addition, CD1c+ mDCs treated with LPS plus Ko143 induced significant higher frequency of IL-10-producing CD4 T cells and lower frequency of IFN-γ-producing CD4 T cells compared to those treated with LPS alone (Figure 4C). Moreover, the majority of IL-10-producing T cells induced by LPS plus Ko143-treated CD1c+ mDCs were FOXP3+ cells, thus possessing the characteristics of Treg cells (Figure 4D).

Bottom Line: Moreover, inhibition of ABCG2 in monocyte-derived DCs (MDDCs) abrogated the up-regulation of co-stimulatory molecules and production of pro-inflammatory cytokines in these cells in response to LPS.Furthermore, CD1c+ mDCs stimulated with LPS plus Ko143 inhibited the proliferation of allogeneic and superantigen-specific syngenic CD4+ T cells and promoted expansion of CD25+FOXP3+ regulatory T (Treg) cells in an IL-10-dependent fashion.These tolerogenic effects of ABCG2 inhibition could be abolished by ERK inhibition.

View Article: PubMed Central - PubMed

Affiliation: Shanghai Public Health Clinical Center, Shanghai Medical College, Fudan University, Shanghai, China.

ABSTRACT
Breast cancer resistance protein (ABCG2), a member of the ATP-binding cassette transporters has been identified as a major determinant of multidrug resistance (MDR) in cancer cells, but ABC transporter inhibition has limited therapeutic value in vivo. In this research, we demonstrated that inhibition of efflux transporters ABCG2 induced the generation of tolerogenic DCs from human peripheral blood myeloid DCs (mDCs). ABCG2 expression was present in mDCs and was further increased by LPS stimulation. Treatment of CD1c+ mDCs with an ABCG2 inhibitor, Ko143, during LPS stimulation caused increased production of IL-10 and decreased production of pro-inflammatory cytokines and decreased expression of CD83 and CD86. Moreover, inhibition of ABCG2 in monocyte-derived DCs (MDDCs) abrogated the up-regulation of co-stimulatory molecules and production of pro-inflammatory cytokines in these cells in response to LPS. Furthermore, CD1c+ mDCs stimulated with LPS plus Ko143 inhibited the proliferation of allogeneic and superantigen-specific syngenic CD4+ T cells and promoted expansion of CD25+FOXP3+ regulatory T (Treg) cells in an IL-10-dependent fashion. These tolerogenic effects of ABCG2 inhibition could be abolished by ERK inhibition. Thus, we demonstrated that inhibition of ABCG2 in LPS-stimulated mDCs can potently induce tolerogenic potentials in these cells, providing crucial new information that could lead to development of better strategies to combat MDR cancer.

Show MeSH
Related in: MedlinePlus