Expression of human NDRG2 by myeloid dendritic cells inhibits down-regulation of activated leukocyte cell adhesion molecule (ALCAM) and contributes to maintenance of T cell stimulatory activity.
ABSTRACT We reported previously that N-myc downstream-regulated gene 2 (NDRG2), a member of a new family of differentiation-related genes, is expressed specifically in dendritic cells (DC) differentiated from monocytes, CD34(+) progenitor cells, and the myelomonocytic leukemic cell line. In this study, we demonstrate that NDRG2 protein expression is detected, not only in in vitro-differentiated DC but also in primary DC from lymph nodes, thymus, and skin when anti-NDRG2 antibodies are used. As predicted from previous studies investigating the mRNA expression pattern of several types of cell lines, progenitor cells, and DC, NDRG2 protein was expressed strongly in DC. Its expression was detected at significant levels after differentiation from progenitor cells. RNA interference of NDRG2 demonstrated that activated leukocyte cell adhesion molecule (ALCAM) expression is down-regulated specifically in DC differentiated from NDRG2 small interfering RNA (siRNA)-transfected monocytes. This was consistent with our observation that U937 cells transfected with NDRG2 became resistant to the GM-CSF/IL-4-induced ALCAM reduction. Furthermore, DC, which had differentiated from NDRG2 siRNA-transfected monocytes, showed a reduced ability to induce T cell proliferation. Taken together, our results indicate that NDRG2 is able to preserve ALCAM expression during DC differentiation from monocytes under cytokine culture conditions and that its expression helps DC maintain costimulatory signals necessary for T cell stimulation.
Article: NDRG2 Promotes GATA-1 Expression through Regulation of the JAK2/STAT Pathway in PMA-stimulated U937 Cells.[show abstract] [hide abstract]
ABSTRACT: N-myc downstream-regulated gene 2 (NDRG2), a member of a newly described family of differentiation-related genes, has been characterized as a regulator of dendritic cells. However, the role of NDRG2 on the expression and activation of transcription factors in blood cells remains poorly understood. In this study, we investigated the effects of NDRG2 overexpression on GATA-1 expression in PMA-stimulated U937 cells. We generated NDRG2-overexpressing U937 cell line (U937-NDRG2) and treated the cells with PMA to investigate the role of NDRG2 on GATA-1 expression. NDRG2 overexpression in U937 cells significantly induced GATA-1 expression in response to PMA stimulation. Interestingly, JAK2/STAT and BMP-4/Smad pathways associated with the induction of GATA-1 were activated in PMA-stimulated U937-NDRG2 cells. We found that the inhibition of JAK2 activation, but not of BMP-4/Smad signaling, can elicit a decrease of PMA-induced GATA-1 expression in U937-NDRG2 cells. The results reveal that NDRG2 promotes the expression of GATA-1 through activation of the JAK2/STAT pathway, but not through the regulation of the BMP-4/Smad pathway in U937 cells. Our findings further suggest that NDRG2 may play a role as a regulator of erythrocyte and megakaryocyte differentiation during hematopoiesis.Immune Network 12/2011; 11(6):348-57.
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ABSTRACT: N-myc downstream regulated gene 2 (NDRG2) is a member of the NDRG gene family. Our previous report indicated a possible role for NDRG2 in regulating the cytokine, interleukin-10 (IL-10), which is an important immunosuppressive cytokine. Several pathways, including p38-MAPK, NF-κB, and JAK/STAT, are used for IL-10 production, and the JAK/STAT pathway can be inhibited in a negative feedback loop by the inducible protein, SOCS3. In the present study, we investigated the effect of NDRG2 gene expression on IL-10 signaling pathway that is modulated via SOCS3 and STAT3. We generated NDRG2-overexpressing U937 cell line (U937-NDRG2) and treated the cells with PMA to investigate the role of NDRG2 in IL-10 production. U937 cells were also transfected with SOCS3- or NDRG2-specific siRNAs to examine whether the knockdown of SOCS3 or NDRG2 influenced IL-10 expression. Lastly, STAT3 and SOCS3 induction was measured to identify the signaling pathway that was associated with IL-10 production. RT-PCR and ELISA assays showed that IL-10 was increased in U937-mock cells upon stimulation with PMA, but IL-10 was inhibited by overexpression NDRG2. After PMA treatment, STAT3 phosphorylation was decreased in a time-dependent manner in U937-mock cells, whereas it was maintained in U937-NDRG2 cells. SOCS3 was markedly reduced in U937-NDRG2 cells compared with U937-mock cells. IL-10 production after PMA stimulation was reduced in U937 cells when SOCS3 was inhibited, but this effect was less severe when NDRG2 was inhibited. NDRG2 expression modulates SOCS3 and STAT3 activity, eventually leading to the inhibition of IL-10 production.Immune Network 12/2010; 10(6):219-29.
Article: A proinflammatory cytokine interleukin-32beta promotes the production of an anti-inflammatory cytokine interleukin-10.[show abstract] [hide abstract]
ABSTRACT: A new proinflammatory cytokine interleukin-32 (IL-32) has six isoforms. Although IL-32 can be detected in sera from patients suffering from Crohn's disease and rheumatoid arthritis, it is unclear which isoforms are involved. To this end, we investigated the functions of the most abundant IL-32beta by generating K562-IL-32beta stable cell lines. This report confirms, using IL-32 small interfering RNA, that IL-32beta induces an anti-inflammatory cytokine IL-10 in K562-IL-32beta cells and U937 promonocytic cells, which express endogenous IL-32beta upon phorbol 12-myristate 13-acetate (PMA) treatment, and monocyte-derived dendritic cells (DC) upon lipopolysaccharide (LPS) treatment. Interleukin-32beta was induced in monocyte-derived macrophages by LPS and in monocyte-derived DC by LPS, poly(I:C), or anti-CD40 antibody, but was not induced by PMA. We showed that IL-32beta expression was increased in a time-dependent manner in monocyte-derived DC upon LPS treatment and peaked at 24 hr. Production of IL-10 was exactly coincident with IL-32beta expression, but IL-1beta and tumour necrosis factor-alpha production peaked at 6 hr after LPS treatment, then steeply declined. Interleukin-12 p40 was induced at 9 hr and gradually increased until 48 hr, at which time IL-32beta and IL-10 were no longer increased. Knock-down of IL-32beta by IL-32 small interfering RNA led to the decrease of IL-10, but the increase of IL-12 in monocyte-derived DC, which means that IL-32beta promotes IL-10 production, but limits IL-12 production. We also showed that IL-10 neutralization increases IL-12, IL-1beta and tumour necrosis factor-alpha production, which implies that IL-10 suppresses such proinflammatory cytokines. Taken together, our results suggest that IL-32beta upregulates the production of an anti-inflammatory cytokine IL-10, and then IL-10 suppresses proinflammatory cytokines.Immunology 09/2009; 128(1 Suppl):e532-40. · 3.32 Impact Factor