Stat3-Dependent Induction of p19INK4D by IL-10 Contributes to Inhibition of Macrophage Proliferation

Abstract

We have previously reported that IL-10 inhibits proliferation of normal bone marrow-derived macrophages and of the monocyte/macrophage cell line J774. Activation of Stat3 was shown to be necessary and sufficient to mediate inhibition of proliferation. To investigate further the mechanism of growth arrest, we examined the effect of IL-10 on expression of cell cycle inhibitors. We found that IL-10 treatment increases expression of the cyclin-dependent kinase inhibitors p19INK4D and p21CIP1 in macrophages. IL-10 cannot induce p19INK4D expression or block proliferation when Stat3 signaling is blocked by a dominant negative Stat3 or a mutant IL-10Rα which does not recruit Stat3 in J774 cells, whereas p21CIP1 induction is not affected. An inducibly active Stat3 (coumermycin-dimerizable Stat3-Gyrase B), which suppresses J774 cell proliferation, also induced p19INK4D expression. Sequencing of the murine p19INK4D promoter revealed two candidate Stat3 binding sites, and IL-10 treatment activated a reporter gene controlled by this promoter. These data suggest that Stat3-dependent induction of p19INK4D mediates inhibition of proliferation. Enforced expression of murine p19INK4D cDNA J774 cells significantly reduced their proliferation. Use of antisense p19INK4D and analysis of p19INK4D-deficient macrophages confirmed that p19INK4D is required for optimal inhibition of proliferation by IL-10, and indicated that additional IL-10 signaling events contribute to this response. These data indicate that Stat3-dependent induction of p19INK4D and Stat3-independent induction of p21CIP1 are important components of the mechanism by which IL-10 blocks proliferation in macrophages.

STAT-1 Mediates the Stimulatory Effect of IL-10 on CD14 Expression in Human Monocytic Cells

“…In general, IL-10-mediated inhibitory effects on cell proliferation and synthesis of proinflammatory cytokines have been shown to be regulated through STAT-3 activation (13,14,16,17). However, in contrast to STAT-3, the role of STAT-1 in regulating IL-10-mediated biological effects remains to be established.…”

“…Interaction of IL-10 with its receptor complex initiates a cascade of events leading to the tyrosine phosphorylation of JAK1 and Tyk2 tyrosine kinases that is followed by the recruitment and phosphorylation of STAT-3 and STAT-1 (13,14). Binding of STAT-3 to the phosphotyrosine residues of the activated IL-10R complex is required for STAT-3 activation and dimerization, and consequent manifestation of a number of biological effects of IL-10 (14,16,17). We provide evidence that IL-10-induced CD14 expression is mediated not through STAT-3, but through STAT-1 activation in monocytic cells.…”

“…IL-10 has been shown to increase the expression of the cyclin-dependent kinase inhibitor p19 INK4D in macrophages. The expression of p19 INK4D was later shown to be dependent on STAT-3 activation (16,17). Recently, the inhibitory effects of IL-10 have also been shown to involve the activation of the suppressor of cytokine signaling (SOCS) 3 proteins, in particular, SOCS1 and SOCS3, in LPS-and IFN-␥-stimulated monocytic cells (18,19).…”

“…IL-10 has been shown to regulate its effects primarily through the activation of STAT-3 (13,14,16,17). Therefore, it was important to determine whether IL-10-induced CD14 expression is regulated by STAT-3 activation directly or indirectly through the ERK/ PI3K-mediated activation.…”

STAT3-mediated differentiation and survival of myeloid cells in response to granulocyte colony-stimulating factor: role for the cyclin-dependent kinase inhibitor p27Kip1

“…For instance, it was shown that STAT5 is involved in both granulocyte colony-stimulating factor (G-CSF)-and interleukin-3 (IL-3)-induced proliferation of Ba/F3 cells (Dong et al, 1998;Mui et al, 1996). In addition, activation of STAT3 is required for IL-6-induced growth arrest and macrophage di erentiation of M1 cells , IL-10-induced inhibition of macrophage proliferation (O'Farrell et al, 1998), and G-CSF-induced neutrophilic di erentiation (Shimozaki et al, 1997). However, the underlying mechanism(s) of STAT-mediated control of growth and di erentiation are still largely unclear.…”