Multiple effects of human recombinant interleukin 4 on human myeloid monocyte cell lines

The effects of recombinant human interleukin 4 (rIL-4) on proliferation and differentiation on human myeloid/monocytic leukemia cell lines were examined. At high concentrations, rIL-4 had a slight enhancing effect on [3H]thymidine incorporation by U937 cells. rIL-4 markedly induced expression of the Fc epsilon receptor (CD23) and the Leu-M3 antigen (CD14) on U937 cells. HL60 and THP-1 cells treated with rIL-4 also showed increased CD23 expression, but little change of CD14 antigen expression. CD23 induction required lower amounts of IL-4 than needed for T cell growth, indicating that CD23 induction on U937 will serve as a sensitive assay for human IL-4. rIL-4 reduced the steady state level of IL-1 beta mRNA in U937.     

Constitutive expression of the nef gene suppresses human immunodeficiency virus type 1 (HIV-1) replication in monocytic cell lines.

In order to study the effect of nef gene expression on viral replication in monocytic cells, we established monocytic (U937 and THP-1) cell transfectants constitutively expressing the human immunodeficiency virus type 1 nef gene. We constructed a plasmid expressing the nef gene derived from an infectious clone, NL432, under the control of SR alpha promoter which can drive a high level of gene expression. We found suppressed viral replication in nef-expressing monocytic cells, although a negative effect of nef was observed, with some variation depending on the virus strain and the cell. We also observed that the expression of the surface CD4 molecule is inversely related to the expression of the nef gene, especially in the U937 transfectants. These results indicate that the suppression of viral replication and the down-modulation of CD4 molecule by nef gene expression occur in monocytic cell lines as in T cell lines.     

Selective DNA-binding activity of interleukin-10-stimulated STAT molecules in human monocytes.

It has been demonstrated that interferon-gamma (IFN-gamma) and interleukin-10 (IL-10) have various reverse effects on macrophages; however, the molecular mechanism of this difference has not been fully understood. In this study, we analyzed the binding activity of IL-10- and IFN-gamma-activated STAT molecules to two kinds of GAS-motif sequences. IL-10-activated STAT1 could bind to the GAS-motif sequence in the promoter region of the Fcgamma receptor, but not to that in the promoter region of the COX-2 gene, whereas IFN-gamma-activated STAT1 and STAT5 could bind to both sequences. IL-10 inhibited IFN-gamma-induced STAT activation without newly synthesized protein. We further demonstrated that aspirin, but not dexamethasone, suppressed IFN-gamma-induced STAT activation. Taken together, these results suggest that IL-10-activated STAT1 has a specificity in binding to the GAS-motif sequences, whereas IFN-gamma-activated STAT1 and STAT5 have a broader spectrum in binding to the GAS-motif sequences. This may explain the difference between IL-10 and IFN-gamma in biological activity, and the inhibitory effect of IL-10 on IFN-gamma activities.     

Regulation of human IL-18 gene expression: interaction of PU.1 with GC-box binding protein is involved in human IL-18 expression in myeloid cells

Interleukin-18 (IL-18) is a pro-inflammatory cytokine which participates in host defense against a variety of infections as well as in chronic inflammation including autoimmune diseases. However, little is known about human IL-18 regulation at the gene level. We have previously demonstrated that sodium butyrate, a bacterial fermentation product, induces IL-18 production via the proximal region of the promoter. In this study we investigated the molecular mechanisms for basal and sodium butyrate-induced expression of IL-18 in human myeloid cells. Two regulatory regions, a consensus binding site for PU.1 and a GC-rich region, are required for basal IL-18 promoter activity in human myeloid cells. PU.1 bound to the PU.1 consensus binding site in electrophoretic mobility shift assays, and overexpression of PU.1 led to activation of the IL-18 promoter through this site. Mutation analysis revealed that the GC-rich region, but not PU.1 site, participates in sodium butyrate-induced transactivation. Furthermore, DNA pull-down experiments and the critical spacing of the two binding sites suggest that formation of a protein complex involving both cis elements and the respective binding proteins might be crucial for human IL-18 expression.     

Tumour necrosis factor-alpha augments the expression of Fc IgE receptor (Fc epsilon RII/CD23) on human monocytic cell lines and down-regulates interleukin-4-driven Fc epsilon RII expression on monocytes.

We investigated the expression of the low affinity Fc IgE receptor (Fc epsilon RII/CD23) on the human monocytic cell lines U937, THP-1, Mono-Mac-6, and cultured human peripheral blood monocytes under stimulation with human tumour necrosis factor-alpha (TNF-alpha) and other cytokines. Fc epsilon RII was demonstrated by flow cytometry analysis employing the anti-Fc epsilon RII monoclonal antibody 3-5. TNF-alpha alone had a weak but significant stimulating effect on the Fc epsilon RII expression on the cell lines U937 and THP-1, and very modestly on Mono-Mac-6 cells. TNF-alpha strongly synergized with interferon-gamma (IFN-gamma) and interleukin-4 (IL-4). IFN-alpha per se was ineffectual, but was able to increase the TNF-alpha effect. Furthermore, the action of TNF-alpha was slightly augmented by human IL-6. Similar effects were noted with TNF-beta alone or in combination with other cytokines. Interestingly, on human monocytes TNF-alpha weakly reduced the basal level of Fc epsilon RII, and markedly diminished the IL-4-induced Fc epsilon RII expression. Our results indicate that several cytokines may interact in a cytokine network to modulate Fc epsilon RII expression on monocytic cell lines. On human blood monocytes, TNF-alpha, like IFN-gamma or IL-6, counteracts the IL-4-induced Fc epsilon RII expression. These data suggest different regulatory pathways of Fc epsilon RII expression on blood monocytes and myelomonocytic cell lines.     

ＩＬ—６在Ｕ９３７细胞中特异、持续激活ＳＴＡＴ３

目的 探讨ＩＬ－６在Ｕ９３７细胞中产生生物学效应的信号转导基础。方法 检测ＩＬ－６对Ｕ９３７细胞生长与分化的影响。并采用凝胶阻滞电泳法,检测ＩＬ－６在Ｕ９３７细胞中激活核因子的情况。结果 ＩＬ－６可诱导Ｕ９３７细胞向巨噬细胞方向分化。分化的细胞酸性醋酸酯酶（ＡＮＡＥ）活性、硝基蓝四唑（ＮＢＴ）还原能力及ＣＤ５４表达增强,ＳＴＡＴ３可被ＩＬ－６显著激活,其活性呈一定的剂量与时间依赖性;而ＮＦ－ＩＬ６、ＮＦ－kB、ＡＰ－１及其它ＳＴＡＴ家族成员则无明显激活。结论 Ｕ９３７细胞的终分化可能是ＪＡＫ－ＳＴＡＴ３通路介导的。     

LPS耐受单核细胞中p50抑制IKKα结合于IL-1β启动子区

目的 研究LPS耐受细胞中IL-1β启动子区p50对IKKct的作用,揭示p50抑制IL-1β mRNA转录的机制.方法 运用人单核细胞系THP-1模拟LPS耐受,使用染色体免疫沉淀(CHIP)和real-time PCR技术定量IL-1β启动子区p50与IKKa的结合情况,并应用基因沉默技术研究p50和/或IKKα沉默后对IC-1βmRNA转录情况的影响.结果 耐受细胞IL-1β启动子区p50结合并不减少,而IKKα结合降低;p50沉默后,IKKα的结合增加,同时IL-1β mRNA转录增加;p50和IKKα双沉默后,IL-1β mRNA转录又降低.结论 在耐受的THP-1细胞中,IL-1β mRNA转录的降低至少部分原因是由于p50抑制了IKKα对IL-1β启动子区的结合而造成的.