Induction of haptoglobin by all-trans retinoic acid in THP-1 human monocytic cell line

Haptoglobin (Hp) is one of the acute-phase proteins and is mainly synthesized in the liver. During our study on the differentiation of leukemia cells, we have found that Hp is synthesized in human monocytic cells by all-trans retinoic acid (ATRA). The synthesis of Hp by ATRA is induced in a dose- and time-dependent manner. Hp cDNA cloned from ATRA-treated THP-1 cells corresponds to the Hp alpha 2(FS)-beta form. Whereas ATRA acted as a strong inducer in THP-1 cells, IL-1 beta, TNF-alpha, IL-6, and LPS had little effect on Hp gene expression in these cells. These findings suggest that THP-1 cells express the Hp gene through a signal pathway different from hepatocytes, and that ATRA is a potent Hp-inducer in these cells.     

Comparison of SP-A and LPS effects on the THP-1 monocytic cell line

Surfactant protein A (SP-A) increases production of proinflammatory cytokines by monocytic cells, including THP-1 cells, as does lipopolysaccharide (LPS). Herein we report differences in responses to these agents. First, polymyxin B inhibits the LPS response but not the SP-A response. Second, SP-A-induced increases in tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta), and IL-8 are reduced by >60% if SP-A is preincubated with Survanta (200 microgram/ml) for 15 min before addition to THP-1 cells. However, the LPS effects on TNF-alpha and IL-8 are inhibited by <20% and the effect on IL-1beta by <50%. Third, at Survanta levels of 1 mg/ml, SP-A-induced responses are reduced by >90%, and although the inhibitory effects on LPS action increase, they still do not reach those seen with SP-A. Finally, we tested whether SP-A could induce tolerance as LPS does. Pretreatment of THP-1 cells with LPS inhibits their response to subsequent LPS treatment 24 h later, including TNF-alpha, IL-1beta, and IL-8. Similar treatment with SP-A reduces TNF-alpha, but IL-1beta and IL-8 are further increased by the second treatment with SP-A rather than inhibited as with LPS. Thus, whereas both SP-A and LPS stimulate cytokine production, their mechanisms differ with respect to inhibition by surfactant lipids and in ability to induce tolerance.     

Coenzyme Q_{10} effects on manganese superoxide dismutase and glutathione peroxidase in the hairless mouse skin induced by ultraviolet B irradiation

Coenzyme Q_{10} (CoQ_{10}) is a naturally occurring antioxidant and a prominent component of mitochondrial electron transport chain. In the present study, we investigated the effect of CoQ_{10} nanoparticle against photoaging using immunohistochemistry and Western blot analysis in the hairless mouse skin induced by ultraviolet B (UVB) irradiation (300 mJ/cm;{2}, 3 min/day for 21 days). In the UVB-irradiated distilled water (DW)-treated group, manganese superoxide dismutase (SOD2) and glutathione peroxidase (GPx) immunoreactivity and their protein levels in the skin were significantly lower than those in the control group. However, SOD2 and GPx immunoreactivity and their protein levels in the skin of the UVB-irradiated CoQ_{10}-treated group were higher than those in the UVB-irradiated DW-treated group. GPx activity in the skin in the UVB-irradiated DW-treated group significantly decreased compared to that in the control group; whereas GPx activity in the UVB-irradiated CoQ_{10}-treated group was similar to that in the control group. These results suggest that CoQ_{10} strongly inhibits oxidative stress in the skin induced by UVB via increasing SOD2 and GPx.     

Toxoplasma gondii triggers secretion of interleukin-12 but low level of interleukin-10 from the THP-1 human monocytic cell line

Previous studies using both in vitro and in vivo mouse models have demonstrated that a subtle balance between pro- and anti-inflammatory cytokines, among which interleukin-12 (IL-12) and interleukin-10 (IL-10), respectively is crucial to control Toxoplasma infection. However, the few studies performed with human cell lines highlighted important host-related differences in the immune response to Toxoplasma gondii. The goal of our work was thus to study the production of both IL-12 and IL-10 by the THP-1 human monocytic cell line in response to Toxoplasma. We demonstrated that infection by live parasites (RH strain) triggers secretion of IL-12, but low level of IL-10. IL-12 secretion appeared within 8 h, up to 48 h. We also showed that infection by live parasites is not mandatory since heat-killed parasites, crude tachyzoite lysate as well as excreted/secreted antigens induced significant, yet reduced production of IL-12.     

Isolation and identification of C-type natriuretic peptide in human monocytic cell line, THP-1.

In a survey for unknown bioactive peptides in mammalian cell lines, we isolated peptides exhibiting a strong relaxant effect on chick rectum from a phorbol ester-supplemented culture medium of the human monocytic cell line, THP-1. The peptide was deduced to be a C-terminal 29-residue peptide derived from a human C-type natriuretic peptide (CNP) precursor. CNP mRNA was also detected in the THP-1 cells, and expression of CNP gene and CNP concentration in the culture medium was found to be highly augmented by the stimulation of phorbol ester. Production of CNP in THP-1 cells suggests that CNP also functions as a local regulator in the blood cell-vascular system, although CNP has previously been recognized as a neuropeptide functioning in the central nervous system.     

Synergistic increases in IL-1 synthesis by the human monocytic cell line THP-1 treated with PAF and endotoxin

The capacity to stimulate cytokine release may be important to the long-term effects of platelet-activating factor (PAF), which has a very short half-life. Previous studies have shown that PAF stimulates interleukin 1 (IL-1) release by human monocytes. IL-1 and other cytokines produced in response to PAF may be important to the long-term effects of this short-lived lipid. The THP-1 human monocytic leukemia cell line, was used to study the mechanism by which PAF stimulates IL-1 release. PAF stimulates the release of IL-1 beta activity into THP-1 cell supernatants with a multiphasic dose-response curve very similar to that for monocytes. When THP-1 cells are treated with PAF and LPS in combination, these two stimuli interact synergistically to greatly increase the release of IL-1 activity. To assess the effect of PAF on IL-1 beta synthesis, THP-1 cell pellet proteins were separated by SDS-PAGE, blotted, and immunostained to detect IL-1 beta. Immunostaining revealed that PAF increases intracellular IL-1 beta precursor and that the combination of PAF and LPS increases IL-1 beta precursor synergistically. PAF increases IL-1 beta release mainly by increasing IL-1 beta synthesis.     

Thrombospondin-1 differentially regulates release of IL-6 and IL-10 by human monocytic cell line U937

We investigated possible regulatory effects of thrombospondin-1 (TSP-1), a multifunctional extracellular matrix protein, on cytokine release from macrophages. Immobilized TSP-1 enhanced IL-6 release from the human monocytic U937 cells stimulated with phorbol myristate acetate and LPS, whereas it inhibited IL-10 release. The 70-kDa fragment of TSP-1 containing the type 1 repeats showed the same regulatory effects. The enhanced IL-6 release by TSP-1 was inhibited by anti-CD36 antibody or antibody against the sequence of the binding site to CD36 in the type 1 repeats of TSP-1. Conversely, the decrease in IL-10 release by TSP-1 was strengthened by the blocking of the interaction between CD36 and TSP-1. Furthermore, the involvement of TGF-beta1 in the inhibition of IL-10 release by TSP-1 was indicated by the facts that (i) TSP-1 induced activation of TGF-beta1 produced by the U937 cells, (ii) exogenously added TGF-beta1 inhibited IL-10 release, and (iii) antibody against TGF-beta1 blocked the inhibition of IL-10 release by TSP-1. Together, the present findings suggest that TSP-1 enhances IL-6 release from macrophages by interaction with CD36, whereas IL-10 release is regulated by the balance between the enhancing effect of TSP-1 via CD36 and the suppressive effect by TSP-1-activated TGF-beta1.     

Novel lectin-like proteins on the surface of human monocytic leukemia cell line THP-1 cells that recognize oxidized cells

Presence of lectin-like receptors on the membranes of human monocytic leukemia cell line THP-1 cells for clustered sialylated poly-N-acetyllactosaminyl sugar chains on the membranes of oxidized erythrocytes and T-lympoid cells was investigated. Membranes of THP-1 cells differentiated into macrophages were solubilized, and the membrane proteins obtained by affinity chromatographies using lactoferrin-Sepharose and band 3-Sepharose were purified by successive DE column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Proteins of 50, 60, and 80 kDa with specificity to bind to sialylated poly-N-acetyllactosaminyl sugar chains were detected in the chromatographic fractions. A 50-kDa protein was isolated in a pure form. N-Terminal amino acid sequence of the protein was Lys-Gln-Lys-Val-Ala-Gly-Lys-Gln-Pro-Val-, which has not been found in the N-terminal regions of the hitherto known proteins. The antibody, raised against the chemially synthesized peptide composed of the N-terminal amino acid sequence, bound to 50-, 60-, and 80-kDa proteins as analyzed by immunoblotting and immunoprecipitation, indicating that these proteins had the same N-terminal amino acid sequence. The results demonstrate that THP-1 cells have novel 50-, 60-, and 80-kDa lectin-like proteins with the same N-terminal amino acid sequence on the cell surface which would bind to clustered sialylated poly-N-acetyllactosaminyl sugar chains generated on oxidized erythrocytes and T-lymphoid cells.     

Effects of Coenzyme Q10 on TNF-alpha secretion in human and murine monocytic cell lines

Studies in humans and cell culture as well as bioinformatics suggested that Coenzyme Q(10) (CoQ10) functions as an anti-inflammatory molecule. Here we studied the influence of CoQ10 (Kaneka Q10) on secretion of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) by using the human and murine monocytic cell lines THP-1 and RAW264.7 expressing human apolipoprotein E3 (apoE3) or pro-inflammatory apoE4. Incubation of cells with physiological (0.1-10 microM) and supra-physiological (> 10 to < 100 microM) concentrations of CoQ10 led to an intracellular accumulation of its reduced form without any cytotoxic effects. Stimulation of cell models with lipopolysaccharide (LPS) resulted in a substantially release of TNF-alpha. When THP-1 cells were pre-incubated with 10 microM CoQ10, the LPS-induced TNF-alpha release was significantly decreased to 72 +/- 32%. This effect is similar to those obtained by 10 microM N-Acetyl-Cysteine, a well known reference antioxidant. In RAW264.7-apoE3 and -apoE4 cells, significant reductions of LPS-induced TNF-alpha secretion to 73.3 +/- 2.8% and 74.7 +/- 8.9% were found with 2.5 microM and 75 microM CoQ10, respectively. In conclusion, CoQ10 has moderate anti-inflammatory effects in two monocytic cell lines which could be mediated by its antioxidant activity.     

α2-Macroglobulin synthesis by the human monocytic cell line THP-1 is differentiation state-dependent

Human α₂-macroglobulin (α₂M) is a broad spectrum proteinase inhibitor and cytokine carrier synthesized by a number of cell types including monocytes and macrophages. In this study, we report on the expression of α₂M by THP-1 cells. This monocytic cell line can be differentiated into a macrophage-like phenotype by treatment with interferon-γ (IFN-γ) or phorbol 12-myristate 13-acetate (PMA). α₂M was synthesized by THP-1 cells at a rate of 75 ng/10⁶ cells/24 h, as determined by Western blot analysis. After treating the cells with 500 U/ml of IFN-γ or with 100 ng/ml PMA, the synthesis rate increased to 219 ng/10⁶ cells/24 h and to 179 ng/10⁶ cells/24 h, respectively. The same agents also increased α₂M expression, as determined by Northern blot analysis. When the α₂M receptor antagonist, receptor associated protein (RAP), was included in the THP-1 medium, the amount of α₂M recovered in the conditioned medium increased. This result suggests that THP-1-secreted proteinases react with secreted α₂M and that the resulting complexes are catabolized by the α₂M receptor, which is also called low density lipoprotein receptor-related protein (LRP). We conclude that α₂M synthesis by THP-1 cells depends on the state of cellular differentiation. Reaction of α₂M with secreted proteinases may have minimized previous estimates of the rate of synthesis of α₂M by certain cells in culture. J. Cell. Biochem. 67:492–497, 1997. © 1997 Wiley-Liss, Inc.     

肺炎克雷伯菌生物膜对单核细胞株THP-1 TLR2受体表达的影响

目的研究肺炎克雷伯菌生物膜对单核细胞株THP-1TLR2受体表达的影响,探索细菌生物膜逃脱宿主免疫防御的机制。方法以体外建立肺炎克雷伯菌生物膜的合成分泌物处理单核细胞株THP－1作为实验组,以等量浮游细菌的合成分泌物处理单核细胞株THP-1作为对照组。RT—PER半定量分析THP－1 TLR2mRNA的表达,流式细胞仪检测分析THP－1 TLR2蛋白的表达。结果实验组THP－1 TLR2 mRNA表达（0．453±0．06）明显低于对照组（4．872±0．36）（P〈0．05）;实验组TLR2蛋白表达率（8．42％±3．74％）明显低于对照组（12．35％±7．36％）（P〈0．05）。结论细菌生物膜与浮游细菌不同的代谢产物能显著下调THP－1 TLR2的表达水平,影响固有免疫进而影响适应性免疫,可能是生物膜细菌逃脱机体免疫防御系统的又一机制。     

Detection of measles virus-induced apoptosis of human monocytic cell line (THP-1) by DNA fragmentation ELISA

Abstract Measles virus (wild strain, Toyoshima strain)-induced cell death is characterized by cell shrinkage, chromatin condensation, and nuclear fragmentation in a human monocytic cell line (THP-1). DNA fragmentation of measles virus-infected THP-1 cells was demonstrated by DNA agarose gel electrophoresis as well as by DNA fragmentation ELISA. When measles virus-infected THP-1 cells were cultured on monolayers of fibroblasts or human umbilical vein endothelial cells (HUVEC), the percentage of measles virus antigen-positive THP-1 cells and DNA fragmentation were significantly decreased. Addition of anti-intercellular adhesion molecule (ICAM)-1 (CD54) monoclonal antibody to culture of measles virus-infected THP-1 cells reduced significantly DNA fragmentation induced by measles virus. These findings suggest that inhibition of virus spread by fibroblasts and HUVEC reduces apoptosis, and ICAM-1 (CD54) may participate in the DNA fragmentation pathway.     

Suppression of cytokine production and cell adhesion molecule expression in human monocytic cell line THP-1 by Tripterygium wilfordii polysaccharide moiety

Extracts of the vine-like plant Tripterygium wilfordii (TW) have been widely used in China as an immunosuppressant and anti-inflammatory drug for the treatments of rheumatoid arthritis, lupus erythematosus and other inflammatory disorders. In this study the molecular mechanisms of action of three TW extracts (ethanol, aqueous, polysaccharide) on the expression of inflammatory cytokines and adhesion molecules were investigated by RT-PCR and immunofluorescence binding techniques. The lipopolysaccharide (LPS)-mediated stimulatory effects of tumor necrosis factor-alpha (TNF-alpha) cytokine production and cell adhesion molecule (CD11c, CD18, CD14, CD54) expression in human monocytic THP-1 cells were modulated by treatments of the TW extracts or tacrolimus (FK506). The TW polysaccharide moiety exhibited more profound immunosuppressive properties than the aqueous and ethanol extracts. Biochemical characterization of the polysaccharide moiety revealed a major molecular weight of 22 kDa (viz. PSP22). The PSP22 was found to be a potential immunosuppressant that manifests the necessary immunomodulating properties.     

A 63 kDa tumor necrosis factor inhibitor released from a human monocytic leukemia cell line, THP-1

A human monocytic cell line, THP-1-S, was cultured in a serum-free medium. The effect of the culture supernatant of THP-1-S on the cytotoxicity of rTNF-alpha to three kinds of cell lines and the binding of rTNF to its receptor were tested. The supernatant inhibited the cytotoxicity of rTNF-alpha when tested by the neutral red uptake method. In addition, the supernatant blocked the binding of 125I-rTNF-alpha to its receptor. Furthermore, following precipitation with PEG we detected complexes between rTNF-alpha and the inhibitory factor which formed during incubation with the culture supernatant from THP-1-S cells. However, the supernatant did not bind to or down-regulate the receptor for TNF-alpha on the cell surface of L-M-2d6 cells. This factor eluted with an apparent molecular mass of 63,000 Da by gel filtration and did not react with antibodies against p55 and p75 TNF receptors. These data suggest that human monocytic cells are capable of releasing an inhibitory factor against rTNF-alpha in serum-free culture conditions.     

Oxysterols induce apoptosis and accumulation of cell cycle at G(2)/M phase in the human monocytic THP-1 cell line

The cytotoxic activity of oxysterols, 7 beta-hydroxycholesterol (7 beta-OHC) and 25-hydroxycholesterol (25-OHC), has been evaluated using various leukemia cell lines. Among the tested cell lines, both oxysterols showed the highest cytotoxicity to THP-1, human monocytic leukemia cell line. These oxysterols induced apoptosis through down-regulation of Bcl-2 expression and activation of caspases. Also, the oxysterols showed the accumulation at G(2)/M phase of cell cycle through down-regulation of cyclin B1 expression. Taken together, these results indicated that both 7 beta-OHC and 25-OHC inhibited the proliferation of THP-1 cells through apoptosis and cell cycle accumulation at G(2)/M phase.     

Influence of length on cytotoxicity of multi-walled carbon nanotubes against human acute monocytic leukemia cell line THP-1 in vitro and subcutaneous tissue of rats in vivo

Carbon nanotubes (CNTs) are single- or multi-cylindrical graphene structures that possess diameters of a few nanometers, while the length can be up to a few micrometers. These could have unusual toxicological properties, in that they share intermediate morphological characteristics of both fibers and nanoparticles. To date, no detailed study has been carried out to determine the effect of length on CNT cytotoxicity. In this paper, we investigated the activation of the human acute monocytic leukemia cell line THP-1 in vitro and the response in subcutaneous tissue in vivo to CNTs of different lengths. We used 220 nm and 825 nm-long CNT samples for testing, referred to as "220-CNTs" and "825-CNTs", respectively. 220-CNTs and 825-CNTs induced human monocytes in vitro, although the activity was significantly lower than that of microbial lipopeptide and lipopolysaccharide, and no activity appeared following variation in the length of CNTs. On the other hand, the degree of inflammatory response in subcutaneous tissue in rats around the 220-CNTs was slight in comparison with that around the 825-CNTs. These results indicated that the degree of inflammation around 825-CNTs was stronger than that around 220-CNTs since macrophages could envelop 220-CNTs more readily than 825-CNTs. However, no severe inflammatory response such as necrosis, degeneration or neutrophil infiltration in vivo was observed around both CNTs examined throughout the experimental period.     

Scavenger receptor of human monocytic leukemia cell line (THP-1) and murine macrophages for nonenzymatically glycosylated proteins

Long-term incubation of proteins with glucose undergo a series of nonenzymatic reactions to form advanced glycosylation end product (AGE) with fluorescence and brown color. The receptor for AGE-proteins was demonstrated in murine macrophages (Vlassara et al. (1985) Proc. Natl. Acad. Sci. USA 82. 5588). Our recent study with rat macrophages revealed that the receptor also recognized proteins modified with aliphatic aldehydes such as formaldehyde or glycolaldehyde, indicating its close identity to a scavenger receptor for aldehyde-modified proteins (Takata, K. et al. (1988) J. Biol. Chem. 263. 14819). This notion was tested in the present study with human monocytic leukemia cell line (THP-1 cells), human monocyte macrophages and murine peritoneal macrophages. Endocytic uptake of AGE-proteins and aldehyde-modified proteins was inhibited in a cross-competitive fashion. The receptor activities of THP-1 cells for AGE-albumin and aldehyde-modified proteins were induced synchronously by phorbol 12-myristate 13-acetate. Furthermore, upon reduction by NaBH4 of the Schiff base formed between proteins and glucose or aldehydes, no ligand activity was generated. However, once the ligand activity was generated, NaBH4 was no longer effective for the ligand activity. Thus, a structure in common between AGE-proteins and aldehyde-modified proteins may be crucial for recognition by the human macrophage receptor.