• The Journal of the Korean Society for Microbiology
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• v.35 no.2
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• pp.149-157
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• 2000

Mycobacterium tuberculosis is capable of growing and survival within macrophage. The purpose of this study was to identify the genes regulated by infection of mycobacteria in human monocytic THP-1 cells. We used the differential display reverse transcriptase polymerase chain reaction (DD RT-PCR) and nothern blot analysis to confirm the differentially expressed genes from THP-1 cells infected with live Mycobacterium tuberculosis H37Rv, heat-killed Mycobacterium tuberculosis H37Rv and live Mycobacterium bovis BCG. Among many up or down-regulated clones, 27 clones were sequenced and compared with known genes on GenBank. Thirteen of over-expressed clones from THP-1 cells infected with live Mycobacterium tuberculosis H37Rv were identical to human prothymosin alpha, eight were novel clones and six clones showed homology with Human ferritin H chain, Esherichia coli bgl, Mouse RNA-dependent EIF-2 alpha kinase, E. coli htrL, Hyaluronan receptor and T cell receptor. Our result suggests that Mycobacterium tuberculosis might regulate prothymosin alpha gene transcription in monocytic THP-1 cell.

• The Korean Journal of Physiology and Pharmacology
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• v.27 no.2
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• pp.157-165
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• 2023

Triple-negative breast cancer (TNBC) is the most aggressive subtype of breast cancer and current therapeutic strategies are limited in their effectiveness. The expressions of Rab5 and the M2 tumor-associated macrophage marker CD163 in tissues were detected by Western blot. The migration and invasion of cells were determined using a Transwell assay. The expressions of the exosome markers were evaluated by Western blot. The polarization of human macrophages (THP-1) was determined by incubation of THP-1 cells with conditioned medium or exosomes collected from MDA-MB-231 cells with indicated transfections or by a coculture system of THP-1 and MDA-MB-231 cells. The M1 and M2 macrophage markers were evaluated by qRT-PCR. The expression of Rab5 in TNBC was significantly higher than that in normal breast tissue. Rab5 expressions in triple-negative and luminal A breast cancer were higher than those in other molecular subtypes. Higher CD163 expression was observed in triple-negative breast cancer and in triple-negative and luminal B subtypes. Rab5 knockdown suppressed but Rab5 overexpression promoted the migration and invasion capacity of MDA-MB-231 cells. The levels of CD63 and CD9 in the medium of Rab5 knockdown cells were lower than those in control cells, whereas higher levels of CD63 and CD9 were observed in Rab5 overexpression cells. Rab5 knockdown decreased the excretion but did not alter the diameter of the exosomes. Knockdown of Rab5 facilitated the anti-tumor polarization of macrophages, which was partially reversed by Rab5 overexpression. Therefore, Rab5 is expected to be a potential therapeutic target for triple-negative breast cancer.

• Journal of Life Science
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• v.32 no.2
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• pp.142-147
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• 2022

Oxysterols are known to be involved in the physiopathology of atherosclerosis. Since 7-ketocholesterol (7-KC) is found in large amounts in oxysterols and in atherosclerotic plaque, the study on how 7-KC may affect monocyte differentiation induced by phorbol myristate acetate (PMA) in the monocytic cell line, THP-1, is essential. 7-KC induced a dose-dependent reduction in cell proliferation without inducing cytotoxicity, and the substantial staining of Nile red demonstrates the increased absorption of intracellular lipids. Although 7-KC itself did not increase cell adhesion, it markedly decreased the adhesion of cells treated with PMA. Furthermore, by observing the effect of 7-KC on phagocytosis using fluorescent-labeled latex beads, 7-KC's ability to abolish phagocytosis in PMA-stimulated macrophages was illustrated. The effect of 7-KC on the expression of selected protein markers on the process of differentiation induced by PMA in THP-1 cells was also examined. 7-KC inhibited expression of ICAM-1, CD11a, SR-A1, and SR-B2 (CD36) in PMA-stimulated THP-1 cells. Conversely, 7-KC drastically increased the expression of SR-D1 (CD68)in PMA-stimulated THP-1 cells. In conclusion, these results suggest that 7-KC modulates monocyte differentiation and activation via the intracellular accumulation of lipid droplets.

• Journal of Physiology & Pathology in Korean Medicine
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• v.36 no.1
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• pp.1-6
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• 2022

The tuber of Pinellia ternata (Thunb.) Brei (TPT) used in traditional Oriental medicine for the treatment of cough, sputum, vomiting, and insomnia, possesses antioxidant, antibacterial, and anti-inflammatory effects. Although recent studies have reported the anticancer effects of TPT in several cancer cells, it is still unclear whether TPT regulates tumor-associated macrophage (TAM) characterized by the immunosuppressive M2 macrophage phenotype. Our results showed that the ethanol extract of TPT (ETPT) suppressed the migration of RAW264.7 mouse macrophage cells and THP-1 human monocytes differentiated into macrophages towards the conditioned media (CM) collected from lung cancer cells, suggesting that ETPT would attenuate the recruitment of macrophages into tumors. In addition, ETPT suppressed the interleukin (IL)-4 or IL-6-induced M2 macrophage polarization in RAW264.7 cells. ETPT treatment not only downregulated the mRNA expression of M2 macrophage markers including arginase-1, mannose receptor C type 1 (MRC-1), and IL-10, but also inhibited the phosphorylation of signal transducer and activator of transcription 3 (STAT3) and STAT6, general regulators of M2 macrophage polarization. Finally, the transwell assay results showed that the CM from M2-polarized RAW264.7 cells increased the migration of mouse lewis lung carcinoma (LLC) cells, while those from RAW264.7 cells co-treated with ETPT and IL-6 significantly reduced the migration of LLC cells. Taken together, our observations clearly demonstrate that ETPT suppressed the cancer cell migration by regulating macrophage recruitment and M2 macrophage polarization.

• Journal of Life Science
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• v.30 no.2
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• pp.113-121
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• 2020

Macrophages are initiators for regulating a host's defenses to eliminate pathogens and trigger tissue repair. Macrophages are classified into two types: classically (M1) activated macrophages and alternatively (M2) activated macrophages. M1-phenotype macrophages directly or indirectly kill infectious organisms and tumor cells via pro-inflammatory responses, whereas M2-phenotype macrophages remodel wounded tissue through anti-inflammatory responses. In this paper, we investigated how Phellinus linteus hot water extract passed from Diaion HP-20 resin (PLEP) regulates polarization of M1-like or M2-like macrophages in human THP-1 cells. PLEP did not have cytotoxicity at a high concentration of 300 ㎍/ml. We observed morphological alteration of the THP-1 cells, which are stimulated by PLEP, LPS/INF-γ (M1 stimulators) or IL-4/IL13 (M2 stimulators). PLEP exposure induced morphology contiguous with LPS/INF-γ. qPCR was also performed to determine whether PLEP influences M1 or M2 polarization-related genes. M1-phenotype macrophage-specific genes, such as TNF-α, IL-1β, IL-6, IL-8, CXCL10 and CCR7, were enhanced by PLEP in a dose-dependent manner similar to LPS/INF-γ. Conversely, M2-phenotype-specific genes, such as MRC-1, DC-SIGN, CCL17 and CCL22, were suppressed by PLEP. PLEP also significantly up-regulated secretory inflammation cytokines related to M1 polarization of macrophages, including TNFα, IL-1β and IL-6, which was similar to the gene expression. Further, MAPK and NF-κB signaling were increased by treatment with PLEP, resulting in enhancement of cytokine secretion. PLEP might therefore be used as a promising booster of pro-inflammatory responses through M1 polarization of human THP-1 cells.

• Archives of Plastic Surgery
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• v.50 no.4
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• pp.432-442
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• 2023

Background Macrophages play a major role in wound healing and prevent infection from the outside. Polarization conversion of macrophages regulates aspects of inflammation, and two macrophages, M1 (classically activated) and M2 (alternatively activated), exist at both ends of broad-spectrum macrophage polarization. Thus, we aimed to investigate whether macrophage polarization can be artificially regulated. To this end, MgSO4 and small-interfering RNA (siRNA) targeting magnesium transport 1 (MAGT1) were used to investigate the effects of intracellular magnesium (Mg2+) concentrations on the differentiation of macrophages in vitro. Methods THP-1 derived macrophages maintained in a culture medium containing 5 mM MgSO4 and siRNA to inhibit the expression of MAGT1. As comparative groups, THP-1 derived macrophages polarized into M1 and M2 macrophages by treatment with M1, M2 inducer cytokine. The polarization status of each group of cells was confirmed by cell surface antigen expression and cytokine secretion. Results We found that MgSO4 treatment increased CD163 and CD206, similar to the effect noted in the M2 group. The expression of CD80 and HLA-DR was increased in the group treated with MAGT1 siRNA, similar to the effect noted in the M1 group. Functional assays demonstrated that the group treated with MgSO4 secreted higher levels of IL-10, whereas the MAGT1 siRNA-treated group secreted higher levels of IL-6 cytokines. Additionally, the conditional medium of the Mg2+ treated group showed enhanced migration of keratinocytes and fibroblasts. Conclusion Mg2+ can help to end the delay in wound healing caused by persistent inflammation in the early stages.

• IMMUNE NETWORK
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• v.7 no.1
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• pp.26-30
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• 2007

Background: Mycobacterial antigens released as PIM, LM, LAM, lipoproteins and other cellular factors may contribute to macrophage and dendritic cell activation through pattern recognition receptors such as TLRs. In this study, we assessed cytokine production and ERK activation with stimulation of several major mycobacterial antigens. Methods: Purified mycobacterial antigens (10, 22, 30, 38kDa) and recombinant antigens (6, 16, 19, 38kDa, Ag85A antigen) were studied. The production of cytokines (TNF-${\alpha}$, IL-12, IL-6) was measured by ELISA. The ERK activation was detected by western blotting. The expression of TLR2 or TLR4 was measured by flow cytometry. Results: Among purified antigens only 30kDa antigen induced production of IL-6 or TNF-${\alpha}$ in THP-1 macrophage cells. When THP-1 macrophage cells were treated with 30kDa antigen, phosphorylation of ERK was detected. ERK activation also occurred in TLR2 transfectant HEK293 cells with 30kDa antigen stimulation. Conclusion: 30kDa antigen is one of the major mycobacterial antigens inducing cytokine production and MAP kinases phosphorylation in macrophages.

• Journal of Physiology & Pathology in Korean Medicine
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• v.23 no.1
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• pp.192-198
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• 2009

Banhabaikchulcheunma-tang (BBCT), a prescription composed of thirteen herbal mixtures, has been widely used in the treatment of brain disorders in Oriental Medicine. However, the mechanisms by which the formula affects on the production of pro-inflammatory cytokines in cerebral infarction (CI) patients remain unknown yet. The levels of secretory protein of pro-inflammatory cytokines, including tumor necrosis factor (TNF)-a, interlukin (IL)-1b, and IL-6, were significantly increased in lipopolysaccharide (LPS)-stimulated THP-1 differentiated macrophage-like (THP-1/M) cells and Peripheral blood mononuclear cells (PBMCs) from CI patients. However, pretreatment with BBCT significantly inhibited the secretion of pro-inflammatory cytokines, including TNF-a, IL-1b, and IL-6, in THP-1/M cells and PBMCs from CI patients with stimulus. Thus, these data indicate that BBCT may be beneficial in the cessation of inflammatory processes of cerebral infarction through suppression on the production of pro-inflammatory cytokines.

• Journal of Life Science
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• v.19 no.12
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• pp.1690-1697
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• 2009

CD11c and costimulatory molecules such as CD80 and CD86 express mainly in dendritic cells (DCs). In this study, we investigated the biologic effects of recombinant Fms-like tyrosine kinase-3 (Flt-3) ligand on the expression of DC surface markers, including CD11c in leukemia cell lines, such as KG-1, HL-60, NB4, and THP-1 cells. The expression of the Flt-3 receptor was found in NB4 and HL-60 cells, as well as KG-1 cells, but not in THP-1 cells. When KG-1 cells were cultured in a medium containing Flt-3 ligand or granulocyte macrophage-colony stimulating factor (GM-CSF) plus tumor necrosis factor (TNF)-$\alpha$, cell proliferation was inhibited and the expression levels of CD11c, major histocompatibility complex (MHC)-I, and MHC-II were increased in the cells. Flt-3 ligand also increased the expression level of CD11c on HL-60 and NB4 cells, but not on THP-1 cells. In comparison with CD11c expression, the expression level of CD11b on KG-1 cells, but not on NB4 and HL-60 cells, was slightly increased by Flt-3 ligand. Flt-3 ligand induced phosphorylation of extracellular signal-regulated kinase-1/2 (ERK-1/2) and p38-mitogen-activated protein kinase (p38-MAPK) in KG-1 cells, and the up-regulation of CD11c expression by Flt-3 ligand in the cells was abrogated by PD98059, an inhibitor of MEK. The results suggest that Flt-3 ligand up-regulates DC surface markers on $CD34^+$ myelomonocytic KG-1 cells, as well as promyelocytic leukemia cells, and that the differentiation of the leukemia cells into DC-like cells by Flt-3 ligand is mediated by ERK-1/2 activity.

• Journal of Acupuncture Research
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• v.40 no.4
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• pp.368-376
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• 2023

Background: Although bee venom (BV) has clinical benefits in osteoarthritis and rheumatoid arthritis, it has not been tested as treatment for gouty arthritis. Moreover, in vitro, BV has been proven to exhibit anti-inflammatory and positive effects on osteoarthritis, but only limited evidence can confirm its beneficial effects on gout. Thus, this study aims to assess the anti-inflammatory effects of BV on monosodium urate (MSU)-induced THP-1 monocytes. Methods: THP-1 monocytes were differentiated into mature macrophages using phorbol 12-myristate 13-acetate (PMA) and pretreated for 6 hours with BV and a Caspase-1 inhibitor in a physiologically achievable range of concentrations (BV, 0.1-1 ㎍/mL; Caspase-1 inhibitor, 1-10 μM), followed by MSU crystal stimulation for 24 hours. The secretions of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), IL-6, IL-8, cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), and nitric oxide (NO) were increased in the MSU crystal-stimulated THP-1 cells. Results: Caspase-1 inhibitors suppressed the production of all mediators in a dose-dependent manner. BV worked on equal terms with Caspase-1 inhibitors and showed more satisfactory effects on TNF-α, PGE2, COX-2, and inducible nitric oxide synthase (iNOS). Moreover, the western blot analysis revealed that BV regulated the transcriptional levels of these mediators via the suppression of extracellular signal-regulated kinase (ERK) pathway activation. Conclusion: The results of the present study clearly suggest that BV inhibits MSU-induced inflammation in vitro, suggesting a possible role for BV in gout treatment.