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Plasma medicine is a rapidly advancing field. As one promising application, cold plasma has been ascribed a beneficial role in wound healing in which monocytes/macrophages play an important part. Wound healing and many other physiological and pathological processes are subject to redox control. Cold plasma expels reactive species of many kinds, thereby altering the redox state and mediating cell and tissue responses. Controlling this species efflux could be a valuable asset to control biological effects. We, therefore, investigated the biological response of a plasma treatment regimen that was gas shielded from the ambient environment. In this gas curtain, several N2:O2 ratios were investigated, and we discovered a modulation within the composition of active compounds ( ${\text {NO}}_{2}^{-}$ , ${\text {NO}}_{3}^{-}$ , and H2O2) in physiological solutions. Using a human monocyte cell line, the cell’s scavenging activity toward these molecules was assessed. With regard to cellular processes important in wound healing, such as cytokines release, O2 shielding provided stronger stimulation compared to N2 shielding. Altogether, the results support the idea that tuning the plasma-derived reactive species composition may be an interesting approach to improve plasma medical applications.

Pectins can modulate the biological responses interacting directly with immune cells. The observed responses can strongly be affected by polysaccharide structural features. We analyzed the intrinsic activation capacity of native and modified sweet pepper pectin on cytokine secretion by THP-1 macrophages as well as compare their effects in the presence of lipopolysaccharide. Modified pectin was obtained by partial acid hydrolysis which promoted the removal of side chains as well as the reduction of molecular weight and the degree of methyl esterification of native pectin. The results showed that both fractions had no effect on THP-1 viability. Native pectin at 300μg/mL increased TNF-α, IL-1β and IL-10 cytokine secretion by THP-1 macrophages. However, in the presence of lipopolysaccharide, it can attenuate the inflammatory response by reducing the production of the pro-inflammatory cytokines TNF-α and IL-1β and increasing the anti-inflammatory cytokine IL-10, as well as decreasing the TNF-α/IL-10 and IL-1β/IL-10 ratios. The structural modifications caused by acid hydrolysis affected the intrinsic activation capacity of native pectin to modulate the cytokines secretion. These results indicate that degree of methyl esterification, molecular weight and presence of side chains are important structural features of pectins involved in the modulation of cytokine secretion by THP-1 macrophages.

Effective therapy of acute myeloid leukemia (AML) remains an unmet need. DNA methylcytosine dioxygenase Ten-eleven translocation 1 (TET1) is a critical oncoprotein in AML. Through a series of data analysis and drug screening, we identified two compounds (i.e., NSC-311068 and NSC-370284) that selectively suppress TET1 transcription and 5-hydroxymethylcytosine (5hmC) modification, and effectively inhibit cell viability in AML with high expression of TET1 (i.e., TET1-high AML), including AML carrying t(11q23)/MLL-rearrangements and t(8;21) AML. NSC-311068 and especially NSC-370284 significantly repressed TET1-high AML progression in vivo. UC-514321, a structural analog of NSC-370284, exhibited a more potent therapeutic effect and prolonged the median survival of TET1-high AML mice over three fold. NSC-370284 and UC-514321 both directly target STAT3/5, transcriptional activators of TET1, and thus repress TET1 expression. They also exhibit strong synergistic effects with standard chemotherapy. Our results highlight the therapeutic potential of targeting the STAT/TET1 axis by selective inhibitors in AML treatment.

Note: BP, MV and LG, KG contributed equally Background Relapsed acute myeloid leukemia (AML) remains the most common reason for allogeneic hematopoietic cell transplant (HCT) failure. Thus, understanding AML immune escape mechanism is important for improving the odds of curing HCT patients with AML. Downregulation of HLA Class I and II expression by AML is one of the potential immune escape mechanisms. Therefore, treatment to restore HLA surface expression is crucial to prevent and treat relapse. Endogenous cytokines, such as IFN-γ, have been shown to stimulate HLA expression but are poorly tolerated by patients. However, two hypomethylating agents (HMA), decitabine (Dec) and azacitadine (Aza), that are routinely used in AML treatment are known to augment HLA expression. For AML, HMAs are often combined with venetoclax (Ven), a drug that blocks the anti-apoptotic B-cell lymphoma-2 (Bcl-2) protein. Thus, while HMAs have been reported to increase HLA expression, what is unknown is whether these agents impact individual HLA loci differently and whether Ven has any impact on HLA expression. To address these questions, we treated the THP-1 cell line with Dec, Aza or Ven and measured changes in cell-surface expression of HLA proteins by flow cytometry using locus-specific HLA mAbs. Methods THP-1 cells were incubated with IFN-γ (500 U/mL), Aza (2µM), Dec (5µM), or Ven (30nM) for 48 hours (drug concentrations were determined by earlier titration experiments). THP-1 cells are a monocytic cell line, derived from the peripheral blood of a childhood case of acute monocytic leukemia (M5 subtype), that express HLA Class I and HLA-DR but not HLA-DQ or -DP under basal conditions, although they are inducible by IFN-γ. Thus, the induction of HLA Class II expression by IFN-γ serves as a positive control. Isotype controls were included to measure background. Data is presented as the difference in MFI (delta MFI) between cells treated with a drug and those treated with diluent only. Results Treatment of THP-1 cells with either IFN-γ or Dec led to increases in Class I HLA-A, -B & -C (Figure 1) compared to untreated cells (a mean fold increase of 1.4 and 1.2, respectively). Notably, Aza did not stimulate additional HLA-C expression and induced less of an increase in HLA-A & -B expression (an increase of 1.1-fold) than IFN-γ or Dec. Treatment of THP-1 cells by Ven did not induce a change in HLA Class I expression. For Class II, IFN-γ or Dec increased HLA-DR, -DQ and -DP expression in comparison to untreated cells (Figure 1). IFN-γ induced greater HLA-DR expression compared to Dec (an increase of 2.3-fold and 1.5-fold, respectively), and both stimulated similar increases in HLA-DQ (increases of 1.5-fold and 1.4-fold, respectively) & -DP (increases of 1.9-fold and 1.5-fold, respectively). However, treatment of cells with either Aza or Ven did not lead to changes in HLA Class II expression. Discussion Previous studies have illustrated the ability of IFN-γ to induce HLA Class II expression in THP-1 cells, however, data for Dec to induce HLA Class II expression was unconfirmed. We report differences in the degree to which IFN-γ and Dec are capable of stimulating HLA-DR with IFN-γ being more potent. The inability of Aza to induce HLA Class II expression in THP-1 cells may be related to the differing drug activating pathways of the two HMAs. Indeed, there are conflicting reports as to whether Aza can stimulate HLA Class II expression. Though Ven treatment of THP-1 cells did not impact HLA expression, because it is given with HMAs, it remains to be seen what effect these drugs may have on HLA expression when administered together. Additional studies to confirm these observations in patient-derived AML blasts are ongoing. Conclusion We report that HMAs increased expression of HLA-A, -B, & -C loci and Dec but not Aza stimulated HLA-DR, -DQ, and -DP expression in THP-1 cells. Given these data, Dec may be superior in increasing HLA Class II expression post-HCT. Figure 1 Figure 1. Disclosures Marcucci: Abbvie: Other: Speaker and advisory scientific board meetings; Agios: Other: Speaker and advisory scientific board meetings; Novartis: Other: Speaker and advisory scientific board meetings. Al Malki: Neximmune: Consultancy; CareDx: Consultancy; Jazz Pharmaceuticals, Inc.: Consultancy; Rigel Pharma: Consultancy; Hansa Biopharma: Consultancy.

17 Páginas.-- 1 Tabla.-- 5 Figuras, Oleanolic acid (OA), a triterpene that is highly present in olive leaves, has been proposed as a component of functional foods for the prevention of metabolic syndrome, due to its anti-inflammatory activity. We analyzed the effects of OA on inflammatory parameters and signaling proteins in LPS-stimulated THP-1 macrophages. Thus, THP-1 macrophages were incubated with LPS for 48 h after pretreatment with OA at different concentrations. Pretreatment with OA was significantly effective in attenuating IL-6 and TNF-α overproduction induced by LPS in macrophages, and also improved the levels of AMPK-α. We also evaluated the effects of human triglyceride-rich lipoproteins (TRLs) derived from individuals consuming an OA-enriched functional olive oil. For this purpose, TRLs were isolated from healthy adolescents before, 2 and 5 h postprandially after the intake of a meal containing the functional olive oil or common olive oil, and were incubated with THP-1 macrophages. THP-1 macrophages incubated with TRLs isolated at 2 h after the consumption of the OA-enriched olive oil showed significant lower levels of IL-6 compared to the TRLs derived from olive oil. Our results suggest that OA might have potential to be used as a lipid-based formulation in functional olive oils to prevent inflammatory processes underlying metabolic syndrome in adolescents., This research was funded by the Andalusia 2014–2020 European Regional Development Fund (ERDF) Operative Program, grant number B-AGR-287-UGR18.

Pyroptosis plays an important role in the pathogenesis of numerous infectious, autoimmune, and inflammatory diseases, which makes it a promising target for intervention. In this study, the effect of luteolin on pyroptosis and the underlying mechanism were investigated using the canonical NLRP3 inflammasome in THP-1 macrophages induced by LPS/ATP. The results showed that luteolin exhibited a potent preventive effect on THP-1 macrophage pyroptosis, as evidenced by the increase in cell viability and the decrease in LDH release. Moreover, luteolin was found to significantly reduce the expression of NLRP3, pro-CASP-1 and CASP-1, which are the key components of NLRP3 inflammasome, as well as the expression of N-GSDMD and IL-1β, and we proved that the inhibition of luteolin on NLRP3 inflammasome activation is ROS-dependent. Furthermore, it was demonstrated that luteolin promoted Nrf2 nuclear translocation, thereby increasing the expression of HO-1 that reduces ROS production, while the anti-pyroptotic effect of luteolin was reversed by a specific Nrf2 inhibitor. Additionally, luteolin inhibited NF-κB p65 phosphorylation and nuclear translocation. In summary, we conclude that luteolin prevents THP-1 macrophage pyroptosis by suppressing ROS production via Nrf2 activation as well as NF-κB inactivation. These results support luteolin as a potential bioactive chemical against pyroptosis-related inflammatory diseases.

The human cell line activation test (h-CLAT) is an OECD approved (Test No. 442E) assay to identify novel skin sensitizers. h-CLAT simulates dendritic cell activation in the skin sensitization pathway and is based on the measurement of CD54 and CD86 overexpression on monocytic, leukemic THP-1 cells. However, the current h-CLAT markers show inconsistent results with moderate and weak sensitizers. Moreover, these markers have accessory roles in cell adhesion and signaling rather than a direct role in cellular inflammation. Therefore, we have explored other inflammation-related markers in this study. PBMCs comprises a mixture of cells that resemble the complex immunological milieu in adults and were primarily used to identify markers. PBMCs (n = 10) and THP-1 cells were treated with 1-chloro-2,4-dinitrobenzene (DNCB, strong) and NiCl2 (Ni, moderate) sensitizers or DMSO (control) and incubated for 24 h. The samples were subjected to RNA sequencing to obtain log2fold change in gene expression. DNCB and NiCl2 significantly upregulated 80 genes in both cell types. Of these, CD109, CD181, CD183, CLEC5A, CLEC8A & CD354 were experimentally validated. DNCB and Ni but not isopropyl alcohol (non-sensitizer) significantly induced the expression of all novel markers except CLEC8A. Moreover, the percentage induction of all novel markers except CLEC8A satisfied the OECD acceptance criteria. In summary, we identified five novel markers that may supplement the current repertoire of h-CLAT markers.

In this study Centaurea behen leaf aqueous extract was used for simple, rapid and eco-friendly synthesis of gold nanoparticles in room temperature. The obtained gold nanoparticles (AuNPs) were analyzed by various analytical technique including UV–vis absorption spectroscopy, Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD) technique, energy dispersive spectroscopy (EDX) and Transmission electron microscopy (TEM). For investigating of cytotoxicity effect and antioxidant properties of C. behen leaf aqueous extract and AuNPs, MTT assay and the DPPH test was used. The result showed that synthesized AuNPs have spherical shape with size below 50 nm and XRD pattern confirmed the crystal structure. Our prepared nanoparticles showed the cytotoxicity effects against THP-1 cell line. The IC50 of synthesized AuNPs for leukemia cell line was calculated to be around 25 µg/mL while the C. behen aqueous extract could not reach the IC50. C. behen extract and AuNPs exhibited a maximum DPPH scavenging activity of 31% and 14%, respectively. In conclusion, these results suggest that AuNPs produced by cost-effective, eco-friendly and green synthesis method, have a suitable anticancer activity against leukemia cell line.

Virosecurinine, a primary alkaloid from Securinega suffruticosa plant is known as a potent differentiation-inducing agent in acute leukemia cells. The present study aimed to investigate the effects and underlying mechanisms of virosecurinine on human leukemia THP-1 cells in vitro. The effects of virosecurinine on cell proliferation were assessed by CCK-8. The effects on apoptosis and cell cycle were assessed by staining with annexin V-fluorescein isothiocyanate and propidium iodide, respectively followed by flow cytometric analysis. The apoptotic cell bodies were observed using a transmission electron microscope, while the mRNA expression of phosphoinositide 3-kinase (PI3K), protein kinase B (AKT), mechanistic target of rapamycin (mTOR) and phosphatase and tensin homolog (PTEN) in THP-1 was evaluated by reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Treatment with virosecurinine was able to decrease the viability of THP-1 cells in a dose- and time-dependent manner. The IC50 values of virosecurinine at 24, 48, and 72 h post-treatment were 68.128, 23.615, and 13.423 µmol/l, respectively. Cell cycle was arrested at the G1/S phase in virosecurinine-treated cells; however, not in untreated control cells. Numerous apoptotic bodies were observed in the THP-1 cells, which were treated with 12.5 µmol/l virosecurinine for 48 h. RT-qPCR indicated that treatment with virosecurinine resulted in upregulated PTEN expression and downregulated expression of PI3K, AKT and mTOR in THP-1 cells. The present study demonstrated that treatment with virosecurinine was able to inhibit proliferation and induce apoptosis in THP-1cells by exerting an inhibitory effect on the activation of PI3K/AKT/mTOR signaling pathways. Therefore, our data suggested that virosecurinine is a promising anti-tumor agent for the treatment of acute monocytic leukemia.

Six new ferrocenyl guanidines were synthesized and characterized by elemental analysis, FT-IR and 1H and 13C NMR. Compounds 1–6 were screened for antioxidant, protein kinase inhibition, lethality of brine shrimp, and cytotoxicity against the human leukemia THP-1 cell line. The compounds demonstrated moderate to significant activities. Antioxidant activity of the synthesized compounds was evaluated by DPPH % inhibition with IC50 values. All compounds 1–6 showed notable antioxidant activity with DPPH having IC50 values between 23.2–15.1 and noteworthy brine shrimp lethality. Compound 6 demonstrated the highest cytotoxicity against brine shrimps with LC50 of 9.09 μg/mL. Protein kinase inhibition activity showed significant hyphea formation inhibition at 100 μg/disc with 10 to 13 mm clear zone of inhibition for the compounds 2–6. The compounds were screened for in vitro cytotoxicity using human leukemia cell line (THP-1 ATCC#TIB-202). Among all compounds, the most significant activity was demonstrated by compounds 6 and 5 with IC50 of 3.88 and 5.59 μg/mL which was comparable with the standard 5-flourouracil and vincristine drugs.

Purpose To investigate the role of phosphorylated JNK in Dectin-1-induced IL-1β production and the role of Dectin-1 in apoptosis in mouse Aspergillus fumigatus ( A. fumigatus ) keratitis. Methods Mice corneas were pretreated with Dectin-1 siRNA or SP600125 (the inhibitor of JNK) before A. fumigatus infection. THP-1 macrophages were preincubated with SP600125 before the stimulation of A. fumigatus conidia. Dectin-1, IL-1β, JNK, Bax, Bcl-2, cytochrome-c (cyt-c), caspase-9, caspase-8 and caspase-3 expressions were tested by PCR, Western blot, or Immunofluorescence staining. Results Pretreatment with Dectin-1 siRNA significantly decreased A. fumigatus -induced IL-1β production and JNK phosphorylation compared with scrambled control in C57BL/6 mice corneas. SP600125 treatment before infection significantly inhibited IL-1β production compared with DMSO control both in mice corneas and THP-1 macrophages. Furthermore, Dectin-1 deficiency resulted in increased ratio of Bax/Bcl-2, release of cyt-c, activation of caspase-9 and caspase-3 in mouse A. fumigatus keratitis . However, Dectin-1 deficiency didn't affect the activation of caspase-8. Conclusions Being an important inflammatory PRR to mediate host inflammatory response, Dectin-1 induced IL-1β production is JNK dependent in mouse A. fumigatus keratitis, and suppressed apoptosis mediated anti-inflammatory response.

Cytomegalovirus (CMV) infection is associated with glucocorticoid resistance in ulcerative colitis (UC) and may exacerbate the disease course. However, the underlying pathogenicity remains unclear. The aim of this study was to explore possible underlying mechanisms during CMV latency and lytic infection in the human mononuclear cell line THP-1. Latent and activated CMV infection cell models were established. We performed real-time PCR and western blotting to examine changes in glucocorticoid receptors (GRs) during CMV latency and activation. Pro-inflammatory and anti-inflammatory cytokines were detected by ELISA. After UV-inactivated CMV infection, GRs and cytokines were also examined. The expression of GRs was elevated in the reactivation group. An increased ratio of GR β/α and phosphorylation of GRα in the CMV reactivation group may explain refractory response to steroids. During CMV lytic infection, pro-inflammatory cytokines IL-6 and TNF-α increased remarkably and anti-inflammatory cytokine IL-5 decreased, which may exacerbate UC. GR and cytokines were unchanged in the UV-inactivated CMV infection group. Changes in the number and function of GRs may account for glucocorticoid resistance in CMV reactivation. The imbalance of pro- and anti-inflammatory cytokines may be related to severe inflammation.

Periodontitis is associated with development of diabetes mellitus. Although lipopolysaccharide (LPS) of Porphyromonas gingivalis (Pg), a major pathogen of periodontitis, may lead the progression of diabetes complications, the precise mechanisms are unclear. We, therefore, investigated the effects of β-carotene on production of Pg LPS-induced inflammatory cytokines in human monocytes cultured high glucose (HG) condition. THP-1 cells were cultured under 5.5 mM or 25 mM glucose conditions, and cells were stimulated with Pg LPS. To investigate the productivity of TNF-α, IL-6 and MCP-1, cell supernatants were collected for ELISA. To examine the effects of NF-kB signals on cytokine production, Bay11-7082 was used. HG enhanced Pg LPS-induced production of TNF-α, IL-6 and MCP-1 via NF-kB signals in THP-1. β-carotene suppressed the enhancement of the Pg LPS-induced cytokine production in THP-1 via NF-κB inactivation. Our results suggest that β-carotene might be a potential anti-inflammatory nutrient for circulating Pg LPS-mediated cytokine production in diabetic patients with periodontitis.

Extracellular Hsp70 (eHsp70) can act as damage-associated molecular pattern (DAMP) via Toll-like receptors TLR2 and TLR4, and stimulate immune and inflammatory responses leading to sterile inflammation and propagation of already existing inflammation. It was found elevated in the blood of patients with chronic obstructive pulmonary disease (COPD), who might suffer occasional bacterial colonizations and infections. We used a monocytic THP-1 cell line as a cellular model of systemic compartment of COPD to assess inflammatory effects of eHsp70 when present alone or together with bacterial products lypopolysaccharide (LPS) and lypoteichoic acid (LTA). THP-1 cells were differentiated into macrophage-like cells and treated with various concentrations of recombinant human Hsp70 protein (rhHsp70), LPS (TLR4 agonist), LTA (TLR2 agonist), and their combinations for 4, 12, 24, and 48 h. Concentrations of IL-1α, IL-6, IL-8, and TNF-α were determined by ELISA. Cell viability was assessed by MTS assay, and mode of cell death by luminometric measurements of caspases-3/7, -8, and -9 activities. rhHsp70 showed cell protecting effect by suppressing caspases-3/7 activation, while LPS provoked cytotoxicity through caspases-8 and -3/7 pathway. Regarding inflammatory processes, rhHsp70 alone induced secretion of IL-1α and IL-8, but had modulatory effects on release of all four cytokines when applied together with LPS or LTA. Combined effect with LPS was mainly synergistic, and with LTA mainly antagonistic, although it was cytokine- and time-dependent. Our results confirmed pro-inflammatory function of extracellular Hsp70, and suggest its possible implication in COPD exacerbations caused by bacterial infection through desensitization or inappropriate activation of TLR2 and TLR4 receptors.

Atlantic sea cucumber (Cucumaria frondosa) is a marine invertebrate that occurs naturally in the North Atlantic. Its collagen rich body walls make the sea cucumber an appreciated food source in many countries. The heteroglycans derived from the cartilage of Atlantic sea cucumbers have shown potential as a source of bioactive compounds, but further investigation is needed to confirm this hypothesis. The objectives of this work were to test the immunomodulating properties of heteroglycan fractions (FCFs) derived from Atlantic sea cucumbers toward phorbol 12-myristate 13-acetate (PMA) differentiated THP-1 macrophages. Oxygen Radical Absorbance Capacity (ORAC) was determined and cell culture media was collected and analyzed for presence of pro-inflammatory and anti-inflammatory cytokines (TNF-α, IL-6, and IL-10). FCFs toxicity toward macrophages was evaluated by light microscopy and confirmed by XTT proliferation assay. Heteroglycans derived from cartilage of Atlantic sea cucumber showed immunomodulatory activity in THP-1 macrophages. FCF-1, −2, and − 3) at concentrations 0.1–100 µg/mL induced levels of TNF-α, IL-6, and IL-10 proofing their bioactivity towards THP-1 macrophages. However, fractions of FCF-1 and FCF-2 may have induced oxidative stress in the macrophages, which possibly could lead to decreased cellular viability.

α7 nicotinic acetylcholine receptors (nAChRs) are ubiquitous in the nervous system and ensure important neurophysiological functionality for many processes. However, they are also found in cells of the immune system, where their role has been less studied. Here we report the pro-inflammatory effect of ImI, a well characterized conotoxin that inhibits α7 nAChRs, on differentiated THP-1 pre-monocyte macrophages (MDM) obtained by phorbol 12-myristate 13 acetate (PMA) treatment. Enzyme-linked immunosorbent assay (ELISA) performed on supernatant fluids of LPS challenged MDM showed ImI-mediated upregulation of pro-inflammatory cytokine TNF-α in an ImI concentration-dependent manner from 0.5 to 5.0 µmol/L and for IL-8 up to 1.0 µmol/L. Levels of anti-inflammatory cytokine TGF-β remained practically unaffected in ImI treated MDMs. Nicotine at 10 µmol/L significantly downregulated the release of TNF-α, but showed a lesser effect on IL-8 secretion and no effect on TGF-β. Fluorescent competitive assays involving ImI, α-bungarotoxin and nicotine using MDM and the murine macrophage RAW 264.7 suggest a common binding site in the α7 receptor. This work extends the application of conotoxins as molecular probes to non-excitatory cells, such as macrophages and supports the involvement of the α7 nAChR in regulating the inflammatory response via the cholinergic anti-inflammatory pathway (CAP).

Two biomaterials obtained by hydrodistilation from an indigenous Satureja hortensis plant (SHEO), and from the rhizoms of Zingiber officinale (ZOEO), purchased from Romanian market, were tested in ...

The purpose of this research project was to infection of human macrophages (THP-1) cell lines by H37Rv strain of Mycobacterium tuberculosis (H37RvMTB) and find out the ratio/dilution of mixture silver (Ag NPs) and zinc oxide nanoparticles (ZnO NPs) whose ability to eliminate phagocytized bacteria compared to rifampicin. The colloidal Ag NPs and ZnO NPs were synthesized and their characteristics were evaluated. The THP-1 cell lines were infected with different concentration of H37RvMTB. Next, the infected cells were treated with different ratios/dilutions of Ag NPs, ZnO NPs and rifampicin. The THP-1 were lysed and were cultured in Lowenstein-Jensen agar medium, for eight weeks. The TEM and AFM images of NPs and H37RvMTB were supplied. It is observed that Ag NPs, 2Ag:8ZnO and 8Ag:2ZnO did not have any anti-tubercular effects on phagocytized H37RvMTB. Conversely, ZnO NPs somehow eliminated 18.7 × 104 CFU ml-1 of H37RvMTB in concentration of ∼ 0.468 ppm. To compare with 40 ppm of rifampicin, ∼ 0.663 ppm of 5Ag:5ZnO had the ability to kill of H37RvMTB, too. Based on previous research, ZnO NPs had strong anti-tubercular impact against H37RvMTB to in-vitro condition, but it was toxic in concentration of ∼ 0.468 ppm to both of THP-1 and normal lung (MRC-5) cell lines. It also seems that 5Ag:5ZnO is justified because in concentration of ∼ 0.663 ppm of 5Ag:5ZnO, phagocytized H37RvMTB into the THP-1 had died without any toxicity effects against THP-1 and also MRC-5 cell lines. It is obvious that the mixture of colloidal silver and zinc oxide NPs with ratio of 5Ag:5ZnO would be trustworthy options as anti-tubercular nano-drugs in future researches.

The role of aryl hydrocarbon receptor (AhR) in carcinogenesis has been studied recently. Indole-3-carbinol (I3C) is an AhR agonist and a potential anticancer agent. Here, we investigated the effects of I3C on cell cycle progression and apoptosis through activation of AhR on THP-1 acute myeloid leukemia (AML) cell line.MTT viability assay was used to measure the cytotoxic effects of I3C on THP-1 cells. Apoptosis and cell cycle assays were investigated using flow cytometry. Real time RT-PCR was conducted to measure the alterations in the expression of AhR gene, key genes associated with AhR activation (IL1β and CYP1A1) and major genes involved in cell cycle regulation and apoptosis including P27, P21, CDK2, P53, BCL2 and FasR.Our findings revealed that I3C inhibits the proliferation of THP-1 cells in a dose- and time-dependent manner with minimal toxicity over normal monocytes. The AhR target genes (CYP1A1, IL1β) were overexpressed upon I3C treatment (p .05 to p .001). The antiproliferative effects of I3C were in association with programed cell death. I3C downregulated BCL2 and upregulated FasR in THP-1 cells (p .05 to p .001). G1 cell cycle arrest was also observed using flow cytometry. G1-acting cell cycle genes (P21, P27 and P53) were overexpressed (p .05 to p .001), while CDK2 was downregulated upon I3C treatment (p .01 to p .001).I3C could exert its antileukemic effects through AhR activation which is associated with programed cell death and G1 cell cycle arrest in a dose- and time-dependent manner. Therefore, AhR could be targeted as a novel treatment possibility in AML.

Macrophages (Mφ) undergo activation to pro-inflammatory (M1) or anti-inflammatory (M2) phenotypes in response to pathophysiologic stimuli and dysregulation of the M1-M2 balance is often associated with diseases. Therefore, studying mechanisms of macrophage polarization may reveal new drug targets. Human Mφ polarization is generally studied in primary monocyte-derived Mφ (PBMC Mφ) and THP-1-derived Mφ (THP-1 Mφ). We compared the polarization profile of THP-1 Mφ with that of PBMC Mφ to assess the alternative use of THP-1 for polarization studies. Cellular morphology, the expression profiles of 18 genes and 4 cell surface proteins, and phagocytosis capacity for apoptotic cells and S. aureus bioparticles were compared between these Mφ, activated towards M1, M2a, or M2c subsets by stimulation with LPS/IFNγ, IL-4, or IL-10, respectively, for 6 h, 24 h and 48 h. The Mφ types are unique in morphology and basal expression of polarization marker genes, particularly CCL22, in a pre-polarized state, and were differentially sensitive to polarization stimuli. Generally, M1 markers were instantly induced and gradually decreased, while M2 markers were markedly expressed at a later time. Expression profiles of M1 markers were similar between the polarized Mφ types, but M2a cell surface markers demonstrated an IL-4-dependent upregulation only in PBMC Mφ. Polarized THP-1 Mφ but not PBMC Mφ showed distinctive phagocytic capacity for apoptotic cells and bacterial antigens, respectively. In conclusion, our data suggest that THP-1 may be useful for performing studies involving phagocytosis and M1 polarization, rather than M2 polarization.

Inflammation is critical for the progression of hyperlipidemia. Although the exact mechanism through which inflammation affects hyperlipidemia is not very clear, evidence suggests that the tumor necrosis factor superfamily member 14 (TNFSF14/LIGHT)LIGHT might regulate lipid metabolism. In this study we investigated the expression of LIGHT in patients with different stages of coronary disease. The expression of lipid metabolism-related enzymes and inflammation-related proteins were further explored in oxidized low-density lipoproteins (oxLDL)-induced THP-1 macrophages. We found that LIGHT is highly expressed and companied with severe inflammations in patients with coronary disease. LIGHT significantly enhanced inflammation response in oxLDL-induced THP-1 macrophages. We further demonstrated that LIGHT markedly decreased the levels of lipolytic genes and increased the expressions of lipogenic genes in oxLDL-induced THP-1 macrophages. In addition, our results showed that LIGHT exerts its pro-inflammatory and pro-lipogenesis roles through activating nuclear factor-kappa B (NF-κB) signaling pathway. Taken together our study has demonstrated that LIGHT NF-κB-dependently exacerbates inflammation response and promotes lipid accumulation, and provided a new potential target for treatment of hyperlipidemia-related disease.

Hyperglycemia leads to diabetes and its diabetic complications. In this study, we investigated the synergistic effects of luteolin and fisetin on proinflammatory cytokine secretion and its underlying epigenetic regulation in human monocytes exposed to hyperglycemic (HG) concentrations. Human monocytic cells (THP-1) were cultured under controlled (14.5 mM mannitol), normoglycemic (5.5 mM glucose), or HG (20 mM glucose) conditions in the absence or presence of the two phytochemicals for 48 h. Whereas HG conditions significantly induced histone acetylation, nuclear factor-kappa B (NF-κB) activation, interleukin 6, and tumor necrosis factor-α release from THP-1 cells; combination treatments with the two phytochemicals (500 nM fisetin, and l μM and 500 nM luteolin) suppressed NF-κB activity and inflammatory cytokine release. Fisetin, luteolin, and their combination treatments also significantly decreased the activity of histone acetyltransferase, a known NF-κB coactivator; inhibited reactive oxygen species production; and activated sirtuin (SIRT)1 and forkhead box O3a (FOXO3a) expressions (P .05). Thus, combination treatments with the two phytochemicals inhibited HG condition-induced cytokine production in monocytes, through epigenetic changes involving NF-κB activation. We, therefore, suggest that combination treatments with luteolin and fisetin may be a potential candidate for the treatment and prevention of diabetes and its complications.

microRNAs (miRNAs) play important roles in the pathogenesis of atherosclerosis. A previous study has reported that miR-497 is elevated in advanced atherosclerotic lesions in an apoE-deficient (apoE-/-) mouse model. The purpose of this study is to test whether miR-497 can modulate macrophage foam cell formation, an initiating event in atherosclerosis. We found that miR-497 was upregulated in THP-1 macrophages after treatment with oxidized low-density lipoprotein (oxLDL). Enforced expression of miR-497 promoted lipid accumulation and decreased cholesterol efflux in oxLDL-exposed THP-1 macrophages. In contrast, downregulation of miR-497 suppressed oxLDL-induced lipid accumulation in THP-1 macrophages. Apelin was identified to be a downstream target of miR-497. Overexpression of miR-497 significantly reduced the expression of apelin in THP-1 macrophages. Interestingly, delivery of a miR-497-resistant variant of apelin significantly inhibited lipid accumulation and enhanced cholesterol efflux in miR-497-overexpressing THP-1 macrophages in response to oxLDL. In addition, miR-497 expression was increased and negatively correlated with apelin protein expression in human atherosclerotic lesions. In conclusion, miR-497 contributes to oxLDL-induced lipid deposition in macrophages largely via targeting of apelin and thus represents a potential therapeutic target for atherosclerosis.

Hemolymph is the circulating fluid of insects and is a key component of their immune system. However, little is known concerning hemocyte identification, development, differentiation and related cellular immune responses. The present study aimed to determine whether a hemocyte extract prepared from Bombyx mori larvae had anti-inflammatory effects; THP-1 (a human monocytic leukemia cell line) cells that had been differentiated into macrophage-like cells by treatment with phorbol myristate acetate (PMA) were used. THP-1 cells were cultured with different concentrations of a B. mori hemocyte extract prior to exposure to lipopolysaccharide (LPS) to induce an inflammatory response. The effects of the B. mori hemocyte extract on anti-inflammatory pathways were determined using reverse transcription-quantitative polymerase chain reaction and western blotting to assess the expression of pro-inflammatory molecules. The B. mori hemocyte extract inhibited the LPS-induced mRNA expression of Toll-like receptor 4 in addition to LPS-induced interleukin (IL)-1β, IL-6, IL-8 and tumor necrosis factor-α. Treatment of PMA-differentiated THP-1 cells with B. mori hemocyte extract also inhibited inducible nitric oxide synthase and cyclooxygenase-2 transcription and translation. Nuclear factor-κB activation and phosphorylation also decreased. Further in-depth functional studies are required to understand the mechanism underlying the anti-inflammatory effects of silkworm hemocyte extract.

The aim of the present study was to examine the apoptotic effect of cordycepin (COR) on human THP-1 acute monocytic leukemia cells. THP-1 cells were exposed to different concentrations of COR for 24, 48, 72 or 96 h. The cell viability and apoptotic rate were analyzed. The gene expression of Akt1, Akt2, Akt3, B-cell lymphoma 2 (Bcl-2) and Bcl-2-associated X protein (Bax) were assessed by reverse-transcription quantitative PCR. Western blot analysis was used to detect the protein levels of phosphorylated (p)-Akt, p-extracellular signal-regulated kinase (ERK) and cleaved caspase-3. It was found that the viability of THP-1 cells was inhibited by COR in a dose- and time-dependent manner. After treatment with 200 µM COR for 24 h, the percentage of apoptotic cells was significantly increased. COR also downregulated the levels of Bcl-2, Akt1, Akt2 and Akt3, and elevated the expression of Bax. The protein levels of p-Akt and p-ERK were suppressed and cleaved caspase-3 was increased after treatment of COR. In conclusion, COR was found to induce apoptosis of THP-1 acute monocytic leukemia cells through downregulation of ERK/Akt signaling.

Mononuclear phogocytic cells and epithelial cells are effective during the initiation and regulation of the innate immune response. They have an active role in mucosal immune response both mechanically and by interaction with other cells with cytokine release. Defensins are microbicidal peptides that are expressed in various cells and are thought to be effective in the first line defense against pathogens. IL-12 and IL-10, showing proinflamatory and antiinflamatory activities, respectively, are actors of the cellular immunity and limit the infection of the host without causing immunopathology. The aim of this study was to observe the differences in the release of IL-12 and IL-10 and the expression of human beta-defensin-3 (hBD-3) inCaco-2 (human colon epidermal adenocarcinoma cell) and THP-1 (human leukemia monocytic cell) cell lines cultured alone or in co-culture, by the stimulation of Toxoplasma gondii tachyzoites either in direct contact with the cells or separated by an insert filter from the cells. Twenty-four hours after the addition of RH strain tachyzoites to the cells, the supernatants were collected from the experiment wells, and commercial ELISA kits (Invitrogen) were used according to the manufacturers instructions to measure IL-12 and IL-10 levels. HBD-3 expression of cells collected from the experiment wells afterfour and 24 hourswere analyzed by using real time PCR. For this procedure, complementary c-DNA was obtained (Transcriptor High Fidelity cDNA Synthesis Kit, Roche Diagnostics, Germany)after the extraction of RNA with a commercial kit (High pure RNA isolation kit, Roche Diagnostics, Germany). IL-12 was higher than IL-10 in all experiment wells. IL-12 was induced more in the co-culture wells where Caco-2 and THP-1 cells were challenged together, than the wells in which the cells infected with T.gondii tachyzoites alone. No differences in respect to cytokine response were observed between the cells with which tachyzoites were in contact and the cells which were separated from the parasites with an insert. In hBD-3 experiments, Caco-2 and THP-1 cells interacted in co-culture wells when infected with tachyzoites and displayed a higher level of hBD-3 expression than the condition when they were infected alone. This study showed that, IL-12 release and hBD-3 expression, which play a role in innate immunity, are greater when various antigens of T.gondii interacted with stimulated mononuclear cell and epithelial cells.

// Jian Chen 1, 2, * , Qitian Mu 1, 2, 3, * , Xia Li 1, 2, * , Xiufeng Yin 1, 2 , Mengxia Yu 1, 2 , Jing Jin 1, 2 , Chenying Li 1, 2 , Yile Zhou 1, 2 , Jiani Zhou 1, 2, 4 , Shanshan Suo 1, 2 , Demin Lu 1, 2 and Jie Jin 1, 2 1 Department of Hematology, The First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China 2 Institute of Hematology, Zhejiang University, Hangzhou, China 3 Laboratory of Stem Cell Transplantation, Ningbo First Hospital, Zhejiang, China 4 Hematology Department of Ningbo Medical Center Lihuili Estern Hospital, Ningbo, China * These authors contributed equally to this work Correspondence to: Jie Jin, email: zjuhematology@163.com Keywords: homoharringtonine, smad3, TGF-β, acute myeloid leukemia Received: April 04, 2016 Accepted: March 29, 2017 Published: April 08, 2017 ABSTRACT Homoharringtonine (HHT) has long and widely been used in China for the treatment of acute myeloid leukemia (AML), the clinical therapeutic effect is significant but the working mechanism is poorly understood. The purpose of this study is to screen the possible target for HHT with virtual screening and verify the findings by cell experiments. Software including Autodock, Python, and MGL tools were used, with HHT being the ligand and proteins from PI3K-Akt pathway, Jak-stat pathway, TGF-β pathway and NK-κB pathway as the receptors. Human AML cell lines including U937, KG-1, THP-1 were cultured and used as the experiment cell lines. MTT assay was used for proliferation detection, flowcytometry was used to detect apoptosis and cell cycle arrest upon HHT functioning, western blotting was used to detect the protein level changes, viral shRNA transfection was used to suppress the expression level of the target protein candidate, and viral mRNA transfection was used for over-expression. Virtual screening revealed that smad3 from TGF-β pathway might be the candidate for HHT binding. In AML cell line U937 and KG-1, HHT can induce the Ser423/425 phosphorylation of smad3, and this phosphorylation can subsequently activate the TGF-β pathway, causing cell cycle arrest at G1 phase in U937 cells and apoptosis in KG-1 cells, knockdown of smad3 can impair the sensitivity of U937 cell to HHT, and over-expression of smad3 can re-establish the sensitivity in both cell lines. We conclude that smad3 is the probable target protein of HHT and plays an important role in the functioning mechanism of HHT.

Zerumbone은 야생 생강의 일종인 Zingiber zerumbet Smith의 정유(essential oil)에 포함되어 있는 주요성분으로, 다양한 연구를 통해 혈액종양을 포함한 암, 염증질환, 활성산소 감소 등에 이용할 가능성이 꾸준히 제기되어왔다. 또한 면역세포들의 증식과 세포주기진행, 사이토카인의 생성ㆍ발현에도 효과를 나타낸다고 알려져 있다. 이외에도 간보호, 통증완화, 항동맥경화, 항미생물 등 다양한 생물학적 기능이 보고되었다. 본 연구에서는 zerumbone이 단구의 활성과 기능에 어떠한 영향을 미치는지 알아보고자 하였다. 우선, 단구세포주 THP-1세포의 활성이 zerumbone에 의해 증가되는 것을 확인하였다. LPS 처리 후 가장 강한 THP-1 증식 증가는 5 μM의 zerumbone 처리시 나타났으며, 세포 단독 증식 증가는 10 μM의 zerumbone 처리 시 가장 증가하였다. 반면에 50 μM 의 zerumbone 처리 시에는 LPS 존재 여부와 관계없이 급격한 증식 감소가 관찰되었다. LPS의 처리에 의해 유도되는 신호전달 단백질 Erk의 인산화도 zerumbone에 의해 증가되었다. 가장 강한 인산화 증가는 증식 감소가 나타난 50 μM의 zerumbone을 처리했을 때 관찰되었다. 전사인자 NF-κB의 활성은 zerumbone 단독 처리 시에는 큰 변화가 없었지만, LPS 와 동시에 처리했을 경우 증가하는 것으로 관찰되었다. 나아가, NFκB에 의해 발현이 조절되는 염증 사이토카인 TNF-α, IL-8의 전사도 zerumbone 에 의해 증가되는 것을 확인하였다. 이와 같은 결과를 통해 zerumbone이 단핵구의 증식과 활성을 강화시킬 수 있음을 확인하였다. 나아가, zerumbone이 LPS 를 보유한 세균 감염 시 단핵구 활성 증가를 통하여 효과적인 탐식과 면역반응을 강화시켜 효과적인 세균 처리에 도움을 줄 수 있을 것으로 여겨진다.

// Nianyi Li 1 , Qinfen Chen 1 , Jingwen Gu 1 , Shuang Li 1 , Guangjie Zhao 1 , Wei Wang 1 , Zhicheng Wang 1 and Xiaoqin Wang 1 1 Department of Haematology, Huashan Hospital, Fudan University, Shanghai, China Correspondence to: Xiaoqin Wang, email: wangxiaoqin@shmu.edu.cn Qinfen Chen, email: chenqinfen@126.com Keywords: synergistic effect, deferasirox, decitabine, iron chelation therapy, demethylation Received: June 19, 2016 Accepted: March 14, 2017 Published: March 27, 2017 ABSTRACT A multi-center study from the French Myelodysplastic Syndrome (MDS) Group confirmed that iron chelation therapy is an independent prognostic factor that can increase the survival rate of patients who are suffering from transfusion-dependent low-risk MDS. In this study, we aimed to explore this clinical phenomena in vitro , by exploring the synergistic effect of the iron chelator Deferasirox (DFX) and the DNA methyl transferase inhibitor Decitabine (DAC) in the leukemia cell lines SKM-1, THP-1, and K-562. Treatment with both DFX or DAC promoted apoptosis, induced cell cycle arrest, and inhibited proliferation in all three of these cell lines. The combination of DFX and DAC was much greater than the effect of using either drug alone. DFX showed a synergistic effect with DAC on cell apoptosis in all three cell lines and on cell cycle arrest at the G0/G1 phase in K-562 cells. DFX decreased the ROS levels to varying degrees. In contrast, DAC increased ROS levels and an increase in ROS was also noted when the two drugs were used in combination. Treatment of cells with DAC induced re-expression of ABAT, APAF-1, FADD, HJV, and SMPD3, presumably through demethylation. However the combination of DAC and DFX just had strong synergistic effect on the re-expression of HJV.

Carbon monoxide (CO), a reaction product of cytoprotective enzyme heme oxygenase-1 (HO-1), is a gaseous messenger with anti-proliferative, anti-apoptotic, and anti-inflammatory actions in many cell types. Here, we investigated the role of CO on the process of monocyte differentiation induced by phorbol 12-myristate 13-acetate (PMA) in human monocytic THP-1 cells. CORM-2 (tricarbonyldichlororuthenium (II) dimer, Ru₂Cl₄ (CO) 6 ), a CO-releasing compound, decreased a marked cell adherence with a slight reduction of proliferation in monocytic THP-1 cells treated with PMA. And, CORM-2 significantly inhibited expression of differentiation markers such as CD14, CD11b plus CD18 (macrophage-1 antigen, Mac-1 or complement receptor 3, CR3) and phagocytosis of carboxylate-modified red fluorescent latex beads, in PMA-stimulated THP-1 cells. For the further experiments, differentiation of PMA-treated cells was enhanced after the initial 2 days stimulus by removing the PMA-containing media then incubating the cells in fresh media for a another 4 days. And, we observed the secretion of inflammatory cytokines and phagocytosis in differentiated macrophages. Treatment with CORM-2 significantly abolished the secretion of IL-6, TNF-α and phagocytosis using fluorescence-conjugated E. coli (K-12 strain) bioparticles in lipopolysaccharide (LPS)-stimulated differentiated macrophages. In conclusion, these results suggest that CO inhibits the differentiation of monocytic THP-1 cells as well as the activation of differentiated macrophages.

Rho GDP-dissociation inhibitor β (RhoGDIβ), a regulator of the Rho family of proteins, is expressed abundantly in the hematopoietic cell lineage. During apoptosis of hematopoietic cells, RhoGDIβ is cleaved by caspase‑3 at Asp19 and this cleaved form (Δ19‑RhoGDIβ) has been implicated in the apoptotic pathway. To clarify the role of RhoGDIβ in hematopoietic cells, the present study performed immunoblotting and immunofluorescence staining to examine the expression of RhoGDIβ and ∆19‑RhoGDIβ during phorbol 12‑myristate 13‑acetate (PMA)‑stimulated differentiation of human THP‑1 monocytic cells to macrophages. During differentiation of the THP‑1 cells to macrophages, the expression of RhoGDIβ remained stable; however, the expression of Δ19‑RhoGDIβ increased, particularly in well‑spreading, non‑apoptotic cells, which differentiated into macrophages. These results suggested that Δ19‑RhoGDIβ has an apoptosis‑independent role in the PMA‑induced differentiation of THP‑1 cells to macrophages.

Macrophages are resistant to cell death and are one of HIV reservoirs. HIV viral protein Vpr has the potential to promote infection of and survival of macrophages, which could be a highly significant factor in the development and/or maintenance of macrophage viral reservoirs. However, the impact of vpr on macrophages resistance to apoptosis is yet to be comprehended. Autophagy is a cell survival mechanism under stress state. In this study, we investigated whether autophagy is involved in macrophages resistant to vpr-induced apoptosis. Using the THP1 macrophages, we studied the interconnection between macrophages resistance to apoptosis and autophagy. We found that vpr is able to trigger autophagy in transfected THP-1 macrophages confirmed by electron microscopy (EM) and western blot analysis, and inhibition of autophagy with 3MA increased vpr-induced apoptosis. The results indicate that autophagy may be responsible for maintenance of macrophage HIV reservoirs.

Background: We have recently found that lipooligosaccharide (LOS) isolated from encapsulated strains of Haemophilus influenzae ( H. influenzae ) has strong adjuvant, but diminished pro-inflammatory ability as compared to Escherichia coli lipopolysaccharide (LPS). In this study, we aimed to determine the immunostimulatory capacity of nontypeable/ non-encapsulated H. influenzae (NTHi) LOS by comparing the effect of killed bacteria with LOS isolated from the same strain. Methods: Following stimulation of human monocytic THP-1 cells with killed NTHi strain 375, or with the corresponding amount of LOS, we studied the protein and gene expression of immunostimulatory and antigen-presenting molecules, cytokines, and innate immune receptors. Results: Stimulation with LOS resulted in lower expression of adhesion (CD54, CD58) as well as costimulatory molecules (CD40, CD86), but in higher expression of antigen-presenting molecules (HLA-DR and HLA-ABC) compared to killed NTHi, whereas killed bacteria induced higher release of both TNF-α and IL-10. The results indicate that while LOS of NTHi has decreased capacity to induce pro-inflammatory responses compared to E. coli LPS or killed NTHi, this LOS has the potential to facilitate antigen presentation. Conclusions: Considering the important role of NTHi as a respiratory pathogen, and its currently increasing significance in the etiology of invasive infections, LOS deserves further attention as a vaccine antigen, which also has potent adjuvant properties. Keywords: Nontypeable Haemophilus influenzae , Lipooligosaccharide, THP-1 cells, innate immune responses

Chronic inflammatory diseases of the airways are associated with gastro-oesophageal reflux (GOR) and aspiration events. The observation of lipid-laden macrophages (LLMs) within the airway may indicate aspiration secondary to GOR. The proposed mechanism, that lipid droplets from undigested or partially digested food are aspirated leading to accumulation in scavenging macrophages, led us to hypothesise that an activated population of LLMs could interact with other immune cells to induce bronchial inflammation.To test this, we generated anin vitromodel using differentiated THP-1 cells, which were treated with a high-fat liquid feed.Here, we show that THP-1 cells can take up lipid from the high-fat feed independent of actin polymerisation or CD36-dependent phagocytosis. These cells did not exhibit M1 or M2 polarisation. Gene array analysis confirmed over 8000 genes were upregulated by at least twofold following high fat exposure, and IL-8 was the most upregulated gene. Pathway analysis revealed upregulation of genes known to be involved in chronic obstructive pulmonary disease (COPD) pathophysiology.We suggest that aspiration and macrophage phagocytosis may be important mechanisms in the aetiology of diseases such as COPD and cystic fibrosis that are characterised by high levels of IL-8 within the airways.

Aim: Atherosclerosis is a kind of chronic inflammatory disease. A crucial pathology change of atherosclerosis is the migration of activated VSMCs to the intima where they interact with leukocytes by expressing adhesion molecules, including intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Moreover, monocyte chemoattractant protein-1 (MCP-1) expressed by VSMCs plays an important role in recruiting monocytes and macrophages. Leech (Whitmania pigra Whitman) is a traditional Chinese medicine to treat cardiovascular diseases including atherosclerosis, however previous research has rarely reported the molecular mechanism for its curative effect. Thus, our study focuses on the effects of leech extracts on the expression of inflammatory factors, adhesion molecules and MCP-1 in rat VSMCs. Methods: In our present study, wound-healing assay and Boyden chamber model were applied to evaluate the anti-migration effect of LEE (Leech Enzyme Extracts) on LPS induced VSMCs. The anti-adhesion effect was assessed using DiI-labeled THP-1 and RAW264.7. Results: LEE suppressed LPS-induced VSMCs migration and decreased the chemotaxis and adhesive capacity of THP-1 and RAW264.7 to LPS-stimulated VSMCs. LEE also attenuated the upregulation of a variety of pro-atherosclerotic factors by inhibiting the phosphorylation of p38 MAPK. LEE was also observed to prevent NF-κB p65 nuclear localization using immune-fluorescent staining. Conclusions: In conclusion, LEE suppresses LPS-induced upregulation of inflammatory factors, adhesion molecules and MCP-1 in rat VSMCs mainly via inhibiting the p38 MAPK/NF-κB pathways, thus partly uncovered LEE's molecular mechanisms for its therapeutic effect on atherosclerosis.

Inflammation caused by the NLRP3 inflammasome has been linked to many diseases. Lunasin is a bioactive peptide from soybeans with reported anti-inflammatory properties. The objective of this work was to determine the effect of pepsin-pancreatin hydrolysis (PPH) on the ability of lunasin-enriched preparation (LEP) to inhibit inflammasome activation in differentiated THP-1 human macrophages. THP-1 macrophages were treated with different concentrations of LEP (0.0625 to 0.25 mg mL-1), primed with 1 μg mL-1 lipopolysaccharide for 6 h and activated by 5 mM adenosine triphosphate for 1 h. LEP reduced secretion of IL-1β and IL-18. In addition, LEP treatment inhibited the production of intracellular reactive oxygen species (ROS) in THP-1 human macrophages without affecting the expressions of NLRP3 and ASC proteins involved in inflammasomes. PPH reduced the ability of LEP to inhibit production of intracellular ROS. Our results showed that LEP inhibited activation of inflammasomes by reducing intracellular ROS in vitro which was reduced by PPH.

It is known that electrochemical energy can be harvested from mammalian cells, more specifically from white blood cells (WBC). This study focuses on an improved biofuel cell operating on phorbol myristate acetate (PMA) activated THP-1 human monocytic cells. Electrochemical investigation showed strong evidence pointing towards hydrogen peroxide being the primary current source, confirming that the current originates from NADPH oxidase activity. Moreover, an adequate substrate for differentiation and activation of THP-1 cells was examined. ITO, gold, platinum and glass were tested and the amount of superoxide anion produced by NADPH oxidase was measured by spectrophotometry through WST-1 reduction at 450nm and used as an indicator of cellular activity and viability. These substrates were subsequently used in a conventional two-compartment biofuel cell where the power density output was recorded. The material showing the highest cell activity compared to the reference cell culture plate and the highest power output was ITO. Under our experimental conditions, a power density of 4.5μW/cm2 was reached. To the best of our knowledge, this is a threefold higher power output than other leukocyte biofuel cells.