Punicalagin Regulates Key Processes Associated with Atherosclerosis in THP-1 Cellular Model

Atherosclerosis may lead to cardiovascular diseases (CVD), which are the primary cause of death globally. In addition to conventional therapeutics for CVD, use of nutraceuticals that prevents cholesterol deposition, reduce existing plaques and hence anti-atherosclerotic effects of nutraceuticals appeared to be promising. As such, in the present study we evaluated the beneficial effects of punicalagin, a phytochemical against an atherosclerotic cell model in vitro. Cytotoxicity assays were examined for 10 µM concentration of punicalagin on THP-1 macrophages. Real-time-polymerase chain reaction (RT-PCR) was used to analyze monocyte chemoattractant protein-1 (MCP-1) and Intercellular adhesion molecule (ICAM-1) expressions. Monocyte migration and cholesterol efflux assays were performed to investigate punicalagin’s further impact on the key steps of atherosclerosis. Cytotoxicity assays demonstrated no significant toxicity for punicalagin (10 µM) on THP-1 macrophages. Punicalagin inhibited the IFN-γ-induced overexpression of MCP-1 and ICAM-1 in macrophages by 10 fold and 3.49 fold, respectively, compared to the control. Punicalagin also reduced the MCP-1- mediated migration of monocytes by 28% compared to the control. Percentages of cellular cholesterol efflux were enhanced in presence or absence of IFN-γ by 88% and 84% compared to control with 58% and 62%, respectively. Punicalagin possesses anti-inflammatory and anti-atherosclerotic effects. Punicalagin also did not exhibit any cytotoxicity and therefore can be considered a safe and potential candidate for the treatment and prevention of atherosclerosis.

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In vitro: The Modulating Effect of Myricetin on the Atherosclerosis Related Processes in THP1 Macrophages

Journal of Pharmaceutical Research International ◽

10.9734/jpri/2020/v32i4731116 ◽

2021 ◽

pp. 62-73

Author(s):

Rehab F. Almassabi ◽

Etimad A. Huwait ◽

Sanaa J. Almowallad ◽

Salma Y. Saddeek ◽

Kalamegam Gauthaman

Keyword(s):

Real Time ◽

Cholesterol Efflux ◽

Control Treatment ◽

Intercellular Adhesion Molecule ◽

Research Centre ◽

Protective Effects ◽

Qrt Pcr ◽

Monocyte Migration ◽

Ifn Γ

Aims: To assess the anti-atherosclerotic effects of Myricetin (pharmaceutical) in human THP-1 macrophages following IFN-γ or MCP-1 stimulation. Study Design: The protective effects of myricetin against atherosclerosis was evaluated using the humanTHP-1 macrophages and studying the following parameters namely, cell viability, cell proliferation, cell migration, inflammation related gene expression and cholesterol efflux in vitro. Place and Duration of Study: THP-1 cell line: Department of biochemistry (faculty of science), Cell Culture Unit, Experimental Biochemistry Unit (King Fahad Medical Research Centre), King Abdul Aziz University, between September 2019 and September 2020. Methodology: The THP-1 cell lines were differentiated into macrophages by incubation with PMA (160 nM) for 24 hours. The viability percentage was determined using Pierce LDH cytotoxicity assay kit, the percentage change in macrophages proliferation was evaluated by crystal violet dye, the RNA was extracted then the cDNA was synthesized and the quantitative real time polymerase chain reaction (qRT-PCR) was done for inflammation-related genes, ICAM-1 and MCP-1. The percentage of monocyte migration and cholesterol efflux were also calculated. Results: Cytotoxicity assays demonstrated no significant toxicity with myricetin at 25μM and 50 μM concentrations on THP-1 macrophages. The quantitative real-time RT-PCR (qRT-PCR) demonstrated a significant increase in interferon gamma (IFN-γ) mediated expression of both intercellular adhesion molecule (ICAM-1) and monocyte chemo-attractant protein-1 (MCP-1) by 2.1 and 7.1 fold respectively, compared to the control. Treatment with myricetin (25 μM and 50 μM) significantly inhibited the IFN-γ induced overexpression of ICAM-1 by 42.86% & 71.34% and MCP-1 by 53.52% & 87.32% respectively. Myricetin (25 μM) significantly reduced the migration of monocytes by 33.66% compared to MCP-1. The cholesterol efflux from THP-1 macrophages treated with myricetin was significantly increased by 47% and 57% in the absence and presence of IFN-γ, respectively compared to the control. Conclusion: Myricetin has anti-inflammatory effects and supports cholesterol efflux, which can help in prevention of atherosclerosis. Furthermore, myricetin did not exhibit any cytotoxic effects and therefore is a safe phytochemical which can complement conventional therapeutics.

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Antiatherogenic Effects of Quercetin in the THP-1 Macrophage Model In Vitro, With Insights Into Its Signaling Mechanisms Using In Silico Analysis

Frontiers in Pharmacology ◽

10.3389/fphar.2021.698138 ◽

2021 ◽

Vol 12 ◽

Author(s):

Etimad A. Huwait ◽

Salma Y. Saddeek ◽

Rehab F. Al-Massabi ◽

Sanaa J. Almowallad ◽

Peter Natesan Pushparaj ◽

...

Keyword(s):

Gene Expression ◽

In Silico ◽

Cholesterol Efflux ◽

Antioxidant Properties ◽

Intercellular Adhesion Molecule ◽

Lipid Uptake ◽

Gene Expression Assay ◽

Monocyte Migration ◽

Signaling Mechanisms

Background: Atherosclerosis (AS), a major risk factor for stroke and brain tissue destruction, is an inflammatory disease of the blood vessels, and the underlying pathology is inflammation mediated by various chemokines and cytokines. Quercetin, a natural flavonol, is reported to have both anti-inflammatory and antioxidant properties. As such, in the present study, we evaluated the antiatherogenic effects of quercetin in a human THP-1 cell line in vitro and also the signaling mechanisms using in silico analysis.Materials and Methods: THP-1 macrophages exposed to different concentrations of quercetin (5–100 μM for 24 h) were tested for cytotoxicity. Real-time gene expression assay for intercellular adhesion molecule-1 (ICAM-1) and monocyte chemoattractant protein-1 (MCP-1) was carried out following treatment with quercetin at 15 and 30 μM for 24 h either in the absence or presence of interferon (IFN-γ) for 3 h to induce inflammation. Monocyte migration and cholesterol efflux were also assessed.Results: Quercetin did not exert any cytotoxic effects on THP-1 cells at the various concentrations tested. The gene expression assay showed a significant decrease in ICAM-1 (by 3.05 and 2.70) and MCP-1 (by 22.71 and 27.03), respectively. Quercetin at 15 µM decreased THP-1 monocyte migration by 33% compared to the MCP-1-treated cells. It also increased cholesterol efflux significantly by1.64-fold and 1.60-fold either alone or in combination with IFN-γ, respectively. Ingenuity Pathway Analysis of the molecular interactions of quercetin identified canonical pathways directly related to lipid uptake and cholesterol efflux. Furthermore, CD36, SR-A, and LXR-α also demonstrated significant increases by 72.16-, 149.10-, and 29.68-fold, respectively.Conclusion: Our results from both in vitro and in silico studies identified that quercetin inhibited the THP-1 monocyte migration, MCP-1, and ICAM-1 and increased cholesterol efflux probably mediated via the LXR/RXR signaling pathway. Therefore, quercetin will help prevent cell infiltration in atherosclerotic plaques and reduce the risk of stroke or brain destruction.

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Folate-modified, curcumin and paclitaxel co-loaded PLA-TPGS nanoparticles: preparation, optimization and in vitro cytotoxicity assays

Advances in Natural Sciences Nanoscience and Nanotechnology ◽

10.1088/2043-6254/aabb5c ◽

2018 ◽

Vol 9 (2) ◽

pp. 025004 ◽

Cited By ~ 1

Author(s):

Hai Doan Do ◽

Hao Le Thi ◽

Thu Huong Le Thi ◽

Hoai Nam Nguyen ◽

Van Khanh Bui ◽

...

Keyword(s):

In Vitro Cytotoxicity ◽

Cytotoxicity Assays

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A Comparison of In Vitro Cytotoxicity Assays and Their Application to Water Samples

Alternatives to Laboratory Animals ◽

10.1177/026119298701500104 ◽

1987 ◽

Vol 15 (1) ◽

pp. 20-29 ◽

Cited By ~ 1

Author(s):

Stephen M. Hunt ◽

Christina Chrzanowska ◽

Christopher R. Barnett ◽

Helen N. Brand ◽

John K. Fawell

Keyword(s):

Cell Growth ◽

Water Samples ◽

Industrial Effluent ◽

Exposure Period ◽

In Vitro Cytotoxicity ◽

Cloning Efficiency ◽

Cytotoxicity Assays ◽

River Water Samples ◽

Vital Dye

A group of 13 compounds were tested for in vitro cytotoxicity in four test systems; MIT-24 test, inhibition of cell growth (protein method), inhibition of cell growth (vital dye method) and cloning efficiency. In general, all four assays tended to rank compounds in a similar order for toxicity. The length of the exposure period appeared to be important for some compounds. The cytotoxicity of a variety of water samples was examined in two tests; inhibition of cell growth (vital dye method) and cloning efficiency. Under the conditions in which the assays were carried out, the latter proved to be the more sensitive test. River water samples gave little or no indication of cytotoxicity, samples of domestic sewage effluent gave some evidence of cytotoxicity, while an industrial effluent was markedly cytotoxic.

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Synthesis and Properties of CurNQ for the Theranostic Application in Ovarian Cancer Intervention

Molecules ◽

10.3390/molecules25194471 ◽

2020 ◽

Vol 25 (19) ◽

pp. 4471

Author(s):

Lara G. Freidus ◽

Pradeep Kumar ◽

Thashree Marimuthu ◽

Priyamvada Pradeep ◽

Viness Pillay ◽

...

Keyword(s):

Ovarian Cancer ◽

Cancer Detection ◽

Ovarian Cancer Cell ◽

In Vitro Cytotoxicity ◽

Cancer Targeting ◽

The Novel ◽

Cytotoxicity Assays ◽

Ovarian Cancer Cell Lines ◽

Cancer Intervention

Synthesis of a novel theranostic molecule for targeted cancer intervention. A reaction between curcumin and lawsone was carried out to yield the novel curcumin naphthoquinone (CurNQ) molecule (2,2′-((((1E,3Z,6E)-3-hydroxy-5-oxohepta-1,3,6-triene-1,7-diyl) bis(2-methoxy-4,1-phenylene))bis(oxy))bis(naphthalene-1,4-dione). CurNQ’s structure was elucidated and was fully characterized. CurNQ was demonstrated to have pH specific solubility, its saturation solubility increased from 11.15 µM at pH 7.4 to 20.7 µM at pH 6.8. This pH responsivity allows for cancer targeting (Warburg effect). Moreover, CurNQ displayed intrinsic fluorescence, thus enabling imaging and detection applications. In vitro cytotoxicity assays demonstrated the chemotherapeutic properties of CurNQ as CurNQ reduced cell viability to below 50% in OVCAR-5 and SKOV3 ovarian cancer cell lines. CurNQ is a novel theranostic molecule for potential targeted cancer detection and treatment.

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CD74 Expression by AML Cell Lines and Bone Marrow Specimens, and Augmented In Vitro Cytotoxicity of Anti-CD74 Antibody After Interferon-Gamma (IFN-γ) Treatment

Blood ◽

10.1182/blood.v116.21.2888.2888 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2888-2888

Author(s):

Abhinav B. Chandra ◽

Jack Burton ◽

Rhona Stein ◽

Susan Chen ◽

Nidhi Mishra ◽

...

Keyword(s):

Bone Marrow ◽

Cell Lines ◽

Conflicts Of Interest ◽

Cellular Signaling ◽

In Vitro Cytotoxicity ◽

Cancer Type ◽

Phosphorylated Akt ◽

Wide Range ◽

Ifn Γ

Abstract Abstract 2888 Background: CD74 (HLA-DR-associated invariant chain) is expressed alone or along with DR in a wide range of hematologic cancers and solid tumors. Humanized anti-CD74 mAb, milatuzumab (Immunomedics, Morris Plains, NJ), exhibits direct cytotoxicity for NHL, CLL and MM cell lines, and is undergoing clinical evaluation for treatment of these malignancies. CD74 is upregulated by interferons in hematologic and epithelial cancer cell lines. Here we present the results of our analysis of CD74 expression and function in AML, and the effect of CD74 upregulation by treatment with IFN-γ on the cytotoxicity of milatuzumab for AML cell lines. Methods: CD74 expression in bone marrow biopsy (BMB) specimens from non-M3 AML patients was evaluated by immunohistochemistry and, for the 3 human AML cell lines, by flow cytometry, with/without permeabilization and with/without IFN-γ (40 and 200 U/mL). These cell lines were also tested in proliferation assays for responses to milatuzumab, with/without IFN-γ. Also, assessment of apoptosis and cellular signaling was performed. Results: In the initial group of AML cases, 13/14 BMB specimens showed moderate to strong CD74 expression by leukemic blasts, which was mostly intracellular, usually with a perinuclear distribution. Three AML cell lines also showed moderate to strong expression of CD74, which was mostly intracellular. Without IFN-γ, surface expression of CD74 was present, but IFN-γ treatment of these 3 lines resulted in upregulation of surface CD74 by 69–117%. Much higher levels of intracellular CD74 were observed in all 3 lines (with and without IFN-γ), with IFN-γ-induced upregulation of intracellular CD74 in all 3 lines (from 85%-868%; P<0.001). In 2/3 lines, IFN-γ increased milatuzumab-mediated growth inhibition (23.7 to 44.8% and -3.9 to 30.9%, P=0.01 and P<0.05, respectively). Cytotoxicity was in part due to apoptosis, as significant increases in Annexin V binding (P=0.01) were observed after treatment with IFN-γ plus milatuzumab. Initial experiments addressing cellular signaling suggest a role for AKT, because phosphorylated AKT levels increased (P=0.06) in response to IFN-γ + milatuzumab. Conclusions: CD74 is expressed in AML patient specimens and in AML cell lines, with the majority of CD74 expression found intracellularly. Cell surface and cytoplasmic expression of CD74 were upregulated in AML lines after IFN-γ exposure. This increased expression resulted in increased cytotoxicity of the anti-CD74 mAb, milatuzumab, in 2/3 AML lines. This effect was through apoptosis and involved the AKT pathway. Thus, AML is another cancer type where combined IFN-γ and milatuzumab treatment may be useful. Supported in part by NIH grant PO1-CA103985 (DMG). Disclosures: No relevant conflicts of interest to declare.

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