HL-60 cells were introduced from Shandong Medical Academy of Science. Hss was obtained from Qingdao Haisheng Oncology Hospital (Shandong, China, batch number 990211). Medium powder RPMI-1640 was purchased from Gibco, further compounded with tri-distilled water and combined with a final concentration of 100 u/ml penicillin and 100 ug/ml of streptomycin. Calf serum was produced by Sijiqing Company, and tretinoin and AS203 were purchased from Sigma. The water-insoluble drugs, tretinoin and AS203 were dissolved by dimethysulfoxide (DMSO from Sigma), filtered, sterilized and diluted into different concentrations via culture solution RPMI-1640 before the experiment. The final concentration of DMSO was less than 0.1%. FTCT labeled mAb CD11b, PE labeled McAb CD15 and its corresponding isotype control were purchased from Sigma. Methyl thiazolyl tetrazolium (MTT) was purchased from Sigma and prepared as 5g/L solution with PBS of pH 7.4 before use. RT-PCR, dNTP, TapDNA polymerase and RNA enzyme inhibitor were purchased from Qiagene. The objective primer, according to the reported reference, was synthesized by the DNA synthesis department of Shanghai Biotechnology Company.
Tretinoin
Manufactured by Merck Group
Sourced in Germany
Tretinoin is a laboratory product manufactured by Merck Group. It is a retinoid compound used in various research and experimental applications.
Automatically generated - may contain errors
Lab products found in correlation
Rpmi 1640, by Thermo Fisher Scientific (2 mentions)
Ciclopirox, by Merck Group (2 mentions)
Fludrocortisone, by Merck Group (2 mentions)
Pentoxifylline, by Merck Group (2 mentions)
Lansoprazole, by Merck Group (2 mentions)
Sirolimus, by Merck Group (2 mentions)
Fluticasone, by Merck Group (2 mentions)
Ciclosporin, by Merck Group (2 mentions)
Dicycloverine, by Merck Group (2 mentions)
Isoxicam, by Merck Group (2 mentions)
Metixene, by Merck Group (2 mentions)
Haloperidol, by Merck Group (2 mentions)
Valproic acid, by Merck Group (2 mentions)
Remdesivir, by Selleck Chemicals (2 mentions)
Clofazimine, by Merck Group (2 mentions)
Bacampicillin, by Merck Group (2 mentions)
Myricetin, by Merck Group (2 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (2 mentions)
Dmem f12, by Thermo Fisher Scientific (2 mentions)
Anti cd4 clone gk1, by BioXCell (1 mentions)
6 protocols using tretinoin
1
HL-60 Cell Culture and Characterization
2
Mouse Chemotherapy and Immune Depletion
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Cyclophosphamide (Endoxan) was obtained from the Sir Charles Gardiner Hospital pharmacy (Perth, Australia) and dosed at 200 mg/kg. Chemotherapy drugs were prepared under sterile conditions and diluted in 0.9% saline solution. Drugs were administered intraperitoneally (i.p) at a maximum volume of 100 µl per 10 g weight of mouse.
All-trans-retinoic acid (≥ 98% by HPLC, powder) was purchased from Sigma-Aldrich (Macquarie Park, NSW) and suspended in dimethyl sulfoxide (DMSO; Sigma-Aldrich) at a concentration of 40 mg/ml for short term storage at − 80˚C. Mice were treated with 10 mg/kg of tretinoin for 9 consecutive days, intraperitoneally. Working concentrations of tretinoin were suspended in PBS and had a final DMSO concentration of 5%.
For all immune cell depletion experiments, antibody administration began three days before tumour inoculation. Anti-CD4 (clone GK1.5) and anti-CD8 (clone YTS 169; both BioXcell, New Hampshire, USA) were dosed i.p at 100 µg in 100 µl of PBS on day 3, day 0 and thereafter weekly until the end of experiment. Depletion was monitored in peripheral blood collected from the tail vein and analysed by flow cytometry.
All-trans-retinoic acid (≥ 98% by HPLC, powder) was purchased from Sigma-Aldrich (Macquarie Park, NSW) and suspended in dimethyl sulfoxide (DMSO; Sigma-Aldrich) at a concentration of 40 mg/ml for short term storage at − 80˚C. Mice were treated with 10 mg/kg of tretinoin for 9 consecutive days, intraperitoneally. Working concentrations of tretinoin were suspended in PBS and had a final DMSO concentration of 5%.
For all immune cell depletion experiments, antibody administration began three days before tumour inoculation. Anti-CD4 (clone GK1.5) and anti-CD8 (clone YTS 169; both BioXcell, New Hampshire, USA) were dosed i.p at 100 µg in 100 µl of PBS on day 3, day 0 and thereafter weekly until the end of experiment. Depletion was monitored in peripheral blood collected from the tail vein and analysed by flow cytometry.
3
Screening Diverse Compounds for Antiviral Efficacy
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Selection of compounds for testing was based on side effect profiles and compound availability. Bacampicillin (B0070000), ciclopirox (SML2011–50MG), ciclosporin (C2163000), clofazimine (1138904–200MG), dicycloverine (D1060000), fludrocortisone (1273003–200MG), fluticasone (1285873–100MG), haloperidol (H1512–5G), isoxicam (I1762–1G), lansoprazole (1356916–150MG), metixene (M1808000), myricetin (M6760–10MG), pentoxifylline (1508901–200MG), sirolimus (S-015–1ML), tretinoin (1674004–5X30MG), and valproic acid (1708707–500MG) were purchased from Sigma-Aldrich. Remdesivir (GS-5734) was purchased from Selleckchem. Compounds were resuspended in DMSO according to manufacturer’s instructions and serially diluted to the relevant concentrations for treatment of infected cells.
4
Directed Differentiation of iPSCs
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The iPSCs were cultured as aforementioned. iPSCs were separated by 0.25% trypsin every 4 days until passage 3 and were incubated with collagenase IV (1% in DMEM/F12; Invitrogen; Thermo Fisher Scientific, Inc.) at 37°C for 15 min in order to separate them from MEFs. After 4 days, iPSCs cultured in stem cell differentiation medium became embryonic bodies. Subsequently, the embryonic bodies were incubated with tretinoin (all-trans-retinoic acid, 500 nM; Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) and purmorphamin (1 µm; Calbiochem; EMD Millipore, Billerica, MA, USA) for 4 days at 37°C. The stem cell differentiation medium consisted of 50% DMEM/F12 (Gibco; Thermo Fisher Scientific, Inc.), 50% Neurobasal medium (Gibco; Thermo Fisher Scientific, Inc.), 1% N2 (Gibco; Thermo Fisher Scientific, Inc.), 1% B27 (Gibco; Thermo Fisher Scientific, Inc.), 10% serum fluid replacement, 0.1 mM mercaptoethanol (Sigma-Aldrich; Merck KGaA), 2 mM Glutamax (Gibco; Thermo Fisher Scientific, Inc.) and 2 µg/ml heparin (Sigma-Aldrich; Merck KGaA). A total of 24 days were required for iPSC preparation and differentiation.
5
Compound Selection and Preparation
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Selection of compounds for testing was based on side effect profiles and compound availability. Bacampicillin (B0070000), ciclopirox (SML2011-50MG), ciclosporin (C2163000), clofazimine (1138904-200MG), dicycloverine (D1060000), fludrocortisone (1273003-200MG), fluticasone (1285873-100MG), haloperidol (H1512-5G), isoxicam (I1762-1G), lansoprazole (1356916-150MG), metixene (M1808000), myricetin (M6760-10MG), pentoxifylline (1508901-200MG), sirolimus (S-015-1ML), tretinoin (1674004-5X30MG), and valproic acid (1708707-500MG) were purchased from Sigma-Aldrich. Remdesivir (GS-5734) was purchased from Selleckchem.
Compounds were resuspended in DMSO according to manufacturer’s instructions and serially diluted to the relevant concentrations for treatment of infected cells.
Compounds were resuspended in DMSO according to manufacturer’s instructions and serially diluted to the relevant concentrations for treatment of infected cells.
6
Immunomodulatory Effects of Chemical Compounds
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Tretinoin, Complete Freund's adjuvant (CFA), Nitro blue tetrazolium, dioxin, dimethyl sulfoxide (DMSO) and 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide (MTT) were obtained from Sigma-Aldrich (St.Louis, MO). RPMI 1640 and fetal calf serum were bought from GIBCO/Life Technologies Inc. (Gaithersburg, MD). The cytokine assay by enzyme-linked immunosorbent assay (ELISA) kits for interferon gamma (IFN-γ), interleukin (IL)-10 and IL-17 were procured from Bender MedSystems (Vienna, Austria). Mouse Regulatory T Cell Staining Kit was purchased from eBioscience (San Diego, CA).
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