Human cervical cancer cell lines (Ca Ski, HeLa, SiHa, ME-180, and MS751) were purchased from the American Type Culture Collection (Manassas, VA, USA). The cell lines were maintained in complete media (RPMI 1640 for Ca Ski; DMEM for HeLa; MEM for SiHa and MS751; McCoy’s 5A for ME-180) supplemented with 10% fetal bovine serum (FBS) and 0.1% gentamicin sulfate (Gemini Bio-Products, Calabasas, CA, USA) in 5% CO2 at 37°C. The SPHK inhibitors SKI-II (2-(p-hydroxyanilino)-4-(p-chlorophenyl) thiazole; Calbiochem, Merck KGaA, Darmstadt, Germany) and FTY720 (Fingolimod; Cayman Chemical, Ann Arbor, MI, USA) were resuspended in dimethyl sulfoxide at a concentration of 100 μg/mL. Cells were seeded at 3 × 103 cells/well in a 96-well microplate in culture media with 10% FBS. Cells were treated with SPHK inhibitors as described in previous reports [32 (link), 33 (link)].
Fty720 fingolimod
Manufactured by Cayman Chemical
Sourced in United States
FTY720 (fingolimod) is a chemical compound used in research applications. It functions as a sphingosine 1-phosphate receptor modulator.
Automatically generated - may contain errors
Lab products found in correlation
Caski, by American Type Culture Collection (1 mentions)
Me 180, by American Type Culture Collection (1 mentions)
Ms751, by American Type Culture Collection (1 mentions)
Ski 2, by Merck Group (1 mentions)
Gentamicin sulfate, by Gemini Bio (1 mentions)
Cl316 243, by Bio-Techne (1 mentions)
Okadaic acid, by Abcam (1 mentions)
17β estradiol e2, by Innovative Research (1 mentions)
60 day release pellets, by Innovative Research (1 mentions)
α tubulin, by Merck Group (1 mentions)
Cell culture medium and supplements, by Thermo Fisher Scientific (1 mentions)
Poly adp ribose polymerase parp, by Cell Signaling Technology (1 mentions)
β actin, by Merck Group (1 mentions)
Sorafenib tosylate, by Bayer (1 mentions)
Ovalbumin (ova), by Merck Group (1 mentions)
Easysep mouse naive cd4 t cell isolation kit, by STEMCELL (1 mentions)
Control sheep serum igg, by Merck Group (1 mentions)
Szx12, by Olympus (1 mentions)
Mrs1523 3 propyl 6 ethyl 5 ethylthio carbonyl 2 phenyl 4 propyl 3 pyridine carboxylate, by Merck Group (1 mentions)
Sew2871, by Cayman Chemical (1 mentions)
10 protocols using fty720 fingolimod
1
Human Cervical Cancer Cell Lines and SPHK Inhibitor Treatment
2
Intracerebroventricular Injection and Ovariectomy Protocol
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Intracerebroventricular injection was performed as previously described (22 (link)). Briefly, mice were anesthetized with isoflurane, a small incision was made in the scalp, and an injection was achieved at a point 1 mm lateral and 1 mm caudal to bregma, at a depth of 2 mm. The volume for all intracerebroventricular injections was 1 μL in a 10-μL syringe (Hamilton). The syringe was left in place for 1 min to allow for infusate diffusion. The proper injection site was verified in pilot experiments by administration and localization of Evans Blue dye. Intracerebroventricular administration of okadaic acid (OA, 20 ng; Abcam) and FTY720 (fingolimod, 2.5 µg; Cayman) was performed twice a week (23 (link),24 (link)).
Ovariectomies were performed in 10-week-old mice, as described previously (25 (link)). For peripheral estrogen treatment, pellets releasing vehicle or 17β-estradiol (E2) (0.25 mg, 60-day release pellets; Innovative Research of America) were implanted 1 week after ovariectomy. The β3-adrenergic receptor agonist CL316243 (0.5 mg/kg; Tocris) was administered daily via i.p. route (26 (link)).
Ovariectomies were performed in 10-week-old mice, as described previously (25 (link)). For peripheral estrogen treatment, pellets releasing vehicle or 17β-estradiol (E2) (0.25 mg, 60-day release pellets; Innovative Research of America) were implanted 1 week after ovariectomy. The β3-adrenergic receptor agonist CL316243 (0.5 mg/kg; Tocris) was administered daily via i.p. route (26 (link)).
3
Sorafenib and Fingolimod Combination Therapy
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Sorafenib tosylate (BAY 54-9085) was provided by Bayer HealthCare Pharmaceuticals (Montville, NJ), FTY720 (Fingolimod) was purchased from Cayman Chemical Ann Arbor, Michigan, USA. (MI). Stock solutions of 100 mmol/L sorafenib and of 20 mmol/L FTY720 dissolved in Dimethyl sulfoxide (DMSO) were aliquoted and deep-frozen. Primary antibodies against poly-ADP ribose polymerase (PARP) and light chain 3II (LC3II) were from Cell Signaling (Cell Signaling Technology Inc., Danvers, MA). p62 from Sigma-Aldrich LLC. (St. Louis, MO), Apoptosis-inducing factor (AIF) from Santa Cruz Biotechnology, Inc. (Texas, USA), cytochrome c oxidase (Cox IV) from Novus Biologicals, LLC, (Littleton, Colorado, USA), Cytochrome c (Cyt c) from BD Bioscience (San Jose, CA), α-Tubulin and β-Actin from Sigma-Aldrich Co. LLC. were used. Secondary fluorescent antibodies, goat anti-rabbit and goat anti-mouse, were purchased from LI-COR® (Nebraska, USA). Cell culture medium and supplements were purchased from Invitrogen (Eugene, OR).
4
HIV Viral Replication Assay with Antiretroviral Drugs
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The following reagents were provided by the AIDS Research and Reference Reagent Program, Division of AIDS, National Institute of Allergy and Infectious Diseases (NIAID, MD): Nelfinavir (cat. # 4621), Raltegravir (cat. # 11680) from Merck & Company (NJ) and HIV-1NLAD8 and HIV-1NL4-3 from Malcolm Martin (MD, cat. #s 11346 and 114). Human αIL-12 and αIL-4 were purchased through PeproTech (NJ). Human rIL-2 was obtained through the BRB/NCI Preclinical Repository (MD). Antibodies were purchased from BD (NJ, Kc57-FITC, CD4-APC), Biolegend (CA, Ki67), and eBiosciences (CA, eF450 fixable viability dye, CXCR4-PE-Cy5.5, CCR5-AF488). FTY720 (Fingolimod) was obtained from Cayman Chemical (MI, cat. # 10006292, CAS#162359-56-0).
5
Modulating CD4+ T Cell Response
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Naive CD4+ T cells were purified from the MLN and spleen of OT-II and OT-II.Il27ra−/− mice using the EasySep Mouse Naive CD4+ T cell isolation Kit (Stem Cell Technologies), according to the manufacturer’s instructions. OT-II cells (1 × 106 cells/mouse) were injected i.v. into C57/BL6 recipients, and recipients were immunized 24 h later by i.p. injection of OVA (0.5 mg, Grade V; Sigma-Aldrich), anti-CD40 (25 μg), and LPS (20 μg). 1 and 3 d after immunization, recipients were injected i.p. with FTY720 (fingolimod, 20 μg; Cayman) to prevent lymphocyte egress from the MLN. Mice were sacrificed 4 d after immunization and MLN isolated for analysis. For antibody treatment studies, anti-mouse IL-27p28 or mouse IgG2a isotype control antibody were injected i.p. (250 μg) at immunization and 2 d after immunization.
6
Synthesis and Neutralization of IL-10
7
Fingolimod Inhibits Glioblastoma Growth
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Wild-type mice were injected intraperitoneally with 1 mg/kg fingolimod (FTY720) (Cayman Chemical, Ann Arbor, MI, USA) in sterile saline solution 1 h before implantation with GL261-FGL2KO tumor cells. The mice were maintained on drinking water containing 2 μg/mL FTY720 for the duration of the experiments22 (link).
8
Combination Immunotherapy for Breast Cancer
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Six-to eight-week-old male BALB/cAJcl mice were purchased from Nihon-Clea (Tokyo, Japan) and maintained in a specific pathogen-free area at Osaka University.
The overall experimental scheme is shown in Figure 2 Tumor rechallenge was conducted for those mice who got cured in the HT þ C4 group by subcutaneously inoculating 1 Â 10 5 4T1 cells into both sides of the legs.
To analyze lung metastasis, we harvested the lungs from mice in the C4 (N ¼ 7) and HT þ C4 (N ¼ 9) groups on day 13 and counted the number of metastatic nodules using a stereoscopic microscope (SZX12; Olympus, Tokyo, Japan) (Figure 2(B)).
Fingolimod (FTY720) (Cayman CHEMICAL, MI, USA), an immunosuppressive agent derived from ISP-1 (myriocin), prevents lymphocyte egress into the peripheral blood [27, 28] . We intraperitoneally administered approximately 5 mg FTY720 dissolved in saline (50 mg/ml) into the mice in the combination therapy group (HT þ C4 þ FTY720 group) every 2 days from day 0 to day 18. Tumor volumes and survival rates were compared between the
The overall experimental scheme is shown in Figure 2 Tumor rechallenge was conducted for those mice who got cured in the HT þ C4 group by subcutaneously inoculating 1 Â 10 5 4T1 cells into both sides of the legs.
To analyze lung metastasis, we harvested the lungs from mice in the C4 (N ¼ 7) and HT þ C4 (N ¼ 9) groups on day 13 and counted the number of metastatic nodules using a stereoscopic microscope (SZX12; Olympus, Tokyo, Japan) (Figure 2(B)).
Fingolimod (FTY720) (Cayman CHEMICAL, MI, USA), an immunosuppressive agent derived from ISP-1 (myriocin), prevents lymphocyte egress into the peripheral blood [27, 28] . We intraperitoneally administered approximately 5 mg FTY720 dissolved in saline (50 mg/ml) into the mice in the combination therapy group (HT þ C4 þ FTY720 group) every 2 days from day 0 to day 18. Tumor volumes and survival rates were compared between the
9
Fingolimod and SEW2871 Receptor Activation
10
Fingolimod Modulation of Neurodegeneration
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Both 3-month-old mice and 12-month-old mice were treated daily with fingolimod (FTY720) (Cayman Chemical, Ann Arbor, Michigan, USA), a sphingosine analog and an S1P receptor modulator for 2 weeks. Before injection, the animals were weighed to assess the correct doses. FTY720 dissolved in 0.9% NaCl, was administered intraperitoneally in a dose of 1 mg/kg b.w., which was selected based on our previous studies [36] . Controls (ACSF + STZ, APP -and APP + ) received the appropriate vehicle. In case of STZ-mice, the administration of the drug started on the same day as injection of STZ. One day after the last treatment, animals were sacrificed by decapitation to isolate brain tissue.
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