Reference genotoxins (mitomycin C, M7949, vinblastine sulphate, V1377, and benzo(a)pyrene, B1760) were purchased from Sigma Aldrich (St Louis, MO). Phenformin HCl (PHR1573, Supelco), purchased from Sigma Aldrich (St Louis, MO) was used as non-genotoxic negative control. Test compound stock solutions were prepared in DMSO (D161802, Sigma Aldrich, St Louis, MO). For the 3D tissues, dilutions were made in complete culture media yielding ≤ 0.1% DMSO final concentration. Media containing 0.1% DMSO (i.e. without test article) were applied to tissues to serve as the vehicle control. All 3D tissues were treated apically every alternate day (from day 0–6) and daily (from day 7–9), while basolateral wells contain untreated culture media to support the tissue growth. TK6 cells were exposed for 24 h with the test materials prior to harvest. S9 liver fraction (11-402L, MolTox, Boone, NC) was added to TK6 cells 3 h prior to benzo(a)pyrene exposure to activate cellular metabolism (Cox et al. 2016 (link)).
B1760
Manufactured by Merck Group
Sourced in Germany, United States
The B1760 is a laboratory equipment product manufactured by Merck Group. It serves as a functional device designed for use in scientific research and analysis activities. The core function of this product is to provide a specific capability required in laboratory settings, without further interpretation of its intended applications.
Automatically generated - may contain errors
Lab products found in correlation
Vinblastine sulphate, by Merck Group (1 mentions)
V1377, by Merck Group (1 mentions)
Mitomycin c, by Merck Group (1 mentions)
Dimethyl sulfoxide (dmso), by Merck Group (1 mentions)
A9525, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by GE Healthcare (1 mentions)
Hepg2, by GE Healthcare (1 mentions)
Dulbecco modified eagle medium (dmem), by GE Healthcare (1 mentions)
7 protocols using b1760
1
Genotoxicity Assay with 3D Tissues
2
Preparing B[a]P in DMSO Solution
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B[a]P (≥96%, B1760, Sigma Aldrich) is not water soluble and was previously dissolved in dimethyl sulfoxide (DMSO) after having determined its solubility limit. Chemical solutions were prepared so that the DMSO concentration in the sea water was 0.001%.
3
SMYAD Alleviates Bap/Ang II-Induced Injury
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After one-week acclimatization, 42 mice were weighed and then randomly divided into two groups (count this day as Day 1): normal control group (n = 6, control for short) and model group (n = 36). From the Day 1 to Day 42, the 6 mice in control received no Bap/Ang II and other treatments; the 36 mice in the model group were injected intraperitoneally with Bap (B1760, Sigma-Aldrich) at a dose of 10 mg/kg body weight weekly and a dose (0.72 mg/kg/day) of Ang II (A9525, Sigma-Aldrich) using subcutaneous osmotic mini-pumps (Model# 2006, Alzet). Current guideline for use of bio-hazardous materials was followed when using Bap. At Day 15, 36 mice in the model group were randomly and equally divided into three groups, namely, Bap-/Ang II-treated + saline group (Bap + Ang II for short, n = 12), Bap-/Ang II-treated + SMYAD-low-dose group (Bap + Ang II + SL for short, n = 12), and Bap-/Ang II-treated + SMYAD-high-dose group (Bap + Ang II + SH for short, n = 12). From Day 15 to Day 42, mice in Bap + Ang II + SL and Bap + Ang II + SH were fed intragastrically 1 ml of SMYAD-low dose or SMYAD-high dose once daily, respectively; meanwhile, mice in Bap + Ang II were fed intragastrically with 1 ml of saline solution once daily. On Day 43, all survival mice were weighed and then euthanized.
4
Hep-G2 Cell Exposure to Benzo(a)Pyrene
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The human hepatoblastoma Hep-G2 cell line was purchased from the Cell Resource Center, Peking Union Medical College (National Infrastructure of Cell Line Resource, Beijing, China). Hep-G2 cells were cultured in Dulbecco's modified Eagle's medium (DMEM) (GE Healthcare Bio-Sciences, Pittsburgh, PA, USA) supplemented with 10% fetal bovine serum (FBS) (GE Healthcare Bio-Sciences) and antibiotics (penicillin 100 U/ml and streptomycin 100 µg/ml) in an incubator with a humidified atmosphere of 5% CO2 at 37°C. For BaP exposure, Hep-G2 cells were treated with different concentrations (0, 2, 4, 8, 16, 32 or 64 µM) of BaP (B1760; >96% HPLC; Sigma-Aldrich; Merck KGaA, Darmstadt, Germany) at different time points (0, 24, 48 or 72 h), as described previously (29 (link),30 (link)). The final concentration of dimethyl sulfoxide (DMSO) used as solvent control was 0.1% (v/v) or less.
5
FITC-LDL Uptake Assay and Treatment
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In brief, 20 μL of FITC (6 mg/mL) and 1 mL of LDL (2 mg/mL) (Yiyuan Biotech Co., Guangzhou, China) were mixed in EP tubes and then incubated at 37 °C for 2 h before being transferred into pre-treated dialysis bags. These bags were dialyzed in PBS for 24 h at 4 °C and protected from light. PBS was changed every other day for three times. Finally, the dialyzed FITC-LDL was transferred into brown EP tubes and stored in a refrigerator at 4 °C before use.
For the cell treatment of LDL uptake, 50 μg/mL FITC-LDL was incorporated with BaP (2.5 μM, B1760, SIGMA, St Louis, MO, USA), methyl β-cyclodextrin (3 mM), aminoguanidine (10 mM), or Hsd (25 μM,50 μM,100 μM, PHR1794, SIGMA, St Louis, MO, USA) as treatment agents. FITC-LDL alone was used as the control. The cells were treated at 37 °C for 3 h. After incubation, the cells were seeded on glass coverslips, incubated at 37 °C for another 3 h, and fixed with 4% paraformaldehyde at room temperature for 15 min). An anti-fluorescence quencher was added to seal the slides (China Beyene Institute of Biotechnology, Nanjing, China). Finally, FITC-LDL uptake was measured by confocal laser scanning microscopy.
For the cell treatment of LDL uptake, 50 μg/mL FITC-LDL was incorporated with BaP (2.5 μM, B1760, SIGMA, St Louis, MO, USA), methyl β-cyclodextrin (3 mM), aminoguanidine (10 mM), or Hsd (25 μM,50 μM,100 μM, PHR1794, SIGMA, St Louis, MO, USA) as treatment agents. FITC-LDL alone was used as the control. The cells were treated at 37 °C for 3 h. After incubation, the cells were seeded on glass coverslips, incubated at 37 °C for another 3 h, and fixed with 4% paraformaldehyde at room temperature for 15 min). An anti-fluorescence quencher was added to seal the slides (China Beyene Institute of Biotechnology, Nanjing, China). Finally, FITC-LDL uptake was measured by confocal laser scanning microscopy.
6
Immortalized Human Fibroblasts: A Versatile Model
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The human diploid VH10tert foreskin fibroblast cell line, immortalized by stable transfection with the telomerase gene (TERT), was kindly provided by Prof. Mullenders (Department of Toxicogenetics at Leiden University Medical Centre, the Netherlands). Human primary bronchial epithelial cells (PBECs) were purchased from Provitro (Berlin) and cultivated in Airway epithelial cell growth medium containing 10% fetal bovine serum. VH10tert cells were cultivated in Dulbecco's minimal essential medium containing 10% fetal bovine serum. MCF7 breast cancer cells were obtained from CLS Cell Lines Service GmbH, Eppelheim, Germany. All cells were regularly checked for mycoplasma contamination and cultivated under nitrogen atmosphere (7% CO2, 5% O2) at 37°C. Experiments involving human individuals (healthy males 25–40 years), were approved by the ethic committee ‘Ethik-Kommision Rheinland Pfalz’ (837.198.15 (9966)) and a written consent from all individuals was obtained. B(a)P was purchased from SIGMA (B1760), activated r-7,t-8-Dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo(a)pyrene (anti-BPDE; CAS no. 58917-67-2) was prepared from trans-7,8-dihydroxy-7,8-dihydrobenzo(a)pyrene (29 ) as described (30 (link)). Commercially available cigarettes used for the in vivo determination of DNA repair genes contained 0.6 mg nicotine, 7 mg tar and 9 mg carbon monoxide.
7
Effects of Benzo[a]pyrene Exposure on Hamster Pregnancy
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Golden Syrian hamster (Mesocricetus auratus), 8-week old, (150 ± 10) g body weight were used. The golden hamsters were paired for mating on a 1:1 male: female ratio per cages overnight. Vaginal smears were made at 8 am of the next day. Sperms were seen under the microscope, which was recorded as the first day after fertilization. Dams were randomly divided into control group, BaP2.5 group and BaP5.0 group, 7 hamsters per group. The dams in the control group were given intraperitoneal injection of normal saline, and dams in the BaP2.5 and BaP5.0 groups were injected intraperitoneally at BaP (Sigma-Aldrich, B1760) 2.5 mg/kg body weight and 5 mg/kg body weight, respectively. The animals were treated once a day from the 4th day after fertilization. All dams were sacrificed after anesthesia on the 14th day after fertilization, fetuses and placenta were separated, and the number of absorbed fetuses, stillborn fetuses and live fetuses were recorded respectively. The rate of adverse pregnancy outcomes = the number of adverse pregnancy outcomes/the total number of fetal rats.
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