The SK-Hep1, Hep3B, Huh-7, HepG2 and SNU449 cell lines were obtained from the Korean Cell Line Bank (Korea). SH-J1 cells were provided by Dr. Dae-Ghon Kim (Medical School, Chonbuk National University) (Kim et al., 2002 (link)). HEK293FT and THLE-2 cells were purchased from Invitrogen and American Type Culture Collection (USA), respectively. HEK293FT and liver cancer cells were cultured at 37°C under 5% CO2 in DMEM supplemented with 10% FBS, 100 units/ml of penicillin and 100 μg/ml streptomycin. THLE-2 cells originated from human primary normal liver cells were plated on culture plates pre-coated with a solution containing 0.01 mg/ml fibronectin, 0.03 mg/ml bovine collagen type I and 0.01 mg/ml bovine serum albumin dissolved in Bronchial Epithelium Basal Medium (BEBM, Lonza). THLE-2 cells were cultured at 37°C under 5% CO2 in BEBM supplemented with Bronchial Epithelium Cell Growth Medium (BEGM) SingleQuots (Lonza). Lipofectamine 2000 (Invitrogen) was used to transfect expression vectors into cells. The same control, VRK1, VRK3 and p53 expression vectors were used as described previously (Kang and Kim, 2006 (link); Kang et al., 2008 (link); Lee et al., 2015b (link)).
Thle 2
Manufactured by Lonza
Sourced in United States, Switzerland
The THLE-2 is a human liver epithelial cell line developed by Lonza. It is a non-transformed, immortalized cell line derived from normal human liver tissue. The THLE-2 cells express several liver-specific functions and can be used for in vitro studies related to liver biology and hepatotoxicity.
Automatically generated - may contain errors
Lab products found in correlation
Thle 2, by American Type Culture Collection (8 mentions)
Bronchial epithelial growth medium (begm), by Lonza (5 mentions)
Begm bullet kit, by Lonza (5 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Hcclm3, by Thermo Fisher Scientific (4 mentions)
Hepg2, by American Type Culture Collection (3 mentions)
Penicillin streptomycin, by Procell (2 mentions)
Hep3b, by Procell (2 mentions)
Rpmi 1640 medium, by Procell (2 mentions)
Hepg2, by Procell (2 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (2 mentions)
Sk hep1, by Thermo Fisher Scientific (2 mentions)
Sk hep1, by American Type Culture Collection (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Mhcc97h, by Thermo Fisher Scientific (2 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (2 mentions)
Snu 182, by American Type Culture Collection (2 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (1 mentions)
Hek293ft, by Thermo Fisher Scientific (1 mentions)
Snu449, by Korean Cell Line Bank (1 mentions)
19 protocols using thle 2
1
Cell Line Culturing and Transfection
2
Hepatocyte Culture and Stability Assays
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A human normal hepatocyte line (THLE-2) was purchased from Ningbo Mingzhou Biotechnology Co., Ltd. (Ningbo, China). THLE-2 cells were cultured in BEBM medium (CC3170, Lonza/Clonetics Corporation, Walkersville, MD, USA) supplemented with 5 ng/mL EGF (SRP3027, Sigma‒Aldrich), 70 ng/mL phosphorylethanolamine (HY-N5034, MedChemExpress, Monmouth Junction, NJ, USA), 10% foetal bovine serum (FBS, 10099141, Gibco Laboratories, Gaithersburg, MD, USA) and 1% penicillin–streptomycin solution (C100C5, NCM Biotech, Suzhou, China) in a 5% CO2 incubator at 37 °C. Human HB cell lines (HepG2 and HuH6) were obtained from the Cell Bank of the Type Culture Collection of the Chinese Academy of Sciences (Shanghai, China). HepG2 and HuH6 cells were separately cultured in MEM (SH30265.01, HyClone, Logan, UT, USA) or DMEM (C11995500BT, Gibco Laboratories) supplemented with 10% FBS and 1% penicillin–streptomycin solution in a 5% CO2 incubator at 37 °C. For RNA decay assays, cells were treated with 5 μg/ml actinomycin D (HY-17559, MedChemExpress) for 0, 30, 60, and 90 min. For protein stability assays, cells were treated with 100 μg/ml cycloheximide (HY-12320, MedChemExpress) for 0, 2, 4, and 8 h. For some assays, cells were treated with culture medium (CM) or succinate (S9512, Sigma‒Aldrich) for the desired time.
3
Assessing KRT15 Expression in Liver Cell Lines
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The normal liver cell line THLE-2 (Cat. No. iCell-h38) cell lines were provided by the iCell Bioscience Inc. and liver cancer cell lines including Huh7 (Cat. No. CL-0120), PLC (Cat. No. CL-0415), Hep3B (Cat. No. CL-0102), HepG2 (Cat. No. CL-0103), and Li-7 (Cat. No. CL-0139) were obtained from Procell Life Science & Technology Co., Ltd. Cells of THLE-2 were maintained in BEGM (Lonza Group, Ltd.), cells including PLC, Hep3B, HepG2 and Li-7 were maintained in RPMI-1640 medium (Procell Life Science & Technology Co., Ltd.) and Huh7 cells were maintained in DMEM (Procell Life Science & Technology Co., Ltd. The medium was supplemented with 10% fetal bovine serum (FBS; Sigma-Aldrich; MilliporeSigma) and 1% penicillin/streptomycin (Procell Life Science & Technology Co., Ltd.). All cells were cultured in a humidity incubator with 5% CO2 at 37°C. The expression level of KRT15 in these cells was assessed using reverse transcription-quantitative PCR (RT-qPCR) and western blot analyses.
4
CircANTXR1 Regulates HCC Progression
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HCC cells (HuH-7 and HCCLM3) and human liver epithelial cell line (THLE-2) were bought from Biovector NTCC (Beijing, China). HCCLM3 and HuH-7 cells were cultured in DMEM (Hyclone, Logan, UT, USA), while THLE-2 cells were grown in BEGM Bullet Kit (Lonza, Walkersville, MD, USA) at 37°C with 5% CO2. All the mediums were additionally supplemented with 10% FBS (Hyclone) and 1% double antibiotics (Invitrogen, Carlsbad, CA, USA). Cell transfection was carried out using Lipofectamine 3000 (Invitrogen). All oligonucleotides and vectors were synthesized from RiboBio (Guangzhou, China), including circANTXR1 small interference RNA, lentiviral short hairpin RNA and overexpression vector (si-circANTXR1#1/#2/#3, sh-circANTXR1 and oe-circANTXR1) or scrambled controls (si-NC, sh-NC and vector), miR-532-5p mimic and inhibitor (miR-532-5p and anti-miR-532-5p) or scrambled controls (miR-NC and anti-miR-NC), pcDNA XRCC5 overexpression vector (pcDNA-XRCC5) and scrambled control (pcDNA-NC).
5
Cultivation of Diverse Liver Cancer Cell Lines
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HCC cell lines (HepG2, HuH-7, HCCLM3, SK-Hep-1, PCL/PRF/5, Hep3B) were obtained from SIBCB (Shanghai Institute of Biochemistry and Cell Biology, Shanghai, China) and Beyotime Biotechnology (Shanghai, China). Human immortalized hepatocytes (THLE-2) were purchased from ATCC (CRL-2706™). Resource Identification Initiative ID (RRID) for each cell lines are shown in Supplementary file 1. HepG2, HuH-7, HCCLM3, and PCL/PRF/5 cells were cultivated in high-glucose DMEM (GIBCO). SK-Hep-1 and Hep3B cells were cultured in MEM (Invitrogen) supplemented with GlutaMAX, NEAA (nonessential amino acids) and sodium pyruvate. THLE-2 cells were cultivated in BEGM BulletKit medium (Lonza). All media were supplemented with 10% fetal bovine serum (FBS) (GIBCO) and 1% penicillin-streptomycin (Pen/Strep) mixture (HyClone, Logan, UT, USA) and deposited at 37 °C with 5% CO2.
6
Cell Line Characterization and Compound Screening
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The THLE2, THLE3, Huh7, SK‐HEP‐1 and PLC/PRC/5 cells were got from American Type Culture Collection, HLE, HLF and Huh1 cells were got from Japanese Collection of Research Bioresources, SNU886, SNU878, SNU761 and SNU739 cells were got from Korean Cell Line Bank, and MHCC‐97H cells were got from the National Collection of Authenticated Cell Cultures (Shanghai, China). Cell lines were determined using MycoBlue Mycoplasma Detection Kit (Vazyme, D101‐01) and confirmed to be free of mycoplasma contamination. All cells were cultured in corresponding medium with 10% fetal bovine serum (FBS; Gbico). The medium were listed as follow: Dulbecco's modified Eagle's medium (DMEM) for MHCC‐97H, Huh1, Huh7, HLE and HLF; Roswell Park Memorial Institute (RPMI) 1640 for SNU761, SNU886, SNU878 and SNU739; Eagle's Minimum Essential Medium (EMEM) for SK‐HEP‐1 and PLC/PRC/5; BEGM (Lonza; CC‐3170) for THLE2 and THLE3. The PLK1 inhibitors BI2536 (S1109) and NMS‐P937 (S7255) as well as the Smad3 inhibitor SIS3 (S7959) were got from Selleck Chemicals. Blasticidin S was got from Beyotime. Puromycin and doxycline (Dox) was purchased from ThermoFisher.
7
Investigating Circular RNA Regulation in Liver Cancer
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HCC cell lines (HCCLM3 and Huh7) and human liver epithelial cell line (THLE-2) were purchased from Procell Life Science & Technology Co., Ltd. (Wuhan, China). HCCLM3 and Huh7 cells were grown in Dulbecco’s modified Eagle medium (DMEM; Invitrogen, Carlsbad, CA, USA), and THLE-2 cells were cultured in BEGM Bullet Kit (Lonza, Walkersville, MD, USA). In addition, all the media should be supplemented with 10% fetal bovine serum (FBS; Hyclone, Logan, UT, USA) and 1% penicillin/streptomycin (Invitrogen). All cells were incubated at 37°C with 5% CO2.
Circ_0046599 small interference RNA and overexpression plasmid (si-circ_0046599#1/#2 and circ_0046599) or negative controls (si-NC and circ-NC), miR-1258 mimic and inhibitor (miR-1258 and anti-miR-1258) or negative controls (NC and anti-NC), Ribophorin II (RPN2) overexpression plasmid (RPN2) and negative control (vector) were synthesized by GenePharma (Shanghai, China). Lentiviral short hairpin RNA against circ_0046599 (sh-circ_0046599) and negative control (sh-NC) were obtained from RiboBio (Guangzhou, China). Lipofectamine 3000 (Invitrogen) was used to transfect the above plasmids and oligonucleotides into cells.
Circ_0046599 small interference RNA and overexpression plasmid (si-circ_0046599#1/#2 and circ_0046599) or negative controls (si-NC and circ-NC), miR-1258 mimic and inhibitor (miR-1258 and anti-miR-1258) or negative controls (NC and anti-NC), Ribophorin II (RPN2) overexpression plasmid (RPN2) and negative control (vector) were synthesized by GenePharma (Shanghai, China). Lentiviral short hairpin RNA against circ_0046599 (sh-circ_0046599) and negative control (sh-NC) were obtained from RiboBio (Guangzhou, China). Lipofectamine 3000 (Invitrogen) was used to transfect the above plasmids and oligonucleotides into cells.
8
Cell Culture of Cancer Cell Lines
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Immortalized liver THLE2 (ATCC CRL-2706), MDA-MB-231, U2OS, HepG2 and Hela cell lines were purchased from ATCC (Rockville, MD). Esophageal cancer cell lines KYSE70 and KYSE140 were purchased from DSMZ (Braunschweig, Germany). The THLE2 cell line was maintained in BEGM from Lonza/Clonetics Corporation (CC3170, Walkersville, MD) and the other cell lines were maintained in RPMI1640, supplemented with 10% FBS and pen–strep at 37 °C in a humidified incubator with 5% CO2.
9
Cultivation of Hepatocellular Carcinoma Cell Lines
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Human THLE-2 and HepG2 cell lines were obtained from the American Type Culture Collection (ATCC, Manassas, VA, United States; Cat #CRL-2706 and HB-8065, respectively). JHH-4 and Huh7 cell lines were obtained from the Japanese Collection of Research Bioresources (JCRB) Cell Bank (Osaka, Japan; Cat #JCRB0435 and JCRB0403, respectively). THLE-2 cells were maintained in bronchial epithelial cell-growth medium (Lonza, Basel, Switzerland; Cat #CC-3171) with growth supplements according to the ATCC-recommended protocol. HepG2 and Huh7 cells were cultured in low glucose DMEM (Gibco, Detroit, MI, United States), supplemented with 10% fetal bovine serum (FBS) (Cytiva, Marlborough, MA, United States) and 1X Gibco™ Antibiotic-Antimycotic (Gibco, Detroit, MI, United States) in a humidified atmosphere of 5% CO2 at 37°C. The culture medium and conditions for JHH-4 cells were similar to those of HepG2 and Huh7 cells, except for using Eagle’s minimal essential medium (EMEM) as the basal medium.
10
HepG2 and THLE2 Cell Culture Protocols
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HepG2 (ATCC HB‐8065) and THLE2 (ATCC CRL‐2706) cell lines were purchased from ATCC. HepG2 cells were cultured in DMEM supplemented with 10% FBS and 100 U mL−1 penicillin/streptomycin. THLE2 cells were cultured by using BEGM Bullet Kit (Lonza/Clonetics Corporation, cat.#CC3170). The culture dish used for THLE2 were precoated with a mixture of 0.01 mg mL−1 fibronectin, 0.03 mg mL−1 bovine collagen type I and 0.01 mg mL−1 bovine serum albumin dissolved in BEBM medium.
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