The MGH-HCC1 cell line (source: male) was derived from a resected metastasis from a patient in this cohort. The NCI-HCC cell line (source: male) was derived from a tumor grown from PDX material obtained from the NCI Patient-Derived Models Repository (model 248138-237-R). The Nthy-ori 3-1 cell line (source: female) was purchased (Sigma; cat. #90011609) and was part of the European Collection of Authenticated Cell Cultures. All cell lines were grown in humidity-controlled incubators at 21% O2, 5% CO2, and 37°C and routinely tested for Mycoplasma (Lonza; cat. #LT07-218; last tested February 16, 2023). Cell lines were authenticated using the ATCC short tandem repeat authentication method (ATCC; cat. #135-XV), and early passage freezer stocks were used for all experiments. MGH-HCC1 and NCI-HCC cells were routinely passaged in DMEM-11995 (Thermo Fisher; cat. #11995073) media with 10% fetal bovine serum (FBS) and 50 mg/mL uridine. Nthy-ori 3-1 and Hela cells (source: female; ATCC; cat. #CCL2) were routinely passed in RPMI 1640 media (Sigma; cat. #R8758) with 10% FBS. All experiments were conducted in DMEM-11995 media with 10% FBS and 50 mg/mL uridine unless otherwise specified.
Nthy ori 3 1 cell line
Manufactured by Merck Group
Sourced in Germany
The Nthy-ori 3-1 cell line is a human thyroid follicular epithelial cell line derived from normal human thyroid tissue. It is commonly used in research applications to study thyroid cell biology and function.
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Lab products found in correlation
Rpmi 1640 medium, by Thermo Fisher Scientific (3 mentions)
Rpmi 1640 media, by Merck Group (2 mentions)
11995 dmem, by Thermo Fisher Scientific (2 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Nthy ori 3 1, by American Type Culture Collection (1 mentions)
Fungizone, by Merck Group (1 mentions)
Rpmi 1640 medium, by Merck Group (1 mentions)
Amphotericin b, by Thermo Fisher Scientific (1 mentions)
Kmh 2, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Ash 3, by Japanese Collection of Research Bioresources Cell Bank (1 mentions)
Tpc 1, by Merck Group (1 mentions)
Fetal calf serum (fcs), by Thermo Fisher Scientific (1 mentions)
Rpmi a, by Capricorn (1 mentions)
Fetal calf serum (fcs), by Harvard Bioscience (1 mentions)
Cell culture flasks, by Sarstedt (1 mentions)
Mem non essential amino acid, by Thermo Fisher Scientific (1 mentions)
Sodium pyruvate, by Thermo Fisher Scientific (1 mentions)
Hepes, by Thermo Fisher Scientific (1 mentions)
Fetal calf serum (fcs), by Corning (1 mentions)
Glutamine, by PanEco (1 mentions)
9 protocols using nthy ori 3 1 cell line
1
Cell Line Establishment and Maintenance
2
Cell Line Culture Protocols
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We used the Nthy-ori 3–1-cell line obtained from Sigma–Aldrich, St. Louis, USA (90011609) and the TPC-1-cell line, kindly provided by Prof. Dr. Valdemar de Jesus Conde Maximo, from the Institute of Pathology and Molecular Immunology - Ipatimup, University of Porto, Portugal. Both cell lines were grown in RPMI 1640 medium containing 10% fetal bovine serum, 1% penicillin–streptomycin and 250 mg/ml fungizone (Sigma–Aldrich, St. Louis, USA). Cells were kept at 37°C in a humidified environment with 5% CO2.
3
Thyroid Cancer Cell Line Culturing
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Three human PTC cell lines, TPC-1, BCPAP, and K1, and one normal human thyroid epithelial cell line (Nthy-ori 3-1) were used. The Nthy-ori 3-1 cell line was purchased from Sigma. BCPAP was obtained from the cell bank at the Chinese Academy of Sciences (Shanghai, People’s Republic of China). The TPC-1 and K1 cell lines were purchased from the Cell Bank of the University of Colorado. All cell lines were cultured in RPMI-1640 medium (GIBCO) supplemented with 10% heat-inactivated fetal bovine serum (FBS; GIBCO) at 37°C in a 5% CO2 chamber.
4
Characterization of Thyroid Cell Lines
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The ATC 8505c, ASH-3 and KMH-2 cell lines were purchased from the Japanese Collection of Research Bioresources Cell Bank, and the normal human thyroid follicular epithelial Nthy-ori 3-1 cell line was purchased from Sigma-Aldrich (Merck KGaA). The 8505c cell line contained a C:G to G:C transversion at codon 248 of the p53 gene, and there were chromosomal abnormalities in the ASH-3 and KMH-2 cell lines at chromosome numbers 79–89 and 92–108, respectively.
The ATC and normal human thyroid follicular epithelial cell lines were grown in RPMI-1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (HyClone; Cytiva), 1% penicillin/streptomycin and 1 µl/ml amphotericin B (Gibco; Thermo Fisher Scientific, Inc.). All cell lines were cultured in a humidified incubator at 37°C with 5% CO2.
The ATC and normal human thyroid follicular epithelial cell lines were grown in RPMI-1640 medium (Gibco; Thermo Fisher Scientific, Inc.) supplemented with 10% FBS (HyClone; Cytiva), 1% penicillin/streptomycin and 1 µl/ml amphotericin B (Gibco; Thermo Fisher Scientific, Inc.). All cell lines were cultured in a humidified incubator at 37°C with 5% CO2.
5
Cell Line Characterization and Cultivation
6
Culturing Thyroid Cancer Cell Lines
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The Nthy-ori3-1 cell line was obtained from Sigma, while the human papillary thyroid cancer cell lines TPC-1 and KTC-1 were acquired from reputable institutes, specifically Shanghai RuiJin Hospital, China. The TPC-1 and KTC-1 cells were cultured in RPMI 1640 medium supplemented with 10% fetal bovine serum (FBS) and subsequently incubated in a humidified environment (5% CO2, 37 °C).
7
Thyroid Follicular Cell Cysteine Peptidase Inhibition
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Normal human thyroid follicular epithelial cells (Nthy-ori 3-1 cell line) (Sigma Aldrich, Taufkirchen, Germnay, #90011609) were cultured in Roswell Park Memorial Institute medium (#RPMI-A, Capricorn Scientific, Hessen, Germany) that contains 10% fetal calf serum (Thermo Fisher Scientific, Darmstadt, Germany, #10270106, origin Brazil). Cell cultures were maintained at 37 °C in a humidified atmosphere at 5% CO2. For cysteine peptidase inhibition by E64 or E64d, confluently grown Nthy-ori 3-1 cells were incubated in serum-free RPMI-A for 48 h during the addition of E64 or E64d. For stimulation, confluently grown Nthy-ori 3-1 cells were incubated in serum-free RPMI-A 1640 for 48 h while being treated with 100 µU/mL TSH (Merck, Darmstadt, Germany, #869006) for up to 4 h and 24 h.
8
Nthy-ori 3-1 Thyroid Cell Culture Protocol
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The Nthy-ori 3-1 cell line (Sigma-Aldrich, Munich, Germany) is a primary human thyroid follicular epithelial cell line. The immortalised cell line exhibits thyroid specific functions, such as iodide-trapping and thyroglobulin synthesis and reveals no signs of tumorigenesis, when transplanted into nude mice [22] .
The Nthy-ori 3-1 cells were cultured under standard cell culture conditions (37°C and 5% CO 2 ). The cells grew in RPMI 1640 medium (Life Technologies, Naerum, Denmark), supplemented with 1 % penicillin/ streptomycin (Life Technologies) and 10 % (v/v) fetal calf serum (FCS) (Biochrom, Berlin, Germany). The cells were cultured in T25 cm 2 vented cell culture flasks (Sarstedt, Newton, USA) for one day prior to RPMexposure. Each flask was seeded with 1 × 10 6 cells. The culture flasks were filled completely with medium without any air bubbles. The flasks (n = 60 per time point) were assigned randomly to use as static controls (1g; n = 30) or as RPM-samples (n = 30). The 1g-controls were positioned next to the RPM in the same incubator. Supernatants and cells were harvested after 4 h, 24 h or 72 h according to the protocol.
The Nthy-ori 3-1 cells were cultured under standard cell culture conditions (37°C and 5% CO 2 ). The cells grew in RPMI 1640 medium (Life Technologies, Naerum, Denmark), supplemented with 1 % penicillin/ streptomycin (Life Technologies) and 10 % (v/v) fetal calf serum (FCS) (Biochrom, Berlin, Germany). The cells were cultured in T25 cm 2 vented cell culture flasks (Sarstedt, Newton, USA) for one day prior to RPMexposure. Each flask was seeded with 1 × 10 6 cells. The culture flasks were filled completely with medium without any air bubbles. The flasks (n = 60 per time point) were assigned randomly to use as static controls (1g; n = 30) or as RPM-samples (n = 30). The 1g-controls were positioned next to the RPM in the same incubator. Supernatants and cells were harvested after 4 h, 24 h or 72 h according to the protocol.
9
Culturing Nthy-ori 3-1 Thyroid Cells
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Nthy-ori 3-1 cell line derived from follicular thyroid epithelium (Sigma -Aldrich, 90011609) was cultured in Cell Culture Treated Flasks at 37℃ in the presence of 5% CO2 in RPMI-1640 medium containing 2mM glutamine (Paneco, Russia) supplemented with 10% fetal calf serum (Corning, USA), 10mM HEPES (GIBCO, USA), 1 mM sodium pyruvate (GIBCO, USA), 100 U/ml penicillin and 100 mg/ml streptomycin (Paneco, Russia), and 1x MEM Non-Essential Amino Acids (GIBCO, USA).
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