Triiodothyronine

Manufactured by Thermo Fisher Scientific

Sourced in United Kingdom, United States

Triiodothyronine is a lab equipment product that measures the level of triiodothyronine, a thyroid hormone, in the body. It is used to assess thyroid function and help diagnose thyroid disorders.

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5 protocols using triiodothyronine

Isolation and Culture of Retinal Ganglion Cells

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P3/P4 retinas were freshly dissected and RGCs were isolated and purified (>99%). For details see Steinmetz et al. (2006) (link); Claudepierre et al., 2008 (link)). Briefly, cells were cultured in Neurobasal medium (Gibco/Invitrogen) supplemented with (all from Sigma, except where indicated) pyruvate (1 mM), glutamine (2 mM; Gibco/Invitrogen), N-acetyl-l-cysteine (60 μg ml⁻¹), putrescine (16 μg ml⁻¹), selenite (40 ng ml⁻¹), bovine serum albumin (100 μg ml⁻¹; fraction V, crystalline grade), streptomycin (100 μg ml⁻¹), penicillin (100 U ml⁻¹), triiodothyronine (40 ng ml⁻¹), holotransferrin (100 μg ml⁻¹), insulin (5 μg ml⁻¹) and progesterone (62 ng ml⁻¹), B27 (1:50, Gibco/Invitrogen), brain-derived neurotrophic factor (BDNF; 25 ng ml⁻¹; PeproTech, London, UK), ciliary neurotrophic factor (CNTF; 10 ng ml⁻¹; PeproTech) and forskolin (10 μm; Sigma). After isolation, RGCs were either treated for RNA extraction or fixed with PFA4% 15' at RT and processed for immunohistochemistry. Stainings were performed and cells were visualized as described above.

Savier E., Eglen S.J., Bathélémy A., Perraut M., Pfrieger F.W., Lemke G, & Reber M. (2017). A molecular mechanism for the topographic alignment of convergent neural maps. eLife, 6, e20470.

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Retinal Ganglion Cell Isolation and Purification

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P1/P2 retinas were freshly dissected and RGCs were isolated and purified (>99%). For details, see Steinmetz et al., 2006 (link) and Claudepierre et al., 2008 (link). Briefly, cells were harvested in neurobasal medium (Gibco/Invitrogen) supplemented with (all from Sigma, except where indicated) pyruvate (1 mM), glutamine (2 mM; Gibco/Invitrogen), N-acetyl-l-cysteine (60 mg/ml), putrescine (16 mg/ml), selenite (40 ng/ml), bovine serum albumin (100 mg/ml; fraction V, crystalline grade), streptomycin (100 mg/ml), penicillin (100 U/ml), triiodothyronine (40 ng/ml), holotransferrin (100 mg/ml), insulin (5 mg/ml) and progesterone (62 ng/ml), B27 (1:50, Gibco/Invitrogen), brain-derived neurotrophic factor (BDNF; 25 ng/ml; PeproTech, London, UK), ciliary neurotrophic factor (CNTF; 10 ng/ml; PeproTech) and forskolin (10 mM; Sigma). After isolation, RGCs were treated for RNA extraction.

Savier E.L., Dunbar J., Cheung K, & Reber M. (2020). New insights on the modeling of the molecular mechanisms underlying neural maps alignment in the midbrain. eLife, 9, e59754.

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Cell Culture Reagents and Suppliers

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The drugs used in the present study were purchased from Sigma-Aldrich (St. Louis, MO, USA), unless indicated otherwise: rapamycin (Cell Signaling Technology, Boston, MA, USA); plasmocin (Fisher Scientific, Hampton, NH, USA); EGF, dexamethasone, triiodothyronine, insulin–transferrin–selenium (ITS, 1X, Invitrogen, Carlsbad, CA, USA); penicillin/streptomycin (1X, Invitrogen); fetal bovine serum (Invitrogen); L-glutamine, HEPES, sodium pyruvate, and 2-mercaptoethanol. Trypsin-ultra, mass spectrometry-grade, restriction enzymes, and other molecular biology reagents were purchased from New England BioLabs (Ipswich, MA, USA).

Armando I., Cuevas S., Fan C., Kumar M., Izzi Z., Jose P.A, & Konkalmatt P.R. (2022). G Protein-Coupled Receptor 37L1 Modulates Epigenetic Changes in Human Renal Proximal Tubule Cells. International Journal of Molecular Sciences, 23(22), 14456.

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Isolation and Culture of Renal Tubular Epithelial Cells

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RTECs were extracted from mice using previously reported methods (Valente et al. 2011 (link)). In brief, mice were euthanized, and the kidneys were immediately excised and placed in ice-cold complete media in HBSS (Hyclone, Marlborough, MA) supplemented with 1% penicillin-streptomycin (Hyclone). Thereafter, the renal capsule was dissected out, and sagittal sections of the renal cortex were obtained and the medulla was discarded. Further, cortical tissue was minced in complete medium with collagenase I (1 mg/mL, Sigma-Aldrich, St. Louis, MO) and soybean trypsin inhibitor (100 μg/mL, Sigma-Aldrich) and incubated at 37°C for 45 min with frequent mixing. The same volume of ice-cold HBSS was added into tubules to terminate the digestion step. The digestion product was sieved successively with 150-μm and 75-μm mesh sieves, and the filtrate was collected and seeded on collagen-treated plates in a selection culture medium comprising DMEM:F12 culture media (Gibco, Waltham, MA) supplemented with 10 mL/L penicillin-streptomycin (Hyclone), 50 nmol/L hydrocortisone (Sigma-Aldrich), epidermal growth factor, insulin/transferring/selenium (10 μg/mL/5.5 μg/mL, Gibco), and tri-iodothyronine (Gibco), in an incubator (37°C, 5% CO₂) and cultured to 80% confluence for subculturing. Cells of the second and third passages were used for subsequent experiments.

Xu H., Song D., Xu R, & He X. (2021). BAFF signaling drives interstitial transformation of mouse renal tubular epithelial cells in a Pin1-dependent manner. In Vitro Cellular & Developmental Biology. Animal, 57(6), 649-659.

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Generating Cystinotic Renal Epithelial Cells

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Lipofectamine 2000 LXT with plus (Life Technologies, Invitrogen, Paisley, UK) into MDCK II or cystinotic PTEC cells. For stable transfection, MDCK II cells expressing cystinosin-LKG-RFP (cys-LKG-RFP) were selected with 200 μg/ml hygromycin B for 14 d (Life Technologies), whereas PTEC expressing cys-LKG-RFP were isolated by cell sorting (FACSAria II; BD Biosciences, San Jose, CA). Analogue procedure was performed for cells expressing cystinosin-RFP (cys-RFP). MDCK II cells were cultured in Dulbecco's modified essential medium (Gibco, Grand Island, NY), supplemented with 10% fetal calf serum (Gibco), 2 mmol/l L-glutamine, 100 units/ml penicillin and 100 μg/ml streptomycin. PTEC were cultured in Dulbecco's modified essential medium/F12 (Gibco), supplemented with 10% fetal calf serum (Gibco), ITS (5 μg/ml insulin, 5 μg/ml transferrin, 5 ng/ml selenium), 36 ng/ml hydrocortisone, 10 ng/ml epidermal growth factor, 40 pg/ml triiodothyronine, 100 units/ml penicillin, and 100 μg/ml streptomycin. These last reagents were supplied by Sigma Aldrich (Milan, Italy).
Cystinotic PTEC were kindly provided by Elena Levtchenko (Department of Pediatrics, University Hospitals Leuven, Belgium). PTEC were isolated from urine of cystinotic and healthy donors, infected with SV40T and hTERT vectors, and characterized with antibodies against epithelial markers (26) .

Taranta A., Bellomo F., Petrini S., Polishchuk E., De Leo E., Rega L.R., Pastore A., Polishchuk R., De Matteis M.A, & Emma F. (2017). Cystinosin-LKG rescues cystine accumulation and decreases apoptosis rate in cystinotic proximal tubular epithelial cells. Pediatric research, 81(1-1).

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