CENFPAA01 was pre-cultured in 5 mL CM medium in 50-mL tubes overnight at 30 °C, 225 rpm. The pre-cultures were diluted into fresh 20 mL CM medium containing 0.5 mM p-coumaric acid substrate to a final OD600 of 0.05. 50 mM pantothenate (Sigma-Aldrich, St. Louis, MO, USA) stock solution was added to the liquid medium to achieve a final pantothenate concentration of 10, 20, and 50 μM, respectively. Fermentation was carried out at 30 °C, 225 rpm for 96 h. Samples were taken from each fermentation broth for naringenin measurement.
Pantothenate
Manufactured by Merck Group
Sourced in United States, Israel
Pantothenate is a lab equipment product that serves as a coenzyme in cellular metabolism. It plays a crucial role in the synthesis of coenzyme A, an essential cofactor involved in various metabolic processes. Pantothenate is a naturally occurring compound found in many foods and is required for the proper functioning of cells.
Automatically generated - may contain errors
Lab products found in correlation
Biotin, by Merck Group (17 mentions)
Dexamethasone (dex), by Merck Group (10 mentions)
Insulin, by Merck Group (5 mentions)
Cortisol, by Merck Group (5 mentions)
3 isobutyl 1 methylxanthine ibmx, by Merck Group (4 mentions)
Rosiglitazone, by Merck Group (4 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (4 mentions)
Recombinant human insulin, by Roche (3 mentions)
Triiodothyronine t3, by Merck Group (3 mentions)
Transferrin, by Merck Group (3 mentions)
3 isobutyl 1 methylxanthine, by Merck Group (3 mentions)
3 isobutyl 1 methyl xanthine (ibmx), by Merck Group (3 mentions)
Penicillin streptomycin, by Merck Group (3 mentions)
Human transferrin, by Merck Group (3 mentions)
Rosiglitazone, by Cayman Chemical (2 mentions)
Human insulin, by Merck Group (2 mentions)
Actrapid, by Novo Nordisk (2 mentions)
Rosiglitazone, by Enzo Life Sciences (2 mentions)
Polybrene, by Merck Group (2 mentions)
D biotin, by Merck Group (2 mentions)
29 protocols using pantothenate
1
Naringenin Production from p-Coumaric Acid
2
Adipogenic Differentiation of ASCs
3
Antibody-based Mitochondrial Protein Analysis
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Monoclonal anti-actin antibody, Prussian Blue, pantothenate, pantethine, vitamin E, L-carnitine, thiamine, Luperox® and trypsin were purchased from Sigma Chemical Co. (St. Louis, MO). Anti-mitochondrial acyl carrier protein (mtACP), anti-NF-Y, anti-FOXN4 and anti-hnRNPA/B were purchased from Invitrogen/Molecular Probes (Eugene, OR). Anti-phospho-PGC1α was purchased from RD systems. NFS1 antibody and Omega 3 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Anti-PANK2, anti-PGC1α, complex 1 activity kit and PDH activity kit were purchased from Abcam (Cambridge, UK). Anti-TFAM was purchased from Cell Signaling. BODIPY® 581/591 C11 was purchased from Thermo-Fisher (Waltham, MA). A cocktail of protease inhibitors (complete cocktail) was purchased from Boehringer Mannheim (Indianapolis, IN). The Immun Star HRP substrate kit was from Bio-Rad Laboratories Inc. (Hercules, CA).
4
EPO Promotes Angiogenesis in Fat Grafts
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Fat graft was harvested using manual suction lipectomy under general anesthesia from 10 women, 41 ± 4 years old, during procedures of reconstructive surgery. The participants gave written informed consent. The fat was harvested using a 14-gauge blunt cannula and centrifuged at 1500 rpm for 2 minutes.
The fat mass was seeded on dishes in appropriate culture on average for 24 hours and then treated for 3 weeks with 3 different doses of EPO (EPREX, Janssen-Cilag, Milan, Italy): 0.15, 0.30, and 0.60 μg/ml. A positive control was performed administering a cocktail of trophic drugs (biotin, T3, pantothenate, hydrocortisone, and insulin; from Sigma, St. Louis, Miss.). The negative control was left without any treatment.
At the end of this phase, the fat mass was fixed in formalin and processed for paraffin inclusion and cutting: the obtained 5-μm-thick slides were stained with CD31 (to identify active vessels and immune system cells) and CD68 (to identify macrophages).
Quantification of cell infiltration in the fat grafts was estimated by counting the number of positive vessels and cells. To avoid counting bias, due to the change of adipocytes diameter, the counting data were referred to tissue fat mass of 80 adipocytes. Parametric statistical analysis was obtained with Prism5.0 (GraphPad Software, San Diego, Calif.).
The fat mass was seeded on dishes in appropriate culture on average for 24 hours and then treated for 3 weeks with 3 different doses of EPO (EPREX, Janssen-Cilag, Milan, Italy): 0.15, 0.30, and 0.60 μg/ml. A positive control was performed administering a cocktail of trophic drugs (biotin, T3, pantothenate, hydrocortisone, and insulin; from Sigma, St. Louis, Miss.). The negative control was left without any treatment.
At the end of this phase, the fat mass was fixed in formalin and processed for paraffin inclusion and cutting: the obtained 5-μm-thick slides were stained with CD31 (to identify active vessels and immune system cells) and CD68 (to identify macrophages).
Quantification of cell infiltration in the fat grafts was estimated by counting the number of positive vessels and cells. To avoid counting bias, due to the change of adipocytes diameter, the counting data were referred to tissue fat mass of 80 adipocytes. Parametric statistical analysis was obtained with Prism5.0 (GraphPad Software, San Diego, Calif.).
5
Differentiation of Preadipocytes into Mature Adipocytes
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Preadipocytes were cultured and differentiated as reported before [22 (link)]. Preadipocytes from passage 1 were thawed at 37°C and expanded into a T-75 flask using preadipocytes medium. Upon reaching 70% confluence, the cells were trypsinized and seeded again into a 12 well plate at density 15,000 cells/cm (passage 2) using preadipocytes media. After reaching confluence, adipogenesis was induced by adding a differentiation cocktail DMEM-F12, 1% PEST, 100 nM insulin, 17 µM pantothenate (Sigma), 33 µM biotin (Sigma), 1 µM dexamethasone (Sigma), 1 µM rosiglitazone (Sigma), 250 µM 3-isobutyl-1-methylxanthine (IBMX, Sigma), 10 µg/ml transferrin (Sigma), 2 nM triiodothyronine (T3, Sigma) for 5 days, changing the medium on day 3. The differentiation continued using maintenance media (composition as that of differentiation cocktail except for omitting IBMX) for 14 days to obtain mature adipocytes. Medium was replenished every 2–3 days. CABLES1 gene expression was measured in cells collected upon confluence (day 0) and days 2, 4, 8 and 14 post induction for temporal profiling, or, for gene editing experiments (see below), on days 0, 7 and 14 for gene edited cells.
6
Metabolic Profiling with HPLC-MS
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DMEM (Cat#11965) was purchased from ThermoFisher (Waltham, MA, USA). Oasis HLB SPE columns were purchased from Waters (Milford, MA, USA). 5-sulfosalicylic acid, etomoxir, pantothenate, dephospho-CoA, and acyl CoA standards were purchased from Sigma-Aldrich (St. Louis, MO, USA). (E)-but-2-enoyl Coenzyme A (crotonoyl CoA) was purchased from Avanti Polar Lipids (Alabaster, Alabama, USA).
7
Adipocyte Differentiation Protocol
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Dexamethasone, 3-isobutyl-1-methylxanthine (IBMX), transferrin, cortisol, triiodothyronine (T3), biotin, pantothenate, dichloromethane, trifluoroacetic anhydride, ethanol, potassium hydroxide (KOH), sodium bicarbonate (NaHCO3), HCL, acetone, Dulbecco’s Modified Eagle Medium (DMEM) powder media, Hams’s F12 powder media, fructose and glucose were all obtained from SIGMA Chemical Company (St. Louis, MO, USA). Fetal bovine serum was obtained from Thermo Fisher Scientific (Waltham, MA, USA). Rosiglitazone was obtained from Cayman Chemicals (Ann Arbor, MI, USA), Insulin was obtained from Novo Nordisk [U-13C6]-d -fructose (>99 % purity, and 99 % isotope enrichment for all carbons) was obtained from Cambridge Isotope Laboratories (Andover, MA, USA), methane and helium (>99.99 % purity), was obtained from PaxAir (Los Angeles, CA, USA), and n-butanol was obtained from Regis Chemical. Company (Chicago, IL, USA).
8
Adipogenic Differentiation of ASCs
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Adipogenic differentiation of ASCs was performed as previously described [49] (link). Briefly, ASCs were cultured in ASC growth medium until the culture reached 90–95% confluency. ASCs were then trypsinized and plated in 24-well plates in ASC growth medium at 30,000 cells/cm2 for 24 hrs. to allow attachment. On day 1 (24 hours after plating), the medium was removed and cells were incubated for three days in ASC adipogenic differentiation medium [Dulbecco's modified Eagle's-Ham's F-12 medium supplemented with 3% or 10% FBS, 15 mM HEPES (pH 7.4), biotin (33 µM), pantothenate (17 µM, Sigma), human recombinant insulin (100 nM, Boehringer Mannheim), dexamethasone (1 µM), 1-methyl-3-isobutylxanthine (IBMX; 0.25 mM), and rosiglitazone (1 µM)]. For the remaining 9 days of the adipocyte differentiation maintenance period, the medium was removed every 3 days and replaced with the same medium that did not contain IBMX and rosiglitazone (maintenance medium). Adipocyte differentiated conditioned medium (ADCM) was collected on day 6 after the switch to ASC adipogenic differentiation medium and stored at 4°C until use.
9
Human Preadipocyte Differentiation Assay
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Primary human preadipocytes (Zenbio, Inc., Research Triangle Park, NC, USA) from donors with body mass inDEXes ⩽24.99 kg m−2 were maintained in Preadipocyte Medium (ZenBio) at 37 °C and 5% CO2. For differentiation, confluent preadipocytes were treated with media containing 33 μM biotin, 17 μM pantothenate (both from Sigma-Aldrich, St Louis, MO, USA) and 100 nM insulin (Roche Applied Science, Laval, QC, Canada) for 14 days. In addition, 500 μM IBMX (Sigma-Aldrich) was also included in the differentiation media from day 0 to day 4. From day 2 until day 14, 5 μM troglitazone (Sigma-Aldrich) and the indicated concentrations of BPA were also included in the differentiation media, which was replenished every 2 days. As a positive control, cells were treated with 1 μM DEX (Sigma-Aldrich) starting on day 2 throughout differentiation with the same media as above instead of BPA. For the ER and GR antagonist studies, 1 nM estradiol, 1 μM ICI-182,780 or 1 μM RU486 (all Sigma-Aldrich) were also added as above with or without BPA.
10
Cell Line Differentiation and Characterization
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Ari (129,722-12-9; Acros Organics), Ola (035M4781V; Sigma-Aldrich), Phorbol 12-myristate 13-acetate (PMA) (Sigma, P1585), Granulocyte-macrophage colony-stimulating factor (GM–CSF– Peprotech # 300–03), human transferrin (Sigma-Aldrich #T8158), human insulin (Sigma-Aldrich #91077C), cortisol (Sigma-Aldrich #H0888), triidothyronine (T3) (Sigma-Aldrich #T6397), dexamethasone (Sigma-Aldrich #D1756), IBMX (Sigma-Aldrich #I5869), rosiglitazone (Cayman #71740), biotin (Sigma-Aldrich #B4501), pantothenate (Sigma-Aldrich #21210), recombinant human IL-1 β (Peprotech # 200-01 B), recombinant human IL-6 (Peprotech # 200–06), recombinant human IL-18 (Peprotech # 200-18BP), recombinant human IL-10 (Peprotech #200–10), p-Akt (Ser473) (Cell Signalling Technology #9271 S, 1:2000), Akt (Cell Signalling Technology #9272, 1:2000), polyclonal rabbit anti-p97 antibody (Abcam #97302, 1:8000), horseradish peroxidase-conjugated secondary antibodies were purchased from Jackson Immunoresearch (West Grove, PA), 2-NBD glucose, fluorescent glucose uptake probe (Abcam #146200) and, human Glut4 Alexa Fluor 488-conjugated antibody (R&D systems # FAB86541).
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