Human adipose-derived stem cells (hASCs) were purchased from Invitrogen (Waltham, MA, USA). hASCs were cultured according to the manufacturer’s instructions. hASCs were grown in low glucose Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Waltham, MA, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS; Gibco) and 1% penicillin-streptomycin (Gibco). The medium was replaced with fresh medium every two to three days, and cells were passaged using 0.25% trypsin-EDTA (Gibco) when they reached about 80% confluency. Cells from passages 6 to 8 were used for experiments.
Hascs
Manufactured by Thermo Fisher Scientific
Sourced in United States
HASCs are a type of laboratory equipment designed for cell culture applications. They provide a controlled environment for the cultivation and maintenance of human adipose-derived stem cells (hASCs). The core function of HASCs is to facilitate the growth and expansion of hASCs under defined conditions.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (6 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (5 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (4 mentions)
α mem, by Thermo Fisher Scientific (2 mentions)
Hbmsc, by Thermo Fisher Scientific (2 mentions)
Hbmscs, by ScienCell (2 mentions)
Trypsin, by Thermo Fisher Scientific (2 mentions)
Hascs, by Lonza (2 mentions)
Trypsin edta, by Thermo Fisher Scientific (1 mentions)
Optima l 90k ultracentrifuge, by Beckman Coulter (1 mentions)
Primary hascs, by ScienCell (1 mentions)
Foetal bovine serum fbs, by ScienCell (1 mentions)
Ethylene diamine tetraacetic acid (edta), by Thermo Fisher Scientific (1 mentions)
Alpha modified eagle s medium, by Thermo Fisher Scientific (1 mentions)
Tryple express without phenol red, by Thermo Fisher Scientific (1 mentions)
L dmem, by Merck Group (1 mentions)
Sodium bicarbonate, by Merck Group (1 mentions)
Minimum essential medium (mem), by Thermo Fisher Scientific (1 mentions)
Hepes, by Merck Group (1 mentions)
Fetal bovine serum (fbs), by Biowest (1 mentions)
15 protocols using hascs
1
Culturing Human Adipose-Derived Stem Cells
2
Isolation and Culture of hASCs and hBMSCs
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Primary hASCs and human bone marrow mesenchymal stem cells (hBMSCs) were obtained from ScienCell Company. Cells were cultured at 37°C in an incubator with 5% CO2 atmosphere and full relative humidity. To minimize the exogenous exosomes, hASCs were cultured in Dulbecco's Modified Eagle Medium (DMEM, Gibco) free of exosomes through ultracentrifugation at 100 000 g overnight with an Optima L‐90K Ultracentrifuge (Beckman Coulter, Inc). For the in vitro experiments, hBMSCs were cultured in proliferation medium (PM), which consisted of minimum essential medium α (α‐MEM, Gibco), 10% (v/v) foetal bovine serum (FBS, ScienCell) and 100 IU/mL antibiotics (Gibco). For osteogenic induction, hBMSCs were cultured in osteogenic medium (OM), which consisted of standard PM supplemented with 10 mmol/L β‐glycerophosphate, 0.2 mmol/L L‐ascorbic acid and 100 nmol/L dexamethasone. All other materials were purchased from Sigma‐Aldrich unless otherwise mentioned, and all experiments conducted with hBMSCs were extracted from three donors (Catalog#15901, #6881, #6890).
3
Characterization and Pooling of Human Adipose-Derived Stem Cells
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Primary human ASCs (hASCs) were purchased from LaCell LLC (New Orleans, LA, USA). Each hASC cell line underwent full characterization individually prior to being pooled together [32 (link),33 (link),34 (link),35 (link),36 (link)]. Pooled hASCs from 5 healthy donors were thawed at passage 3 and maintained in complete culture medium (CCM) consisting of minimum essential medium alpha (Cat #: 12561; Gibco, Grand Island, NY, USA) supplemented with 10% heat-inactivated fetal bovine serum (FBS, Cat #: SH30396.03; Thermo Fisher, Waltham, MA, USA), and 1% penicillin-streptomycin (Cat #: 15140122; 10,000 U/mL, Thermo Fisher) in a humidified 5% CO2 incubator. Media was changed every 2–3 days until cells achieved 80% confluence then harvested with 0.25% trypsin/1 mM EDTA (Cat #: 25200056; Thermo Fisher) and passaged. For all experiments in this study, cells were used at passage 5. Complete donor information is listed in Table 1 .
4
Micropatterning of hASCs on Heparin Hydrogels
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We used hASCs (R7788115; Thermo Fisher Scientific, Waltham, MA, USA) for cell-loaded micropatches. hASCs were cultured in a 10 cm2 culture dish containing Alpha modified Eagle’s medium (Gibco, Grand Island, NY, USA), 10% fetal bovine serum (FBS; Gibco), and 1% penicillin–streptomycin (Gibco) at 37 °C in a humidified atmosphere of 5% CO2 as manufactures protocol. When cells reached 70–80% confluence, they were trypsinized with TrypLE™ Express without phenol red (Gibco), and the seeding density was 5000 cells/cm2. The medium was replaced every 3–4 days with fresh subculture. Cells were used for transplantation at passages 5–6.
Before seeding hASCs on the micropatterned heparin-based hydrogel, PEG-SH in PBS (pH 8.0) was grafted on the remaining area of the electrode without hydrogel pattern to prevent the non-selective cell attachment on the substrate. Then, hASCs (2 × 105 cells) were seeded on the electrode with hydrogel micropattern in a 6-well plate (ultra-low attachment). The seeded cells on the micropatterned hydrogels were cultured for 7 days in DMEM containing 10% FBS and 1% penicillin/streptomycin.
Before seeding hASCs on the micropatterned heparin-based hydrogel, PEG-SH in PBS (pH 8.0) was grafted on the remaining area of the electrode without hydrogel pattern to prevent the non-selective cell attachment on the substrate. Then, hASCs (2 × 105 cells) were seeded on the electrode with hydrogel micropattern in a 6-well plate (ultra-low attachment). The seeded cells on the micropatterned hydrogels were cultured for 7 days in DMEM containing 10% FBS and 1% penicillin/streptomycin.
5
Fabrication of Minispheroid-Loaded Liver Tissues
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The present study used hASCs (Lonza, Basel, Switzerland) to establish an E-MS-printing process. Dulbecco's modified Eagle's medium low glucose (DMEM-L; Sigma-Aldrich, USA) containing 10% fetal bovine serum (FBS; Biowest, France) and 1% penicillin/streptomycin (PS; Gibco, USA) was used to culture the hASCs. HepG2 cells (Korean Cell Line Bank, South Korea) and HUVECs (Lonza, USA) were fabricating minispheroid-loaded liver tissues. Minimum essential medium (MEM) (Gibco, USA) containing 10% FBS, 1% PS, 25 mM HEPES (Sigma-Aldrich, USA), 25 mM Sodium Bicarbonate (Sigma-Aldrich, USA), and EBM™-2 Bullet Kit™ (EBM; Lonza, USA) containing 10% FBS and 1% PS were used to culture HepG2 cells and HUVECs, respectively. The cells were incubated at 37 °C with 5% CO2, and the culture medium was changed every two days.
6
Evaluating Bone Graft Biocompatibility with hASCs
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Human adipose-derived stem cells (hASCs) were purchased from Lonza (PT-5006, Lot.0000605220). hASCs were maintained in Dulbecco’s Modified Eagle’s Medium (DMEM, Gibco®, Thermo Fisher Scientific, lnc., Walthan, MA, USA) with 10% Fetal Bovine Serum (FBS, Gibco®, Thermo Fisher Scientific, lnc., Walthan, MA, USA) and 100 units/mL of penicillin–streptomycin (Gibco®) in a humidified 37 °C incubator with 5% CO2. hASCs (passage 5) were seeded at a density of 1 × 105 cells/well with culture media onto a 12-well plate. After 24 h of culture, the culture medium was replaced with an extract medium which was eluted for 24 h by soaking each bone graft. The control groups were exposed only to standard culture medium. After 24 or 48 h of incubation, cell viability was evalutated using a CCK-8 assay (Dojindo Molecular Technologies, Inc., Rockville, MD, USA). CCK reagent was added per well and incubated at 37 °C for 3 h. The absorbance values were measured at 450 nm using a microplate reader (Epoch™ Microplate Spectrophotometer, BioTek Instruments, Inc., Winooski, VT, USA). All experiments were performed in triplicate wells for each group.
7
3D Spheroid Formation of hASCs
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hASCs (CEFO, Seoul, Korea) were grown in CEFOgro media (CEFO, Seoul, Korea) according to manufacturer’s instruction. Fifth passage (P5) hASCs were used for all experiments. For 3D spheroid (FECS-Ad) formation of hASCs, cells were trypsinized and suspended in StemPro® MSC serum-free media (SFM) (Gibco, MA), and plated with a density of 1 × 104 cells per well in MBP-FGF2 coated 384-well microplates. Cells were cultured for 24 h at 37 °C incubator with 5% CO2. Formation of FECS-Ad was determined using an Axio Vert A1 phase contrast microscope (Zeiss, Oberkochen, Germany).
8
Comparison of Adipose-Derived and Bone Marrow Stem Cells
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The human adipose‐derived stem cells (hASCs) and human bone marrow mesenchymal stem cells (hBMSCs) used in our study were obtained from ScienCell Research Laboratory (Carlsbad, CA). Cells in this study were between three and five passages and obtained from three healthy adult donors. All materials used in cell culture were bought from Sigma–Aldrich (St. Louis, MO). For the in vitro experiments, proliferation medium (PM) for hASCs consisted of fetal bovine serum (FBS; 10%, vol/vol), penicillin G (100 U/ml), and streptomycin (100 mg ml) into Dulbecco's modified Eagle's medium; the PM for hBMSCs consisted of Minimum Essential Medium α (α‐MEM, Gibco, Grand island, USA), 10% (vol/vol) FBS, penicillin G (100 U/ml), and streptomycin (100 mg/ml). Dexamethasone (100 nM), l ‐ascorbic acid (200 mM), and β‐glycerophosphate (10 mM) were added into PM to make osteogenic medium (OM). The cell culture conditions were 95% air, 5% CO2, 100% relative humidity, and 37°C.
9
Adipose-Derived Stem Cell Signaling
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Human adipose-derived stem cells (hASCs) were purchased from Invitrogen (USA). Primary antibodies against p44/42 MAPK (ERK1/2), Phospho-p44/42 MAPK (p-ERK1/2), acetyl CoA carboxylase (ACC), phospho-ACC (p-ACC), Src homology region 2 domain-containing phosphatase-1 (SHP1), carnitine palmitoyltransferase-1a (CPT-1a), fatty acid synthetase (FAS), sterol regulatory element-binding protein-1c (SREBP-1c), or β-actin and horseradish peroxidase (HRP)-conjugated secondary antibody were purchased from Abcam (Cambridge, UK). All cell culture related reagents were obtained from Shanghai Sangon Biotechnology (China). All mRNA generation related reagents were obtained from Shanghai Yeasen Biotechnology (China). All other reagents were purchased from Abcam (Cambridge, UK) or Sigma-Aldrich (Germany) unless otherwise stated. Pioglitazone with purity over 99.5% was purchased from MedChemExpress（USA）
Healthy male Sprague Dawley (SD) rats, weighing 200 ± 20 g, were purchased from Shanghai SLAC Laboratory Animal Co., Ltd. (China, Shanghai). Before the experiment, the animals were adaptively fed with standard rat diet for 1 week at room temperature 23 ± 2 °C. All animal experiments were approved by the Animal Protection and Utilization Committee of Fudan University.
Healthy male Sprague Dawley (SD) rats, weighing 200 ± 20 g, were purchased from Shanghai SLAC Laboratory Animal Co., Ltd. (China, Shanghai). Before the experiment, the animals were adaptively fed with standard rat diet for 1 week at room temperature 23 ± 2 °C. All animal experiments were approved by the Animal Protection and Utilization Committee of Fudan University.
10
Cultivation of Human Adipose Stem Cells
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Human adipose-derived stem cells (hASCs) were purchased from Invitrogen (Waltham, MA, USA). hASCs were cultured according to the manufacturer’s instructions. We used Dulbecco’s modified Eagle’s medium (DMEM; Gibco, Waltham, MA, USA) containing 10% fetal bovine serum (FBS; Gibco) and 1% antibiotics (Gibco). The media were changed every 3 days, and cells from passages 7 to 10 were used for the experiments.
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