hAFS were cultured for 24 hours in serum‐free (SF) medium (Minimum Essential Medium Eagle alpha, with 1% l ‐glutamine and 1% penicillin/streptomycin) under normoxic (20% O2 and 5% CO2 at 37°C) or hypoxic (1% O2 and 5% CO2 at 37°C in a hypoxic incubator, Eppendorf, Hamburg, Germany; https://www.eppendorf.com/ ) conditions. The hAFS‐conditioned medium (hAFS‐CM) was collected and processed for hAFS‐EV isolation.
Hypoxic incubator
Manufactured by Eppendorf
Sourced in Germany, United States
The Hypoxic Incubator is a laboratory equipment designed to provide a controlled environment for cell culture and research applications. The device maintains a reduced oxygen concentration, simulating hypoxic conditions commonly found in certain biological systems.
Automatically generated - may contain errors
Lab products found in correlation
Fetal bovine serum (fbs), by Thermo Fisher Scientific (3 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (2 mentions)
Htr 8 svneo, by Thermo Fisher Scientific (1 mentions)
Rpmi 1640 medium, by Thermo Fisher Scientific (1 mentions)
Htr 8 svneo, by American Type Culture Collection (1 mentions)
Hep3b, by Korean Cell Line Bank (1 mentions)
Snu449, by Korean Cell Line Bank (1 mentions)
Hep3b, by Welgene (1 mentions)
Snu449, by Welgene (1 mentions)
Penicillin streptomycin p s, by Welgene (1 mentions)
Fgm 2 fibroblast growth medium 2 bulletkit, by Lonza (1 mentions)
Hct116, by Korean Cell Line Bank (1 mentions)
Mccoy s 5a medium, by Thermo Fisher Scientific (1 mentions)
8 protocols using hypoxic incubator
1
Hypoxic Conditioning of hAFS Cells
2
hBMSCs Hypoxia Culture Protocol
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hBMSCs were cultured until 60% confluence. Then, the medium was discarded and, after three PBS 1X washes, replaced with α-MEM-GlutaMax medium with only 100 U/mL of Penicillin-100 mg/mL Streptomycin (basal medium). Cells were maintained under normoxic (20% O2 and 5% CO2 at 37 °C) or hypoxic (1% O2 and 5% CO2 at 37 °C in a hypoxic incubator, Eppendorf, Hamburg, Germany) conditions, for 72 h. Conditioned media (CM) were collected and processed as described below.
3
Hypoxia-Conditioned MSC Secretome Enhances HUVEC
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Hypoxia is a pathophysiologic consequence of vascular and perfusion deficits [14 (link), 15 (link)]. Hypoxia has also been shown to enhance the proangiogenic properties of the MSC secretome [16 (link)–19 (link)]. Thus, we determined whether sbMSCs conditioned media in the setting of hypoxia could also promote human umbilical vein endothelial cell (HUVEC) survival and migration. To generate hypoxic conditioned media, sbMSCs (n = 4 patients) were cultured in T75 flask with 6 mL of basal EGM-2 media at 1% O2 using a hypoxic incubator (Eppendorf Inc., Enfield, CT). Conditioned media was then collected at 48 hours and stored at −80°C until used in downstream experiments.
4
Hypoxia Response of Extravillous Trophoblasts
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The human extravillous trophoblast cell line HTR‐8/SVneo was purchased from the American Type Culture Collection (ATCC, Manassas, VA). HTR‐8/SVneo cells were cultured in RPMI 1640 medium (Gibco Life Technologies, Carlsbad, CA), supplemented with 10% foetal bovine serum (FBS, Gibco Life Technologies) and 1% penicillin/streptomycin, in humidified air at 37°C with 5% CO2. Hypoxic culturing was performed by incubating HTR‐8/SVneo cells in a hypoxic incubator (Eppendorf, Hamburg, Germany) for 48 hours at oxygen concentrations of 10%, 5% and 2%.
5
Hypoxic culture of human HCC cell lines
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Human HCC cell lines, Hep3B, SNU449, and Huh7 cells were obtained from the Korean Cell line Bank (Seoul, Republic of Korea). Hep3B and SNU449 cells were grown in Roswell Park Memorial Institute-1640 medium (Welgene, Daegu, Republic of Korea) supplemented with 10% fetal bovine serum (FBS) (Welgene) and 1% penicillin streptomycin (PS) (Welgene). Huh7 cells were cultured in Dulbecco’s modified Eagle’s medium containing 10% FBS and 1% PS. For hypoxic incubation, cells were incubated in a hypoxic incubator (New Brunswick Scientific, Edison, NJ, USA) with a humidified environment consisting of 1% O2, 5% CO2, and 94% N2.
6
Hypoxia Effects on Lung Cells
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The human alveolar epithelial cell line A549 and diseased human lung fibroblasts from patients with idiopathic pulmonary fibrosis (DHLF-IPF cells) were obtained from the Korean Cell Line Bank (Seoul, South Korea) and Lonza (Basel, Switzerland), respectively. A549 cells were cultured in RPMI-1640 medium (Gibco Cell Culture, Carlsbad, CA, USA) containing 10% fetal bovine serum (FBS; Gibco) and 1% penicillin–streptomycin (Gibco), and DHLF-IPF cells were grown using an FGM-2 Fibroblast Growth Medium-2 BulletKit (Lonza). For hypoxic incubation, cells were incubated in a hypoxic incubator (New Brunswick Scientific, Edison, NJ, USA) with a humidified environment consisting of 1% O2, 5% CO2, and 94% N2.
7
Hypoxic Culture of Colon Cancer Cells
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HT29 and HCT116 human colon cancer cells were obtained from the Korean Cell Line Bank (Seoul, South Korea). The cells were cultured in McCoy's 5A medium (Gibco, Carlsbad, CA, USA) with 1% penicillin streptomycin (Gibco) and 10% fetal bovine serum (FBS; Gibco), at 37°C in a 5% CO2 humidified incubator. To generate hypoxic conditions, the cells were incubated in a hypoxic incubator (New Brunswick Scientific, Edison, NJ, USA) with 1% O2 and 5% CO2 balanced with 94% N2.
8
Hypoxia Exposure Impacts Cell Function
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