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2 protocols using srebf1

1

HepG2 Cell Culture and Characterization

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The HepG2 human
hepatocellular liver cell line was obtained from the Korea Research
Institute of Bioscience and Biotechnology (Daejeon, South Korea) and
grown in Eagle’s minimum essential medium (EMEM) containing
10% fetal bovine serum and 100 U/mL penicillin/streptomycin sulfate.
Cells were incubated in a humidified 5% CO2 atmosphere
at 37 °C. EMEM, penicillin, and streptomycin were purchased from
Hyclone (Logan, UT, USA). Bovine serum albumin (BSA) and 25-hydroxycholesterol
were purchased from Sigma-Aldrich (St. Louis, MO, USA). 4′,6-Diamidino-2-phe-nylindole
(DAPI) and DiI-LDL were purchased from Thermo Fisher Scientific (Waltham,
MA, USA). Antibodies against LSS, SREBP1, SREBP2, HMGCR, SQLE, and
β-actin were purchased from Abcam, Inc. (Cambridge, MA, USA).
Antibodies against PCSK9 were purchased from Cell Signaling Technology.
(Beverly, MA, USA). SREBF1, SREBF2, HMGCR, PCSK9, IDI1, SQLE, ABCA1,
ACSL6, DHCR7, FDFT1, FDPS, SOAT1, and glyceraldehyde-3-phosphate dehydrogenase
(GAPDH) oligonucleotide primers were purchased from Bioneer Corp.
(Daejeon, South Korea). α-Mangostin was isolated from the chloroform
fraction of G. mangostana, as previously
described, and confirmed by a spectroscopic analysis. The purity of
α-mangostin was determined to be over 95% by high-performance
liquid chromatography using an ultraviolet detector.4 (link)
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2

HepG2 Cell Culture and Molecular Analysis

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The HepG2 human
hepatocellular liver cell line (provided by the Korea Research Institute
of Bioscience and Biotechnology, South Korea) was cultured according
to the previous publication.48 (link) Eagle’s
minimum essential medium, penicillin, streptomycin of Hyclone (Logan,
UT, USA), and bovine serum albumin from Sigma-Aldrich (St. Louis,
MO, USA) were used for cell culture. Antibodies (PCSK9, LDLR, and
β-actin) were purchased from Abcam, Inc. (Cambridge, MA, USA).
Primers for PCSK9, LDLR, HNF1a, SREBF1, SREBF2, and glyceraldehyde-3-phosphate
dehydrogenase (GAPDH) were purchased from Bioneer Corp. (Daejeon,
South Korea). Solvents for extraction and isolation (ethanol, methanol,
chloroform, etc.) were provided by SK Chemicals (Seoul, Korea). HPLC-grade
solvents, acetonitrile and methanol (SK Chemicals, Seoul, South Korea),
were used for HPLC. NMR solvents were purchased from Cambridge Isotope
Laboratories, Inc. (Andover, MA, USA).
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