Protocol to track the biosynthesis of cholesterol in cultured HCC cells using 13C compound-specific stable isotopic tracers

Jonathan D. Cybulski; Kit Sum Leung; Carmen Oi Ning Leung; David M. Baker; Terence Kin Wah Lee

doi:10.1016/j.xpro.2023.102506

Protocol to track the biosynthesis of cholesterol in cultured HCC cells using ¹³C compound-specific stable isotopic tracers

Jonathan D. Cybulski
, Kit Sum Leung
, Carmen Oi Ning Leung
, David M. Baker
, Terence Kin Wah Lee

Research output: Journal article publication › Journal article › Academic research › peer-review

Abstract

Cholesterol biosynthesis supports proliferation and drives resistance to tyrosine kinase inhibitor (TKI) therapy in hepatocellular carcinoma (HCC). Here, we present a protocol for using stable isotopic tracers to track the biosynthesis of cholesterol in cultured HCC cells. We describe steps for cell preparation, incubation, separation, and homogenization. We then detail lipid extraction and compound-specific isotope analysis for comparing and quantifying cholesterol synthesis between TKI-resistant HCC cells and their mock counterparts. This protocol can be expanded for use with other shorter-chained lipids.

Original language	English
Article number	102506
Journal	STAR Protocols
Volume	4
Issue number	3
DOIs	https://doi.org/10.1016/j.xpro.2023.102506
Publication status	Published - 15 Sept 2023

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

SDG 3 Good Health and Well-being

Keywords

Cancer
Cell Culture
Chemistry
Metabolism

ASJC Scopus subject areas

General Neuroscience
General Biochemistry,Genetics and Molecular Biology
General Immunology and Microbiology

More information

10.1016/j.xpro.2023.102506

http://hdl.handle.net/10397/108488

Cite this

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title = "Protocol to track the biosynthesis of cholesterol in cultured HCC cells using 13C compound-specific stable isotopic tracers",

abstract = "Cholesterol biosynthesis supports proliferation and drives resistance to tyrosine kinase inhibitor (TKI) therapy in hepatocellular carcinoma (HCC). Here, we present a protocol for using stable isotopic tracers to track the biosynthesis of cholesterol in cultured HCC cells. We describe steps for cell preparation, incubation, separation, and homogenization. We then detail lipid extraction and compound-specific isotope analysis for comparing and quantifying cholesterol synthesis between TKI-resistant HCC cells and their mock counterparts. This protocol can be expanded for use with other shorter-chained lipids.",

keywords = "Cancer, Cell Culture, Chemistry, Metabolism",

author = "Cybulski, \{Jonathan D.\} and Leung, \{Kit Sum\} and Leung, \{Carmen Oi Ning\} and Baker, \{David M.\} and Lee, \{Terence Kin Wah\}",

note = "Publisher Copyright: {\textcopyright} 2023 The Author(s)",

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T1 - Protocol to track the biosynthesis of cholesterol in cultured HCC cells using 13C compound-specific stable isotopic tracers

AU - Cybulski, Jonathan D.

AU - Leung, Kit Sum

AU - Leung, Carmen Oi Ning

AU - Baker, David M.

AU - Lee, Terence Kin Wah

PY - 2023/9/15

Y1 - 2023/9/15

N2 - Cholesterol biosynthesis supports proliferation and drives resistance to tyrosine kinase inhibitor (TKI) therapy in hepatocellular carcinoma (HCC). Here, we present a protocol for using stable isotopic tracers to track the biosynthesis of cholesterol in cultured HCC cells. We describe steps for cell preparation, incubation, separation, and homogenization. We then detail lipid extraction and compound-specific isotope analysis for comparing and quantifying cholesterol synthesis between TKI-resistant HCC cells and their mock counterparts. This protocol can be expanded for use with other shorter-chained lipids.

AB - Cholesterol biosynthesis supports proliferation and drives resistance to tyrosine kinase inhibitor (TKI) therapy in hepatocellular carcinoma (HCC). Here, we present a protocol for using stable isotopic tracers to track the biosynthesis of cholesterol in cultured HCC cells. We describe steps for cell preparation, incubation, separation, and homogenization. We then detail lipid extraction and compound-specific isotope analysis for comparing and quantifying cholesterol synthesis between TKI-resistant HCC cells and their mock counterparts. This protocol can be expanded for use with other shorter-chained lipids.

KW - Cancer

KW - Cell Culture

KW - Chemistry

KW - Metabolism

UR - https://www.scopus.com/pages/publications/85167980480

U2 - 10.1016/j.xpro.2023.102506

DO - 10.1016/j.xpro.2023.102506

M3 - Journal article

C2 - 37594893

AN - SCOPUS:85167980480

SN - 2666-1667

VL - 4

JO - STAR Protocols

JF - STAR Protocols

IS - 3

M1 - 102506

ER -