Altered mitochondrial function and cholesterol synthesis influences protein synthesis in extended HepG2 spheroid cultures.
Cultures of hepatocytes and HepG2 cells provide useful in vitro models of liver specific function. In this study, we investigated metabolic and biosynthetic function in 3-D HepG2 spheroid cultures, in particular to characterise changes on prolonged culture. We show that HepG2 cells cultured in spheroids demonstrate a reduction in mitochondrial membrane potential and respiration following 10 days of culture. This coincides with a modest reduction in glycolysis but an increase in glucose uptake where increased glycogen synthesis occurs at the expense of the intracellular ATP pool. Lowered biosynthesis coincides with and is linked to mitochondrial functional decline since low glucose-adapted spheroids, which exhibit extended mitochondrial function, have stable biosynthetic activity during extended culture although biosynthetic function is lower. This indicates that glucose is required for biosynthetic output but sustained mitochondrial function is required for the maintenance of biosynthetic function. Furthermore, we show that cholesterol synthesis is markedly increased in spheroids cf. monolayer culture and that inhibition of cholesterol synthesis by lovastatin extends mitochondrial and biosynthetic function. Therefore, increased cholesterol synthesis and/or its derivatives contributes to mitochondrial functional decline in extended HepG2 spheroid cultures.
Damelin
Leonard H
LH
Department of Medicine, Royal Free Campus, Royal Free and UCL Medical School, Rowland Hill Street, Hampstead, London, NW3 2PF, United Kingdom. l.damelin@rfc.ucl.ac.uk
Coward
Sam
S
Choudhury
Sarah F
SF
Chalmers
Sherri-Ann
SA
Cox
I Jane
IJ
Robertson
Nicola J
NJ
Revial
Gilbert
G
Miles
Marianne
M
Tootle
Rosemary
R
Hodgson
Humphrey J F
HJ
Selden
Clare
C
D000418:Albumins / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods*; D019069:Cell Respiration / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002784:Cholesterol / Q000096:biosynthesis*; D004777:Environment; D005947:Glucose / Q000378:metabolism; D022781:Hepatocytes / Q000187:drug effects / Q000502:physiology*; D008148:Lovastatin / Q000494:pharmacology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D008863:Microspheres; D008930:Mitochondria, Liver / Q000187:drug effects / Q000502:physiology*; D024101:Mitochondrial Proteins / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D015398:Signal Transduction / Q000502:physiology*; D023822:Tissue Engineering / Q000379:methods*
D000418:Albumins; D024101:Mitochondrial Proteins; D002784:Cholesterol; D008148:Lovastatin; D005947:Glucose; D010100:Oxygen