Nonphysiological overexpression of low-density lipoprotein receptors causes pathological intracellular lipid accumulation and the formation of cholesterol and cholesteryl ester crystals in vitro

Tools

Item Type:	Article
Title:	Nonphysiological overexpression of low-density lipoprotein receptors causes pathological intracellular lipid accumulation and the formation of cholesterol and cholesteryl ester crystals in vitro
Creators Name:	Heeren, J., Steinwaerder, D.S., Schnieders, F., Cichon, G., Strauss, M. and Beisiegel, U.
Abstract:	Recent therapeutic strategies for the treatment of familial hypercholesterolemia have been based on liver-directed gene transfer of a functional low-density lipoprotein (LDL) receptor cDNA under control of viral or strong housekeeping promoters. Strong viral promoters including cytomegalovirus, Rous sarcoma virus, and simian virus 40 promoters are commonly employed to reach significant physiological effects. These promoters mediate constitutive and nonphysiological overexpression in every transduced cell, while the endogenous LDL receptor expression is controlled by a complex feedback mechanism based on intracellular cholesterol concentration. To investigate intracellular consequences of persistent LDL receptor overexpression we constructed a recombinant adenovirus encoding the human LDL receptor under control of the Rous sarcoma virus promoter. The metabolic and morphological effects of LDL receptor expression were characterized by uptake experiments with human hepatoma cells using fluorescent and radiolabeled LDL. We observed that large amounts of LDL accumulate within LDL receptor transduced cells, which eventually lead to massive intracellular lipid deposition. Kinetic experiments with LDL-supplemented medium resulted in numerous crystal shaped structures in the cytosol of transduced cells as visualized by digital interference contrast optic within 60 min after LDL supplementation. Thin layer chromatography analyses of cellular lipids suggested these crystalline structures to be dependent on intracellular cholesterol and cholesterol ester levels. Mock-infected cells showed neither cholesterol lipid accumulation nor crystal formation. In conclusion, our data demonstrate that nonphysiological overexpression of the LDL receptor can cause massive lipid accumulation, which cannot be compensated by the hepatoma cell metabolism. This phenomenon may result in negative selection of LDL receptor overexpressing cells in vitro and in vivo.
Keywords:	Gene Therapy, Low-Density Lipoprotein Receptor, Regulated Gene Expression, Cholesterol Deposition, Familial Hypercholesterolemia, Animals, Cattle
Source:	Journal of Molecular Medicine
ISSN:	0946-2716
Publisher:	Springer
Volume:	77
Number:	10
Page Range:	735-743
Date:	October 1999
Official Publication:	https://doi.org/10.1007/s001099900045
PubMed:	View item in PubMed

Repository Staff Only: item control page