Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biology of Lipids
Effects of apolipoprotein B-100 on the metabolism of a lipid microemulsion model in rats
Introduction
LDL is a cholesterol-rich lipoprotein produced in vivo via very-low-density lipoprotein (VLDL) delipidation cascade [1], [2]. Apolipoprotein (apo) B-100, the protein moiety of low-density lipoprotein (LDL), plays an important role in the intracellular assembly of the precursor VLDL [3], [4]. In contrast with the other apolipoproteins, which exchange intensively among lipoprotein particles, apo B-100 remains part of the lipoprotein structure during the catabolic process, until the particles are removed from the bloodstream [2]. In this last step, apo B-100 is the ligand for the cellular recognition and uptake of LDL by the specific LDL receptor (LDLR) on the cell plasma membrane [3], [5].
In rats, apo B-100 is synthesized predominantly by the liver, whereas apo B-48, a shorter isoform, is mainly produced by the intestine [3]. The latter actually results from an editing mechanism of the apo B-100 mRNA [6], [7]. Apo B-100 is a large molecule with polar and apolar domains allowing both the association of the protein with lipids in LDL particle, as well as the interaction of the lipoprotein with LDLR [8], [9]. Previous investigations have used protein-free microemulsions resembling LDL, designated LDE, to study LDL metabolism [10], [11]. It has been shown that LDE can be removed from circulation through interaction of the apo E, adsorbed by LDE particles, with LDLR [11], [12].
In this study, we have evaluated the influence of apo B-100 on LDL metabolism, by modeling the LDL lipid structure with LDE. LDE labeled with radioactive lipid analogues and associated with apo B-100 (LDE-apoB) was injected into the bloodstream of control rats and rats pre-treated with 17α-ethinylestradiol. Administration of this hormone is known to increase LDLR activity [13]. Following injection, plasma kinetics and organ uptake of the LDE-apoB were determined and compared with the metabolic parameters of the LDE and native LDL.
Section snippets
Animals and treatments
Male Wistar rats, weighing 250–350 g, were fed on a standard commercial chow, with free access to food and water. One group of rats was injected subcutaneously a daily dose of 17α-ethinylestradiol (5 mg/kg body wt.) for 5 days before the experiment [13]. Control rats received 0.5 ml of propyleneglycol in equivalent periods.
Preparation of LDE
LDE was prepared from lipid mixture composed of 33% cholesteryl oleate (Nu Check Prep, Elysian, USA), 66% egg phosphatidylcholine (Lipid Products, Surrey, UK) and 1% glyceryl
Results
The chemical analysis of the microemulsions showed that lipid composition of LDE-apoB was similar to that of LDE (Table 1). Both LDE and LDE-apoB microemulsions and native LDL presented compositional data comparable to that previously reported [10], [11], [12], [17], [29], [30]. Cholesteryl ester (CE) label of both LDE and LDE-apoB presented a similar distribution profile on density gradient ultracentrifugation. About 80% of particles floated between 1.020 g/ml and 1.065 g/ml (Fig. 1). Protein
Discussion
The lipid structure of LDL can be artificially modeled by very well defined procedures [10], [11], [31], [32]. These protein-free microemulsion models mimic the lipid physical behavior of native LDL. In previous studies, we had demonstrated that the metabolism of LDE had some similarities with the metabolism of endogenous LDL in rats [11] and in human subjects [12]. We have hypothesized that in the plasma compartment, LDE is capable of assimilating apolipoproteins from the plasma lipoproteins,
Acknowledgements
The authors are grateful to Ricardo Omoto, Edson L. Silva and Edilberto P. Oliveira for help with the experiments, and Prof. Sonia Q. Doi (USUHS, USA) for revising the manuscript. This study was supported by grants from FAPESP, São Paulo, Brazil. M.H.H. and R.C.M. are recipients of award research scholarships from CNPq, Brasilia, Brazil.
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