Isolation and characterization of apolipoprotein B48-containing lipoproteins with a monoclonal antibody against apolipoprotein B48.
OBJECTIVE
Remnant lipoproteins are well known to play a pivotal role in atherosclerosis. In patients with postprandial dyslipidemia, metabolic pathways for exogenous lipoproteins are generally disturbed, resulting in accumulation of chylomicron remnants. Although it has been difficult to make a specific antibody against apolipoprotein B48 (apoB48) , a constituent of exogenous lipoproteins, we succeeded in creating a specific monoclonal antibody against apoB48. In this study, we isolated apoB48-containing lipoproteins from lipoproteins with a density less than 1.019 g/mL using this anti-apoB48 monoclonal antibody (4C8).


METHODS
Apolipoproteins and lipids were analyzed to confirm whether apoB48-containing lipoproteins are isolated from other lipoproteins. The characteristics of apoB48-containing lipoproteins in the plasma of patients were compared with type 2 diabetes mellitus and non-diabetic patients. Furthermore, the uptake of apoB48-containing lipoproteins by THP-1 cells (a human acute monocytic leukemia cell line) , HepG2 cells (a human hepatoma cell line) , and human umbilical vein endothelial cells (HUVEC) was investigated. Also, the expression of apoB48 receptors in these cells was tested with RT-PCR.


RESULTS
Apolipoprotein analysis of 4C8-bound lipoproteins indicated the isolation of apoB48-containing lipoproteins, because the content of apoB100 was quite low (less than 5%) . Compared with lipoproteins that were not bound to the antibody, apoB48-containing lipoproteins had a higher content of triglycerides. There was no significant difference in the composition of apoB48-containing lipo-proteins between patients with and without type 2 diabetes. Uptake of fluorescence-labeled apoB48-containing lipoproteins by THP-1-derived macrophages and HepG2 cells, but not by HUVEC, was observed. The specificity of this uptake was confirmed because the fluorescent signal was competed out by an excess amount of the same unlabeled lipoproteins. RT-PCR revealed the expression of apoB48 receptors in THP-1 and HepG2 cells but not in HUVEC. These results suggest that specific uptake of apoB48-containing lipoproteins may occur via apoB48 receptors.


CONCLUSIONS
ApoB48-containing lipoproteins have a higher triglyceride content and are taken up into THP-1-derived macrophages and HepG2 cells by a specific pathway.
Yoshimura
Nakayuki
N
Department of Internal Medicine, Teikyo University School of Medicine, Tokyo, Japan.
Kinoshita
Makoto
M
Teramoto
Tamio
T
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D053283:Apolipoprotein B-48 / Q000737:chemistry* / Q000302:isolation & purification*; D045744:Cell Line, Tumor; D002914:Chylomicrons / Q000737:chemistry; D003924:Diabetes Mellitus, Type 2 / Q000097:blood* / Q000276:immunology; D050171:Dyslipidemias / Q000378:metabolism / Q000473:pathology; D004730:Endothelium, Vascular / Q000378:metabolism; D005786:Gene Expression Regulation; D015972:Gene Expression Regulation, Neoplastic; D056945:Hep G2 Cells; D006801:Humans; D008074:Lipoproteins / Q000737:chemistry*; D019518:Postprandial Period; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000911:Antibodies, Monoclonal; D053283:Apolipoprotein B-48; D002914:Chylomicrons; D008074:Lipoproteins