Apolipoprotein M (apoM) is a novel apolipoprotein with unknown function. In this study, we established a method for isolating apoM-containing lipoproteins and studied their composition and the effect of apoM on HDL function. ApoM-containing lipoproteins were isolated from human plasma with immunoaffinity chromatography and compared with lipoproteins lacking apoM. The apoMcontaining lipoproteins were predominantly of HDL size; z5% of the total HDL population contained apoM. Mass spectrometry showed that the apoM-containing lipoproteins also contained apoJ, apoA-I, apoA-II, apoC-I, apoC-II, apoC-III, paraoxonase 1, and apoB. ApoM-containing HDL (HDL apoM1 ) contained significantly more free cholesterol than HDL lacking apoM (HDL apoM2 ) (5.9 6 0.7% vs. 3.2 6 0.5%; P , 0.005) and was heterogeneous in size with both small and large particles. HDL apoM1 inhibited Cu 21 -induced oxidation of LDL and stimulated cholesterol efflux from THP-1 foam cells more efficiently than HDL apoM2 . In conclusion, our results suggest that apoM is associated with a small heterogeneous subpopulation of HDL particles. Nevertheless, apoM designates a subpopulation of HDL that protects LDL against oxidation and stimulates cholesterol efflux more efficiently than HDL lacking apoM.
Apolipoprotein M (apoM) is a 188 amino acid, 25 kDa protein belonging to the lipocalin protein superfamily. Although predominantly associated with high density lipoprotein, apoM is found in all major lipoprotein classes. To facilitate clinical studies of apoM, we have developed a sandwich ELISA for the measurement of apoM in human plasma. This method has been used to investigate normal apoM variation and to establish reference values for healthy individuals through the measurement of 598 samples from the Nordic Reference Interval Project Bio-bank and Database (NOBIDA) biobank. For women 18-49 years old, the reference interval for apoM was 0.58-1.18 mmol/l, whereas for women 501 years and for men, the reference range was 0.61-1.30 mmol/l. Correlation studies of apoM with 26 common clinical chemical analytes from the NOBIDA database revealed a marked positive correlation with plasma total cholesterol (r 5 0.52) and LDL and HDL cholesterol (r 5 0.43 and 0.36, respectively). There was no statistically significant correlation with HDL/total cholesterol ratio or body mass index. In conclusion, a sandwich ELISA for the measurement of apoM in human plasma shows that apoM concentration is strongly correlated to total cholesterol in healthy individuals.-Axler, O., J. Ahnström, and B. Dahlbäck. An ELISA for apolipoprotein M reveals a strong correlation to total cholesterol in human plasma.
Lipoproteins consist of lipids solubilized by apolipoproteins. The lipid-binding structural motifs of apolipoproteins include amphipathic ␣-helixes and -sheets. Plasma apolipoprotein (apo) M lacks an external amphipathic motif but, nevertheless, is exclusively associated with lipoproteins (mainly high density lipoprotein). Uniquely, however, apoM is secreted to plasma without cleavage of its hydrophobic NH 2 -terminal signal peptide. To test whether the signal peptide serves as a lipoprotein anchor for apoM in plasma, we generated mice expressing a mutated apoM Q22A cDNA in the liver (apoM Q22A -Tg mice (transgenic mice)) and compared them with mice expressing wild-type human apoM (apoM-Tg mice). The substitution of the amino acid glutamine 22 with alanine in apoM Q22A results in secretion of human apoM without a signal peptide. The human apoM mRNA level in liver and the amount of human apoM protein secretion from hepatocytes were similar in apoM-Tg and apoM Q22A -Tg mice. Nevertheless, human apoM was not detectable in plasma of apoM Q22A -Tg mice, whereas it was easily measured in the apoM-Tg mice. To examine the plasma metabolism, recombinant apoM lacking the signal peptide was produced in Escherichia coli and injected into wild-type mice. The apoM without signal peptide did not associate with lipoproteins and was rapidly cleared in the kidney. Accordingly, ligation of the kidney arteries in apoM Q22A -Tg mice resulted in rapid accumulation of human apoM in plasma. The data suggest that hydrophobic signal peptide sequences, if preserved upon secretion, can anchor plasma proteins in lipoproteins. In the case of apoM, this mechanism prevents rapid loss by filtration in the kidney.Lipoproteins consist of lipids (mainly cholesterol, phospholipids, and triglycerides) solubilized by apolipoproteins. The apolipoproteins play essential roles in controlling plasma and tissue lipid homeostasis by interacting with cellular lipoprotein receptors (the low density lipoprotein receptor, the scavenger receptor class B type-I, the ATP-binding cassette transporter AI, etc.) and enzymes (e.g. lecithin-cholesterol acyltransferase, hepatic lipase, and lipoprotein lipase) and lipid transfer protein (cholesteryl ester transfer protein and phospholipid transfer protein). However, several apolipoproteins have roles beyond lipid metabolism. Indeed, a recent shotgun proteomic approach revealed that HDL 3 on average contains Ͼ40 proteins, of which many affect complement activation or protease inhibition (1). Also, several well known HDL apolipoproteins have roles in inflammation and host defense against microbial infections (1). For instance, apoL is involved in fighting Trypanosoma infections (2), and apoJ (clusterin) has even been proposed to play a significant role in tumorigenesis (3). For apolipoproteins without any (known) effect on plasma lipid metabolism, it is conceivable that the host HDL particle mainly serves as a carrier that prevents rapid removal of its associated apolipoproteins (e.g. by filtration in the kidney) or perh...
Apolipoprotein M (apoM) is a plasma protein associated mainly with HDL.
A novel human apolipoprotein [apolipoprotein M (apoM)] was recently described and demonstrated to be a lipocalin. We have now examined apoM in wild-type mice and mice with genetically altered lipoprotein metabolism. Liver and kidney showed high mRNA expression, whereas spleen, heart, brain, and testis demonstrated low expression. ApoM gene expression was initiated on embryonic day 10. Western blot analysis of plasma suggested that mouse apoM, like its human counterpart, is secreted with a retained signal peptide, but unlike human apoM it is not glycosylated. Gel filtration of plasma showed apoM to be associated with HDL-sized particles in wild-type and apoA-Ideficient mice and with HDL-and LDL-sized particles in LDL receptor-deficient mice, whereas apoM was mainly found in VLDL-sized particles in high-fat, high-cholesterolfed apoE-deficient mice. The plasma concentration of apoM was similar in wild-type, LDL receptor-deficient, and apoE-deficient mice but was reduced to 33% in apoA-I-deficient compared with wild-type mice ( P ؍ 0.007). These data suggest that apoM mainly associates with HDL in normal mice but also with the pathologically increased lipoprotein fraction in genetically modified mice. The substantially decreased apoM levels in apoA-I-deficient mice suggest a connection between apoM and apoA-I metabolism.
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