Combined statin and niacin therapy remodels the high-density lipoprotein proteome

Abstract

Background: Boosting low high-density lipoprotein (HDL) levels is a current strategy for preventing clinical events that result from cardiovascular disease. We previously showed that HDL(3) of subjects with coronary artery disease is enriched in apolipoprotein E and that the lipoprotein carries a distinct protein cargo. This observation suggests that altered protein composition might affect the antiatherogenic and antiinflammatory properties of HDL. We hypothesized that an intervention that increases HDL levels-combined statin and niacin therapy-might reverse these changes.

Methods and results: HDL(3) isolated from 6 coronary artery disease subjects before and 1 year after combination therapy was analyzed by liquid chromatography-Fourier transform-mass spectrometry. Alterations in protein composition were detected by spectral counting and confirmed with extracted ion chromatograms. We found that combination therapy decreased the abundance of apolipoprotein E in HDL(3) while increasing the abundance of other macrophage proteins implicated in reverse cholesterol transport. Treatment-induced decreases in apolipoprotein E levels of HDL(3) were validated biochemically in a second group of 18 coronary artery disease subjects. Interestingly, the changes in HDL(3) proteome with niacin/statin treatment resulted in a protein composition that more closely resembled that of HDL(3) in healthy control subjects.

Conclusions: Combined statin and niacin therapy partially reverses the changes in the protein composition seen in HDL(3) in coronary artery disease subjects. Our observations raise the possibility that quantifying the HDL proteome could provide insights into the therapeutic efficacy of antiatherosclerotic interventions.

Figures

Figure 1. Niacin/statin treatment remodels the HDL3 proteome in six subjects with CAD

HDL3 was isolated from 6 CAD subjects before and after 1 year of therapy with sustained-release niacin and atorvastatin and tryptic digests of HDL3 analyzed by LC-MS/MS. Proteins identified as differentially expressed before (black bars) and during treatment (empty bars) by spectral counting were quantified by extracted ion chromatograms, using the area under the curve (AUC) of the chromatograms. Average AUCs for two representative peptides from each protein are shown (left column) along with a sample chromatograph for a peptide from each protein (right column). The peptides analyzed were: (A,B) PLTP, AGA (AGALQLLLVGDK3+) and SVV (SSVDELVGIDYSLM3+) (C,D) apoF, SGV (SGVQQLIQYYQDQK3+) and SYD (SYDLDPGAGSLEI); (E,F) apoJ, LFD (LFDSDPITVTVPVEVSR2+) and VTT (VTTVASHTSDSDVPSGVTEVVVK3+); (G,H) apoE, AAT (AATVGSLAGQPLQER2+) and GEV (GEVQAMLGQSTEELRVR3+). The effect of treatment on protein levels was determined by using all peptides quantified for each protein by repeated measures two-way analysis of variance. P<0.05 for all proteins other than apoA-I (P=0.44) and apoC-II (P=0.06). Shown are mean±SEM.

Figure 2. The HDL3 proteomes of healthy subjects and subjects with CAD differ

The peptides listed in Supplementary Table 2 were quantified by extracted ion chromatogram, using AUCs of the chromatograms from HDL3 isolated from six healthy subjects and six CAD subjects. The average normalized change for all peptides examined for each protein is shown. *P<0.01 for all proteins other than apoF (P=0.08) and apoA-I (P=0.39). Shown are mean±SEM

Figure 3. ApoE levels measured by nephelometry correlate with apoE levels quantified by extracted ion chromatogram (XIC)

The peptides used for the extracted ion chromatograms were (A) AAT (AATVGSLAGQPLQER2+, m/z [M+2H]2+ 749.405) and (B) LKS (LKSWFEPLVEDMQR3+, m/z [M+3H]3+ 593.303). Nephelometry and mass spectrometry were performed on two different preparations of HDL3 from the same plasma sample. The solid line represents the fit, and the dashed lines show the 95% confidence intervals in linear regression analysis.

Figure 4. Niacin/statin treatment reduces apoE levels in HDL3

ApoE levels from 18 subjects (Table 1, Group 2) were measured by nephelometry in HDL3 (A) and plasma (B) before and after 1 year of combined niacin/statin (N/S) therapy. *P=0.02. Shown is the correlation between baseline (C) and changes with N/S therapy (D) in apoE between plasma and HDL3. Shown are mean±SEM