Differential expression of double-band apolipoprotein(a) phenotypes in healthy Spanish subjects detected by SDS–agarose immunoblotting

Abstract

A sodium dodecyl sulphate-agarose apolipoprotein(a) [apo(a)] phenotyping method was set up to attain accurate scanning densitometry of proteins. Serum samples from 99 healthy Spanish men were analysed and twenty-five different apo(a) isoforms (12 to 37 kringle 4 repeats) were detected. Double-band phenotypes accounted for 39.4% ( n=39) and three different patterns of protein expression were identified: pattern A (20.5% of double-band phenotyped samples) predominantly expressed the highest molecular weight isoform; pattern B (53.9%) mainly the lowest molecular weight isoform, and pattern AB (25.6%), expressed both isoforms equally. A significant linear association between expression pattern and lipoprotein(a) [Lp(a)] concentration ≥0.30 g/l was observed. Single-band phenotyped samples ( n=60) were stratified according to apo(a) kringle 4 repeat categories and showed that 90% of isoforms <20 K4 repeats had high Lp(a) concentrations (≥0.30 g/l), whereas isoforms with 20 to 24 or more than 24 kringle 4 repeats had Lp(a) concentrations ≥0.30 g/l in 47% and 14%, respectively. A logistic regression model was fitted to test the association between apo(a) size, expression pattern and Lp(a) concentration. In this model, apo(a) isoform <25 kringle 4 repeats was significantly associated with serum Lp(a) concentration ≥0.30 g/l in both single and double-band phenotyped samples (odds ratio=8.9, p<0.001). In the latter, a differential expression pattern with respect to smaller size isoforms (pattern AB vs A) was significantly associated with Lp(a) concentration ≥0.30 g/l (odds ratio=17.97, P=0.045). Heterogeneity in protein apo(a) size expressed according to kringle 4 repeat number could be categorized in heterozygous phenotypes as three patterns. When small-sized isoform was expressed (pattern B) or both isoforms were equally expressed (pattern AB), the probability of having Lp(a)≥0.30 g/l is higher.