Objective-The purpose of this study was to identify rare APOA5 variants in 130 severe hypertriglyceridemic patients by sequencing, and to test their functionality, since no patient recall was possible. Methods and Results-We studied the impact in vitro on LPL activity and receptor binding of 3 novel heterozygous variants, apoAV-E255G, -G271C, and -H321L, together with the previously reported -G185C, -Q139X, -Q148X, and a novel construct -⌬139 to 147. Using VLDL as a TG-source, compared to wild type, apoAV-G255, -L321 and -C185 showed reduced LPL activation (Ϫ25% [Pϭ0.005], Ϫ36% [PϽ0.0001], and Ϫ23% [Pϭ0.02]), respectively). ApoAV-C271, -X139, -X148, and ⌬139 to 147 had little affect on LPL activity, but apoAV-X139, -X148, and -C271 showed no binding to LDL-family receptors, LR8 or LRP1. Although the G271C proband carried no LPL and APOC2 mutations, the H321L carrier was heterozygous for LPL P207L. The E255G carrier was homozygous for LPL W86G, yet only experienced severe hypertriglyceridemia when pregnant. Key Words: apolipoprotein AV Ⅲ LDL-R family Ⅲ LR8 Ⅲ LRP1 Ⅲ HSPG-bound LPL P remature truncations of APOA5, Q139X, Q148X, and the IVS3ϩ3gϾc are associated with severe hypertriglyceridemia (HyperTG), 1-3 behaving as phenocopies of lipoprotein lipase (LPL) deficiency. These rare APOA5 variants do not always lead to a deficiency in circulating plasma apoAV, and carriers present with a range of apoAV levels (reviewed in 4 ). Kao et al 5 identified a common polymorphism G185C in a Taiwanese study which occurs at a minor allele frequency of 0.04 in controls but at a 6.3-fold higher frequency of 0.27 in hypertriglyceridemic patients (PϽ0.001).
Conclusion-TheApoAV is present on chylomicrons, VLDL, and HDL, but not on IDL or LDL, suggesting that VLDL-containing apoAV is cleared before the lipolytic cascade. 6 One function of apoAV is to activate LPL, and Apoa5 knockout mice have 4-fold higher TG levels than wild type. [7][8][9] Whereas LPL transgenic mice can rescue Apoa5 knockout mice from HyperTG, this is not entirely reciprocal 8 suggesting that the effect of apoAV on plasma TG is dependent on heparinsulfate proteoglycan (HSPGs)-bound LPL. 7,8 In addition, apoAV-dependent TG catabolism acts by enhancing receptormediated endocytosis via members of the LDL-receptor (LDLR) family. 10,11 We have identified 3 novel APOA5 missense variants (E255G, G271C, H321L) in patients with TG levels Ͼ10 mmol/L. Although LPL and APOC2 variants had been excluded for the G271C carrier, the coding exons of LPL and APOC2 were sequenced in the 2 other probands. Because family studies were not possible, the APOA5 variants were expressed in vitro together with G185C, Q139X, Q148X. 1,2,5 In addition we designed a deletion construct ⌬139 to 147, to
MethodsSee supplemental methods (available online at http://atvb.ahajournals. org) for full details.
PatientsIn total 130 patients with TG levels Ͼ10 mmol/L were recruited into the study; seven patients from the UK, 28 patients from the Netherlands with LPL and APOC2 mutations excluded, and 95 p...
