Abstract
One of the leading causes of global mortality and morbidity is cardiovascular diseases (CVD). A well-known risk factor for the development of CVD is elevated plasma levels of lipoprotein (a) [Lp(a)]. Lp(a) consists of an low density lipoprotein (LDL) like lipoprotein attached to the plasminogen homologue, apolipoprotein (a) [apo(a)]. Despite many years of research, the exact pathway for the clearance of Lp(a) is still uncertain. Recently, it has been observed that Lp(a) was endocytosed into the liver via macropinocytosis and that macropinocytotic uptake of Lp(a) was enhanced by anti-depressants that inhibit serotonin and/or norepinephrine uptake. Increased Lp(a) uptake has been attributed to serotonin-induced upregulation in plasminogen receptor expression, thereby enhancing the surface binding of apo(a). Lp(a) levels are primarily genetically determined by the LPA gene which encodes apo(a) and determines its expression levels. Variants in the SLC22A3 gene, including the rs3088442 variant, have also been linked to Lp(a) levels, suggesting a separate genetic determinant of circulating Lp(a). Interestingly, the SLC22A3 transporter is involved in transporting serotonin and norepinephrine suggesting a potential mechanistic link to the involvement of serotonin in Lp(a) clearance. Thus, this study aimed to investigate the efficacy of different anti-depressants on Lp(a) uptake and possible involvement of SLC22A3.
Initially, the efficacy of the anti-depressants citalopram, imipramine, paroxetine and atomoxetine on Lp(a) uptake in liver cells (HepG2) were compared. Western blotting and confocal microscopy indicated that both citalopram and imipramine significantly increased Lp(a) uptake. Next, linkage disequilibrium (LD) analysis was performed to establish if the SLC22A3 rs3088442 variant was linked to variation in LPA or whether it was independently associated with plasma Lp(a) levels. Results from linkage analysis showed that rs3088442 was not in LD with any SNPs in the LPA gene, but it was in linkage with several other SNPs in the SLC22A3 gene. Analysis through the Genotype-Tissue Expression (GTEx) portal demonstrated that rs3088442 was associated with changes in gene expression of both LPA and SLC22A3 in the liver. Although rs3088442 was not likely in a regulatory region according to ENCODE, it was in linkage with other SNPs associated with Lp(a) levels in SLC22A3 that were in possible regulatory regions. This implies that the SLC22A3 transporter itself may be involved in Lp(a) catabolism. Following this analysis, structural investigation into coding variants was performed to explore possible functional consequences. Functional studies were carried out on these variants; however, it was apparent that optimisation was needed to produce robust reliable results.