Immortalized Mouse WildType Aortic Endothelial Cells (iMAEC-WT)

Synthetic LXR and RXR Agonists, but Not Endogenous LXR and RXR Agonists, Increase ABCA1 Protein Expression in iMAEC

Atherosclerosis, leading to heart attacks and strokes, involves arterial cholesterol buildup. Statins lack effectiveness in directly reducing arterial cholesterol. Recent studies suggest targeting endothelial cell cholesterol with LXR/RXR agonists might benefit treatment, as they boost ABCA1 expression and enhance cholesterol efflux. Huang's team tests synthetic and endogenous LXR/RXR agonists on immortalized mouse aortic endothelial cells (iMAEC).

The researchers chose 22?-hydroxycholesterol and 9-cis-retinoic acid as natural activators of LXR and RXR because these compounds increase ABCA1 mRNA levels within endothelial cells. The synthetic LXR agonist GW3965 was selected for its function in increasing ABCA1 levels and promoting cholesterol efflux while T0901317 operates as an LXR agonist but also targets pregnane X receptors. SR11237 was selected as the synthetic RXR agonist for its specificity. Their findings demonstrated that 22?-hydroxycholesterol or 9-cis-retinoic acid alone failed to increase ABCA1 protein in iMAECs, but GW3965 or SR11237 alone significantly increased it (Fig. 1A–D). This suggests synthetic LXR or RXR agonists alone effectively upregulate ABCA1 protein, unlike their endogenous counterparts.

Fig. 1. Effect of LXR and RXR agonist treatment on ABCA1 protein expression in immortalized mouse aortic endothelial cells (iMAEC) (Huang K, Jo H, et al., 2021).

LPS-Challenged iMAECs Exhibit Robust Plasmid DNA Transfection Efficiency

Cholesterol and inflammation lead to atherosclerosis which commonly affects endothelial cells making them a significant focus for researchers. The functions of miR-33a-3p in biological processes have yet to be defined even though miR-33a is recognized for its role in managing cholesterol levels. Through plasmid transfection, Huang's team reduced miR-33a-3p levels in cultured iMAECs to investigate if miR-33a-3p inhibition boosts cholesterol efflux by increasing ABCA1 expression in pro-inflammatory endothelial cells.

To determine if miR-33a-3p inhibition can upregulate ABCA1 protein in pro-inflammatory endothelium, they first confirmed successful transfection of pA3p into these cells. Transfecting immortalized cell lines faces challenges, especially when inflammation affects transfection efficiency. We used endpoint RT-PCR to verify transgenic transcripts in LPS-challenged iMAECs transfected with pA3p or pScr (Fig. 2A). Flow cytometry evaluated transfection efficiency, showing high efficiency with both plasmids (Fig. 2B and C). miR-33a-3p expression was significantly reduced in LPS-challenged iMAECs transfected with pA3p compared to pScr (Fig. 2D). These results confirm that pA3p effectively inhibits miR-33a-3p in cultured pro-inflammatory endothelial cells.

Fig. 2. Transfecting pro-inflammatory endothelial cells with pA3p decreases miR-33a-3p expression (Huang K, Pokhrel A, et al., 2025).