# Bmal1 is involved in the regulation of macrophage cholesterol homeostasis

## TL;DR

This study shows that Bmal1 in macrophages controls cholesterol balance, and its absence worsens atherosclerosis by affecting cholesterol uptake, transport, and removal.

## Contribution

The study identifies Bmal1 as a master regulator of macrophage cholesterol homeostasis and atherosclerosis through direct and indirect gene regulation.

## Key findings

- Bmal1-deficient macrophages have higher cholesterol content and increased atherosclerosis.
- Bmal1 regulates cholesterol metabolism genes like Cd36, Abca1, Abcg1, Npc1, and Npc2.
- Bmal1 controls cholesterol efflux, reverse cholesterol transport, and lysosomal cholesterol egress in macrophages.

## Abstract

Atherosclerotic cardiovascular disease is a major contributor to the global disease burden. We previously demonstrated that Clock-mutant mice, and mice with global or liver-specific Bmal1 deficiency, exhibit enhanced atherosclerosis because of overproduction of lipoproteins and sustained hyperlipidemia. Atherosclerosis initiation depends on cholesterol accumulation in subendothelial macrophages (Mφs). To clarify the role of Bmal1 in Mφ function and atherosclerosis, we used several global and myeloid-specific Bmal1-deficient mouse models. Mφ-specific Bmal1-deficient mice had similar plasma lipid levels, higher Mφ cholesterol content, and displayed greater atherosclerosis compared with controls. We attempted to understand molecular mechanisms for increased cellular cholesterol levels. Bmal1-deficient Mφs exhibited: (a) elevated expression of Cd36 and uptake of oxLDL; (b) diminished expression of Abca1 and Abcg1, and decreased cholesterol efflux and reverse cholesterol transport; and (c) reduced Npc1 and Npc2 expression and diminished cholesterol egress from lysosomes. Molecular studies revealed that Bmal1 directly regulates basal and cyclic expression of Npc1 and Npc2 by binding the E-box motif (CANNTG) sequence recognized by Bmal1 in their promoters and indirectly regulates the basal and temporal regulation of Cd36 and Abca1/Abcg1 involving Rev-erbα and Znf202 repressors, respectively. In conclusion, Mφ Bmal1 is a key regulator of the uptake of modified lipoproteins, cholesterol efflux, lysosomal cholesterol egress, and atherosclerosis and, therefore, may be a master regulator of cholesterol metabolism in Mφs. Restoration of Mφ Bmal1 expression or blocking of factors that decrease its activity may be effective in preventing atherosclerosis.

We provide evidence that macrophage Bmal1 regulates uptake of modified lipoproteins, intracellular cholesterol trafficking and cholesterol efflux.