Lantation on atherosclerosis in Apoe-/- mice To figure out whether or not macrophage dysfunction contributes to enhanced atherosclerosis independent of hyperlipidemia, we transplanted bone marrow cells obtained from Clk19/19Apoe-/- or Apoe-/- mice into lethally irradiated Apoe-/- mice. Bone marrow transplantation slightly reduced total plasma cholesterol in Apoe-/- mice (Fig 7A). However,NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptCirculation. Author manuscript; accessible in PMC 2014 October 15.Pan et al.PageApoe-/- mice that received bone marrow cells from Clk19/19Apoe-/- mice had 2- to two.7fold greater atherosclerotic plaques in the ascending aortas and three major branching arteries (Fig 7B) and at the cardiac/aortic junctions (Fig 7C). Further, there was 3-fold larger lipid staining within the aorta (Fig 7D). In addition, macrophages obtained from mice transplanted with bone marrow cells from Clk19/19Apoe-/- were defective in cholesterol efflux to ApoAI and HDL (Fig 7E). Gene expression analysis showed that macrophages isolated from Apoe-/- mice transplanted with Clk19/19Apoe-/- bone marrow cells had low mRNA levels of ABCA1/ABCG1 and larger levels of CD36/SR-A1 (Fig 7F). Additional evaluation of transcription elements that regulate ABCA1 revealed that these macrophages had higher levels of USF2 (Fig 7G). As a result, macrophage dysfunction as a consequence of the expression of Clock19/19 protein contributes to atherosclerosis in Apoe-/- mice.NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptDiscussionUsing 3 distinct mouse models and three diverse diets we show for the initial time that Clock dysfunction as a consequence of the expression of a dominant adverse Clock19/19 protein increases atherosclerosis in mice.Methyl dec-9-enoate Order Different mouse models carrying Clock19/19 protein had higher cholesterol in apoB-containing non-HDL lipoproteins.Azido-PEG4-C2-acid Chemical name Mechanistic research revealed that Clock19/19 protein enhances cholesterol absorption by enterocytes and uptake of modified lipoproteins by macrophages in Apoe-/- mice.PMID:24118276 In contrast, it reduces cholesterol efflux from macrophages. Hence, Clock plays a vital and novel part in the regulation cholesterol metabolism in enterocytes and macrophages to prevent hypercholesterolemia and atherosclerosis. Biochemical analysis showed that hypercholesterolemia in Clk19/19Apoe-/- mice was on account of accumulation of cholesteryl ester-rich ApoB48containing lipoproteins. Physiologic research showed that enterocytes expressing Clock19/19 protein take up a lot more cholesterol from the intestinal lumen and secrete additional cholesterol with chylomicrons. Molecular research demonstrated that enhanced cholesterol uptake was related with enhanced expression of NPC1L1with no significant modifications in cholesterol exporters ABCG1/ABCG8. After uptake cholesterol is transported to plasma involving HDL and chylomicrons. HDL pathway was not impacted, but chylomicron pathway was up regulated in Clk19/19Apoe-/- mice. Two proteins, ACAT2 and MTP, involved in the assembly of chylomicrons had been elevated in Clk19/19Apoe-/- mice. ACAT2 converts free of charge cholesterol into cholesteryl esters and MTP transfers these cholesteryl esters to nascent lipoproteins to assist within the assembly and secretion of ApoB48-containing chylomicrons. As a result, Clock regulates cholesterol absorption by modulating cholesterol uptake, cholesterol esterification and chylomicron assembly. This study shows that Clock19/19 protein disrupts various macrophage functions; secretion of cytokines,.