Fig. 2. Hepatic cholesterol levels in response to hepatic EL expression in wild-type mice (A), SR-BI knockout mice (B) and mice with hepatic SR-BI overexpression (C). On day 5 following injection of the respective mouse models with either an adenovirus expressing … In SR-BI Axitinib melanoma knockout mice, hepatic total cholesterol content increased significantly by 32% in response to EL overexpression (16.8 �� 1.3 versus 12.7 �� 0.8 ��mol/liver, respectively; P < 0.05) (Fig. 2B), while hepatic free cholesterol (8.3 �� 1.3 versus 6.1 �� 0.7 ��mol/liver, respectively, NS) as well as cholesterol ester content (8.5 �� 1.0 versus 6.6 �� 0.9 ��mol/liver, respectively, NS) were higher in AdhEL injected mice, however, not significantly. Hepatic triglyceride content as well as phospholipid content remained unchanged.
In mice receiving AdSR-BI together with AdhEL or AdNull, hepatic total cholesterol content was already higher due to hepatic SR-BI overexpression (P < 0.001 compared with wild-type AdNull injected mice) and increased even more in response to EL overexpression (24.4 �� 1.8 versus 17.2 �� 2.1 ��mol/liver, respectively; P < 0.05) (Fig. 2C). Hepatic free cholesterol content was significantly increased in mice receiving AdEL (13.2 �� 2.1 versus 7.8 �� 1.6 ��mol/liver, respectively; P < 0.05), while cholesterol ester content (11.2 �� 2.9 versus 9.4 �� 2.3 ��mol/liver, respectively, NS) increased, however, not significantly. Hepatic triglyceride content as well as phospholipid content remained unchanged.
EL modification of the HDL particle results in increased selective uptake via SR-BI and in increased holoparticle uptake independent of SR-BI in primary mouse hepatocytes in vitro To further investigate the underlying mechanism of increased hepatic cholesterol content in response to EL overexpression in all experimental models used, a series of in vitro studies using primary mouse hepatocytes was performed. In wild-type hepatocytes infected with AdNull (i.e., expressing endogenous SR-BI), EL modification of the HDL particle resulted in increased uptake of 125I-TC-labeled HDL apolipoproteins (15.6 �� 0.6 versus 31.9 �� 3.5%; P < 0.01) and, to an even further extent, increased uptake of 3H-cholesteryl linoleyl ether, suggesting increased uptake of cholesteryl esters (28.5 �� 2.8 versus 58.6 �� 6.2%; P < 0.01) leading to an increased net selective uptake (12.9 �� 2.
3 versus 26.6 �� 3.2%; P < 0.05) (Figs. 3A, 3B). SR-BI overexpression in wild-type hepatocytes increased 3H-cholesteryl linoleyl ether uptake from control HDL (52.7 �� 3.4%; P < 0.01) without affecting 125I-TC-labeled apolipoprotein uptake (19.9 �� 0.8%, NS), resulting in increased selective uptake (32.8 �� 2.7%; Cilengitide P < 0.05) (Figs. 3A, 3B). In addition, SR-BI overexpression in hepatocytes did not affect 125I-TC-labeled HDL protein uptake from EL-modified HDL (31.8 �� 3.