We employed homologous recombination in embryonic stem cells to produce mice lacking functional LDL receptor genes. Homozygous male and female mice lacking LDL receptors (LDLR-'-mice) were viable and fertile. Total plasma cholesterol levels were twofold higher than those of wild-type littermates, owing to a seven-to ninefold increase in intermediate density lipoproteins (IDL) and LDL without a significant change in HDL. Plasma triglyceride levels were normal. The half-lives for intravenously administered 1251-VLDL and 1251_ LDL were prolonged by 30-fold and 2.5-fold, respectively, but the clearance of '251-HDL was normal in the LDLR-'-mice. Unlike wild-type mice, LDLR-1-mice responded to moderate amounts of dietary cholesterol (0.2% cholesterol/ 10% coconut oil) with a major increase in the cholesterol content of IDL and LDL particles. The elevated IDL/ LDL level ofLDLR-I-mice was reduced to normal 4 d after the intravenous injection of a recombinant replication-defective adenovirus encoding the human LDL receptor driven by the cytomegalovirus promoter. The virus restored expression of LDL receptor protein in the liver and increased the clearance of 125I-VLDL. We conclude that the LDL receptor is responsible in part for the low levels of VLDL, IDL, and LDL in wild-type mice and that adenovirusencoded LDL receptors can acutely reverse the hypercholesterolemic effects of LDL receptor deficiency. (J. Clin. Invest. 1993. 92:883-893.)

Macrophage type-I and type-II class-A scavenger receptors (MSR-A) are implicated in the pathological deposition of cholesterol during atherogenesis as a result of receptor-mediated uptake of modified low-density lipoproteins (mLDL). MSR-A can bind an extraordinarily wide range of ligands, including bacterial pathogens, and also mediates cation-independent macrophage adhesion in vitro. Here we show that targeted disruption of the MSR-A gene in mice results in a reduction in the size of atherosclerotic lesions in an animal deficient in apolipoprotein E. Macrophages from MSR-A-deficient mice show a marked decrease in mLDL uptake in vitro, whereas mLDL clearance from plasma occurs at a normal rate, indicating that there may be alternative mechanisms for removing mLDL from the circulation. In addition, MSR-A-knockout mice show an increased susceptibility to infection with Listeria monocytogenes or herpes simplex virus type-1, indicating that MSR-A may play a part in host defence against pathogens.

Cholesterol in the mammalian brain is a risk factor for certain neurodegenerative diseases, raising the question of its normal function. In the mature brain, the highest cholesterol content is found in myelin. We therefore created mice that lack the ability to synthesize cholesterol in myelin-forming oligodendrocytes. Mutant oligodendrocytes survived, but CNS myelination was severely perturbed, and mutant mice showed ataxia and tremor. CNS myelination continued at a reduced rate for many months, and during this period, the cholesterol-deficient oligodendrocytes actively enriched cholesterol and assembled myelin with >70% of the cholesterol content of wild-type myelin. This shows that cholesterol is an indispensable component of myelin membranes and that cholesterol availability in oligodendrocytes is a rate-limiting factor for brain maturation.

Hormone-sensitive lipase (HSL) is known to mediate the hydrolysis not only of triacylglycerol stored in adipose tissue but also of cholesterol esters in the adrenals, ovaries, testes, and macrophages. To elucidate its precise role in the development of obesity and steroidogenesis, we generated HSL knockout mice by homologous recombination in embryonic stem cells. Mice homozygous for the mutant HSL allele (HSL؊͞؊) were superficially normal except that the males were sterile because of oligospermia. HSL؊͞؊ mice did not have hypogonadism or adrenal insufficiency. Instead, the testes completely lacked neutral cholesterol ester hydrolase (NCEH) activities and contained increased amounts of cholesterol ester. Many epithelial cells in the seminiferous tubules were vacuolated. NCEH activities were completely absent from both brown adipose tissue (BAT) and white adipose tissue (WAT) in HSL؊͞؊ mice. Consistently, adipocytes were significantly enlarged in the BAT (5-fold) and, to a lesser extent in the WAT (2-fold), supporting the concept that the hydrolysis of triacylglycerol was, at least in part, impaired in HSL؊͞؊ mice. The BAT mass was increased by 1.65-fold, but the WAT mass remained unchanged. Discrepancy of the size differences between cell and tissue suggests the heterogeneity of adipocytes. Despite these morphological changes, HSL؊͞؊ mice were neither obese nor cold sensitive. Furthermore, WAT from HSL؊͞؊ mice retained 40% of triacylglycerol lipase activities compared with the wild-type WAT. In conclusion, HSL is required for spermatogenesis but is not the only enzyme that mediates the hydrolysis of triacylglycerol stored in adipocytes.