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The steroid 11beta-hydroxylase inhibitor metyrapone is able to effectively reverse the hypercortisolemia detected in human Cushing's Syndrome patients. In this current preclinical study, we investigated whether metyrapone monotherapy can also reverse the hypercortisolemia-associated increase in atherosclerotic cardiovascular disease risk. In this instance, female low-density lipoprotein receptor knockout mice fed a cholic acid-containing high cholesterol/high fat diet to induce the development of hypercorticosteronemia and atherosclerotic lesions were treated twice daily with 100 mg/kg metyrapone for 4 weeks. Metyrapone effectively protected against hypercorticosteronemia development with endpoint plasma corticosterone levels remaining 43% lower than in controls (p < 0.01). Gene expression analysis in livers and adrenals validated that glucocorticoid receptor signaling was also reduced. Importantly, metyrapone treatment did not impact plasma cholesterol levels or alter atherosclerotic plaque areas or lesional collagen contents. However, metyrapone induced significant systemic lymphocytopenia as evident from marked decreases in splenic white pulp contents and thymus weights (−48% and −41%, respectively; p < 0.001). In conclusion, we have shown that treatment with metyrapone diminishes hypercorticosteronemia without affecting atherosclerosis susceptibility in cholic acid-containing high cholesterol/high fat diet-fed low-density lipoprotein receptor knockout mice. These preclinical findings highlight that restoring plasma glucocorticoid levels to normal is not necessarily sufficient to overcome the cardiovascular co-morbidities associated with human Cushing's disease. [ABSTRACT FROM AUTHOR]

Chronic glucocorticoid overexposure predisposes to the development of atherosclerotic cardiovascular disease in humans. Cholestatic liver disease is associated with increased plasma glucocorticoid levels. Here, we determined - in a preclinical setting - whether the chronic presence of cholestatic liver disease also induces a concomitant negative impact on atherosclerosis susceptibility. Hereto, regular chow diet-fed atherosclerosissusceptible hypercholesterolemic apolipoprotein E (APOE)-knockout mice were treated with the bile duct toxicant alpha-naphthylisothiocyanate (ANIT) for 8 weeks. ANIT exposure induced the development of fibrotic cholestatic liver d isease as evident from collagen deposits and compensatory bile duct hyperproliferation within the liver and the rise in plasma levels of bilirubin (+60%; P < 0.01) and bile acids (10-fold higher; P < 0.01). Adrenal weights (+22%; P < 0.01) and plasma corticosterone levels (+72%; P < 0.01) were increased in ANIT-treated mice. In contrast, athe rosclerosis susceptibility was not increased in response to ANIT feeding, despite the concomitant increase in plasma free cholesterol (+30%; P < 0.01) and cholesteryl ester (+42%; P < 0.001) levels. The ANIT-induced hypercorticosteronemia coincided with marked immunosuppression as judged from the 50% reduction (P < 0.001) in circulating lymphocyte numbers. However, hepatic glucocorticoid signaling was not enhanced after ANIT treatment. It thus appears that the immunosuppressive effect of glucocorticoids is uncoupled from their metabolic effect under cholestatic disease conditions. In conclu sion, we have shown that cholestatic liver disease-associated endogenous glucocorticoid overexposure does not increase atherosclerosis susceptibility in APOE-knockout mice. Our studies provide novel preclinical evidence for the observations that the hypercholesterolemia seen in cholestatic human subjects does not translate into a higher risk for atherosclerotic cardiovascular disease. [ABSTRACT FROM AUTHOR]

Abstract Background and aims Although studies in mice have suggested that lesion regression is feasible, the underlying mechanisms remain largely unknown. Here we determined the impact of high-density lipoprotein (HDL) on atherosclerosis regression outcome. Methods Atherosclerotic lesion dynamics were studied upon bone marrow transplantation-mediated re-introduction of apolipoprotein E (Apoe) in Apoe knockout mice. Probucol was used to pharmacologically deplete HDL. Results Restoration of Apoe function was associated with an initial growth of atherosclerotic lesions and parallel decrease in lesional macrophage foam cell content (47 ± 4% at 4 weeks versus 72 ± 2% at baseline: p < 0.001), despite the fact that cholesterol levels were markedly reduced. Notably, significant lesion regression was detected from 4 weeks onwards, when plasma cholesterol levels had returned to the normolipidemic range. As a result, lesions were 41% smaller (p < 0.05) at 8 weeks than at 4 weeks after bone marrow transplantation. Regressed lesions contained an even lower level of macrophage foam cells (33 ± 5%: p < 0.001) and were rich in collagen. Probucol co-treatment was associated with a 3.2-fold lower (p < 0.05) plasma HDL-cholesterol level and a more pro-inflammatory (CCR2+) monocyte phenotype. Importantly, probucol-treated mice exhibited atherosclerotic lesions that were larger than those of regular chow diet-fed bone marrow transplanted mice at 8 weeks (186 ± 15*103 μm2 for probucol-treated versus 120 ± 19*103 μm2 for controls: p < 0.05). Conclusions We have shown that probucol-induced HDL deficiency impairs the ability of established lesions to regress in response to reversal of the genetic hypercholesterolemia in Apoe knockout mice. Our studies thus highlight a crucial role for HDL in the process of atherosclerosis regression. Highlights • Reversal of hypercholesterolemia reduces lesional foam cell content in Apoe KO mice. • Normalization of cholesterol levels induces lesion regression in Apoe KO mice. • Probucol treatment is associated with a more pro-inflammatory monocyte phenotype. • Probucol-induced HDL lowering impairs the ability of established lesions to regress. [ABSTRACT FROM AUTHOR]

The pituitary-derived hormone prolactin has been suggested to stimulate the development of atherosclerosis and cardiovascular disease through its effects on metabolism and inflammation. In this study, we aimed to challenge the hypothesis that inhibition of prolactin function may beneficially affect atherosclerosis burden. Hereto, atherosclerosissusceptible LDL receptor (Ldlr) knockout mice were transplanted with bone marrow from transgenic mice expressing the pure prolactin receptor antagonist Del1-9-G129R-hPRL or their non-transgenic littermates as control. Recipient mice expressing Del1-9-G129R-hPRL exhibited a decrease in plasma cholesterol levels (--29%; P<0.05) upon feeding a Westerntype diet (WTD), which could be attributed to a marked decrease (--47%; P<0.01) in the amount of cholesterol esters associated with pro-atherogenic lipoproteins VLDL/LDL. By contrast, Del1-9-G129R-hPRL-expressing mice did not display any change in the susceptibility for atherosclerosis after 12 weeks of WTD feeding. Both the absolute atherosclerotic lesion size (223±33X10³ μm² for Del1-9-G129R-hPRL vs 259±32X10³ μm² for controls) and the lesional macrophage and collagen contents were not different between the two groups of bone marrow recipients. Importantly, Del1-9-G129R-hPRL exposure increased levels of circulating neutrophils (+91%; P<0.05), lymphocytes (+55%; P<0.05), and monocytes (+43%; P<0.05), resulting in a 49% higher (P<0.01) total blood leukocyte count. In conclusion, we have shown that prolactin receptor signaling inhibition uncouples the plasma atherogenic index from atherosclerosis susceptibility in Ldlr knockout mice. Despite an associated decrease in VLDL/LDL cholesterol levels, application of the prolactin receptor antagonist Del1-9-G129R-hPRL does not alter the susceptibility for initial development of atherosclerotic lesions probably due to the parallel increase in circulating leukocyte concentrations. [ABSTRACT FROM AUTHOR]

Abstract: Long-term changes in the secretion of immunosuppressive adrenal-derived glucocorticoid hormones influence cardiovascular disease risk. Here we determined the consequences of changes in adrenal steroid metabolism for the development of atherosclerotic lesions in mice. Atherosclerosis-susceptible low-density-lipoprotein (LDL) receptor knockout mice were subjected to adrenalectomy (ADX) or a control (SHAM) operation and subsequently fed an atherogenic diet for 4 weeks. Atherogenic diet feeding raised plasma corticosterone levels in SHAM mice, but not adrenalectomized mice, resulting in an 83% lower (P <0.01) corticosterone level in adrenalectomized mice. Adrenalectomy was associated with a respectively 22% and 29% lower plasma level of cholesterol and triglycerides. In contrast, white blood cell counts were increased 2-fold (P <0.01) in adrenalectomized mice, which could be attributed to a significant 2.1- to 2.6-fold rise in lymphocyte (P <0.05) and monocyte (P <0.05) numbers. Probably as a result of the enhanced systemic inflammatory status, adrenalectomy was associated with a higher susceptibility for diet-induced atherosclerosis (321±18×103 μm2 for ADX vs 240±31×103 μm2 for SHAM; P <0.05) not withstanding the lowered cholesterol levels. Restoring adrenocortical steroid secretion – but not adrenal medulla function – and the associated downstream glucocorticoid receptor signaling in adrenalectomized mice through adrenal transplantation induced a reversal of the adrenalectomy-associated rise in white blood cell numbers, plasma monocyte chemoattractant protein 1 (MCP-1) levels, and atherosclerotic lesion development (lesion size in transplanted mice: 258±34×103 μm2; P <0.05 vs ADX). In conclusion, our studies show that adrenal-derived steroids protect against the development of initial atherosclerotic lesions in LDL receptor knockout mice. [Copyright &y& Elsevier]

Adrenal-derived glucocorticoids mediate the physiological response to stress. Chronic disturbances in glucocorticoid homeostasis, i.e. in Addison's and Cushing's disease patients, predispose to the development of atherosclerotic cardiovascular disease. Here we review preclinical and clinical findings regarding the relation between changes in plasma glucocorticoid levels and the atherosclerosis extent. It appears that, although the altered glucocorticoid function can in most cases be restored in the different patient groups, current therapies do not necessarily reverse the associated risk for atherosclerotic cardiovascular disease. In our opinion much attention should therefore be given to the development of a Cushing's disease mouse model that can (1) effectively replicate the effect of hypercortisolemia on atherosclerosis outcome observed in humans and (2) be used to investigate, in a preclinical setting, the relative impact on atherosclerosis susceptibility of already available (e.g. metyrapone) and potentially novel (i.e. SR-BI activity modulators) therapeutic agents that target the adrenal glucocorticoid output. • Glucocorticoids impact both metabolic and inflammatory processes. • Chronic changes in glucocorticoid status predispose to cardiovascular disease. • Glucocorticoids elicit variable effects on atherosclerosis in preclinical models. • Current Cushing's treatments do no effectively reverse the cardiovascular risk. • Adrenal cholesterol metabolism may serve as novel target in Cushing's syndrome. [ABSTRACT FROM AUTHOR]

Hyperlipidemic apolipoprotein E (APOE) knockout mice show an enhanced level of adrenal-derived anti-inflammatory glucocorticoids. Here we determined in APOE knockout mice the impact of total removal of adrenal function through adrenalectomy (ADX) on two inflammation-associated pathologies, endotoxemia and atherosclerosis. ADX mice exhibited 91% decreased corticosterone levels (P<0.001), leukocytosis (WBC count: 10.0 ± 0.4 x 10E9/L vs 6.5 ± 0.5 x 10E9/L; P<0.001) and an increased spleen weight (P<0.01). FACS analysis on blood leukocytes revealed increased B-lymphocyte numbers (55 ± 2% vs 46 ± 1%; P<0.01). T-cell populations in blood appeared to be more immature (CD62L+: 26 ± 2% vs 19 ± 1% for CD4+ T-cells, P<0.001 and 58 ± 7% vs 47 ± 4% for CD8+ T-cells, P<0.05), which coincided with immature CD4/CD8 double positive thymocyte enrichment. Exposure to lipopolysaccharide failed to increase corticosterone levels in ADX mice and was associated with a 3-fold higher (P<0.05) TNF-alpha response. In contrast, the development of initial fatty streak lesions and progression to advanced collagen-containing atherosclerotic lesions was unaffected. Plasma cholesterol levels were decreased by 35% (P<0.001) in ADX mice. This could be attributed to a decrease in pro-atherogenic very-low-density lipoproteins (VLDL) as a result of a diminished hepatic VLDL secretion rate (-24%; P<0.05). In conclusion, our studies show that adrenalectomy induces leukocytosis and enhances the susceptibility for endotoxemia in APOE knockout mice. The adrenalectomy-associated rise in white blood cells, however, does not alter atherosclerotic lesion development probably due to the parallel decrease in plasma levels of pro-atherogenic lipoproteins. [ABSTRACT FROM AUTHOR]

Abstract: While numerous studies have aimed to develop strategies to inhibit the development and progression of atherosclerosis, recent attention has focussed on the regression of pre-existing atherosclerotic plaques. As important regulator of total body cholesterol homeostasis, the liver X receptor (LXR) could possibly be an important target to induce regression. Here, we describe the effect of LXR activation by the synthetic agonist T0901317 on lesion regression in different mouse models with early fatty streak lesions or advanced collagen-rich lesions. Although T0901317 caused a dramatic increase in plasma (V)LDL levels in low-density lipoprotein (LDL) receptor knockout mice, no further increase in lesion size was observed, which points to beneficial LXR activity in the vascular wall. In normolipidemic C57BL/6 mice with cholate diet-induced atherosclerotic lesions, T0901317 treatment improved plasma lipoprotein levels and induced lesion regression (−43%, p <0.05). Apolipoprotein E (APOE) reconstitution in APOE knockout mice by means of bone marrow transplantation dramatically improved plasma lipoprotein profiles and resulted in a marked regression of initial (−45%, p <0.001) and advanced lesions (−23%, p <0.01). Atherosclerosis regression was associated with a decrease in the absolute macrophage content (−84%, p <0.001). T0901317 supplementation further decreased the size of early (−71%, p <0.001 vs baseline; −48%, p <0.01 vs chow diet alone) and more advanced atherosclerotic lesions (−36%, p <0.001 and −17%, p =0.06 respectively). In conclusion, our study highlights the potential of LXR agonist T0901317 to stimulate removal of macrophages from atherosclerotic lesions ultimately leading to a highly significant plaque regression of both early and advanced atherosclerotic lesions. [Copyright &y& Elsevier]

Scavenger receptor BI (SR-BI) has been suggested to modulate adipocyte function. To uncover the potential relevance of SR-BI for the development of obesity and associated metabolic complications, we compared the metabolic phenotype of wild-type and SR-BI deficient mice fed an obesogenic diet enriched in fat. Both male and female SR-BI knockout mice gained significantly more weight as compared to their wild-type counterparts in response to 12 weeks high fat diet feeding (1.5-fold; P < .01 for genotype). Plasma free cholesterol levels were ~2-fold higher (P < .001) in SR-BI knockout mice of both genders, whilst plasma cholesteryl ester and triglyceride concentrations were only significantly elevated in males. Strikingly, the exacerbated obesity in SR-BI knockout mice was paralleled by a better glucose handling. In contrast, only SR-BI knockout mice developed atherosclerotic lesions in the aortic root, with a higher predisposition in females. Biochemical and histological studies in male mice revealed that SR-BI deficiency was associated with a reduced hepatic steatosis degree as evident from the 29% lower (P < .05) liver triglyceride levels. Relative mRNA expression levels of the glucose uptake transporter GLUT4 were increased (+47%; P < .05), whilst expression levels of the metabolic PPARgamma target genes CD36, HSL, ADIPOQ and ATGL were reduced 39%-58% (P < .01) in the context of unchanged PPARgamma expression levels in SR-BI knockout gonadal white adipose tissue. In conclusion, we have shown that SR-BI deficiency is associated with a decrease in adipocyte PPARgamma activity and a concomitant uncoupling of obesity development from hepatic steatosis and glucose intolerance development in high fat diet-fed mice. [ABSTRACT FROM AUTHOR]