Objective-Enhanced endothelial permeability leading to intimal accumulation of low-density lipoproteins (LDL) stimulates the formation of atherosclerotic lesions. Histamine is known to increase vascular permeability. Whether this affects the formation of atherosclerotic lesions, however, remains elusive. Methods and Results-Apolipoprotein E-null (ApoE Ϫ/Ϫ ) mice treated with a histamine H1 receptor but not an H2 receptor antagonist developed 40% fewer atherosclerotic lesions in the aorta than placebo-treated controls. Similarly, genetic deletion of the H1 but not the H2 receptor resulted in a 60% reduction of lesions compared with ApoE Ϫ/Ϫ controls. The H1 receptor enhanced LDL permeability and lipid accumulation in the aorta, whereas plasma lipoprotein levels remained unaltered. In contrast, the H1 receptor did not affect proliferation and migration of vascular smooth muscle cells. Bone marrow transplantation confirmed that the formation of atherosclerotic lesions depended on the H1 receptor in vascular cells, whereas its presence in bone marrow-derived cells was irrelevant for plaque development. Mice expressing the H1 receptor exhibited higher levels of the chemokine (C-C motif) ligand 5 and higher numbers of macrophages and T-helper lymphocytes in plaques, higher numbers of circulating lymphocytes, and larger spleens.

AimsSirtuin 3 (Sirt3) is a mitochondrial, nicotinamide adenine dinucleotide (NAD+)-dependent deacetylase that reduces oxidative stress by activation of superoxide dismutase 2 (SOD2). Oxidative stress enhances arterial thrombosis. This study investigated the effects of genetic Sirt3 deletion on arterial thrombosis in mice in an inflammatory setting and assessed the clinical relevance of these findings in patients with ST-elevation myocardial infarction (STEMI).Methods and resultsUsing a laser-induced carotid thrombosis model with lipopolysaccharide (LPS) challenge, in vivo time to thrombotic occlusion in Sirt3−/− mice (n = 6) was reduced by half compared to Sirt3+/+ wild-type (n = 8, P < 0.01) controls. Ex vivo analyses of whole blood using rotational thromboelastometry revealed accelerated clot formation and increased clot stability in Sirt3−/− compared to wild-type blood. rotational thromboelastometry of cell-depleted plasma showed accelerated clotting initiation in Sirt3−/− mice, whereas overall clot formation and firmness remained unaffected. Ex vivo LPS-induced neutrophil extracellular trap formation was increased in Sirt3−/− bone marrow-derived neutrophils. Plasma tissue factor (TF) levels and activity were elevated in Sirt3−/− mice, whereas plasma levels of other coagulation factors and TF expression in arterial walls remained unchanged. SOD2 expression in bone marrow -derived Sirt3−/− neutrophils was reduced. In STEMI patients, transcriptional levels of Sirt3 and its target SOD2 were lower in CD14+ leukocytes compared with healthy donors (n = 10 each, P < 0.01).Conclusions Sirt3 loss-of-function enhances experimental thrombosis in vivo via an increase of neutrophil extracellular traps and elevation of TF suggesting thrombo-protective effects of endogenous Sirt3. Acute coronary thrombosis in STEMI patients is associated with lower expression levels of SIRT3 and SOD2 in CD14+ leukocytes. Therefore, enhancing SIRT3 activity by pan-sirtuin activating NAD+-boosters may provide a novel therapeutic target to prevent or treat thrombotic arterial occlusion in myocardial infarction or stroke.

Atherosclerosis is an inflammatory disease characterized by accumulation of leukocytes in the arterial intima. Members of the selectin family of adhesion molecules are important mediators of leukocyte extravasation. However, it is unclear whether L-selectin (L-sel) is involved in the pathogenesis of atherosclerosis. In the present study, mice deficient in L-selectin (L-sel −/−) animals were crossed with mice lacking Apolipoprotein E (ApoE −/−). The development of atherosclerosis was analyzed in double-knockout ApoE/L-sel (ApoE −/− L-sel −/−) mice and the corresponding ApoE −/− controls fed either a normal or a high cholesterol diet (HCD). After 6 weeks of HCD, aortic lesions were increased two-fold in ApoE −/− L-sel −/− mice as compared to ApoE −/− controls (2.46%±0.54% vs 1.28%±0.24% of total aortic area; p<0.05). Formation of atherosclerotic lesions was also enhanced in 6-month-old ApoE −/− L-sel −/− animals fed a normal diet (10.45%±2.58% vs 1.87%±0.37%; p<0.05). In contrast, after 12 weeks of HCD, there was no difference in atheroma formation between ApoE −/− L-sel −/− and ApoE −/− mice. Serum cholesterol levels remained unchanged by L-sel deletion. Atherosclerotic plaques did not exhibit any differences in cellular composition assessed by immunohistochemistry for CD68, CD3, CD4, and CD8 in ApoE −/− L-sel −/− as compared to ApoE −/− mice. Leukocyte rolling on lesions in the aorta was similar in ApoE −/− L-sel −/− and ApoE −/− animals. ApoE −/− L-sel −/− mice exhibited reduced size and cellularity of peripheral lymph nodes, increased size of spleen, and increased number of peripheral lymphocytes as compared to ApoE −/− controls. These data indicate that L-sel does not promote atherosclerotic lesion formation and suggest that it rather protects from early atherosclerosis.

Background: The HIV drugs lopinavir and ritonavir have recently been reported to cause transient adrenal insufficiency in preterm newborns. We, therefore, considered HIV drugs as a cause of transiently elevated 17-hydroxyprogesterone (17OHP) levels in a neonatal screening test for congenital adrenal hyperplasia in a preterm girl exposed to zidovudine, efavirenz, tenofovir, and emtricitabine. Objective: So far, HIV drugs have not been tested for their effect on steroidogenesis and the steroidogenic enzyme activity of CYP21A2 specifically in an in vitro system. Methods: We tested the effect of efavirenz, tenofovir, emtricitabine, and zidovudine on steroidogenesis of human adrenal H295R cells. Cells were treated with the drugs at different concentrations including concentrations in therapeutic use. The effect on CYP21A2 activity was assessed by testing the conversion of radiolabeled 17OHP to 11-deoxycortisol. Cell viability was tested by an MTT assay. In addition, recombinant human CYP21A2 protein was used to assess direct drug effects on CYP21A2 activity. Results: We observed significantly decreased CYP21A2 activity in both in vitro testing systems after treatment with efavirenz at therapeutic concentrations. Moreover, efavirenz affected cell viability. By contrast, the other test drugs did not affect steroidogenesis. Follow-up of our patient revealed elevated 17OHP and androgen levels during the first weeks of life, but values normalized spontaneously. Genetic testing for CYP21A2 mutations was negative. Thus, it remains unsettled whether the transient 17OHP elevation in this baby was due to a drug effect. Conclusion: The HIV drug efavirenz inhibits CYP21A2 activity in vitro through direct interaction with enzyme catalysis at therapeutic concentrations. This may have clinical implications for HIV treatment in children and adults. However, so far, clinical data are scarce, and further studies are needed to be able to draw clinical conclusions.