Karsten Grote

Mycoplasmas and their membranes are potent activators of macrophages, the active principle being lipoproteins and lipopeptides. Two stereoisomers of the mycoplasmal lipopeptide macrophage-activating lipopeptide-2 (MALP-2) differing in the configuration of the lipid moiety were synthesized and compared in their macrophage-activating potential, the R-MALP being >100 times more active than the S-MALP in stimulating the release of cytokines, chemokines, and NO. To assess the role of the Toll-like receptor (TLR) family in mycoplasmal lipopeptide signaling, the MALP-2-mediated responses were analyzed using macrophages from wild-type, TLR2-, TLR4-, and MyD88-deficient mice. TLR2- and MyD88-deficient cells showed severely impaired cytokine productions in response to R- and S-MALP. The MALP-induced activation of intracellular signaling molecules was fully dependent on both TLR2 and MyD88. There was a strong preference for the R-MALP in the recognition by its functional receptor, TLR2.

Mechanical stretch is a hallmark of arterial hypertension and leads to vessel wall remodeling, which involves matrix metalloproteinases (MMPs). Because mechanical stretch is further capable of inducing reactive oxygen species (ROS) formation via the NAD(P)H oxidase, we assessed whether mechanical stretch enhances MMP expression and activity in a NAD(P)H oxidase-dependent manner. Therefore, vascular smooth muscle cells (VSMCs) isolated from C57BL/6 mice were exposed to cyclic mechanical stretch. The impact of ROS was assessed using VSMCs isolated from p47phox-/- mice, deficient for a NAD(P)H oxidase subunit responsible for ROS formation. Transcript levels were investigated by cDNA array and confirmed by RT-PCR. ROS formation was determined by DCF fluoroscopy and MMP-2 activity by zymography. Mechanical stretch of wild-type VSMCs resulted in a rapid ROS formation and p47phox membrane translocation that is followed by an increase in Nox-1 transcripts. ROS formation was completely abrogated in p47phox-/- VSMCs. cDNA array further revealed an increase of MMP-2 mRNA in response to mechanical stretch, which was validated by RT-PCR. Using p47phox-/- VSMCs, this increase in MMP-2 mRNA was completely blunted. mRNA expression of tissue inhibitor of MMP-2 TIMP-1 and TIMP-2 and membrane-type 1 MMP was unaffected by mechanical stretch. Gelatinolytic activity of pro-MMP-2 has been increased rapidly in wild-type VSMCs and was completely abolished in p47phox-/- VSMCs. These results indicate that mechanical stretch induces ROS formation via the NAD(P)H oxidase and thereby enhances MMP-2 mRNA expression and pro-MMP-2 release. These results are consistent with the notion that in arterial hypertension, reactive oxygen species are involved in vascular remodeling via MMP activation. The full text of this article is available online at http://www.circresaha.org.

In utero exposure to a single low dose of 2,2 ,4,4 ,5-pentabromodiphenyl ether (PBDE-99) disrupts neurobehavioral development and causes permanent effects on the rat male reproductive system apparent in adulthood. PBDEs, a class of flame retardants, are widely used in every sector of modern life to prevent fire. They are persistent in the environment, and increasing levels of PBDEs have been found in biota and human breast milk. In the present study we assessed the effects of developmental exposure to one of the most persistent PBDE congeners (PBDE-99) on juvenile basal motor activity levels and adult male reproductive health. Wistar rat dams were treated by gavage on gestation day 6 with a single low dose of 60 or 300 microg PBDE-99/kg body weight (bw). In offspring, basal locomotor activity was evaluated on postnatal days 36 and 71, and reproductive performance was assessed in males at adulthood. The exposure to low-dose PBDE-99 during development caused hyperactivity in the offspring at both time points and permanently impaired spermatogenesis by the means of reduced sperm and spermatid counts. The doses used in this study (60 and 300 microg/kg bw) are relevant to human exposure levels, being approximately 6 and 29 times, respectively, higher than the highest level reported in human breast adipose tissue. This is the lowest dose of PBDE reported to date to have an in vivo toxic effect in rodents and supports the premise that low-dose studies should be encouraged for hazard identification of persistent environmental pollutants.

BACKGROUND: Vascular lipid accumulation and inflammation are hallmarks of atherosclerosis and perpetuate atherosclerotic plaque development. Mediators of inflammation, ie, interleukin (IL)-6, are elevated in patients with acute coronary syndromes and may contribute to the exacerbation of atherosclerosis. METHODS AND RESULTS: To assess the role of IL-6 in atherosclerosis, ApoE-/--IL-6-/- double-knockout mice were generated, fed a normal chow diet, and housed for 53+/-4 weeks. Mortality and blood pressure were unaltered. However, serum cholesterol levels and subsequent atherosclerotic lesion formation (oil red O stain) were significantly increased in ApoE-/--IL-6-/- mice compared with ApoE-/-, wild-type (WT), and IL-6-/- mice. Plaques of ApoE-/--IL-6-/- mice showed significantly reduced transcript and protein levels of matrix metalloproteinase-9, tissue inhibitor of metalloproteinase-1, collagen I and V, and lysyl oxidase (by reverse transcriptase-polymerase chain reaction and immunohistochemistry). Recruitment of macrophages and leukocytes (Mac3- and CD45-positive staining) into the atherosclerotic lesion was significantly reduced in ApoE-/--IL-6-/- mice. The transcript and serum protein (ELISA) levels of IL-10 were significantly reduced. CONCLUSIONS: Thus, a lifetime IL-6 deficiency enhances atherosclerotic plaque formation in ApoEK-/--IL-6-/- mice and leads to maladaptive vascular developmental processes. These observations are consistent with the notion that baseline levels of IL-6 are required to modulate lipid homeostasis, vascular remodeling, and plaque inflammation in atherosclerosis.

The importance of inflammation as a driver of pathology is no longer confined to autoimmune and infectious diseases. In line with convincing experimental data as well as abundant clinical findings the current view of atherosclerosis points to inflammation as a critical regulator of atherosclerotic plaque formation and progression leading to the fatal clinical endpoints myocardial infarction, stroke or sudden cardiac death. The underlying mechanisms have been a matter of intense research during the last decades. In this regard, the interleukin-6 (IL-6) cytokines and their signalling events have been shown to contribute to both, atherosclerotic plaque development and plaque destabilisation via a variety of mechanisms. These involve the release of other pro-inflammatory cytokines, oxidation of lipoproteins by phospholipases, stimulation of acute phase protein secretion, the release of prothrombotic mediators, and the activation of matrix metalloproteinases. Moreover, the formation of reactive oxygen species generated by vascular enzyme systems may play a critical role in the regulation of IL-6 indicating a cross talk between vasoactive substances i.e. angiotensin II or adrenalin and pro-inflammatory cytokines such as IL-6. In this review we will summarise and discuss the underlying molecular and cellular mechanisms how IL-6 as an early and central regulator of inflammation contributes to atherosclerosis and how this knowledge can be integrated into the clinical context.