Conditional ablation of macrophages disrupts ovarian vasculature

Macrophages are the most abundant immune cell within the ovary. Their dynamic distribution throughout the ovarian cycle and heterogenic array of functions suggest the involvement in various ovarian processes, but their functional role has yet to be fully established. The aim was to induce conditional macrophage ablation to elucidate the putative role of macrophages in maintaining the integrity of ovarian vasculature. Using the CD11b-diphtheria toxin receptor (DTR) mouse, in which expression of human DTR is under the control of the macrophage-specific promoter sequence CD11b, ovarian macrophages were specifically ablated in adult females by injections of diphtheria toxin (DT). CD11b-DTR mice were given DT treatment or vehicle and ovaries collected at 2, 8, 16, 24 and 48 h. Histochemical stains were employed to characterise morphological changes, immunohistochemistry for F4/80 to identify macrophages and the endothelial cell marker CD31 used to quantify vascular changes. In normal ovaries, macrophages were detected in corpora lutea and in the theca layer of healthy and atretic follicles. As macrophage ablation progressed, increasing amounts of ovarian haemorrhage were observed affecting both luteal and thecal tissue associated with significant endothelial cell depletion, increased erythrocyte accumulation and increased follicular atresia by 16 h. These events were followed by necrosis and profound structural damage. Changes were limited to the ovary, as DT treatment does not disrupt the vasculature of other tissues likely reflecting the unique cyclical nature of the ovarian vasculature and heterogeneity between macrophages within different tissues. These results show that macrophages play a critical role in maintaining ovarian vascular integrity

11β-Hydroxysteroid Dehydrogenase type 1 is expressed in neutrophils and restrains an inflammatory response in male mice

Endogenous glucocorticoid action within cells is enhanced by prereceptor metabolism by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts intrinsically inert cortisone and 11-dehydrocorticosterone into active cortisol and corticosterone, respectively. 11β-HSD1 is highly expressed in immune cells elicited to the mouse peritoneum during thioglycollate-induced peritonitis and is down-regulated as the inflammation resolves. During inflammation, 11β-HSD1-deficient mice show enhanced recruitment of inflammatory cells and delayed acquisition of macrophage phagocytic capacity. However, the key cells in which 11β-HSD1 exerts these effects remain unknown. Here we have identified neutrophils (CD11b(+),Ly6G(+),7/4(+) cells) as the thioglycollate-recruited cells that most highly express 11β-HSD1 and show dynamic regulation of 11β-HSD1 in these cells during an inflammatory response. Flow cytometry showed high expression of 11β-HSD1 in peritoneal neutrophils early during inflammation, declining at later states. In contrast, expression in blood neutrophils continued to increase during inflammation. Ablation of monocytes/macrophages by treatment of CD11b-diphtheria-toxin receptor transgenic mice with diphtheria toxin prior to thioglycollate injection had no significant effect on 11β-HSD1 activity in peritoneal cells, consistent with neutrophils being the predominant 11β-HSD1 expressing cell type at this time. Similar to genetic deficiency in 11β-HSD1, acute inhibition of 11β-HSD1 activity during thioglycollate-induced peritonitis augmented inflammatory cell recruitment to the peritoneum. These data suggest that neutrophil 11β-HSD1 increases during inflammation to contribute to the restraining effect of glucocorticoids upon neutrophil-mediated inflammation. In human neutrophils, lipopolysaccharide activation increased 11β-HSD1 expression, suggesting the antiinflammatory effects of 11β-HSD1 in neutrophils may be conserved in humans

Modulation of 11β-hydroxysteroid dehydrogenase as a strategy to reduce vascular inflammation

Atherosclerosis is a chronic inflammatory disease in which initial vascular damage leads to extensive macrophage and lymphocyte infiltration. Although acutely glucocorticoids suppress inflammation, chronic glucocorticoid excess worsens atherosclerosis, possibly by exacerbating systemic cardiovascular risk factors. However, glucocorticoid action within the lesion may reduce neointimal proliferation and inflammation. Glucocorticoid levels within cells do not necessarily reflect circulating levels due to pre-receptor metabolism by 11β-hydroxysteroid dehydrogenases (11β-HSDs). 11β-HSD2 converts active glucocorticoids into inert 11-keto forms. 11β-HSD1 catalyses the reverse reaction, regenerating active glucocorticoids. 11β-HSD2-deficiency/ inhibition causes hypertension, whereas deficiency/ inhibition of 11β-HSD1 generates a cardioprotective lipid profile and improves glycemic control. Importantly, 11β-HSD1-deficiency/ inhibition is atheroprotective, whereas 11β-HSD2-deficiency accelerates atherosclerosis. These effects are largely independent of systemic risk factors, reflecting modulation of glucocorticoid action and inflammation within the vasculature. Here, we consider whether evidence linking the 11β-HSDs to vascular inflammation suggests these isozymes are potential therapeutic targets in vascular injury and atherosclerosis

11β-hydroxysteroid dehydrogenase type 1 deficiency in bone marrow-derived cells reduces atherosclerosis

11β-Hydroxysteroid dehydrogenase type-1 (11β-HSD1) converts inert cortisone into active cortisol, amplifying intracellular glucocorticoid action. 11β-HSD1 deficiency improves cardiovascular risk factors in obesity but exacerbates acute inflammation. To determine the effects of 11β-HSD1 deficiency on atherosclerosis and its inflammation, atherosclerosis-prone apolipoprotein E-knockout (ApoE-KO) mice were treated with a selective 11β-HSD1 inhibitor or crossed with 11β-HSD1-KO mice to generate double knockouts (DKOs) and challenged with an atherogenic Western diet. 11β-HSD1 inhibition or deficiency attenuated atherosclerosis (74–76%) without deleterious effects on plaque structure. This occurred without affecting plasma lipids or glucose, suggesting independence from classical metabolic risk factors. KO plaques were not more inflamed and indeed had 36% less T-cell infiltration, associated with 38% reduced circulating monocyte chemoattractant protein-1 (MCP-1) and 36% lower lesional vascular cell adhesion molecule-1 (VCAM-1). Bone marrow (BM) cells are key to the atheroprotection, since transplantation of DKO BM to irradiated ApoE-KO mice reduced atherosclerosis by 51%. 11β-HSD1-null macrophages show 76% enhanced cholesterol ester export. Thus, 11β-HSD1 deficiency reduces atherosclerosis without exaggerated lesional inflammation independent of metabolic risk factors. Selective 11β-HSD1 inhibitors promise novel antiatherosclerosis effects over and above their benefits for metabolic risk factors via effects on BM cells, plausibly macrophages.—Kipari, T., Hadoke, P. W. F., Iqbal, J., Man, T. Y., Miller, E., Coutinho, A. E., Zhang, Z., Sullivan, K. M., Mitic, T., Livingstone, D. E. W., Schrecker, C., Samuel, K., White, C. I., Bouhlel, M. A., Chinetti-Gbaguidi, G., Staels, B., Andrew, R., Walker, B. R., Savill, J. S., Chapman, K. E., Seckl, J. R. 11β-hydroxysteroid dehydrogenase type 1 deficiency in bone marrow-derived cells reduces atherosclerosis

Pregnancy in Obese Mice Protects Selectively against Visceral Adiposity and Is Associated with Increased Adipocyte Estrogen Signalling

Maternal obesity is linked with increased adverse pregnancy outcomes for both mother and child. The metabolic impact of excessive fat within the context of pregnancy is not fully understood. We used a mouse model of high fat (HF) feeding to induce maternal obesity to identify adipose tissue-mediated mechanisms driving metabolic dysfunction in pregnant and non-pregnant obese mice. As expected, chronic HF-feeding for 12 weeks preceding pregnancy increased peripheral (subcutaneous) and visceral (mesenteric) fat mass. However, unexpectedly at late gestation (E18.5) HF-fed mice exhibited a remarkable normalization of visceral but not peripheral adiposity, with a 53% reduction in non-pregnant visceral fat mass expressed as a proportion of body weight (P<0.001). In contrast, in control animals, pregnancy had no effect on visceral fat mass proportion. Obesity exaggerated glucose intolerance at mid-pregnancy (E14.5). However by E18.5, there were no differences, in glucose tolerance between obese and control mice. Transcriptomic analysis of visceral fat from HF-fed dams at E18.5 revealed reduced expression of genes involved in de novo lipogenesis (diacylglycerol O-acyltransferase 2--Dgat2) and inflammation (chemokine C-C motif ligand 20--Ccl2) and upregulation of estrogen receptor α (ERα) compared to HF non pregnant. Attenuation of adipose inflammation was functionally confirmed by a 45% reduction of CD11b+CD11c+ adipose tissue macrophages (expressed as a proportion of all stromal vascular fraction cells) in HF pregnant compared to HF non pregnant animals (P<0.001). An ERα selective agonist suppressed both de novo lipogenesis and expression of lipogenic genes in adipocytes in vitro. These data show that, in a HF model of maternal obesity, late gestation is associated with amelioration of visceral fat hypertrophy, inflammation and glucose intolerance, and suggest that these effects are mediated in part by elevated visceral adipocyte ERα signaling

Novel role for endogenous mitochondrial formylated peptide-driven formyl peptide receptor 1 signalling in acute respiratory distress syndrome.

BACKGROUND: Acute respiratory distress syndrome (ARDS) is an often fatal neutrophil-dominant lung disease. Although influenced by multiple proinflammatory mediators, identification of suitable therapeutic candidates remains elusive. We aimed to delineate the presence of mitochondrial formylated peptides in ARDS and characterise the functional importance of formyl peptide receptor 1 (FPR1) signalling in sterile lung inflammation. METHODS: Mitochondrial formylated peptides were identified in bronchoalveolar lavage fluid (BALF) and serum of patients with ARDS by liquid chromatography-tandem mass spectrometry. In vitro, human neutrophils were stimulated with mitochondrial formylated peptides and their effects assessed by flow cytometry and chemotaxis assay. Mouse lung injury was induced by mitochondrial formylated peptides or hydrochloric acid. Bone marrow chimeras determined the contribution of myeloid and parenchymal FPR1 to sterile lung inflammation. RESULTS: Mitochondrial formylated peptides were elevated in BALF and serum from patients with ARDS. These peptides drove neutrophil activation and chemotaxis through FPR1-dependent mechanisms in vitro and in vivo. In mouse lung injury, inflammation was attenuated in Fpr1-/- mice, effects recapitulated by a pharmacological FPR1 antagonist even when administered after the onset of injury. FPR1 expression was present in alveolar epithelium and chimeric mice demonstrated that both myeloid and parenchymal FPR1 contributed to lung inflammation. CONCLUSIONS: We provide the first definitive evidence of mitochondrial formylated peptides in human disease and demonstrate them to be elevated in ARDS and important in a mouse model of lung injury. This work reveals mitochondrial formylated peptide FPR1 signalling as a key driver of sterile acute lung injury and a potential therapeutic target in ARDS