Ceramides are sphingolipids that modulate a variety of cellular processes via 2 major mechanisms: functioning as second messengers and regulating membrane biophysical properties, particularly lipid rafts, important signaling platforms. Altered sphingolipid levels have been implicated in many cardiovascular diseases, including hypertension, atherosclerosis, and diabetes mellitus–related conditions; however, molecular mechanisms by which ceramides impact endothelial functions remain poorly understood. In this regard, we generated mice defective of endothelial sphingolipid de novo biosynthesis by deleting the Sptlc2 (long chain subunit 2 of serine palmitoyltransferase)—the first enzyme of the pathway. Our study demonstrated that endothelial sphingolipid de novo production is necessary to regulate (1) signal transduction in response to NO agonists and, mainly via ceramides, (2) resting eNOS (endothelial NO synthase) phosphorylation, and (3) blood pressure homeostasis. Specifically, our findings suggest a prevailing role of C16:0-Cer in preserving vasodilation induced by tyrosine kinase and GPCRs (G-protein coupled receptors), except for Gq-coupled receptors, while C24:0- and C24:1-Cer control flow-induced vasodilation. Replenishing C16:0-Cer in vitro and in vivo reinstates endothelial cell signaling and vascular tone regulation. This study reveals an important role of locally produced ceramides, particularly C16:0-, C24:0-, and C24:1-Cer in vascular and blood pressure homeostasis, and establishes the endothelium as a key source of plasma ceramides. Clinically, specific plasma ceramides ratios are independent predictors of major cardiovascular events. Our data also suggest that plasma ceramides might be indicative of the diseased state of the endothelium.

BackgroundMany asthmatic women complain of symptom exacerbations in particular periods, i.e. during pregnancy and menstrual cycles (perimenstrual asthma: PMA)". The goal of this study was to study the effect of the luteal and follicular phases of the menstrual cycle on bronchial reactivity (BR) in a group of asthmatic women.MethodsFor this purpose, 36 pre-menopausal women were enrolled and underwent testing for resting pulmonary function, measurement of the diffusing capacity of the lung for carbon monoxide (DLCO), and airway responsiveness to methacholine in the follicular and luteal phases of their menstrual cycles. We also measured plasma hormone levels and levels of cyclic adenosine monophosphate (cAMP; a mediator of bronchial smooth muscle contraction) and testosterone in induced sputum samples.ResultsOur study showed that about 30% of the asthmatic women had decreased PC20FEV1.0 in the follicular phase of menstrual cycle with a significant correlation between PC20FEV1.0 and serum testosterone levels. Moreover, marked increases in sputum testosterone levels (mean = 2.6-fold increase) together with significant increases in sputum cAMP concentrations (mean = 3.6-fold increases) were observed during the luteal phase of asthmatic patients, suggesting that testosterone contributes to the pathophysiology of PMA. We excluded the possibility that testosterone directly inhibits phosphodiesterase (PDE) activity as incubating PDE with testosterone in vitro did not reduce PDE catalytic activity.ConclusionsIn conclusion, our data show that PC20FEV1.0 was decreased in the follicular phase of the menstrual cycle in about 30% of women and was associated with lower cAMP levels in sputum samples, which may contribute to bronchoconstriction. Our results also suggest a link between PMA and testosterone levels. However, whether these findings are of clinical significance in terms of the management of asthma or asthma worsening during the menstrual cycle needs further investigation.

Ultrastructural and light microscopic cytochemical methods were used to study the distribution and changes in distribution of three phosphatase enzymes: 5'-nucleotidase (5N); thiamine pyrophosphatase (TPP); and adenosine triphosphatase (ATP) in the rat endometrium during early pregnancy up to the time of blastocyst attachment. The authors were particularly interested in changes in the apical plasma membrane and reaction product for all three enzymes was clearly localized along this membrane especially on day 1 of pregnancy. However, the three enzymes showed markedly different patterns of organization of reaction product at later times during early pregnancy. 5N, while showing a continuous lining along the microvilli on day 1 was virtually undetectable by day 6. TPP was also strongly present apically on day 1, but reaction product was not always found as a continuous lining. Again, by day 6, there was no presence of this enzyme along the apical surface. ATP differed from the other two in that it produced a strong, and relatively unchanged reaction product along the apical plasma membrane from day 1 through to day 6 of pregnancy. The changes in distribution of these enzymes was particularly obvious at the electron microscopic level and we consider their contribution to the process of 'plasma membrane transformation' of early pregnancy.

Ultrastructural and light microscopic catalytic histochemical methods were used to study the distribution and changes in distribution of four phosphatase enzymes; alkaline phosphatase, 5'-nucleotidase, thiamine pyrophosphatase and adenosine triphosphatase in uterine epithelial cells in response to the ovarian hormones, oestrogen, progesterone or a combination of both used in different regimes on ovariectomised rats. Reaction product for all four enzymes was clearly localised in the epithelial cells, especially with oestrogen priming. However, the four enzymes showed markedly different patterns of organisation of reaction product in response to other hormonal treatments. Our findings clearly show that the expression of these enzymes is under ovarian hormonal control. However, while all of the enzymes are upregulated by oestrogen, the response to progesterone is variable, which can upregulate or downregulate different enzymes. The findings are particularly obvious at the electron microscopic level on the apical plasma membrane of the uterine epithelial cells, which was the main focus of our study.

Ultrastructural and light microscopic cytochemical methods were used to study the distribution and changes in distribution of alkaline phosphatase in the apical plasma membrane of rat uterine epithelial cells during different stages of early pregnancy up to the time of attachment of the blastocyst. Reaction product generated by alkaline phosphatase (AP) was located along the apical plasma membrane at each stage investigated. However, a very different organization of reaction product was observed depending on the time during early pregnancy with a continuous pattern appearing all along the microvilli on day 1. This pattern was subsequently converted into a clumped and highly 'patchy' appearance around the time of blastocyst attachment by day 6 of pregnancy. This change in pattern and distribution was only seen on the luminal epithelial cells with glandular epithelial cells and blood vessels displaying an unchanging distribution.