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Chronic kidney disease (CKD) frequently leads to hyperphosphatemia and hyperparathyroidism, mineral bone disorder (CKD-MBD), ectopic calcifications and cardiovascular mortality. PTH activates the osteoanabolic Gαs/PKA and the Gαq/11/PKC pathways in osteoblasts, the specific impact of the latter in CKD-MBD is unknown. We generated osteoblast specific Gαq/11 knockout (KO) mice and established CKD-MBD by subtotal nephrectomy and dietary phosphate load. Bone morphology was assessed by micro-CT, osteoblast function by bone planar scintigraphy at week 10 and 22 and by histomorphometry. Osteoblasts isolated from Gαq/11 KO mice increased cAMP but not IP3 in response to PTH 1-34, demonstrating the specific KO of the PKC signaling pathway. Osteoblast specific Gαq/11 KO mice exhibited increased serum calcium and reduced bone cortical thickness and mineral density at 24 weeks. CKD Gαq/11 KO mice had similar bone morphology compared to WT, while CKD Gαq/11-KO on high phosphate diet developed decreased metaphyseal and diaphyseal cortical thickness and area, as well as a reduction in trabecular number. Gαq/11-KO increased bone scintigraphic tracer uptake at week 10 and mitigated tracer uptake in CKD mice at week 22. Histological bone parameters indicated similar trends. Gαq/11-KO in osteoblast modulates calcium homeostasis, bone formation rate, bone morphometry, and bone mineral density. In CKD and high dietary phosphate intake, osteoblast Gαq/11/PKC KO further aggravates mineral bone disease.

The present work compares intravenous (IV) and subcutaneous (SC) routes for contrast-enhanced MRI (CE-MRI) in mice. For that purpose, we selected two contrast media used in clinical practice. MRI acquisitions were performed at 1.5 T on five adult mice (Swiss, 41 g +/- 3 g). On each animal, four acquisitions were achieved with IV and SC administration of either Gd-DOTA or MS-325 (1 acquisition per week). For each route, 0.1 mL of NaCl and 0.1 mL of contrast agent were injected. For each acquisition, 200 T1-weighted images were acquired in a 2 h 34 min time lapse. For each route and contrast medium, dynamic contrast enhancement (DCE) curves were obtained. Time-to-peak (TTP), uptake, and washout constant-time values and contrast-to-noise ratio (CNR) were extracted. IV route TTP value was 4.9 min with Gd-DOTA and 5.4 min with MS-325. SC route TTP was 43.3 min with Gd-DOTA and 45.0 min with MS-325. Despite slower uptake constant-time, we show that SC is a potentially valuable alternative to the IV route in mouse preclinical CE-MRI.

Despite DOAs not being able to undergo laboratory monitoring regarding their pharmacokinetics, the delay in starting treatment after surgery, age, weight, liver function, renal function and concomitant treatments are all factors that may alter their safety profile. Dabigatran is to be avoided by patients presenting with renal impairment, and also when co-prescribed with P-gp inhibitors/inducers. Anti-Xas are the medicinal products with the most reliable safety profile for the elderly population, if interactions with medicines such as CYP3A4 inhibitors/inducers are controlled. The power of anti-Xas lies in its quick, direct, competitive and reversible inhibition plus the strong affinity of the Xa factor enzyme activity, whether the Xa factor is free or within the prothrombinase complex, preventing the generation of thrombin. Tests for modified coagulation include the prothrombin ratio, aPTT (less sensitive), and chromogenic tests. With dabigatran, this product directly inhibits free or bound thrombin, reversibly, rapidly and competitively. The thrombin and ecarin clotting times are the two coagulation tests that reflect the activity of dabigatran. Dabigatran also modifies the aPTT, but not in a concentration dependent manner. Currently, there are no commercially available antidotes to reverse the effects of anticoagulants, however, some are in the process of being developed.

International audience; We use high-resolution [Formula: see text] data in multiple experiments to estimate the sources of error during coregistration of images acquired on separate preclinical instruments. In combination with experiments with phantoms, we completed in vivo imaging on mice, aimed at identifying the possible sources of registration errors, caused either by transport of the animal, movement of the animal itself, or methods of coregistration. The same imaging cell was used as a holder for phantoms and animals. For all procedures, rigid coregistration was carried out using a common landmark coregistration system, placed inside the imaging cell. We used the fiducial registration error and the target registration error to analyze the coregistration accuracy. We found that moving an imaging cell between two preclinical devices during a multimodal procedure gives an error of about [Formula: see text] at most. Therefore, it could not be considered a source of coregistration errors. Errors linked to spontaneous movements of the animal increased with time, to nearly 1 mm at most, excepted for body parts that were properly restrained. This work highlights the importance of animal intrinsic movements during a multiacquisition procedure and demonstrates a simple method to identify and quantify the sources of error during coregistration.

The mitochondrial permeability transition (mPTP) is a key feature of cardiac cell death in ischemia-reperfusion injury (I/R). The mPTP blocker, cyclosporine A (CsA), has been shown to give protection against reperfusion-induced myocardial necrosis and troubles generated by acute coronary artery repermeabilization. Nevertheless, the results of the CIRCUS trial (Does Cyclosporine Improve Clinical Outcome in ST-Elevation Myocardial Infarction Patients) seem to go against this hypothesis. Pharmacological reasons linked to CsA pharmacokinetics and pharmacodynamics could be suggested. First, it could be explained by a limited diffusion of the drug in the area at risk, due to the only inclusion of patients with a TIMI 0 or 1 coronary blood flow in the anterior territory and the absence of collateral perfusion. Second, to explain a low tissue diffusion of the compound, blood cell capture and high metabolism could be suggested. Moreover, CsA is highly metabolized by cytochrome P450 3A4 (CYP3A4), a polymorphic enzyme leading to variations of Cmax and AUC between 10-20% in patients using CsA. Finally, CsA blocks calcineurin, a protein implied in I/R damage but calcineurin inhibition could contribute to protection towards I/R damage only when Rcan1, a calcineurin natural inhibitor, expression is low. The results of the CIRCUS trial are disappointing and could contribute to the withdrawal of the mPTP blockade pharmacological strategy as a way to protect the myocardium from I/R lesions. Nevertheless, many pharmacological insights could have contributed to an increased variability and, as a consequence, an important reduction of the pharmacological power of the study. This article is protected by copyright. All rights reserved.

International audience; Serotonergic dysfunction is mainly associated with neuropsychiatric and cardiovascular disorders but has also been linked with many other pathological conditions. Serotonin (5-hydroxytryptamine, 5-HT) mediates numerous physiological functions in the brain and the periphery by activating a variety of receptors. 5-HT receptors are divided into four classes, three of which belong to the G protein-coupled receptor family. This review provides an overview of the recent pharmacological developments involving the Gq-coupled 5-HT2 receptor subfamily as well as the pathological implications of this receptor subfamily with regard to fibrosis, the central nervous system, cardiovascular disorders, and cancer. The final section highlights new therapeutic opportunities and emerging research revealing unexplored medical opportunities for this class of 5-HT receptors. The development of biased 5-HT2 receptor ligands appears to be an interesting topic in various areas. In light of recent discoveries, the need for the development of new and safer drugs should take into account the risk of cardiovascular side effects such as pulmonary hypertension and heart valve disease.

National audience

Introduction The causal role of sympathetic nervous system in the development of metabolic disorders has not been clearly defined. The aim of the study was to determine if a chronic increase of the sympathetic nervous activity can induce and/or accelerate the development of glucose metabolism disorders. For that purpose, we used a transgenic mouse model, in which the gene encoding for the reuptake norepinephrine transporter (NET) has been deleted; these animals display increased urinary catecholamine levels. Methods Carbohydrate homeostasis (intraperitoneal glucose tolerance test, IPGTT, insulin tolerance test, ITT, insulin rates) was studied in heterozygous NET knockout mice (±) or wild-type mice (+/+) at 10, 20 and 25 weeks old. Mice were then fed either with normal chow or 30% w/v fructose in drinking water; glucose metabolism was assessed again after 15 weeks of such diet. Results Heterozygous mice displayed glucose intolerance already at 10 weeks old (area under the curve of the IPGTT (AUC; %) = 24859 vs 18016, p=0.004); similar results were obtained at 25 weeks old, i.e, just before starting the fructose diet: AUC = 24524 vs 20573, p=0.005 for ± and +/+ mice respectively. Moreover sensibility to insulin seemed to be diminished in ± mice compared to +/+ mice (insulin rate = 1.12 vs 0.86 μg/L and HOMAIR= 11.1 vs 8.4). Heterozygous animals were also much more sensitive to high fructose, since a 20% increase in the AUC of the IPGTT was obtained at the end of the high fructose diet, compared to the 2% increase in +/+ mice. This was associated with a decrease of insulin sensibility (HOMA-IR of ± mice = 11.1 vs 18.1, p=0.01, before and after fructose diet respectively). Conclusion Our data show that constitutive chronic sympathetic hyperactivity can induce the early development of carbohydrate metabolism disorders. Moreover, it seems to represent a major factor of susceptibility to diet-induced metabolic dysfunction. The author hereby declares no conflict of interest

Metabolic syndrome (MetS) is defined as a cluster of cardiovascular and metabolic disorders. Previous studies have demonstrated that ligands selective for I 1 imidazoline receptor (LNPs) ameliorate lipid metabolism (plasma total cholesterol and triglycerides levels) in rat models of metabolic syndrome. The objective of this study was to explore possible direct actions on hepatocytes which are involved in the regulation of lipid metabolism. Experiments were carried out in HepG2 cells, a cell line of hepatocytes. In order to evaluate the effect of LNP509 on lipid metabolism, the activation (i.e. phosphorylation) of a key actor of this signaling pathway, AMPK, was evaluated by measuring the ratio pAMPK/AMPK by Western Blot. AMPK is the main regulator of Acetyl-CoA-carboxylase (ACC) which is known to provide the malonyl-CoA substrate for the biosynthesis of fatty acids and to play a major role in hepatic steatosis. The effect of LNP509 on ACC phosphorylation (inactivation of ACC) was evaluated by measuring pACC/ACC by Western Blot. Hepatic steatosis, induced by treating HepG2 cells with oleic acid, was quantified by Oil Red O coloration and colorimetric measurements after oil Red O extraction. LNP509 (1 μM, 60 min) induced the M, phosphorylation of AMPK (pAMPK/ AMPK=1.15±0.27) compared to control (pAMPK/AMPK=0.37±0.06). Interestingly, treatment by LNP509 during 60 min also increased the phosphorylation (inhibition) of ACC (untreated cells: pACC/ACC=0.24±0.06; LNP 509 pACC/ACC=0.70±0.22). Concerning hepatic steatosis, LNP509 seemed to prevent lipid accumulation in HepG2 cell (cells alone=100%, cells+oleic acid=138.54%±10.34, cells+oleic acid+LNP509=112.81%±9.29). These data suggest that LNPs act directly on hepatic cells to activate AMPK pathway and prevent lipid accumulation. This mechanism probably accounts, at least partly, for the beneficial effects of LNPs on dyslipidemia observed in vivo. The author hereby declares no conflict of interest

Introduction Determination of adipose tissue proportion, as well as its spatial distribution, is a major part of metabolic syndrome assessment. Various methods, commonly used in mouse models, are either indirect measurement, invasive (destructive) or imaging techniques. Indirect measurement techniques, such as deuterium dilution (considered as gold standard) or bioimpedance are based on the measurement of body composition. They give access to the measure of total body fat, without distinction between visceral and subcutaneous fat volumes. Sacrifice and ex-vivo tissue weight allow both total fat measurement and partition between subcutaneous and visceral fat pad. It is a direct method that does not allow for longitudinal studies. Non-invasive micro-imaging modalities (magnetic resonance imaging and μCT) add, over others, the advantage of spatial localization of fat, in vivo. Aim Of our study was to compare results of measurements of total body fat done by μCT and bioimpedance in mice. Methods μCT was done using a dedicated in vivo imaging instrument (eXplore Vision 120, GE, USA) on 30 anesthetized mice. Each acquisition lasted approximately 12 min. Just after μCT, measurement of body composition was performed by bioimpedance (ImpediVET, ImpediMed, USA). Results The average value of total body fat by μCT is 5% higher than that obtained by bioimpedance. The standard deviation calculated from μCT measurements is lower than that obtained with bioimpedance (4.8% vs 6.8% respectively). The linear correlation of μCT and bioimpedance measure of total adipose tissue is moderate (r2 = 0.55). Conclusion It is the first time, to our knowledge, that a comparison of total fat volume measurement between μCT and bioimpedance is performed in mice. Surprisingly, the correlation appears not to be so trivial between the 2 methods. Before a deeper exploration, comparison of results obtained with these 2 methods should be made with caution.

Introduction If sympathetic hyperactivity seems to be associated to several components of the metabolic syndrome, its involvement in the development of metabolic syndrome has not been clearly defined. Aim It was to determine if a chronic increase of the sympathetic nervous activity is a sufficient condition for the development of metabolic disorders. For that purpose, we used a transgenic mouse model, in which the gene coding for the reuptake norepinephrine transporter (NET) has been deleted; these animals display increased plasma catecholamine rates, associated to slight increases in blood pressure and heart rate. We focused on the consequences of this sympathetic hyperactivity on glucose and lipid metabolism in this mouse model. Methods Heterozygous NET knockout mice (+/-) or wild-type mice (+/+) were fed either with a normal chow or a 30% w/v fructose in drinking water during a period of 15 weeks to induce metabolic disorders. Metabolic parameters (plasma triglycerides and total cholesterol, intraperitoneal glucose tolerance test, IPGTT) were measured after a 4h fasting period before treatment and then repeated every 5 weeks. Results No significant difference on triglycerides or cholesterol rates could be detected after the 15 weeks period. Heterozygous mice displayed clear glucose intolerance already before starting the fructose diet (area under the curve (%)=25434 vs 20822, p=0.013 for +/- and +/+ mice respectively). These animals were also much more sensitive to high fructose, since a 50% increase in the area under the curve of the IPGTT was obtained at the end of the high fructose diet, compared to the 11% increase in +/+ mice. Conclusion Our data suggest that constitutive chronic sympathetic hyperactivity can induce the early development of carbohydrate metabolism disorders. Moreover, it seems to represent a major factor of susceptibility to dietinduced metabolic dysfunction.