Protein kinase C inhibitors stimulate arachidonic and docosahexaenoic acids release from uterine stromal cells through a Ca2+-independent pathway1Enzymes: Phospholipase A2 (EC 3.1.1.4).1

AbstractThe mechanisms underlying arachidonic acid (AA) release by uterine stromal (UIII) cells were studied. Stimulation of AA release by calcium ionophore and PMA are inhibited by various PKC inhibitors and by calcium deprivation. These results suggest the involvement of an AA-specific cPLA2 as the release of docosahexaenoic acid (DHA) from prelabelled cells is much lower than the release of AA. The results also show a more original stimulation of AA and DHA release induced by PKC inhibitors, which is insensitive to calcium deprivation. This stimulation is not due to acyltransferase inhibition, suggesting the participation of a Ca2+-independent PLA2 (iPLA2). However, iPLA2 activity measured in UIII cells is inhibited by the specific iPLA2 inhibitor, BEL, and is not stimulated by PKC inhibitors, in contrast with the AA and DHA release. It seems therefore that this iPLA2 cannot be involved in this mechanism. The participation of another iPLA2, BEL-insensitive, is discussed

Anti-bis(monoacylglycero)phosphate antibody accumulates acetylated LDL-derived cholesterol in cultured macrophages.

International audienceBis(monoacylglycero)phosphate (BMP), also called lysobisphosphatidic acid, is a phospholipid highly enriched in the internal membranes of multivesicular late endosomes, in which it forms specialized lipid domains. It has been suggested that BMP-rich membranes regulate cholesterol transport. Here, we examine the effects of an anti-BMP antibody on cholesterol metabolism and transport in two macrophage cell lines, RAW 264.7 and THP-1, during loading with acetylated low density lipoprotein (AcLDL). Anti-BMP antibody was internalized and accumulated in both macrophage cell types. Cholesterol staining with filipin and mass measurements indicate that AcLDL-stimulated accumulation of free cholesterol (FC) was enhanced in macrophages that had accumulated the antibody. Unlike the hydrophobic amine U18666A (3-beta-[2-(diethylamino)ethoxy]androst-5-en-17-one), esterification of AcLDL-derived cholesterol by ACAT was not modified after anti-BMP treatment. AcLDL loading led to an increase of FC in the plasma membrane. This increase was further enhanced in anti-BMP-treated macrophages. However, cholesterol efflux to HDL was reduced in antibody-treated cells. These results suggest that the accumulation of anti-BMP antibody alters cholesterol homeostasis in AcLDL-loaded macrophages

Mast cell- and dendritic cell-derived exosomes display a specific lipid composition and an unusual membrane organization.

Exosomes are small vesicles secreted from multivesicular bodies, which are able to stimulate the immune system leading to tumour cell eradication. We have analysed lipids of exosomes secreted either upon stimulation from rat mast cells (RBL-2H3 cells), or constitutively from human dendritic cells. As compared with parent cells, exosomes displayed an enrichment in sphingomyelin, but not in cholesterol. Phosphatidylcholine content was decreased, but an enrichment was noted in disaturated molecular species as in phosphatidylethanolamines. Lyso(bis)phosphatidic acid was not enriched in exosomes as compared with cells. Fluorescence anisotropy demonstrated an increase in exosome-membrane rigidity from pH 5 to 7, suggesting their membrane reorganization between the acidic multivesicular body compartment and the neutral outer cell medium. NMR analysis established a bilayer organization of exosome membrane, and ESR studies using 16-doxyl stearic acid demonstrated a higher flip-flop of lipids between the two leaflets as compared with plasma membrane. In addition, the exosome membrane exhibited no asymmetrical distribution of phosphatidylethanolamines. Therefore exosome membrane displays a similar content of the major phospholipids and cholesterol, and is organized as a lipid bilayer with a random distribution of phosphatidylethanolamines. In addition, we observed tight lipid packing at neutral pH and a rapid flip-flop between the two leaflets of exosome membranes. These parameters could be used as a hallmark of exosomes

Five‐year outcomes in liver transplant patients receiving everolimus with or without a calcineurin inhibitor: Results from the CERTITUDE study

BACKGROUND AND AIMS: To report 5‐year outcomes of the CERTITUDE study. METHODS: An observational study in patients with liver transplantation (LTx) compared the long‐term impact of immunosuppression (with/without a calcineurin inhibitor) on renal function, cancers, major cardiovascular events (MACEs) and other safety parameters. All patients completing the 6‐month SIMCER study were recruited and analysed according to treatment received at randomization and actual treatment received during the follow‐up. RESULTS: Of the 143 enrolled patients, 119 completed the 5‐year follow‐up (everolimus [EVR], n = 55; tacrolimus [TAC], n = 64). The mean absolute change in estimated glomerular filtration rate was not statistically different between both groups (TAC, −15.53 ml/min/1.73 m(2) and EVR, –14.56 ml/min/1.73 m(2)). In the treatment subgroups based on actual treatment received, renal function was preserved better in the EVR subgroup compared with other subgroups (p = .051). Treated biopsy‐proven acute rejection was higher in the EVR group (15.4% vs. 6.4%); however, the majority of events were mild in severity. MACE occurred in 9.2% vs. 14.1% of patients in the EVR and TAC groups respectively (p = .370). De novo cancer was reported in 14 and 5 patients in EVR and TAC groups respectively. Hepatocellular carcinoma (HCC) recurrence was observed in the TAC group alone (n = 4). Adverse events and treatment discontinuation owing to an adverse event were higher in the EVR group. CONCLUSIONS: The CERTITUDE study demonstrated that EVR‐ and TAC‐based regimens have comparable efficacy, safety and tolerability up to 5 years post‐LTx

Sofosbuvir-based antiviral therapy in hepatitis C virus patients with severe renal failure

IF 4.470International audienceBackgroundChronic hepatitis C virus (HCV) infection is the most common chronic liver disease in patients with end-stage renal disease (ESRD). Over the last few years, second-generation direct-acting antivirals have been revolutionary in the treatment of hepatitis C, and sofosbuvir (SOF) is the backbone of most modern treatment strategies. Since SOF is eliminated through the kidney, the aim of this multicentre retrospective study was to assess its antiviral efficacy and safety in HCV-infected patients with severe renal failure [including haemodialysis (HD) patients].MethodsFifty patients (36 males, mean age ± standard deviation 60.5 ± 7.5 years) with chronic HCV infection (G1: 28/56%, cirrhosis: 27/54%) and severe renal failure [i.e. MDRD estimated glomerular filtration rate (eGFR) <35 mL/min], including 35 on HD, were enrolled. Antiviral treatment consisted of SOF/ribavirin (RBV) (n = 7), SOF/RBV/pegylated interferon (n = 2), SOF/daclatasvir ± RBV (n = 30) or SOF/simeprevir ± RBV (n = 11) for 12 or 24 weeks. A reduced dose of SOF (400 mg three times a week or 400 mg every other day) was given to all HD patients. Initial dose of RBV (n = 12) ranged from 400 to 4200 mg/week.ResultsOn an intent-to-treat-based analysis, sustained virological response rate was 86% at 12 weeks. During therapy, haemoglobin levels were not significantly modified, but recombinant erythropoietin (rEPO) dose significantly increased in patients treated with RBV. Two patients (4%) required blood transfusion. No patient had treatment discontinuation due to side effects. Dose of RBV was reduced in two patients (16.7%) during antiviral therapy. Dose of SOF was reduced in two non-HD patients because of side effects. In non-HD patients, median eGFR was not significantly modified during treatment.ConclusionsOur results strongly suggest that SOF-based antiviral therapy, with a reduced dose of SOF, is safe and effective for the treatment of HCV patients with ESRD, including HD patients

Sorafenib alone vs. sorafenib plus GEMOX as 1st-line treatment for advanced HCC: the phase II randomised PRODIGE 10 trial

International audienceBACKGROUND:Sorafenib remains one major first-line therapeutic options for advanced hepatocellular carcinoma (aHCC), with modest efficacy. We investigated the addition of gemcitabine and oxaliplatin (GEMOX) to sorafenib in aHCC patients.METHODS:Our multicentre phase II trial randomised aHCC first-line patients to sorafenib (400 mg BID) or sorafenib-GEMOX every 2 weeks (1000 mg/m2 gemcitabine; 100 mg/m2 oxaliplatin). Primary endpoint was the 4-month progression-free survival (PFS) rate.RESULTS:Ninety-four patients were randomised (sorafenib-GEMOX: n = 48; sorafenib: n = 46). Median age was 64 years, PS 0 (69%) or 1 (31%), 63% patients had cirrhosis, 29% portal vein thrombosis and 70% extra-hepatic disease. Median duration of sorafenib treatment was 4 months (1-51); median number of GEMOX cycles was 7 (1-16). The 4-month PFS rates were 64% and 61% in the sorafenib-GEMOX and sorafenib arms, respectively; median PFS and OS were 6.2 (95% CI: 3.8-6.8) and 13.5 (7.5-16.2) months, and 4.6 (3.9-6.2) months and 14.8 (12.2-22.2), respectively. The ORR/DCR were 9%/70% and 15%/77% in the sorafenib-GEMOX and sorafenib alone arms, respectively. Main toxicities were (sorafenib-GEMOX/sorafenib) neutropenia (23%/0), thrombocytopenia (33%/0), diarrhoea (18%/9), peripheral neuropathy (5%/0) and hand-foot syndrome (5%/18).CONCLUSIONS:Addition of GEMOX had an inpact on ORR and was well-tolerated as frontline systemic therapy. The benefit on PFS seems moderate; no subsequent study was planned