L Di Renzo

Cardiolipin and its metabolites move from mitochondria to other cellular membranes during death receptor-mediated apoptosis

AbstactWe previously reported that during death receptor-mediated apoptosis, cardiolipin (CL) relocates to the cell surface, where it reacts with autoantibodies from antiphospholipid syndrome sera. Here, we analysed the intracellular distribution of CL and its metabolites during the early phase of cell death signalling triggered by Fas stimulation in U937 cells and mouse liver. We found a redistribution of mitochondrial CL to the cell surface by using confocal microscopy and flow cytometry. Mass spectrometry revealed that CL and its metabolites relocated from mitochondria to other intracellular organelles during apoptosis, with a conversion into non-mitochondrial lipids. Concomitantly, cytosolic Bid relocated to the light membranes comprised in fraction P100, including the plasma membrane and associated vesicular systems. A direct Bid–CL interaction was demonstrated by the observation that CL and monolysoCL coimmunoprecipitated with Bid especially after Fas stimulation, suggesting a dynamic interaction of the protein with CL and its metabolites.

Persistent production of platelet thromboxane A2 in patients chronically treated with aspirin.

Summary.  Background: Patients treated with aspirin may have a reduced sensitivity to its antiplatelet effect. The mechanism accounting for such a reduced sensitivity might involve an impaired interaction of aspirin with cyclooxygenase‐1 (COX)‐1. Objective: We sought to investigate whether platelets from patients under chronic treatment with aspirin still produce TxA2 and whether there is any relationship between the eventual persistent TxA2 formation and platelet aggregation. Finally, whether platelet‐derived TxA2 can be inhibited by in vitro addition of aspirin. Methods: Collagen‐induced platelet aggregation and thromboxane‐A2 (TxA2) were measured in 196 patients treated with aspirin (100–330 mg day−1) because of previous vascular events or presence of risk factors of atherosclerosis. Results: Collagen‐induced TxA2 production of the entire cohort was 128.7 ± 21.6 pg 10−8 cells, and was significantly correlated with platelet aggregation (Spearmans correlation coefficient = 0.44; P < 0.0001). Patients in the highest quartile of TxA2 showed higher platelet response to collagen (P < 0.0001) when compared with those in the lowest quartile. In a subgroup of 96 patients, platelets were treated in vitro with a TxA2 receptor antagonist (13‐azaprostanoic acid) or aspirin before stimulation with collagen. 13‐APA acid significantly inhibited platelet aggregation. Aspirin reduced (−72.9%) TxA2 production in patients with TxA2 values above the median but it was ineffective in those with TxA2 values below the median. Conclusion: In some patients chronically treated with aspirin platelet production of TxA2 may persist and account for enhanced platelet aggregation. Incomplete inhibition of COX‐1 seems to be implicated in persistent TxA2 production.

α-Lipoic Acid Supplementation: A Tool for Obesity Therapy?

Lipid peroxidation has supposed as the major biochemical alteration underling oxidant-induced cell injury in stress including numerous diseases. One of the natural molecules know to prevent or retard oxidation is alpha-lipoic acid (LA) and, therefore, the lipoic acid/dihydrolipoic acid (LA/DHLA) redox couple has received considerable attention. Recent studies have highlighted the potential of free LA and DHLA as powerful metabolic antioxidants that are able to scavenge the reactive oxygen species, to recycle other antioxidants. Our aim was to investigate the beneficial effects of LA in the treatment of Italian pre-obese and obese subjects. We screened 1612 subjects for enrollment; of these, 1127 subjects (445 men and 682 women, 18-60 age) met enrolment criteria and were enrolled in the study. According to body mass index (BMI) the 53% was obese and the 43% was pre-obese. The subjects were treated for 4 month with 800 mg/day of LA. In pre-obese subject significant reduction (p<0.001) of weight (8%, both gender), BMI (2 points), blood pressure, and abdominal circumference (female 6 cm, male 7 cm) were observed. In obese subjects significant reductions (p<0.001) of weight (9%, both gender), BMI (female 3 point, male 4 point), blood pressure and abdominal circumference (female 9 cm, male 11 cm) were observed. Our study indicated that LA is an ideal antioxidant candidate for the therapy of obesity related diseases. Further clinical studies should be considered to highlight the role and efficacy of LA treatment.

JNK2 is activated during ER stress and promotes cell survival.

Adaptation to endoplasmic reticulum (ER) stress relies on activation of the unfolded protein response (UPR) and induction of autophagy. Indeed, cells die if ER stress is not countered by the UPR. Here we show in U937 cells that the ER stressors tunicamycin and thapsigargin cause increased expression of c-Jun N-terminal kinase 2 (JNK2), which allows regulation of the UPR, whose silencing or pharmacological inhibition delays BiP (immunoglobulin heavy-chain binding protein) upregulation, and causes earlier and greater expression of CCAAT/enhancer-binding protein-homologous protein (CHOP). Furthermore, we show that pharmacological inhibition or silencing of JNK2 causes accumulation of both p62 and the acidic compartment, caspase 3 activation and apoptosis. Our results reveal that JNK2 prevents accumulation of the acidic compartment in U937 cells undergoing autophagic flux and, by this mechanism, it keeps stressed cells alive. Our findings highlight a potential role for JNK2 in tumor cell survival, senescence and neurodegenerative diseases, in which ER stress, autophagy and lysosome activity are known to interplay.

HSP70 inhibition by 2-phenylethynesulfonamide induces lysosomal cathepsin D release and immunogenic cell death in primary effusion lymphoma

Heat-shock protein (HSP) 70 is aberrantly expressed in different malignancies and has a cancer-specific cell-protective effect. As such, it has emerged as a promising target for anticancer therapy. In this study, the effect of the HSP70-specific inhibitor (PES), also Pifitrin-μ, on primary effusion lymphoma (PEL) cell viability was analyzed. PES treatment induced a dose- and time-dependent cytotoxic effect in BC3 and BCBL1 PEL cells by inducing lysosome membrane permeabilization, relocation of cathepsin D in the cytosol, Bid cleavage, mitochondrial depolarization with release and nuclear translocation of apoptosis-activating factor. The PES-induced cell death in PEL cells was characterized by the appearance of Annexin-V/propidium iodide double-positive cells from the early times of treatment, indicating the occurrence of an additional type of cell death other than apoptosis, which, accordingly, was not efficiently prevented by the pan-caspase inhibitor Z-VAD-fmk. Conversely, PES-induced cell death was robustly reduced by pepstatin A, which inhibits Bid and caspase 8 processing. In addition, PES was responsible for a block of the autophagic process in PEL cells. Finally, we found that PES-induced cell death has immunogenic potential being able to induce dendritic cell activation.

Body composition changes after laparoscopic adjustable gastric banding: what is the role of -174G>C interleukin-6 promoter gene polymorphism in the therapeutic strategy?

Background:There is growing evidence that interleukin-6 (IL-6) is linked to the regulation of fat mass (FM). Our previous data define the common −174G>C IL-6 polymorphism as a marker for ‘vulnerable’ individuals at risk of age- and obesity-related diseases. An association between −174G>C IL-6 polymorphism and weight loss after bariatric surgery has been demonstrated.Objective:We investigated the impact of −174G>C IL-6 polymorphism on weight loss, body composition, fluid distribution and cardiometabolic changes in obese subjects, after laparoscopic adjustable gastric banding (LAGB) surgery.Design and Outcome measures:A total of 40 obese subjects were studied at baseline and at 6 months follow-up after LAGB surgery. Cardiometabolic and genetic assessment of −174G>C IL-6 polymorphism, anthropometric, body composition and fluid distribution analysis were performed.Results:After LAGB surgery, significant reductions in weight (Δ%=−11.66±7.78, P<0.001), body mass index (P<0.001), total and trunk FM (kg, %) (Δ% of total FM=−22.22±12.15, P<0.01), bone mineral density (T-score) (P<0.001), resting metabolic rate (RMR) (P<0.01), and total body water and intracellular water (TBW, ICW) (P<0.05) were observed. At baseline, C(−) carriers of IL-6 polymorphism had a significantly higher RMR (P<0.05), free FM (kg), but less total and trunk FM (%), higher body cell mass (BCM), content of TBW (L) and ECW (extracellular water)/ICW ratio compared with C(+) carriers (P<0.001). After LAGB, C(+) carriers had a significantly stronger reduction of total FM (kg), but lower bone density, compared with C(−) carriers (P<0.05).Conclusions:Beyond the relationship between −174G>C IL-6 polymorphism and body composition, this study provides first evidence about the association of IL-6 variant with fluid distribution, at baseline, and FM and bone density loss in obese subjects at 6 months follow-up after LAGB surgery. LAGB was less effective if the subjects were carrying risk genotypes, C(−) carriers, for obesity, suggesting a role of genetic variations on bariatric surgery outcomes.

Dendritic cell differentiation blocked by primary effusion lymphoma-released factors is partially restored by inhibition of P38 MAPK.

To better understand the molecular mechanisms underlying the dendritic cell (DC) defects in cancer, we analyzed which signaling pathway is implicated in the abnormal monocyte differentiation into DC determined by the presence of Primary effusion lymphoma (PEL) released factors. Our results indicate that the DC, obtained in this condition, together with phenotypic abnormalities and reduced allostimulatory function, showed hyperphosphorylation of signal transducer and activator of transcription 3 (STAT3) and p38 mitogen-activated protein kinase (MAPK) molecules, in comparison to the DC differentiated in the absence of PEL-released factors. The inhibition of p38 MAPK but not of STAT3 phosphorylation, with specific inhibitors, was able to revert the effect of the PEL-released factors on the DC phenotype. This study suggests that p38 MAPK signaling pathway is an important contributor to the abnormal differentiation of DC in PEL.

Role of interleukin-15 receptor α polymorphisms in normal weight obese syndrome.

Previous published studies have identified a class of women, Normal Weight Obese women (NWO) with normal BMI and high fat content. An important role of Interleukin-15 (IL-15) has been documented in facilitating muscle proliferation and promoting fat depletion. Indeed the presence of three types of IL-15 receptor subunits in fat tissue suggests a direct effect on adipose tissue. We studied three single nucleotide polymorphisms (SNP) of IL-15Rα receptor gene and investigated their relationship with NWO phenotype. We considered two classes of women according to their BMI and percent fat mass (%FAT), class 1: including 72 overweight-obese women (high BMI-high fat mass) and class 2: including 36 NWO (normal BMI, high fat mass). Three sites of Interleukin-15 receptor subunit α gene were examined, located respectively in exon4, exon5 intron-exon border and exon7. Genotyping of the identified polymorphisms was performed by restriction fragment length polymorphism. Haplotype frequency estimation was performed by using the Mendel-University of Chicago program. Odds ratio analyses were calculated by EPISTAT program. Highly significant differences were observed for exon 7-exon5 intron-exon border and exon 4-exon 7 haplotype distribution between class 1 and class 2 women. These results strongly support the hypothesis that genetic variability of the IL-15 receptor has an important role in body fat composition. Our data underscore previous findings that suggest a potential role of IL-15 cytokine in NWO syndrome.

A new predictive equation for evaluating women body fat percentage and obesity-related cardiovascular disease risk.

BackgroundObesity represents a global public health problem due to its association with cardiovascular diseases and reduced lifespan. The most widely used classification of obesity is expressed as Body Mass Index (BMI); however, this formula is an imprecise adiposity measurement that ignores several important factors involved. Body Adiposity Index (BAI) was more recently proposed as an indirect evaluation of percentage body fat (PBF). PBF is a direct measure of person’s relative body fat and a better predictor of obesity-related risk diseases than BMI and BAI. Since obesity and consequent diseases are considered epidemic, new accurate formulas for epidemiological studies are of interest to the scientific community. Because direct measurement of body composition could be quite expensive, the aims of our work were to analyse the distributions of PBF by Dual X-ray absorptiometry, and the creation of new predictive equation using only anthropometric measures that could be helpful to clinicians to assess easily body fat of female patients.Methods/resultsA sample of 1,031 Caucasian Italian women was recruited and BMI, BAI and PBF were evaluated. With the aim of developing a predictive model of PBF a multivariate regression model was fitted to observed data.ConclusionsThe definition of universally recognized PBF by gender and age could have public health implications. In this study, we developed a new predictive PBF equation that does not require the use of medical instruments or skilled measurement techniques and that may be easily applicable to Italian women.

Stimulation of macrophages with IFNγ or TNFα shuts off the suppressive effect played by PGE2

PGE2 has been shown to be able to interfere with various lymphocyte and macrophage functions, but its effects on macrophage activation are still unclear. In this study, carried out on peritoneal macrophages obtained from healthy, tumour-bearing and Corynebacterium parvum-treated mice, we demonstrated that PGE2 is involved in the down-regulation of macrophage activation, but it cannot exert its inhibiting effect when macrophages are further stimulated with activating cytokines, such as IFNγ and TNFα. Our findings provide new insight into how macrophage tumoricidal activity may be induced and maintained even in presence of significant levels of PGE2.