E007 Differential effects of microparticles from human apoptotic T lymphocytes and from human apoptotic monocytes in endothelial cells

During cell activation or apoptosis, cells release vesicles, also called microparticles (MPs) from the plasma membrane. Since composition of MPs is dependent on cell origin and the type of stimulation, we compared the effects of MPs generated from both apoptotic T lymphocytes and apoptotic monocytes on endothelial function with respect to both nitric oxide (NO) pathway and reactive oxygen species (ROS). MPs were produced by treatment of either human T lymphocytes with the apoptotic agent actinomycin D or human monocytic cell line THP-1 with the apoptotic agent, the etoposide VP-16. Human Eahy 926 endothelial cells were incubated with 10μg/ml MPs for 24h. Apoptotic MPs from human T lymphocytes decreased NO production that was associated with overexpression and phosphorylation of endothelial NOsynthase (eNOS). Also, T lymphocytes MPs enhanced expression of caveolin-1 and decreased its phosphorylation. T lymphocytes MPs enhanced ROS by a mechanism sensitive to xanthine oxidase and P-IkappaBalpha inhibitors. PI3-kinase inhibition reduced the effects of T lymphocytes MPs on eNOS, but not on caveolin-1, whereas it enhanced the effects of MPs on ROS production. Inhibition of MEK reversed eNOS phosphorylation but it had no effect on ROS production induced by T lymphocytes MPs. By contrast, apoptotic MPs from human monocytes increased both NO production and in much less extent ROS. These effects were associated with a decrease of caveolin-1 expression and increased its phosphorylation, without affecting eNOS expression and phosphorylation. The inhibitor of the PI-3kinase, LY294002, reversed the effects of monocyte MPs on caveolin-1 expression but not on its phosphorylation. The MEK1/2 inhibitor, U0126, reversed the decrease of caveolin-1 expression induced by MPs from monocytes. Interestingly, U0126 potentiated ROS production induced by monocyte MPs. Whereas in vivo injection of T lymphocytes MPs in mice impaired endothelial function, apoptotic MPs from human monocytes did not affect endothelium-dependent relaxation. In addition, monocyte MPs were able to promote in vitro angiogenesis. In summary, these results highlight differential effects of apoptotic MPs from different origins by activating diverse multiple pathways related to NO and ROS productions

The Mechanism of Action of Cytokines to Control the Release of Hypothalamic and Pituitary Hormones in Infection

Abstract: During infection, bacterial and viral products, such as bacterial lipopolysaccharide (LPS), cause the release of cytokines from immune cells. These cytokines can reach the brain by several routes. Furthermore, cytokines, such as interleukin‐1 (IL‐1), are induced in neurons within the brain by systemic injection of LPS. These cytokines determine the pattern of hypothalamic‐pituitary secretion that characterizes infection. IL‐2, by stimulation of cholinergic neurons, activates neural nitric oxide synthase (nNOS). The nitric oxide (NO) released diffuses into corticotropin‐releasing hormone (CRH)‐secreting neurons and releases CRH. IL‐2 also acts in the pituitary to stimulate adrenocorticotropic hormone (ACTH) secretion. On the other hand, IL‐1α blocks the NO‐induced release of luteinizing hormone‐releasing hormone (LHRH) from LHRH neurons, thereby blocking pulsatile LH but not follicle‐stimulating hormone (FSH) release and also inhibiting sex behavior that is induced by LHRH. IL‐1α and granulocyte macrophage colony‐stimulating factor (GMCSF) block the response of the LHRH terminals to NO. The mechanism of action of GMCSF to inhibit LHRH release is as follows. It acts on its receptors on γ‐aminobutyric acid (GABA)ergic neurons to stimulate GABA release. GABA acts on GABAa receptors on the LHRH neuronal terminal to block NOergic stimulation of LHRH release. IL‐1α inhibits growth hormone (GH) release by inhibiting GH‐releasing hormone (GHRH) release, which is mediated by NO, and stimulating somatostatin release, also mediated by NO. IL‐1α‐induced stimulation of PRL release is also mediated by intra‐hypothlamic action of NO, which inhibits release of the PRL‐inhibiting hormone dopamine. The actions of NO are brought about by its combined activation of guanylate cyclase‐liberating cyclic guanosine monophosphate (cGMP) and activation of cyclooxygenase (COX) and lipoxygenase (LOX) with liberation of prostaglandin E2 and leukotrienes, respectively. Thus, NO plays a key role in inducing the changes in release of hypothalamic peptides induced in infection by cytokines. Cytokines, such as IL‐1β, also act in the anterior pituitary gland, at least in part via induction of inducible NOS. The NO produced inhibits release of ACTH. The adipocyte hormone leptin, a member of the cytokine family, has largely opposite actions to those of the proinflammatory cytokines, stimulating the release of FSHRF and LHRH from the hypothalamus and FSH and LH from the pituitary directly by NO.Fil: McCann, Samuel M.. Pennington Biomedical Research Center; Estados UnidosFil: Kimura, M.. Medical and Dental University; JapónFil: Karanth, S.. Pennington Biomedical Research Center; Estados UnidosFil: Yu, W. H.. Pennington Biomedical Research Center; Estados UnidosFil: Mastronardi, C. A.. Pennington Biomedical Research Center; Estados UnidosFil: Besuhli, Valeria. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Houssay. Centro de Estudios Farmacológicos y Botánicos. Universidad de Buenos Aires. Facultad de Medicina. Centro de Estudios Farmacológicos y Botánicos; Argentin

Whole exome sequencing of extreme morbid obesity patients: translational implications for obesity and related disorders

Whole-exome sequencing (WES) is a new tool that allows the rapid, inexpensive and accurate exploration of Mendelian and complex diseases, such as obesity. To identify sequence variants associated with obesity, we performed WES of family trios of one male teenager and one female child with severe early-onset obesity. Additionally, the teenager patient had hypopituitarism and hyperprolactinaemia. A comprehensive bioinformatics analysis found de novo and compound heterozygote sequence variants with a damaging effect on genes previously associated with obesity in mice (LRP2) and humans (UCP2), among other intriguing mutations affecting ciliary function (DNAAF1). A gene ontology and pathway analysis of genes harbouring mutations resulted in the significant identification of overrepresented pathways related to ATP/ITP (adenosine/inosine triphosphate) metabolism and, in general, to the regulation of lipid metabolism. We discuss the clinical and physiological consequences of these mutations and the importance of these findings for either the clinical assessment or eventual treatment of morbid obesity.Gilberto Paz-Filho, Margaret C.S. Boguszewski, Claudio A. Mastronardi, Hardip R. Patel, Angad S. Johar, Aaron Chuah, Gavin A. Huttley, Cesar L. Boguszewski, Ma-Li Wong, Mauricio Arcos-Burgos and Julio Licini

Comparison of perioperative outcomes between standard laparoscopic and robot-assisted approach in patients with rectosigmoid endometriosis

Introduction: Robot-assisted laparoscopic surgery (RALS) has gained widespread application in several surgical specialties. Previous studies on the feasibility and safety of RALS vs standard laparoscopy (S-LPS) for rectosigmoid endometriosis are limited and reported conflicting data. This study aims to compare S-LPS and RALS in patients with rectosigmoid endometriosis in terms of perioperative surgical and clinical data. Material and methods: This is a multicentric, observational, prospective cohort study including 44 patients affected by rectosigmoid endometriosis referred to two tertiary referral centers for endometriosis from September 2018 to September 2019. Patients were divided into two groups: 22 patients underwent S-LPS, and 22 underwent RALS. Our primary outcome was to compare operative time (from skin incision to suture) between the two groups. Secondary outcomes included: operative room time (patient entry into operative room and patient out), estimated blood loss, laparotomic conversion rate, length of hospital stay, perioperative complications, and evaluation of endometriosis-related symptoms at 12-month follow up. Results: The two groups were comparable regarding preoperative and surgical data, except for higher rates of hysterectomies and bilateral uterosacral ligament removal procedures in the RALS group. Also after adjusting for these discrepancies, operative time was similar between S-LPS and RALS. Operative room time was statistically longer in the RALS group compared with that of S-LPS. No statistically significant difference was found concerning other study outcomes. Pain and bowel symptoms improved in both groups at 12-month follow up. Conclusions: If performed by expert teams, RALS provides similar perioperative outcomes compared with S-LPS in rectosigmoid endometriosis surgical treatment, except for longer operative room time

Efficient synthesis of kainic acid analogues

The present paper deals with an improved synthesis of two molecular hybrids of AMPA and KA, compounds CIP-A and CIP-B, and their transformation into CIOP-A and CIOP-B, the corresponding amido derivatives. Exploiting the continuous-flow technology, a significant improvement in the synthesis of the glutamate agonists CIP-A and CIP-B was accomplished, in terms of overall yield, time, and excess of ethyl chlorooximinoacetate. Moreover, we find out the HPLC conditions suitable to separate, at a preparative level, the three intermediates formed in the 1,3-dipolar cycloaddition step

Smart materials based on DNA aptamers: Taking aptasensing to the next level

"Smart" materials are an emerging category of multifunctional materials with physical or chemical properties that can be controllably altered in response to an external stimulus. By combining the standard properties of the advanced material with the unique ability to recognize and adapt in response to a change in their environment, these materials are finding applications in areas such as sensing and drug delivery. While the majority of these materials are responsive to physical or chemical changes, a particularly exciting area of research seeks to develop smart materials that are sensitive to specific molecular or biomolecular stimuli. These systems require the integration of a molecular recognition probe specific to the target molecule of interest. The ease of synthesis and labeling, low cost, and stability of DNA aptamers make them uniquely suited to effectively serve as molecular recognition probes in novel smart material systems. This review will highlight current work in the area of aptamer-based smart materials and prospects for their future applications

Lipopolysaccharide-induced leptin release is neurally controlled

Our hypothesis is that leptin release is controlled neurohormonally. Conscious, male rats bearing indwelling, external, jugular catheters were injected with the test drug or 0.9% NaCl (saline), and blood samples were drawn thereafter to measure plasma leptin. Anesthesia decreased plasma leptin concentrations within 10 min to a minimum at 120 min, followed by a rebound at 360 min. Administration (i.v.) of lipopolysaccharide (LPS) increased plasma leptin to almost twice baseline by 120 min, and it remained on a plateau for 360 min, accompanied by increased adipocyte leptin mRNA. Anesthesia largely blunted the LPS-induced leptin release at 120 min. Isoproterenol (β-adrenergic agonist) failed to alter plasma leptin but reduced LPS-induced leptin release significantly. Propranolol (β-receptor antagonist) produced a significant increase in plasma leptin but had no effect on the response to LPS. Phentolamine (α-adrenergic receptor blocker) not only increased plasma leptin (P < 0.001), but also augmented the LPS-induced increase (P < 0.001). α-Bromoergocryptine (dopaminergic-2 receptor agonist) decreased plasma leptin (P < 0.01) and blunted the LPS-induced rise in plasma leptin release (P < 0.001). We conclude that leptin is at least in part controlled neurally because anesthesia decreased plasma leptin and blocked its response to LPS. The findings that phentolamine and propranolol increased plasma leptin concentrations suggest that leptin release is inhibited by the sympathetic nervous system mediated principally by α-adrenergic receptors because phentolamine, but not propranolol, augmented the response to LPS. Because α-bromoergocryptine decreased basal and LPS-induced leptin release, dopaminergic neurons may inhibit basal and LPS-induced leptin release by suppression of release of prolactin from the adenohypophysis

Synthesis of L-tricholomic acid analogues and pharmacological characterization at ionotropic glutamate receptors

The synthesis of analogues of the natural compound ltricholomic acid and of its threo diastereoisomer was accomplished in order to explore their affinity for glutamate ionotropic receptors. In this study, fourteen new unnatural amino acids, characterized by a 3-hydroxy-~2 -isoxazoline or 3- hydroxy-~2 -pyrazoline-skeleton, were obtained exploiting, as key reaction, a 1,3-dipolar cycloaddition or an intramolecular cyclization

Phosphatidylinositol 3-Kinase and Xanthine Oxidase Regulate Nitric Oxide and Reactive Oxygen Species Productions by Apoptotic Lymphocyte Microparticles in Endothelial Cells

Microparticles (MPs) are membrane vesicles released during cell activation and apoptosis. We have previously shown that MPs from apoptotic T cells induce endothelial dysfunction, but the mechanisms implicated are not completely elucidated. In this study, we dissect the pathways involved in endothelial cells with respect to both NO and reactive oxygen species (ROS). Incubation of endothelial cells with MPs decreased NO production that was associated with overexpression and phosphorylation of endothelial NO synthase (eNOS). Also, MPs enhanced expression of caveolin-1 and decreased its phosphorylation. Microparticles enhanced ROS by a mechanism sensitive to xanthine oxidase and P-IκBα inhibitors. PI3K inhibition reduced the effects of MPs on eNOS, but not on caveolin-1, whereas it enhanced the effects of MPs on ROS production. Microparticles stimulated ERK1/2 phosphorylation via a PI3K-depedent mechanism. Inhibition of MEK reversed eNOS phosphorylation but had no effect on ROS production induced by MPs. In vivo injection of MPs in mice impaired endothelial function. In summary, MPs activate pathways related to NO and ROS productions through PI3K, xanthine oxidase, and NF-κB pathways. These data underscore the pleiotropic effects of MPs on NO and ROS, leading to an increase oxidative stress that may account for the deleterious effects of MPs on endothelial function

The use of FDG-PET in the initial staging of 142 patients with follicular lymphoma: a retrospective study from the FOLL05 randomized trial of the Fondazione Italiana Linfomi

BACKGROUND: The role of [(18)F] fluorodeoxyglucose (FDG)-positron emission tomography (PET) in follicular lymphoma (FL) staging is not yet determined. PATIENTS AND METHODS: The aim of the present study was to investigate the role of PET in the initial staging of FL patients enrolled in the FOLL05-phase-III trial that compared first-line regimens (R-CVP, R-CHOP and R-FM). Patients should have undergone conventional staging and have available PET baseline to be included. RESULTS: A total of 142 patients were analysed. PET identified a higher number of nodal areas in 32% (46 of 142) of patients and more extranodal (EN) sites than computed tomography (CT) scan. Also, the Follicular Lymphoma International Prognostic Index (FLIPI) score increased in 18% (26 of 142) and decreased in 6% (9 of 142) of patients. Overall, the impact of PET on modifying the stage was highest in patients with limited stage. Actually, 62% (15 of 24) of cases with limited disease were upstaged with PET. CONCLUSIONS: The inclusion of PET among staging procedures makes the evaluation of patients with FL more accurate and has the potential to modify therapy decision and prognosis in a moderate proportion of patients. Further prospective clinical trials on FL should incorporate PET at different moments, and the therapeutic criteria to start therapy should be re-visited in the views of this new tool