The adjuvant activity of two urea derivatives on cytokinins: an example of serendipitous dual effect

The aim of this study was to investigate the action spectrum of two urea derivatives, the 1,3-di(benzo[d]oxazol-5-yl)urea (5-BDPU) and the 1,3-di(benzo[d]oxazol-6-yl)urea (6-BDPU). In order to evaluate a possible adjuvant activity on cytokinins the compounds alone or in the simultaneous presence of different cytokinins were assayed either on in vitro typical cytokinin-related bioassays, or on in planta interaction with cytokinin signal transduction pathway. The compounds ability to activate the cytokinin receptor CRE1/AHK4 was studied either by a heterologous bacterial assay or by a competitive binding assay and docking simulations were performed with the crystal structure of the same receptor. Then, owing to their chemical structure which resembles that of urea-type cytokinins, the ability of 5- and 6-BDPU to inhibit the activity of cytokinin oxidase/dehydrogenase of Zea mays (ZmCKX1) was investigated and docking simulations were performed as well. Accordingly to the experimental results, we speculate that BDPUs could show a dual activity: the blocking of the conformational re-adaption of CRE1/AHK4 receptor maintaining the cytokinin inside its binding pocket, thus possibly enhancing its kinase action; the inhibition of cytokinin oxidase/dehydrogenase activity thus possibly preventing its cleavage of natural cytokinins with isoprenoid side chain. Graphic abstract: [Figure not available: see fulltext.

Homocysteine induces cell death in H9C2 cardiomyocytes through the generation of peroxynitrite

Homocysteine (HCY) is toxic on blood vessels, but a potential direct toxicity of HCY on the heart is unknown. We addressed this issue by exposing H9C2 cardiomyocytes to HCY (0.1-5 mM) for up to 6h. At these concentrations, HCY reduced cell viability, induced necrosis and apoptosis and triggered the cleavage of caspase-3 and poly(ADP-ribose) polymerase (PARP). This was associated with the intracellular generation of the potent oxidant peroxynitrite. Removing peroxynitrite by the decomposition catalyst FeTPPS considerably reduced LDH release, DNA fragmentation, cleavage of caspase-3 and PARP, and restored normal cell morphology. In additional experiments performed in primary rat ventricular cardiomyocytes, HCY (1 mM, 6h) activated the phosphorylation of the MAP kinases ERK and JNK, two essential stress signaling kinases regulating myocardial apoptosis, hypertrophy and remodeling. These results provide the first demonstration that HCY kills cardiomyocytes through the generation of peroxynitrite and can activate key signaling cascades in the myocardium

Peroxynitrite is a potent inhibitor of NF-{kappa}B activation triggered by inflammatory stimuli in cardiac and endothelial cell lines.

Peroxynitrite is a potent oxidant and nitrating species proposed as a direct effector of myocardial damage in numerous cardiac pathologies. Whether peroxynitrite also acts indirectly, by modulating cell signal transduction in the myocardium, has not been investigated. Therefore, we examined a possible role for peroxynitrite on the activation of NF-kappaB, a crucial pro-inflammatory transcription factor, in cultured H9C2 cardiomyocytes. H9C2 cells were stimulated with tumor necrosis factor-alpha or lipopolysaccharide following a brief (20-min) exposure to peroxynitrite. NF-kappaB activation (phosphorylation and degradation of its inhibitor IkappaBalpha, nuclear translocation of NF-kappaB p65, and NF-kappaB DNA binding) triggered by lipopolysaccharide or tumor necrosis factor-alpha was abrogated by peroxynitrite. Peroxynitrite also inhibited NF-kappaB in two human endothelial cell lines activated with tumor necrosis factor-alpha or interleukin-1beta. These effects were related to oxidative but not nitrative chemistry and were still being observed while nitration was suppressed by epicatechin. The mechanism of NF-kappaB inhibition by peroxynitrite was a complete blockade of phosphorylation and activation of the upstream kinase IkappaB kinase (IKK) beta, required for canonical, pro-inflammatory NF-kappaB activation. At the same time, peroxynitrite activated phosphorylation of NF-kappaB-inducing kinase and IKKalpha, considered as part of an alternative, noncanonical NF-kappaB activation pathway. Suppression of IKKbeta-dependent NF-kappaB activation translated into a marked inhibition of the transcription of NF-kappaB-dependent genes by peroxynitrite. Thus, peroxynitrite has a dual effect on NF-kappaB, inhibiting canonical IKKbeta-dependent NF-kappaB activation while activating NF-kappaB-inducing kinase and IKKalpha phosphorylation, which suggests its involvement in an alternative pathway of NF-kappaB activation. These findings offer new perspectives for the understanding of the relationships between redox stress and inflammation

Trimester-specific reference intervals for thyroid function parameters in pregnant Caucasian women using Roche platforms: a prospective study

Background: Standard thyroid function parameters reference intervals (RI) are unsuitable during pregnancy, potentially resulting in incongruous treatments that may cause adverse effects on pregnancy outcomes. We aimed at defining trimester-specific TSH, FT4 and FT3 RI, using samples longitudinally collected from healthy Caucasian women. Materials and methods: Blood samples from 150 healthy Caucasian women, who had a physiological gestation and a healthy newborn at term, were collected in each trimester and at around six months post-partum. They showed mild iodine deficiency. After excluding women with overt TSH abnormalities (> 10 mU/L) and/or TPO antibodies, data from 139 pregnant women were analyzed by means of widely used Roche platforms, and TSH, FT4 and FT3 trimester-specific RI were calculated. Post-partum data were available for 55 subjects. Results: Serum TSH RI were 0.34-3.81 mU/L in the first trimester, and changed slightly to 0.68-4.07 U/L and 0.63-4.00 mU/L in the second and third trimester, respectively. Conversely, both FT4 and FT3 concentrations progressively decreased during pregnancy, the median values in the third trimester being 14.8% and 13.2% lower, respectively, than in the first trimester. Thyroid function parameters in the first trimester were similar to those measured after the end of pregnancy. Conclusions: This study calculates trimester-specific RI for thyroid function parameters in pregnancy, and proposes the reference limits that should be adopted when using Roche platforms in Caucasian women

Different pioneer plant species select specific rhizosphere bacterial communities in a high mountain environment

The rhizobacterial communities of 29 pioneer plants belonging to 12 species were investigated in an alpine ecosystem to assess if plants from different species could select for specific rhizobacterial communities. Rhizospheres and unvegetated soils were collected from a floristic pioneer stage plot at 2,400 m a.s.l. in the forefield of Weisskugel Glacier (Matsch Valley, South Tyrol, Italy), after 160 years of glacier retreat. To allow for a culture-independent perspective, total environmental DNA was extracted from both rhizosphere and bare soil samples and analyzed by Automated Ribosomal Intergenic Spacer Analysis (ARISA) and Denaturing Gradient Gel Electrophoresis (DGGE). ARISA fingerprinting showed that rhizobacterial genetic structure was extremely different from bare soil bacterial communities while rhizobacterial communities clustered strictly together according to the plant species. Sequencing of DGGE bands showed that rhizobacterial communities were mainly composed of Acidobacteria and Proteobacteria whereas bare soil was colonized by Acidobacteria and Clostridia. UniFrac significance calculated on DGGE results confirmed the rhizosphere effect exerted by the 12 species and showed different bacterial communities (P < 0.05) associated with all the plant species. These results pointed out that specific rhizobacterial communities were selected by pioneer plants of different species in a high mountain ecosystem characterized by oligotrophic and harsh environmental conditions, during an early primary succession

Proceedings of the Workshop on Monte Carlo's, Physics and Simulations at the LHC PART II

These proceedings collect the presentations given at the first three meetings of the INFN "Workshop on Monte Carlo's, Physics and Simulations at the LHC", held at the Frascati National Laboratories in 2006. The first part of these proceedings contains pedagogical introductions to several basic topics of both theoretical and experimental high pT LHC physics. The second part collects more specialised presentations.Comment: 157 pages, 136 figures; contribution by M. Grazzini has been adde

Safe-site effects on rhizosphere bacterial communities in a high-altitude alpine environment

The rhizosphere effect on bacterial communities associated with three floristic communities (RW, FI, and M sites) which differed for the developmental stages was studied in a high-altitude alpine ecosystem. RW site was an early developmental stage, FI was an intermediate stage, M was a later more matured stage. The N and C contents in the soils confirmed a different developmental stage with a kind of gradient from the unvegetated bare soil (BS) site through RW, FI up to M site. The floristic communities were composed of 21 pioneer plants belonging to 14 species. Automated ribosomal intergenic spacer analysis showed different bacterial genetic structures per each floristic consortium which differed also from the BS site. When plants of the same species occurred within the same site, almost all their bacterial communities clustered together exhibiting a plant species effect. Unifrac significance value (P < 0.05) on 16S rRNA gene diversity revealed significant differences (P < 0.05) between BS site and the vegetated sites with a weak similarity to the RW site. The intermediate plant colonization stage FI did not differ significantly from the RW and the M vegetated sites. These results pointed out the effect of different floristic communities rhizospheres on their soil bacterial communities

Lung carcinoid tumours: histology and Ki-67, the eternal rivalry

WHO classification of Thoracic Tumours defines lung carcinoid tumours (LCTs) as well-differentiated neuroendocrine neoplasms (NENs) classified in low grade typical (TC) and intermediate grade atypical carcinoids (AC). Limited data exist concerning protein expression and morphologic factors able to predict disease aggressiveness. Though Ki-67 has proved to be a powerful diagnostic and prognostic factor for Gastro-entero-pancreatic NENs, its role in lung NENs is still debated. A retrospective series of 370 LCT from two oncology centers was centrally reviewed. Morphology and immunohistochemical markers (Ki-67, TTF-1, CD44, OTP, SSTR-2A, Ascl1, and p53) were studied and correlated with Overall Survival (OS), Cancer-specific survival (CSS) and Disease-free survival (DFS). Carcinoid histology was confirmed in 355 patients: 297 (83.7%) TC and 58 (16.3%) AC. Ki-67 at 3% was the best value in predicting DFS. Ki-67 ≥ 3% tumours were significantly associated with AC histology, stage III-IV, smoking, vascular invasion, tumour spread through air spaces OTP negativity, and TTF-1, Ascl1 and p53 positivity. After adjustment for center and period of diagnosis, both Ki-67 (≥3 versus <3) and histology (AC versus TC) alone significantly added prognostic information to OS and CSS multivariable model with age, stage and OTP; addition of both variables did not provide further prognostic information. Conversely, an improved significance of the DFS prediction model at multivariate analysis was seen by adding Ki-67 (≥3 versus <3, P adj = 0.01) to TC and AC histological distinction, age, lymph node involvement, residual tumour and OTP. Ki-67 ≥ 3% plays a potentially pivotal role in LCT prognosis, irrespective of histological grade