Intensive nitrogen loss over the Omani Shelf due to anammox coupled with dissimilatory nitrite reduction to ammonium

A combination of stable isotopes (¹⁵N) and molecular ecological approaches was used to investigate the vertical distribution and mechanisms of biological N₂ production along a transect from the Omani coast to the central–northeastern (NE) Arabian Sea. The Arabian Sea harbors the thickest oxygen minimum zone (OMZ) in the world''s oceans, and is considered to be a major site of oceanic nitrogen (N) loss. Short (<48 h) anoxic incubations with ¹⁵N-labeled substrates and functional gene expression analyses showed that the anammox process was highly active, whereas denitrification was hardly detectable in the OMZ over the Omani shelf at least at the time of our sampling. Anammox was coupled with dissimilatory nitrite reduction to ammonium (DNRA), resulting in the production of double-¹⁵N-labeled N₂ from ¹⁵NO₂⁻, a signal often taken as the lone evidence for denitrification in the past. Although the central–NE Arabian Sea has conventionally been regarded as the primary N-loss region, low potential N-loss rates at sporadic depths were detected at best. N-loss activities in this region likely experience high spatiotemporal variabilities as linked to the availability of organic matter. Our finding of greater N-loss associated with the more productive Omani upwelling region is consistent with results from other major OMZs. The close reliance of anammox on DNRA also highlights the need to take into account the effects of coupling N-transformations on oceanic N-loss and subsequent N-balance estimates.     

Identification,biological characterization and pharmacophoric analysis of a new potent and selective NK1 receptor antagonist clinical candidate

The last two decades have provided a large weight of preclinical data implicating the neurokinin-1 receptor (NK₁) and its cognate ligand substance P (SP) in a broad range of both central and peripheral disease conditions. However, to date, only the NK₁ receptor antagonist aprepitant has been approved as a therapeutic and this is to prevent chemotherapy-induced nausea & vomiting (CINV). The belief remained that the full therapeutic potential of NK₁ receptor antagonists had yet to be realized; therefore clinical evidence that NK₁ receptor antagonists may be effective in major depression disorder, resulted in a significant further investment in discovering novel CNS penetrant druggable NK₁ receptor antagonists to address this condition. At GlaxoSmithKline after the discovery of casopitant, that went on to demonstrate efficacy as a novel antidepressant in the clinic, additional novel analogues of this NK₁ receptor antagonist were designed to further enhance its drug developability characteristics. Herein, we therefore describe the discovery process and the vivo pharmacological and pharmacokinetic profile of the new NK₁ receptor antagonist 3a (also called orvepitant), selected as clinical candidate and further progressed into clinical studies for major depressive disorder. Moreover, molecular modeling studies enabled us to improve the pharmacophore model of the NK₁ receptor antagonists with the identification of a region able to accommodate a variety of heterocycle moieties.     

Highly efficient production of anti-HER2 scFv antibody variant for targeting breast cancer cells

In the present study, cultivation conditions and medium components were optimized using statistical design and analysis to enhance the production of Chi21702, a cold-active extracellular chitinase from the Antarctic bacterium Sanguibacter antarcticus KOPRI 21702. Identification of significant carbon sources and other key elements was performed using a statistical design technique. Chitin and glycerol were selected as main carbon sources, and the ratio of complex nitrogen sources to carbon sources was determined to be 0.5. Among 15 mineral components included in basal medium, NaCl, Fe(C₆H₅O₇), and MgCl₂ were found to have the most influence on Chi21702 production. The optimal parameters of temperature, initial pH, and dissolved oxygen level were found to be 25°C, 6.5, and above 30% of air saturation, respectively. The maximum Chi21702 activity obtained under the optimized conditions was 90 U/L. Through statistical optimization methods, a 7.5-fold increase in Chi21702 production was achieved over unoptimized conditions. Chi21702 showed relatively high activity, even at low temperatures close to 0°C. The information obtained in the present study could be applied to the production of cold-active endochitinase on a large scale, suitable for a process at low temperature in industry.     

Down-regulation of zeta chain and zeta-associated protein 70 (Zap 70) expression in circulating T lymphocytes in laryngeal squamous cell carcinoma

OBJECTIVE: To evaluate zeta chain and Zap 70 expression in T lymphocytes of patients with laryngeal cancer in relation to surgical treatment. STUDY DESIGN: This study investigated, by dual-color flow cytometry, zeta chain and Zap 70 expression in the circulating T lymphocytes of 13 patients with laryngeal cancer patients before and after surgical treatment. RESULTS: Patients exhibited a significant lower expression of both zeta chain and Zap 70 compared to healthy normal controls; no statistical differences were observed after surgical treatment. CONCLUSION: The results of this investigation seem to indicate that both the zeta chain and the Zap 70 expression in circulating T lymphocytes are down-regulated in patients with laryngeal cancer and that these changes do not immediately return to normal after surgery. Flow cytometry analysis may represent an easy-to-use procedure for monitoring the immune status of patients with laryngeal cancer.     

Naphthoquinone-mediated Inhibition of Lysine Acetyltransferase KAT3B/p300, Basis for Non-toxic Inhibitor Synthesis

Hydroxynaphthoquinone-based inhibitors of the lysine acetyltransferase KAT3B (p300), such as plumbagin, are relatively toxic. Here, we report that free thiol reactivity and redox cycling properties greatly contribute to the toxicity of plumbagin. A reactive 3rd position in the naphthoquinone derivatives is essential for thiol reactivity and enhances redox cycling. Using this clue, we synthesized PTK1, harboring a methyl substitution at the 3rd position of plumbagin. This molecule loses its thiol reactivity completely and its redox cycling ability to a lesser extent. Mechanistically, non-competitive, reversible binding of the inhibitor to the lysine acetyltransferase (KAT) domain of p300 is largely responsible for the acetyltransferase inhibition. Remarkably, the modified inhibitor PTK1 was a nearly non-toxic inhibitor of p300. The present report elucidates the mechanism of acetyltransferase activity inhibition by 1,4-naphthoquinones, which involves redox cycling and nucleophilic adduct formation, and it suggests possible routes of synthesis of the non-toxic inhibitor.     

HIV-1 Subtype F1 Epidemiological Networks among Italian Heterosexual Males Are Associated with Introduction Events from South America

About 40% of the Italian HIV-1 epidemic due to non-B variants is sustained by F1 clade, which circulates at high prevalence in South America and Eastern Europe. Aim of this study was to define clade F1 origin, population dynamics and epidemiological networks through phylogenetic approaches. We analyzed pol sequences of 343 patients carrying F1 subtype stored in the ARCA database from 1998 to 2009. Citizenship of patients was as follows: 72.6% Italians, 9.3% South Americans and 7.3% Rumanians. Heterosexuals, Homo-bisexuals, Intravenous Drug Users accounted for 58.1%, 24.0% and 8.8% of patients, respectively. Phylogenetic analysis indicated that 70% of sequences clustered in 27 transmission networks. Two distinct groups were identified; the first clade, encompassing 56 sequences, included all Rumanian patients. The second group involved the remaining clusters and included 10 South American Homo-bisexuals in 9 distinct clusters. Heterosexual modality of infection was significantly associated with the probability to be detected in transmission networks. Heterosexuals were prevalent either among Italians (67.2%) or Rumanians (50%); by contrast, Homo-bisexuals accounted for 71.4% of South Americans. Among patients with resistant strains the proportion of clustering sequences was 57.1%, involving 14 clusters (51.8%). Resistance in clusters tended to be higher in South Americans (28.6%) compared to Italian (17.7%) and Rumanian patients (14.3%). A striking proportion of epidemiological networks could be identified in heterosexuals carrying F1 subtype residing in Italy. Italian Heterosexual males predominated within epidemiological clusters while foreign patients were mainly Heterosexual Rumanians, both males and females, and South American Homo-bisexuals. Tree topology suggested that F1 variant from South America gave rise to the Italian F1 epidemic through multiple introduction events. The contact tracing also revealed an unexpected burden of resistance in epidemiological clusters underlying the need of public interventions to limit the spread of non-B subtypes and transmitted drug resistance.     

Towards molecular computing: co-development of microfluidic devices and chemical reaction media

Microfluidics provides a powerful technology for both the production of molecular computing components and for the implementation of molecular computing architectures. The potential commercial applications of microfluidics drive rapid progress in this field-but at the same time focus interest on materials that are compatible with physiological aqueous conditions. For engineering applications that consider a broader range of physico-chemical conditions the narrow set of established materials for microfluidics can be a challenge. As a consequence of the large surface to volume ratio inherent in microfluidic technology the material of the device can greatly affect the chemistry in the channels of the device. In practice it is necessary to co-develop the chemical medium to be used in the device together with the microfluidic devices. We describe this process for a molecular computing architecture that makes use of excitable lipid-coated droplets of Belousov-Zhabotinsky reaction medium as its active processing components. We identify fluoropolymers with low melting temperature as a suitable substrate for microfluidics to be used in conjunction with Belousov-Zhabotinsky droplets in decane.     

Circulating cytokines present in the serum of peripheral arterial disease patients induce endothelial dysfunction

Peripheral arterial disease (PAD) is a chronic condition caused by atherosclerosis and is a severe complication of type 2 diabetes (T2D). We hypothesised that chronic condition of arterial disease engenders inflammation and endothelial damage in response to circulating cytokines released in the blood stream of PAD patients. We explored the levels of circulating cytokines in PAD patients with and without diabetes by multiplex cytokine array compared with non-PAD controls. Serum from PAD patients with or without diabetes showed high levels of VEGF, IFN-gamma, TNF-alpha, MCP-1, and EGF. VEGF levels correlated with TNF-alpha and IFN-gamma, significantly. Endothelial cells (ECs) were exposed to the different altered cytokines to evaluate changes in cell growth, migration and tubule-like formation, displaying impairment on proliferation, migration and tubule formation. Our findings demonstrate that a set of cytokines is significantly increased in the serum of PAD patients. These cytokines act to induce endothelial dysfunction synergistically. VEGF strongly correlated with TNF-alpha and IFN-gamma, opening new therapeutic perspectives.