Activation of GPER-1 estradiol receptor downregulates production of testosterone in isolated rat Leydig cells and adult human testis.

PURPOSE: Estradiol (E2) modulates testicular functions including steroidogenesis, but the mechanisms of E2 signaling in human testis are poorly understood. GPER-1 (GPR30), a G protein-coupled membrane receptor, mediates rapid genomic and non-genomic response to estrogens. The aim of this study was to evaluate GPER-1 expression in the testis, and its role in estradiol dependent regulation of steroidogenesis in isolated rat Leydig cells and human testis. MATERIALS AND METHODS: Isolated Leydig cells (LC) from adult rats and human testicular tissue were used in this study. Expression and localization studies of GPER-1 were performed with qRT-PCR, immunofluorescence, immunohistochemistry and Western Blot. Luteinizing Hormone (LH) -stimulated, isolated LC were incubated with estradiol, G-1 (GPER-1-selective agonist), and estrogen receptor antagonist ICI 182,780. Testosterone production was measured with radioimmunoassay. LC viability after incubation with G-1 was measured using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay. RESULTS: GPER-1 mRNA is abundantly expressed in rat LC and human testis. Co-localization experiments showed high expression levels of GPER-1 protein in LC. E2-dependent activation of GPER-1 lowers testosterone production in isolated rats LCs and in human testis, with statistically and clinically significant drops in testosterone production by 20-30% as compared to estradiol-naïve LC. The exposure to G-1 does not affect viability of isolated LCs. CONCLUSIONS: Our results indicate that activation of GPER-1 lowers testosterone levels in the rat and human testis. The expression of GPER-1 in human testis, which lack ERα, makes it an exciting target for developing new agents affecting testosterone production in men

Architecture of the nuclear pore complex coat

The nuclear pore complex (NPC) constitutes the sole gateway for bidirectional nucleocytoplasmic transport. Despite half a century of structural characterization, the architecture of the NPC remains unknown. Here, we present the crystal structure of a reconstituted ~400 kDa coat nucleoporin complex (CNC) from S. cerevisiae at a 7.4-Å resolution. The crystal structure revealed a curved Y-shaped architecture and the molecular details of the coat nucleoporin interactions forming the central “triskelion” of the Y. A structural comparison of the yeast CNC with an electron microscopy reconstruction of its human counterpart suggested the evolutionary conservation of the elucidated architecture. Moreover, 32 copies of the CNC crystal structure docked readily into a cryoelectron tomographic reconstruction of the fully-assembled human NPC, thereby accounting for ~16 MDa of its mass

Recognition of an α-helical hairpin in P22 large terminase by a synthetic antibody fragment.

The genome-packaging motor of tailed bacteriophages and herpesviruses is a multisubunit protein complex formed by several copies of a large (TerL) and a small (TerS) terminase subunit. The motor assembles transiently at the portal protein vertex of an empty precursor capsid to power the energy-dependent packaging of viral DNA. Both the ATPase and nuclease activities associated with genome packaging reside in TerL. Structural studies of TerL from bacteriophage P22 have been hindered by the conformational flexibility of this enzyme and its susceptibility to proteolysis. Here, an unbiased, synthetic phage-display Fab library was screened and a panel of high-affinity Fabs against P22 TerL were identified. This led to the discovery of a recombinant antibody fragment, Fab4, that binds a 33-amino-acid α-helical hairpin at the N-terminus of TerL with an equilibrium dissociation constant Kd of 71.5 nM. A 1.51 Å resolution crystal structure of Fab4 bound to the TerL epitope (TLE) together with a 1.15 Å resolution crystal structure of the unliganded Fab4, which is the highest resolution ever achieved for a Fab, elucidate the principles governing the recognition of this novel helical epitope. TLE adopts two different conformations in the asymmetric unit and buries as much as 1250 Å2 of solvent-accessible surface in Fab4. TLE recognition is primarily mediated by conformational changes in the third complementarity-determining region of the Fab4 heavy chain (CDR H3) that take place upon epitope binding. It is demonstrated that TLE can be introduced genetically at the N-terminus of a target protein, where it retains high-affinity binding to Fab4

Ozone-based eye drops activity on ocular epithelial cells and potential pathogens infecting the front of the eye

Confirmation of the biological effectiveness of new ophthalmic preparations introduced in the market is an important element in maintaining the safety of using this type of medications. This study aimed to investigate the activity of Ozodrop® on human corneal and conjunctival epithelial cells, as well as its antibacterial and antifungal activity. Cytotoxicity analyses of ocular surface epithelial cells were performed in vitro by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide) and Neutral Red uptake assays. The level of nitric oxide released by the cells was assessed by the Griess method. The reduction of the DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical by the tested formulation was analyzed. Microbiological tests were also performed. It was found that the Ozodrop® preparation exhibited biological activity, but was less active than the reference antibiotics and the anti-yeast agent. The cytotoxic activity of the Ozodrop® formulation was dependent on the time of cell exposure to it. No toxic effect was observed in the short-term, for up to 3 h. It appeared after 24 h of exposure of the cells to the preparation. The drops showed antioxidant activity in the specified concentration range. They also stimulated the release of nitric oxide, mainly by corneal epithelial cells. The Ozodrop® formulation exhibits biological activity that can be considered useful in the treatment of infections in the front part of the eye

Dense Plasma Focus: physics and applications (radiation material science, single-shot disclosure of hidden illegal objects, radiation biology and medicine, etc.)

The paper presents some outcomes obtained during the year of 2013 of the activity in the frame of the International Atomic Energy Agency Co-ordinated research project "Investigations of Materials under High Repetition and Intense Fusion-Relevant Pulses". The main results are related to the effects created at the interaction of powerful pulses of different types of radiation (soft and hard X-rays, hot plasma and fast ion streams, neutrons, etc. generated in Dense Plasma Focus (DPF) facilities) with various materials including those that are counted as perspective ones for their use in future thermonuclear reactors. Besides we discuss phenomena observed at the irradiation of biological test objects. We examine possible applications of nanosecond powerful pulses of neutrons to the aims of nuclear medicine and for disclosure of hidden illegal objects. Special attention is devoted to discussions of a possibility to create extremely large and enormously diminutive DPF devices and probabilities of their use in energetics, medicine and modern electronics

Architecture of the fungal nuclear pore inner ring complex

The nuclear pore complex (NPC) constitutes the sole gateway for bidirectional nucleocytoplasmic transport. We present the reconstitution and interdisciplinary analyses of the ~425-kDa inner ring complex (IRC), which forms the central transport channel and diffusion barrier of the NPC, revealing its interaction network and equimolar stoichiometry. The Nsp1•Nup49•Nup57 channel nucleoporin hetero-trimer (CNT) attaches to the IRC solely through the adaptor nucleoporin Nic96. The CNT•Nic96 structure reveals that Nic96 functions as an assembly sensor that recognizes the three dimensional architecture of the CNT, thereby mediating the incorporation of a defined CNT state into the NPC. We propose that the IRC adopts a relatively rigid scaffold that recruits the CNT to primarily form the diffusion barrier of the NPC, rather than enabling channel dilation

Recent results of studies of magnetic field distribution and neutron scaling on PF-1000 and PF-3 facilities

The recent results of studies of the magnetic field distribution and the neutron yield scaling in two largest plasma focus facilities, PF-3 and PF-1000 is done. The power-law dependence of the neutron yield on the current in the imploding plasma sheath has been demonstrated experimentally. For the first time the presence of the Вz magnetic field components is experimentally shown. In the compression stage, the axial component of the magnetic field reaches several kG that comprises ~10 % of the azimuthal component. The presence of the Bz field is a powerful argument in favor of the existence of closed magnetic configurations, which play an important role in the generating of neutrons.Представлены результаты последних исследований распределения магнитного поля и скейлинга нейтронного выхода на двух крупнейших плазмофокусных установках ПФ-3 и ПФ-1000. Экспериментально показана степенная зависимость нейтронного выхода от величины тока в сжимающейся плазменной оболочке. Впервые экспериментально показано наличие Вz-компоненты магнитного поля. В стадии сжатия величина аксиальной компоненты магнитного поля достигает нескольких килограмм-сил, что составляет ~ 10% от величины азимутальной компоненты. Наличие Bz-поля является весомым аргументом в пользу существования замкнутых магнитных конфигураций, играющих важную роль в механизме генерации нейтронов.Представлено результати останніх досліджень розподілу магнітного поля і скейлінга нейтронного вихoдy на двох найбільших плазмофокусних установках ПФ-3 та ПФ-1000. Експериментально показана ступенева залежність нейтронного виходу від величини струму в плазмовїй оболонці, що стикається. Вперше експериментально показано наявність Вz-компоненти магнітного поля. У стадії стиснення величина аксіальної компоненти магнітного поля досягає декількох кілограм-сил, що складає ~ 10 % від величини азимутальної компоненти. Наявність Вz-поля є вагомим аргументом на користь існування замкнутих магнітних конфігурацій, що мають неаби яку роль у механізмі генерації нейтронів