Characterization of EN-1078D, a poorly differentiated human endometrial carcinoma cell line: a novel tool to study endometrial invasion in vitro

Background  

To date, tools to study metastasis in endometrial cancers are insufficiently developed. The aim of this study was to characterize the cell line EN-1078D, a new endometrial carcinoma cell line derived from a metastasis to the ovary.     

Elementary Energy Transfer Pathways in Allochromatium vinosum Photosynthetic Membranes

Allochromatium vinosum (formerly Chromatium vinosum) purple bacteria are known to adapt their light-harvesting strategy during growth according to environmental factors such as temperature and average light intensity. Under low light illumination or low ambient temperature conditions, most of the LH2 complexes in the photosynthetic membranes form a B820 exciton with reduced spectral overlap with LH1. To elucidate the reason for this light and temperature adaptation of the LH2 electronic structure, we performed broadband femtosecond transient absorption spectroscopy as a function of excitation wavelength in A. vinosum membranes. A target analysis of the acquired data yielded individual rate constants for all relevant elementary energy transfer (ET) processes. We found that the ET dynamics in high-light-grown membranes was well described by a homogeneous model, with forward and backward rate constants independent of the pump wavelength. Thus, the overall B800→B850→B890→ Reaction Center ET cascade is well described by simple triexponential kinetics. In the low-light-grown membranes, we found that the elementary backward transfer rate constant from B890 to B820 was strongly reduced compared with the corresponding constant from B890 to B850 in high-light-grown samples. The ET dynamics of low-light-grown membranes was strongly dependent on the pump wavelength, clearly showing that the excitation memory is not lost throughout the exciton lifetime. The observed pump energy dependence of the forward and backward ET rate constants suggests exciton diffusion via B850→ B850 transfer steps, making the overall ET dynamics nonexponential. Our results show that disorder plays a crucial role in our understanding of low-light adaptation in A. vinosum.     

Phosphate and ADP Differently Inhibit Coordinated Smooth Muscle Myosin Groups

Actin filaments propelled in vitro by groups of skeletal muscle myosin motors exhibit distinct phases of active sliding or arrest, whose occurrence depends on actin length (L) within a range of up to 1.0 μm. Smooth muscle myosin filaments are exponentially distributed with ≈150 nm average length in vivo—suggesting relevance of the L-dependence of myosin group kinetics. Here, we found L-dependent actin arrest and sliding in in vitro motility assays of smooth muscle myosin. We perturbed individual myosin kinetics with varying, physiological concentrations of phosphate (P_i, release associated with main power stroke) and adenosine diphosphate (ADP, release associated with minor mechanical step). Adenosine triphosphate was kept constant at physiological concentration. Increasing [P_i] lowered the fraction of time for which actin was actively sliding, reflected in reduced average sliding velocity (ν) and motile fraction (f_mot, fraction of time that filaments are moving); increasing [ADP] increased the fraction of time actively sliding and reduced the velocity while sliding, reflected in reduced ν and increased f_mot. We introduced specific P_i and ADP effects on individual myosin kinetics into our recently developed mathematical model of actin propulsion by myosin groups. Simulations matched our experimental observations and described the inhibition of myosin group kinetics. At low [P_i] and [ADP], actin arrest and sliding were reflected by two distinct chemical states of the myosin group. Upon [P_i] increase, the probability of the active state decreased; upon [ADP] increase, the probability of the active state increased, but the active state became increasingly similar to the arrested state.     

Treatment with a sphingosine analog after the inception of house dust mite-induced airway inflammation alleviates key features of experimental asthma

Background

In vivo phosphorylation of sphingosine analogs with their ensuing binding and activation of their cell-surface sphingosine-1-phosphate receptors is regarded as the main immunomodulatory mechanism of this new class of drugs. Prophylactic treatment with sphingosine analogs interferes with experimental asthma by impeding the migration of dendritic cells to draining lymph nodes. However, whether these drugs can also alleviate allergic airway inflammation after its onset remains to be determined. Herein, we investigated to which extent and by which mechanisms the sphingosine analog AAL-R interferes with key features of asthma in a murine model during ongoing allergic inflammation induced by Dermatophagoides pteronyssinus.

Methods

BALB/c mice were exposed to either D. pteronyssinus or saline, intranasally, once-daily for 10 consecutive days. Mice were treated intratracheally with either AAL-R, its pre-phosphorylated form AFD-R, or the vehicle before every allergen challenge over the last four days, i.e. after the onset of allergic airway inflammation. On day 11, airway responsiveness to methacholine was measured; inflammatory cells and cytokines were quantified in the airways; and the numbers and/or viability of T cells, B cells and dendritic cells were assessed in the lungs and draining lymph nodes.

Results

AAL-R decreased airway hyperresponsiveness induced by D. pteronyssinus by nearly 70%. This was associated with a strong reduction of IL-5 and IL-13 levels in the airways and with a decreased eosinophilic response. Notably, the lung CD4⁺ T cells were almost entirely eliminated by AAL-R, which concurred with enhanced apoptosis/necrosis in that cell population. This inhibition occurred in the absence of dendritic cell number modulation in draining lymph nodes. On the other hand, the pre-phosphorylated form AFD-R, which preferentially acts on cell-surface sphingosine-1-phosphate receptors, was relatively impotent at enhancing cell death, which led to a less efficient control of T cell and eosinophil responses in the lungs.

Conclusion

Airway delivery of the non-phosphorylated sphingosine analog, but not its pre-phosphorylated counterpart, is highly efficient at controlling the local T cell response after the onset of allergic airway inflammation. The mechanism appears to involve local induction of lymphocyte apoptosis/necrosis, while mildly affecting dendritic cell and T cell accumulation in draining lymph nodes.     

Substrate and inhibitor specificities differ between human cytosolic and mitochondrial thioredoxin reductases: Implications for development of specific inhibitors

The cytosolic and mitochondrial thioredoxin reductases (TrxR1 and TrxR2) and thioredoxins (Trx1 and Trx2) are key components of the mammalian thioredoxin system, which is important for antioxidant defense and redox regulation of cell function. TrxR1 and TrxR2 are selenoproteins generally considered to have comparable properties, but to be functionally separated by their different compartments. To compare their properties we expressed recombinant human TrxR1 and TrxR2 and determined their substrate specificities and inhibition by metal compounds. TrxR2 preferred its endogenous substrate Trx2 over Trx1, whereas TrxR1 efficiently reduced both Trx1 and Trx2. TrxR2 displayed strikingly lower activity with dithionitrobenzoic acid (DTNB), lipoamide, and the quinone substrate juglone compared to TrxR1, and TrxR2 could not reduce lipoic acid. However, Sec-deficient two-amino-acid-truncated TrxR2 was almost as efficient as full-length TrxR2 in the reduction of DTNB. We found that the gold(I) compound auranofin efficiently inhibited both full-length TrxR1 and TrxR2 and truncated TrxR2. In contrast, some newly synthesized gold(I) compounds and cisplatin inhibited only full-length TrxR1 or TrxR2 and not truncated TrxR2. Surprisingly, one gold(I) compound, [Au(d2pype)(2)]Cl, was a better inhibitor of TrxR1, whereas another, [(iPr(2)Im)(2)Au]Cl, mainly inhibited TrxR2. These compounds also inhibited TrxR activity in the cytoplasm and mitochondria of cells, but their cytotoxicity was not always dependent on the proapoptotic proteins Bax and Bak. In conclusion, this study reveals significant differences between human TrxR1 and TrxR2 in substrate specificity and metal compound inhibition in vitro and in cells, which may be exploited for development of specific TrxR1- or TrxR2-targeting drugs.     

Human diurnal preference and circadian rhythmicity are not associated with the CLOCK 3111C/T gene polymorphism

Genetic association studies of the CLOCK 3111C/T polymorphism and diurnal preference have yielded conflicting results since the first report that the 3111C allele was associated with eveningness. The goal of the present study was to investigate the association of this polymorphism with diurnal preference and circadian physiology in a group of 179 individuals, by comparing the frequency of the 3111C allele to diurnal preference, habitual sleep timing, circadian phase markers, and circadian period. We did not find a significant association between this allele and morningness/eveningness or any circadian marker.     

Signaling and metabolic predispositions linked to the colorectal cancer

The setting up and the progression of the colorectal cancer (CCR) follow a sequence of events that are spatio-temporally rigorously orchestrated. The failures that specifically target the signaling pathways responsible for the cancerization of the colorectal mucosa have been well described and among these it seems that a dysregulation of the Wnt/β-catenin pathway is involved in the triggering of near 90 % of the cases. It has been also described that several risk factors linked to metabolic disorders (feeding, insulin resistance, metabolic syndrome, etc.) predispose individuals to CCR but no rational explanations were given. We propose that, since it is implicated in the control of the insulin pathway among other actions, the nutritional sensor O-GlcNAcylation may be the element linking these metabolic disorders to CCR.     

Saponins from the traditional medicinal plant Momordica charantia stimulate insulin secretion in vitro

The antidiabetic activity of Momordica charantia (L.), Cucurbitaceae, a widely-used treatment for diabetes in a number of traditional medicine systems, was investigated in vitro. Antidiabetic activity has been reported for certain saponins isolated from M. charantia. In this study insulin secretion was measured in MIN6 β-cells incubated with an ethanol extract, saponin-rich fraction, and five purified saponins and cucurbitane triterpenoids from M. charantia, 3β,7β,25-trihydroxycucurbita-5,23(E)-dien-19-al (1), momordicine I (2), momordicine II (3), 3-hydroxycucurbita-5,24-dien-19-al-7,23-di-O-β-glucopyranoside (4), and kuguaglycoside G (5). Treatments were compared to incubation with high glucose (27 mM) and the insulin secretagogue, glipizide (50 μM). At 125 μg/ml, an LC-ToF-MS characterized saponin-rich fraction stimulated insulin secretion significantly more than the DMSO vehicle, p=0.02. At concentrations 10 and 25 μg/ml, compounds 3 and 5 also significantly stimulated insulin secretion as compared to the vehicle, p≤0.007, and p=0.002, respectively. This is the first report of a saponin-rich fraction, and isolated compounds from M. charantia, stimulating insulin secretion in an in vitro, static incubation assay.     

Comparative population genetic structure in a plant-pollinator/seed predator system

Comparative analyses of spatial genetic structure of populations of plants and the insects they interact with provide an indication of how gene flow, natural selection and genetic drift may jointly influence the distribution of genetic variation and potential for local co‐adaptation for interacting species. Here, we analysed the spatial scale of genetic structure within and among nine populations of an interacting species pair, the white campion Silene latifolia and the moth Hadena bicruris, along a latitudinal gradient across Northern/Central Europe. This dioecious, short‐lived perennial plant inhabits patchy, often disturbed environments. The moth H. bicruris acts both as its pollinator and specialist seed predator that reproduces by laying eggs in S. latifolia flowers. We used nine microsatellite markers for S. latifolia and eight newly developed markers for H. bicruris. We found high levels of inbreeding in most populations of both plant and pollinator/seed predator. Among populations, significant genetic structure was observed for S. latifolia but not for its pollinator/seed predator, suggesting that despite migration among populations of H. bicruris, pollen is not, or only rarely, carried over between populations, thus maintaining genetic structure among plant populations. There was a weak positive correlation between genetic distances of S. latifolia and H. bicruris. These results indicate that while significant structure of S. latifolia populations creates the potential for differentiation at traits relevant for the interaction with the pollinator/seed predator, substantial gene flow in H. bicruris may counteract this process in at least some populations.