Adaptation to Shift Work: Physiologically Based Modeling of the Effects of Lighting and Shifts’ Start Time

Shift work has become an integral part of our life with almost 20% of the population being involved in different shift schedules in developed countries. However, the atypical work times, especially the night shifts, are associated with reduced quality and quantity of sleep that leads to increase of sleepiness often culminating in accidents. It has been demonstrated that shift workers’ sleepiness can be improved by a proper scheduling of light exposure and optimizing shifts timing. Here, an integrated physiologically-based model of sleep-wake cycles is used to predict adaptation to shift work in different light conditions and for different shift start times for a schedule of four consecutive days of work. The integrated model combines a model of the ascending arousal system in the brain that controls the sleep-wake switch and a human circadian pacemaker model. To validate the application of the integrated model and demonstrate its utility, its dynamics are adjusted to achieve a fit to published experimental results showing adaptation of night shift workers (n = 8) in conditions of either bright or regular lighting. Further, the model is used to predict the shift workers’ adaptation to the same shift schedule, but for conditions not considered in the experiment. The model demonstrates that the intensity of shift light can be reduced fourfold from that used in the experiment and still produce good adaptation to night work. The model predicts that sleepiness of the workers during night shifts on a protocol with either bright or regular lighting can be significantly improved by starting the shift earlier in the night, e.g.; at 21∶00 instead of 00∶00. Finally, the study predicts that people of the same chronotype, i.e. with identical sleep times in normal conditions, can have drastically different responses to shift work depending on their intrinsic circadian and homeostatic parameters.     

Excision of 8-oxoguanine from methylated CpG dinucleotides by human 8-oxoguanine DNA glycosylase

CpG dinucleotides are targets for epigenetic methylation, many of them bearing 5-methylcytosine (mCyt) in the human genome. Guanine in this context can be easily oxidized to 8-oxoguanine (oxoGua), which is repaired by 8-oxoguanine-DNA glycosylase (OGG1). We have studied how methylation affects the efficiency of oxoGua excision from damaged CpG dinucleotides. Methylation of the adjacent cytosine moderately decreased the oxoGua excision rate while methylation opposite oxoGua lowered the rate of product release. Cytosine methylation abolished stimulation of OGG1 by repair endonuclease APEX1. The OGG1 S326C polymorphic variant associated with lung cancer showed poorer base excision and lost sensitivity to the opposite-base methylation. The overall repair in the system reconstituted from purified proteins decreased for CpG with mCyt in the damaged strand.     

Non-targeted Identification of Prions and Amyloid-forming Proteins from Yeast and Mammalian Cells

The formation of amyloid aggregates is implicated both as a primary cause of cellular degeneration in multiple human diseases and as a functional mechanism for providing extraordinary strength to large protein assemblies. The recent identification and characterization of several amyloid proteins from diverse organisms argues that the amyloid phenomenon is widespread in nature. Yet identifying new amyloid-forming proteins usually requires a priori knowledge of specific candidates. Amyloid fibers can resist heat, pressure, proteolysis, and denaturation by reagents such as urea or sodium dodecyl sulfate. Here we show that these properties can be exploited to identify naturally occurring amyloid-forming proteins directly from cell lysates. This proteomic-based approach utilizes a novel purification of amyloid aggregates followed by identification by mass spectrometry without the requirement for special genetic tools. We have validated this technique by blind identification of three amyloid-based yeast prions from laboratory and wild strains and disease-related polyglutamine proteins expressed in both yeast and mammalian cells. Furthermore, we found that polyglutamine aggregates specifically recruit some stress granule components, revealing a possible mechanism of toxicity. Therefore, core amyloid-forming proteins as well as strongly associated proteins can be identified directly from cells of diverse origin.     

The Local Dinucleotide Preference of APOBEC3G Can Be Altered from 5′-CC to 5′-TC by a Single Amino Acid Substitution

APOBEC3A and APOBEC3G are DNA cytosine deaminases with biological functions in foreign DNA and retrovirus restriction, respectively. APOBEC3A has an intrinsic preference for cytosine preceded by thymine (5′-TC) in single-stranded DNA substrates, whereas APOBEC3G prefers the target cytosine to be preceded by another cytosine (5′-CC). To determine the amino acids responsible for these strong dinucleotide preferences, we analyzed a series of chimeras in which putative DNA binding loop regions of APOBEC3G were replaced with the corresponding regions from APOBEC3A. Loop 3 replacement enhanced APOBEC3G catalytic activity but did not alter its intrinsic 5′-CC dinucleotide substrate preference. Loop 7 replacement caused APOBEC3G to become APOBEC3A-like and strongly prefer 5′-TC substrates. Simultaneous loop 3/7 replacement resulted in a hyperactive APOBEC3G variant that also preferred 5′-TC dinucleotides. Single amino acid exchanges revealed D317 as a critical determinant of dinucleotide substrate specificity. Multi-copy explicitly solvated all-atom molecular dynamics simulations suggested a model in which D317 acts as a helix-capping residue by constraining the mobility of loop 7, forming a novel binding pocket that favorably accommodates cytosine. All catalytically active APOBEC3G variants, regardless of dinucleotide preference, retained human immunodeficiency virus type 1 restriction activity. These data support a model in which the loop 7 region governs the selection of local dinucleotide substrates for deamination but is unlikely to be part of the higher level targeting mechanisms that direct these enzymes to biological substrates such as human immunodeficiency virus type 1 cDNA.     

How to measure top–down vs bottom–up effects: a new population metric and its calibration on Daphnia

Research on the role of top–down (predation) and bottom–up (food) effects in food webs has led to the understanding that the variability of these effects in space and time is a fundamental feature of natural systems. Consequently, our measurement tools must allow us to evaluate the effects from a dynamical perspective. A population‐dynamics approach may be appropriate to the task. More specifically, because food and predators both affect birth rate, birth rate dynamics may be a key to understanding their impact on the population of interest. Based on the Edmondson–Paloheimo model for birth rate, we propose a new population metric to assess the relative strength of top–down vs bottom–up effects. The metric is the ratio of contributions of changes in proportion of adults and fecundity to change in birth rate. Proportion of adults reflects a top–down effect (predators are assumed to be size‐selective), fecundity reflects a bottom–up effect, and birth rate appears as a common currency with which to compare the former and the latter. Using microcosm experiments and computer simulations on the cladoceran Daphnia, we calibrate the metric and show that, in both types of tests, the ratio of contributions is typically 0.5–0.7 under a strong bottom–up effect and 2.0–2.2 under a strong top–down effect. This provides experimental evidence that the ratio of contributions may allow one to distinguish a strong top–down effect from a strong bottom–up effect.     

The antileukemic activity of modified fibrinogen–methotrexate conjugate

Background

The search for new, innovative methods to treat all types of diseases, especially cancer-related ones, is a challenge taken by pharmaceutical companies and academic institutions. The use of conjugates containing widely-known and widely-used bioactive substances is one of the ways to solve this problem. Research into drug binding with macromolecular carrier systems has joined the search for new therapeutic strategies.

Methods

The main goal of this paper is the potential offered by the use of fibrinogen derivatives as an antileukemic drug carrier. Physicochemical properties of the obtained conjugate were analyzed, characterizing alterations in relation to the starting carrier and analyzing biological implications. The intraperitoneally (i.p.) inoculated P388 mouse leukemia model for in vivo studies was used.

Results and conclusions

Conjugates consisting of a fibrinogen derivative with a covalently bound anticancer drug were developed. Carrier preparation and a conjugate synthesis in aqueous solution were formulated, as well as purification of the conjugate was performed. The study showed that the survival of leukemia mice treated with FH–MTX conjugate was indeed significantly longer than survival in both untreated animals (control) and mice treated with unbound MTX. A significant increase in the antileukemic activity of MTX conjugated with hydrolysed fibrinogen was observed as compared with the unconjugated drug. Reported data suggest that hydrolysed fibrinogen can serve as a carrier molecule for the MTX drug with the aim of enhancing its antileukemic activity.

General significance

Conjugates consisting of a fibrinogen derivative with a covalently bound anticancer drug seem to be a promising anticancer drug.     

Therapeutic regulation of myeloid-derived suppressor cells and immune response to cancer vaccine in patients with extensive stage small cell lung cancer

Myeloid-derived suppressor cells (MDSC) are one of the major factors limiting the efficacy of immune therapy. In a clinical trial of patients with extensive stage small cell lung cancer (SCLC), we tested the possibility that targeting MDSC can improve the induction of immune responses by a cancer vaccine. Forty-one patients with extensive stage SCLC were randomized into three arms: arm A—control, arm B—vaccination with dendritic cells transduced with wild-type p53, and arm C—vaccination in combination with MDSC targeted therapy with all-trans-retinoic acid (ATRA). Interim results of the ongoing clinical trial are presented. Pre-treatment levels of MDSC populations in patients from all three arms were similar. Vaccine alone did not affect the proportion of MDSC, whereas in patients treated with ATRA, the MDSC decreased more than twofold (p = 0.02). Before the start of treatment, no patients had detectable p53-specific responses in IFN-γ ELISPOT. Sequential measurements did not show positive p53 responses in any of the 14 patients from arm A. After immunization, only 3 out of 15 patients (20 %) from arm B developed a p53-specific response (p = 0.22). In contrast, in arm C, 5 out of 12 patients (41.7 %) had detectable p53 responses (p = 0.012). The proportion of granzyme B-positive CD8⁺ T cells was increased only in patients from arm C but not in arm B. Depletion of MDSC substantially improved the immune response to vaccination, suggesting that this approach can be used to enhance the effect of immune interventions in cancer.     

A new species of Aspidistra (Ruscaceae s.l., Asparagales) from southern Vietnam, field observations on its flowering and possible pollination by flies (Phoridae)

The genus Aspidistra with almost 100 species is the most diverse in southern China and northern Vietnam. We describe a new species Aspidistra phanluongii N.Vislobokov from southern Vietnam. Species of Aspidistra are herbs characteristic of Southeast Asian forests. Flowering of these plants can be considered cryptic. There are no recorded field observations on phenology, biology of flowering or pollination of Aspidistra. Previously proposed pollinators of Aspidistra include collembolans, amphipods, fungus gnats (Diptera: Mycetophilidae, Sciaridae) and even slugs. We present observation data on the flowering of A. phanluongii in the wild. Flowers were visited by flies of Megaselia (Phoridae) and ants. Megaselia flies, well-known pollinators of numerous tropical plant species, are shown to be likely pollinators of Aspidistra phanluongii. The present study does not provide evidence of pollination by fungus gnats.