Checks and Balances between Autophagy and Inflammasomes during Infection

Autophagy and inflammasome complex assembly are physiological processes that control homeostasis, inflammation, and immunity. Autophagy is a ubiquitous pathway that degrades cytosolic macromolecules or organelles, as well as intracellular pathogens. Inflammasomes are multi-protein complexes that assemble in the cytosol of cells upon detection of pathogen- or danger-associated molecular patterns. A critical outcome of inflammasome assembly is the activation of the cysteine protease caspase-1, which activates the pro-inflammatory cytokine precursors pro-IL-1β and pro-IL-18. Studies on chronic inflammatory diseases, heart diseases, Alzheimer's disease, and multiple sclerosis revealed that autophagy and inflammasomes intersect and regulate each other. In the context of infectious diseases, however, less is known about the interplay between autophagy and inflammasome assembly, although it is becoming evident that pathogens have evolved multiple strategies to inhibit and/or subvert these pathways and to take advantage of their intricate crosstalk. An improved appreciation of these pathways and their subversion by diverse pathogens is expected to help in the design of anti-infective therapeutic interventions.     

Structure-based design and synthesis of macrocyclic human rhinovirus 3C protease inhibitors

The design and synthesis of macrocyclic inhibitors of human rhinovirus 3C protease is described. A macrocyclic linkage of the P1 and P3 residues, and the subsequent structure-based optimization of the macrocycle conformation and size led to the identification of a potent biochemical inhibitor 10 with sub-micromolar antiviral activity.     

Coping with shift work-related circadian disruption: A mixed-methods case study on napping and caffeine use in Australian nurses and midwives

ABSTRACT

Introduction: Two of the most ubiquitous fatigue countermeasures used by shift-working nurses are napping and caffeine. This mixed-methods case study investigated the ways nurses and midwives utilised napping and caffeine countermeasures to cope with shift work, and associated sleep, physical health and psychological health outcomes.

Materials and Methods: N = 130 Australian shift-working nurses and midwives (mean age = 44 years, range = 21–67, 115F, 15M) completed the Standard Shiftwork Index. A sub-set of 22 nurses and midwives completed an in-depth interview.

Results: Nearly 70% of participants reported napping. Those who napped during night shifts had significantly less total sleep time before (F_2,75 = 5.5, p < 0.01) and between days off (F_2,82 = 3.9, p < 0.05). By the end of the night shift, average hours of time awake were significantly less for prophylactic and on-shift nappers compared to non-nappers (F_2,85 = 97.2, p < 0.001). Since starting shift work, the percentage of high caffeine consumers (>400 mg/day) increased from 15% to 33% of the sample and an average of 4 (SD = 2) caffeinated beverages per day was reported. Increased caffeine consumption was associated with greater sleep disturbance (r = 0.26, p < 0.05), psychological distress (r = 0.37, p < 0.001), abdomen pain (r = 0.27, p < 0.05) and weight gain since starting shift work (r = 0.25, p < 0.05). Interviews confirmed these relationships and revealed that caffeine consumption on night shift was common, whereas napping on night shift was dependent on a number of factors including ability to sleep during the day.

Conclusion: This study identified reasons shift workers chose to engage in or abstain from napping and consuming caffeine, and how these strategies related to poor sleep and health outcomes. Further research is required to help develop recommendations for shift workers regarding napping and caffeine consumption as fatigue countermeasures, whilst taking into account the associated hazards of each strategy.     

A validation study of Fitbit Charge 2™ compared with polysomnography in adults

We evaluated the performance of a consumer multi-sensory wristband (Fitbit Charge 2?), against polysomnography (PSG) in measuring sleep/wake state and sleep stage composition in healthy adults.

In-lab PSG and Fitbit Charge 2? data were obtained from a single overnight recording at the SRI Human Sleep Research Laboratory in 44 adults (19—61 years; 26 women; 25 Caucasian). Participants were screened to be free from mental and medical conditions. Presence of sleep disorders was evaluated with clinical PSG. PSG findings indicated periodic limb movement of sleep (PLMS, > 15/h) in nine participants, who were analyzed separately from the main group (n = 35). PSG and Fitbit Charge 2? sleep data were compared using paired t-tests, Bland–Altman plots, and epoch-by-epoch (EBE) analysis.

In the main group, Fitbit Charge 2? showed 0.96 sensitivity (accuracy to detect sleep), 0.61 specificity (accuracy to detect wake), 0.81 accuracy in detecting N1+N2 sleep (“light sleep”), 0.49 accuracy in detecting N3 sleep (“deep sleep”), and 0.74 accuracy in detecting rapid-eye-movement (REM) sleep. Fitbit Charge 2? significantly (p < 0.05) overestimated PSG TST by 9 min, N1+N2 sleep by 34 min, and underestimated PSG SOL by 4 min and N3 sleep by 24 min. PSG and Fitbit Charge 2? outcomes did not differ for WASO and time spent in REM sleep. No more than two participants fell outside the Bland–Altman agreement limits for all sleep measures. Fitbit Charge 2? correctly identified 82% of PSG-defined non-REM–REM sleep cycles across the night. Similar outcomes were found for the PLMS group.

Fitbit Charge 2? shows promise in detecting sleep-wake states and sleep stage composition relative to gold standard PSG, particularly in the estimation of REM sleep, but with limitations in N3 detection. Fitbit Charge 2? accuracy and reliability need to be further investigated in different settings (at-home, multiple nights) and in different populations in which sleep composition is known to vary (adolescents, elderly, patients with sleep disorders).     

Evaluation of promoter sequences for the secretory production of a Clostridium thermocellum cellulase in Paenibacillus polymyxa

Due to their high secretion capacity, Gram-positive bacteria from the genus Bacillus are important expression hosts for the high-yield production of enzymes in industrial biotechnology; however, to date, strains from only few Bacillus species are used for enzyme production at industrial scale. Herein, we introduce Paenibacillus polymyxa DSM 292, a member of a different genus, as a novel host for secretory protein production. The model gene cel8A from Clostridium thermocellum was chosen as an easily detectable reporter gene with industrial relevance to demonstrate heterologous expression and secretion in P. polymyxa. The yield of the secreted cellulase Cel8A protein was increased by optimizing the expression medium and testing several promoter sequences in the expression plasmid pBACOV. Quantitative mass spectrometry was used to analyze the secretome in order to identify promising new promoter sequences from the P. polymyxa genome itself. The most abundantly secreted host proteins were identified, and the promoters regulating the expression of their corresponding genes were selected. Eleven promoter sequences were cloned and tested, including well-characterized promoters from Bacillus subtilis and Bacillus megaterium. The best result was achieved with the promoter for the hypothetical protein PPOLYM_03468 from P. polymyxa. In combination with the optimized expression medium, this promoter enabled the production of 5475 U/l of Cel8A, which represents a 6.2-fold increase compared to the reference promoter P_aprE. The set of promoters described in this work covers a broad range of promoter strengths useful for heterologous expression in the new host P. polymyxa.