Effects of User Experience in Automated Information Processing on Perceived Usefulness of Digital Contact-Tracing Apps: Cross-Sectional Survey Study.

BACKGROUND: In pandemic situations, digital contact tracing (DCT) can be an effective way to assess one's risk of infection and inform others in case of infection. DCT apps can support the information gathering and analysis processes of users aiming to trace contacts. However, users' use intention and use of DCT information may depend on the perceived benefits of contact tracing. While existing research has examined acceptance in DCT, automation-related user experience factors have been overlooked. OBJECTIVE: We pursued three goals: (1) to analyze how automation-related user experience (ie, perceived trustworthiness, traceability, and usefulness) relates to user behavior toward a DCT app, (2) to contextualize these effects with health behavior factors (ie, threat appraisal and moral obligation), and (3) to collect qualitative data on user demands for improved DCT communication. METHODS: Survey data were collected from 317 users of a nationwide-distributed DCT app during the COVID-19 pandemic after it had been in app stores for >1 year using a web-based convenience sample. We assessed automation-related user experience. In addition, we assessed threat appraisal and moral obligation regarding DCT use to estimate a partial least squares structural equation model predicting use intention. To provide practical steps to improve the user experience, we surveyed users' needs for improved communication of information via the app and analyzed their responses using thematic analysis. RESULTS: Data validity and perceived usefulness showed a significant correlation of r=0.38 (P<.001), goal congruity and perceived usefulness correlated at r=0.47 (P<.001), and result diagnosticity and perceived usefulness had a strong correlation of r=0.56 (P<.001). In addition, a correlation of r=0.35 (P<.001) was observed between Subjective Information Processing Awareness and perceived usefulness, suggesting that automation-related changes might influence the perceived utility of DCT. Finally, a moderate positive correlation of r=0.47 (P<.001) was found between perceived usefulness and use intention, highlighting the connection between user experience variables and use intention. Partial least squares structural equation modeling explained 55.6% of the variance in use intention, with the strongest direct predictor being perceived trustworthiness (ß=.54; P<.001) followed by moral obligation (ß=.22; P<.001). Based on the qualitative data, users mainly demanded more detailed information about contacts (eg, place and time of contact). They also wanted to share information (eg, whether they wore a mask) to improve the accuracy and diagnosticity of risk calculation. CONCLUSIONS: The perceived result diagnosticity of DCT apps is crucial for perceived trustworthiness and use intention. By designing for high diagnosticity for the user, DCT apps could improve their support in the action regulation of users, resulting in higher perceived trustworthiness and use in pandemic situations. In general, automation-related user experience has greater importance for use intention than general health behavior or experience.

Energy Consumption Displays in Electric Vehicles: Differential Effects on Estimating Consumption and Experienced Energy Dynamics Awareness.

OBJECTIVE: The effects of three prototypical designs of energy consumption displays on energy-specific situation awareness were examined. BACKGROUND: Energy efficiency is crucial for the sustainability of technical systems. However, without accurate situation awareness of energy dynamics (energy dynamics awareness, EDA) it can be challenging for humans to optimize the use of energy resources of electric vehicles (EVs) through their behavior. METHOD: We examined three prototypical energy display designs that varied by their informational value to support EDA. Furthermore, we investigated the differential effects on EDA measured by (1) a newly constructed scale (experienced EDA), (2) estimating energy consumption, and (3) identifying efficient trips in an online experiment. Participants (N = 82) watched standardized driving scenes (videos) of EV trips presenting the energy displays. RESULTS: We found a strong effect of display type on experienced EDA, with the trace display being the most supportive. The EDA scale showed excellent internal consistency. The consumption estimation and efficient trip identification indicators were not affected by the display type. CONCLUSION: The study indicates that experienced EDA is immediately affected by displays with higher information value, but performance might need more time and training. More research is needed to investigate the cognitive processes related to EDA and to examine how distinct display elements enhance EDA. APPLICATION: Results from this research can be used as guidance for the design of energy displays, especially in EVs. The EDA scale can be used as an evaluation measure in the human-centered design process of energy displays.

AI based image analysis of red blood cells in oscillating microchannels.

The flow dynamics of red blood cells in vivo in blood capillaries and in vitro in microfluidic channels is complex. Cells can obtain different shapes such as discoid, parachute, slipper-like shapes and various intermediate states depending on flow conditions and their viscoelastic properties. We use artificial intelligence based analysis of red blood cells (RBCs) in an oscillating microchannel to distinguish healthy red blood cells from red blood cells treated with formaldehyde to chemically modify their viscoelastic behavior. We used TensorFlow to train and validate a deep learning model and achieved a testing accuracy of over 97%. This method is a first step to a non-invasive, label-free characterization of diseased red blood cells and will be useful for diagnostic purposes in haematology labs. This method provides quantitative data on the number of affected cells based on single cell classification.

Advances, Challenges, and Future Perspectives of Microwave Reflectometry for Plasma Position and Shape Control on Future Nuclear Fusion Devices.

Providing energy from fusion and finding ways to scale up the fusion process to commercial proportions in an efficient, economical, and environmentally benign way is one of the grand challenges for engineering. Controlling the burning plasma in real-time is one of the critical issues that need to be addressed. Plasma Position Reflectometry (PPR) is expected to have an important role in next-generation fusion machines, such as DEMO, as a diagnostic to monitor the position and shape of the plasma continuously, complementing magnetic diagnostics. The reflectometry diagnostic uses radar science methods in the microwave and millimetre wave frequency ranges and is envisaged to measure the radial edge density profile at several poloidal angles providing data for the feedback control of the plasma position and shape. While significant steps have already been given to accomplish that goal, with proof of concept tested first in ASDEX-Upgrade and afterward in COMPASS, important, ground-breaking work is still ongoing. The Divertor Test Tokamak (DTT) facility presents itself as the appropriate future fusion device to implement, develop, and test a PPR system, thus contributing to building a knowledge database in plasma position reflectometry required for its application in DEMO. At DEMO, the PPR diagnostic's in-vessel antennas and waveguides, as well as the magnetic diagnostics, may be exposed to neutron irradiation fluences 5 to 50 times greater than those experienced by ITER. In the event of failure of either the magnetic or microwave diagnostics, the equilibrium control of the DEMO plasma may be jeopardized. It is, therefore, imperative to ensure that these systems are designed in such a way that they can be replaced if necessary. To perform reflectometry measurements at the 16 envisaged poloidal locations in DEMO, plasma-facing antennas and waveguides are needed to route the microwaves between the plasma through the DEMO upper ports (UPs) to the diagnostic hall. The main integration approach for this diagnostic is to incorporate these groups of antennas and waveguides into a diagnostics slim cassette (DSC), which is a dedicated complete poloidal segment specifically designed to be integrated with the water-cooled lithium lead (WCLL) breeding blanket system. This contribution presents the multiple engineering and physics challenges addressed while designing reflectometry diagnostics using radio science techniques. Namely, short-range dedicated radars for plasma position and shape control in future fusion experiments, the advances enabled by the designs for ITER and DEMO, and the future perspectives. One key development is in electronics, aiming at an advanced compact coherent fast frequency sweeping RF back-end [23-100 GHz in few µs] that is being developed at IPFN-IST using commercial Monolithic Microwave Integrated Circuits (MMIC). The compactness of this back-end design is crucial for the successful integration of many measurement channels in the reduced space available in future fusion machines. Prototype tests of these devices are foreseen to be performed in current nuclear fusion machines.

The DZHK research platform: maximisation of scientific value by enabling access to health data and biological samples collected in cardiovascular clinical studies.

The German Centre for Cardiovascular Research (DZHK) is one of the German Centres for Health Research and aims to conduct early and guideline-relevant studies to develop new therapies and diagnostics that impact the lives of people with cardiovascular disease. Therefore, DZHK members designed a collaboratively organised and integrated research platform connecting all sites and partners. The overarching objectives of the research platform are the standardisation of prospective data and biological sample collections among all studies and the development of a sustainable centrally standardised storage in compliance with general legal regulations and the FAIR principles. The main elements of the DZHK infrastructure are web-based and central units for data management, LIMS, IDMS, and transfer office, embedded in a framework consisting of the DZHK Use and Access Policy, and the Ethics and Data Protection Concept. This framework is characterised by a modular design allowing a high standardisation across all studies. For studies that require even tighter criteria additional quality levels are defined. In addition, the Public Open Data strategy is an important focus of DZHK. The DZHK operates as one legal entity holding all rights of data and biological sample usage, according to the DZHK Use and Access Policy. All DZHK studies collect a basic set of data and biosamples, accompanied by specific clinical and imaging data and biobanking. The DZHK infrastructure was constructed by scientists with the focus on the needs of scientists conducting clinical studies. Through this, the DZHK enables the interdisciplinary and multiple use of data and biological samples by scientists inside and outside the DZHK. So far, 27 DZHK studies recruited well over 11,200 participants suffering from major cardiovascular disorders such as myocardial infarction or heart failure. Currently, data and samples of five DZHK studies of the DZHK Heart Bank can be applied for.

Acoustic sorting of microfluidic droplets at kHz rates using optical absorbance.

Droplet microfluidics allows one to address the ever-increasing demand to screen large libraries of biological samples. Absorbance spectroscopy complements the golden standard of fluorescence detection by label free target identification and providing more quantifiable data. However, this is limited by speed and sensitivity. In this paper we increase the speed of sorting by including acoustofluidics, achieving sorting rates of target droplets of 1 kHz. We improved the device design for detection of absorbance using fibre-based interrogation of samples with integrated lenses in the microfluidic PDMS device for focusing and collimation of light. This optical improvement reduces the scattering and refraction artefacts, improving the signal quality and sensitivity. The novel design allows us to overcome limitations based on dielectrophoresis sorting, such as droplet size dependency, material and dielectric properties of samples. Our acoustic activated absorbance sorter removes the need for offset dyes or matching oils and sorts about a magnitude faster than current absorbance sorters.

Active single cell encapsulation using SAW overcoming the limitations of Poisson distribution.

We demonstrate the use of an acoustic device to actively encapsulate single red blood cells into individual droplets in a T-junction. We compare the active encapsulation with the passive encapsulation depending on the number of loaded cells as well as the created droplet volumes. This method overcomes the Poisson limitation statistical loading of cells for the passive encapsulation. In our experiments we reach a single cell encapsulation efficiency of 97.9 ± 2.1% at droplet formation rates exceeding 15 Hz.

Barriers to and Facilitators for Using Nutrition Apps: Systematic Review and Conceptual Framework.

BACKGROUND: Nutrition apps are effective in changing eating behavior and diet-related health risk factors. However, while they may curb growing overweight and obesity rates, widespread adoption is yet to be achieved. Hence, profound knowledge regarding factors motivating and hindering (long-term) nutrition app use is crucial for developing design guidelines aimed at supporting uptake and prolonged use of nutrition apps. OBJECTIVE: In this systematic review, we synthesized the literature on barriers to and facilitators for nutrition app use across disciplines including empirical qualitative and quantitative studies with current users, ex-users, and nonusers of nutrition apps. METHODS: A systematic literature search including 6 databases (PubMed, Web of Science, PsychINFO, PSYNDEX, PsycArticles, and SPORTDiscus) as well as backward and forward citation search was conducted. Search strategy, inclusion and exclusion criteria, and the planned data extraction process were preregistered. All empirical qualitative and quantitative studies published in German or English were eligible for inclusion if they examined adolescents (aged 13-18) or adults who were either current users, ex-users, and nonusers of nutrition apps. Based on qualitative content analysis, extracted individual barriers and facilitators were grouped into categories. RESULTS: A total of 28 publications were identified as eligible. A framework with a 3-level hierarchy was designed which grouped 328 individual barriers and facilitators into 23 subcategories, 12 categories, and 4 clusters that focus on either the individual user (goal setting and goal striving, motivation, routines, lack of awareness of knowledge), different aspects of the app and the smartphone (features, usability of the app or food database, technical issues, data security, accuracy/trustworthiness, costs), positive and negative outcomes of nutrition app use, or interactions between the user and their social environment. CONCLUSIONS: The resulting conceptual framework underlines a pronounced diversity of reasons for (not) using nutrition apps, indicating that there is no "one-size-fits-all" approach for uptake and prolonged use of nutrition apps. Hence, tailoring nutrition apps to needs of specific user groups seems promising for increasing engagement.

Record Linkage in Clinical Cancer Registration: Experiences and Findings from Lower Saxony.

In cancer registries, record linkage procedures are used to link records of the same patient from different health care providers. In the Clinical Cancer Registry of Lower Saxony, a multi-level combination of exact assignment using the statutory health insurance number and a probabilistic procedure with control numbers and address data is applied. The procedure implemented in the register application assigns the incoming messages in this way as far as possible automatically. The aim of the observation carried out was to check the efficiency of the match variables and threshold values used, above which manual assignment is required. Weak points were identified and approaches to solutions were developed.

Biased energy efficiency perception based on instantaneous consumption displays - Indication for heuristic energy information processing.

Instantaneous consumption displays (ICDs) can be used as central information source to perceive the energy efficiency of manoeuvre-level driving. A key question is whether drivers who use ICDs can accurately derive energy efficiency differences of different driving strategies based on ICDs. There is reason to assume that drivers' consumption judgements may be biased, similar to driving-related phenomena like the time-saving bias. Therefore, the aim of the present research was to examine drivers' accuracy in deriving average consumption from dynamic ICD sequences. Participants viewed videos of a schematic ICD in a controlled experiment where the maximum instantaneous consumption systematically varied over time. Participants (N = 55) overestimated the average consumption values. The empirical ranking of the sequences did significantly correlate with the heuristic but not with the correct efficiency ranking. The current study incorporated multilevel modelling due to the nested structure of the data. The estimation difference was greater with higher peak height and shorter peak duration. The effect of peak height on estimation difference weakened with longer peak duration. In sum, the results indicate that ICDs can create biased perceptions of energy efficiency and that drivers seem to use simplifying heuristics. Knowledge and affinity for technology interaction appear to relate to biased estimations, whereas the intensity of prior experience with consumption displays seems irrelevant. Further studies should test other interfaces with debiasing potential such as manoeuvre-based aggregation or fading-trace approaches. Moreover, studies are needed that enable modelling of the effects of more natural temporal-spatial visual attention distribution (e.g. in a driving simulator setting).

Degradation of the low-calorie sugar substitute 5-ketofructose by different bacteria.

There is an increasing public awareness about the danger of dietary sugars with respect to their caloric contribution to the diet and the rise of overweight throughout the world. Therefore, low-calorie sugar substitutes are of high interest to replace sugar in foods and beverages. A promising alternative to natural sugars and artificial sweeteners is the fructose derivative 5-keto-D-fructose (5-KF), which is produced by several Gluconobacter species. A prerequisite before 5-KF can be used as a sweetener is to test whether the compound is degradable by microorganisms and whether it is metabolized by the human microbiota. We identified different environmental bacteria (Tatumella morbirosei, Gluconobacter japonicus LMG 26773, Gluconobacter japonicus LMG 1281, and Clostridium pasteurianum) that were able to grow with 5-KF as a substrate. Furthermore, Gluconobacter oxydans 621H could use 5-KF as a carbon and energy source in the stationary growth phase. The enzymes involved in the utilization of 5-KF were heterologously overproduced in Escherichia coli, purified and characterized. The enzymes were referred to as 5-KF reductases and belong to three unrelated enzymatic classes with highly different amino acid sequences, activities, and structural properties. Furthermore, we could show that 15 members of the most common and abundant intestinal bacteria cannot degrade 5-KF, indicating that this sugar derivative is not a suitable growth substrate for prokaryotes in the human intestine. KEY POINTS: • Some environmental bacteria are able to use 5-KF as an energy and carbon source. • Four 5-KF reductases were identified, belonging to three different protein families. • Many gut bacteria cannot degrade 5-KF.

An alternative pentose phosphate pathway in human gut bacteria for the degradation of C5 sugars in dietary fibers.

The microbial degradation of pentoses in the human gut is a crucial factor for the utilization of plant-based dietary fibers. A vast majority of gut microbes are able to use these C5-sugars as a carbon and energy source. However, the underlying metabolic pathways are not fully understood. Bioinformatic analysis showed that a large number of abundant gut bacteria lack genes encoding a transaldolase as a key enzyme of the pentose phosphate pathway. Among them was the important human gut microbe Prevotella copri, which was able to grow in minimal media containing xylose or hemicelluloses as the sole carbon source. Therefore, we looked for an alternative pathway for pentose conversion in P. copri using bioinformatics, enzyme activity assays, and the detection of intermediates of pentose metabolism. It became evident that the organism converted C5-sugars via the sedoheptulose-1,7-bisphosphate pathway (SBPP) to connect pentose metabolism with glycolysis. To circumvent the transaldolase reaction, P. copri uses the combined catalysis of a pyrophosphate-dependent phosphofructokinase and a fructose-bisphosphate aldolase. Furthermore, we present strong evidence that the SBPP is widely distributed in important gut bacteria, including members of the phyla Bacteroides, Firmicutes, Proteobacteria, Verrucomicrobia, and Lentisphaerae.

Rapid additive-free bacteria lysis using traveling surface acoustic waves in microfluidic channels.

We report an additive-free method to lyse bacteria and extract nucleic acids and protein using a traveling surface acoustic wave (TSAW) coupled to a microfluidic device. We characterize the effects of the TSAW on E. coli by measuring the viability of cells exposed to the sound waves and find that about 90% are dead. In addition, we measure the protein and nucleic acids released from the cells and show that we recover about 20% of the total material. The lysis method should work for all types of bacteria. These results demonstrate the feasibility of using TSAW to lyse bacteria in a manner that is independent of the type of bacteria.

High-Throughput Triggered Merging of Surfactant-Stabilized Droplet Pairs Using Traveling Surface Acoustic Waves.

We present an acoustofluidic device for fluorescently triggered merging of surfactant-stabilized picoliter droplet pairs at high throughput. Droplets that exceed a preset fluorescence threshold level are selectively merged by a traveling surface acoustic wave (T-SAW) pulse. We characterize the operation of our device by analyzing the merging efficiency as a function of acoustic pulse position, duration, and acoustic pressure amplitude. We probe droplet merging at different droplet rates and find that efficient merging occurs above a critical acoustic power level. Our results indicate that the efficiency of acoustically induced merging of surfactant stabilized droplets is correlated with acoustic streaming velocity. Finally, we discuss how both time-averaged and instantaneous acoustic pressure fields can affect the integrity of surfactant layers. Our technique, by allowing the merging of up to 105 droplets per hour, shows great potential for integration into microfluidic systems for high-throughput and high-content screening applications.

A Privacy Preserving Approach to Feasibility Analyses on Distributed Data Sources in Biomedical Research.

Funding agencies and field experts promote reuse of scientific data and biomaterial beyond the scope of the original project. The availability of research data, however, is limited by the interest of original authors as well as the privacy rights of the study participants, especially in the biomedical sciences. On the other hand, for an available data set to be a useful contribution to the scientific community, it has to be findable and accessible with reasonable effort. Therefore, using the R Shiny library, we designed and implemented a software for data discovery and feasibility analyses with compliance to regulatory and contractual regulations. Due to its genericity, it was successfully tested with heterogeneous data sets and ultimately applied to the data and biomaterial of the German Center for Cardiovascular Research (DZHK). The resulting tool - named the Feasibility Explorer - is publicly available and can be used by researchers to get an overview of data and biomaterial available in the DZHK and to select collectives in the process of submitting a usage application. To implement the rights of participants and original authors, data is integrated by querying the informed consent and not persistently stored. All calculations on the data are performed server-sided and only aggregated information is send to a client, whereas the extent of information was strictly limited to a necessary minimum that allows an applicant to assess whether an application is worthwhile.

Automated segmentation of changes in FLAIR-hyperintense white matter lesions in multiple sclerosis on serial magnetic resonance imaging.

Longitudinal analysis of white matter lesion changes on serial MRI has become an important parameter to study diseases with white-matter lesions. Here, we build on earlier work on cross-sectional lesion segmentation; we present a fully automatic pipeline for serial analysis of FLAIR-hyperintense white matter lesions. Our algorithm requires three-dimensional gradient echo T1- and FLAIR- weighted images at 3 Tesla as well as available cross-sectional lesion segmentations of both time points. Preprocessing steps include lesion filling and intrasubject registration. For segmentation of lesion changes, initial lesion maps of different time points are fused; herein changes in intensity are analyzed at the voxel level. Significance of lesion change is estimated by comparison with the difference distribution of FLAIR intensities within normal appearing white matter. The method is validated on MRI data of two time points from 40 subjects with multiple sclerosis derived from two different scanners (20 subjects per scanner). Manual segmentation of lesion increases served as gold standard. Across all lesion increases, voxel-wise Dice coefficient (0.7) as well as lesion-wise detection rate (0.8) and false-discovery rate (0.2) indicate good overall performance. Analysis of scans from a repositioning experiment in a single patient with multiple sclerosis did not yield a single false positive lesion. We also introduce the lesion change plot as a descriptive tool for the lesion change of individual patients with regard to both number and volume. An open source implementation of the algorithm is available at http://www.statistical-modeling.de/lst.html.

Designing meaningful outcome parameters using mobile technology: a new mobile application for telemonitoring of patients with heart failure.

AIMS: Health data captured by commercially available smart devices may represent meaningful patient-reported outcome measures (PROMs) in heart failure (HF) patients. The purpose of this study was to test this hypothesis by evaluating the feasibility of a new telemonitoring concept for patients following initial HF hospitalization. METHODS AND RESULTS: We designed a cardio patient monitoring platform (CPMP) that comprised mobile iOS-based applications for patients' smartphone/smartwatch and the equivalent application on a physicians' tablet. It allowed for safe and continuous data transmission of self-measured physiological parameters, activity data, and patient-reported symptoms. In a prospective feasibility trial with 692 patient days from 10 patients hospitalized for newly diagnosed HF with reduced ejection fraction (mean left ventricular ejection fraction (LVEF) 26.5 ± 9.8%), we examined the CPMP during the first 2 months following discharge (69 ± 15 observation days per patient). The mean daily step count recorded by the mobile devices emerged as a promising new PROM. Its 14 day average increased over the study period (3612 ± 3311 steps/day at study inclusion and 7069 ± 5006 steps/day at end of study; P < 0.0001). It is unique for continuously reflecting real-life activity and correlated significantly with traditional surrogate parameters of cardiac performance including LVEF (r = 0.44; 95% CI 0.07-0.71; P = 0.0232), 6 min walk test (r = 0.67; 95% CI 0.38-0.84; P = 0.0002), and scores in health-related quality of life questionnaires. CONCLUSIONS: We provide the first patient monitoring platform for HF patients that relies on commercially available iOS/watchOS-based devices. Our study suggests it is ready for implementation as a tool for recording meaningful PROMs in future HF trials and telemonitoring.

Know-how or know-why? The role of hybrid electric vehicle drivers' acquisition of eco-driving knowledge for eco-driving success.

Hybrid electric vehicles (HEVs) can contribute to sustainable transport. Yet, their real-world energy efficiency depends on HEV drivers' eco-driving behaviour. Eco-driving knowledge is key for successful eco-driving. The present research focused on the role of perceived strategy knowledge (know-how) versus technical system knowledge (know-why) in a study with 121 HEV drivers. The relationship between knowledge components and knowledge acquisition processes, as well as fuel efficiency, were examined. Structural equation modelling results indicated that perceived strategy knowledge was related to acquisition by testing (i.e., interacting with the vehicle and its interfaces) and reading (i.e., manuals, books and websites) while technical system knowledge was only related to acquisition by reading. In contrast to technical system knowledge, perceived strategy knowledge was no significant predictor of fuel efficiency. The results indicated that emphasis should be put into promoting technical system knowledge (e.g., by tutoring systems) to support motivated drivers' in achieving higher fuel efficiency.

Physiology and central carbon metabolism of the gut bacterium Prevotella copri.

The human gut microbiota is a crucial factor for the host's physiology with respect to health and disease. Metagenomic shotgun sequencing of microbial gut communities revealed that Prevotella copri is one of the most important players in the gastrointestinal tract of many individuals. Because of the importance of this bacterium we analyzed the growth behavior and the central metabolic pathways of P. copri. Bioinformatic data, transcriptome profiling and enzyme activity measurements indicated that the major pathways are based on glycolysis and succinate production from fumarate. In addition, pyruvate can be degraded to acetate and formate. Electron transport phosphorylation depends on fumarate respiration with NADH and reduced ferredoxin as electron donors. In contrast to Bacteroides vulgatus, P. copri showed a more pronounced dependency on the addition of CO2 or bicarbonate for biomass formation, which is a remarkable difference between P. copri and Bacteroides spp. with important implication in the context of gut microbial competition. The analysis of substrate consumption and product concentrations from many P. copri cultures with different optical densities allowed a prediction of the carbon and electron flow in the central metabolism and a detailed calculation of growth yields as well as carbon and redox balances.

Context-dependent expression of a conditionally-inducible form of active Akt.

Akt kinases are key signaling components in proliferation-competent and post-mitotic cells. Here, we sought to create a conditionally-inducible form of active Akt for both in vitro and in vivo applications. We fused a ligand-responsive Destabilizing Domain (DD) derived from E. coli dihydrofolate reductase to a constitutively active mutant form of Akt1, Akt(E40K). Prior work indicated that such fusion proteins may be stabilized and induced by a ligand, the antibiotic Trimethoprim (TMP). We observed dose-dependent, reversible induction of both total and phosphorylated/active DD-Akt(E40K) by TMP across several cellular backgrounds in culture, including neurons. Phosphorylation of FoxO4, an Akt substrate, was significantly elevated after DD-Akt(E40K) induction, indicating the induced protein was functionally active. The induced Akt(E40K) protected cells from apoptosis evoked by serum deprivation and was neuroprotective in two cellular models of Parkinson's disease (6-OHDA and MPP+ exposure). There was no significant protection without induction. We also evaluated Akt(E40K) induction by TMP in mouse substantia nigra and striatum after neuronal delivery via an AAV1 adeno-associated viral vector. While there was significant induction in striatum, there was no apparent induction in substantia nigra. To explore the possible basis for this difference, we examined DD-Akt(E40K) induction in cultured ventral midbrain neurons. Both dopaminergic and non-dopaminergic neurons in the cultures showed DD-Akt(E40K) induction after TMP treatment. However, basal DD-Akt(E40K) expression was 3-fold higher for dopaminergic neurons, resulting in a significantly lower induction by TMP in this population. Such findings suggest that dopaminergic neurons may be relatively inefficient in protein degradation, a property that could relate to their lack of apparent DD-Akt(E40K) induction in vivo and to their selective vulnerability in Parkinson's disease. In summary, we generated an inducible, biologically active form of Akt. The degree of inducibility appears to reflect cellular context that will inform the most appropriate applications for this and related reagents.