Proteome census upon nutrient stress reveals Golgiphagy membrane receptors.

During nutrient stress, macroautophagy degrades cellular macromolecules, thereby providing biosynthetic building blocks while simultaneously remodelling the proteome1,2. Although the machinery responsible for initiation of macroautophagy has been well characterized3,4, our understanding of the extent to which individual proteins, protein complexes and organelles are selected for autophagic degradation, and the underlying targeting mechanisms, is limited. Here we use orthogonal proteomic strategies to provide a spatial proteome census of autophagic cargo during nutrient stress in mammalian cells. We find that macroautophagy has selectivity for recycling membrane-bound organelles (principally Golgi and endoplasmic reticulum). Through autophagic cargo prioritization, we identify a complex of membrane-embedded proteins, YIPF3 and YIPF4, as receptors for Golgiphagy. During nutrient stress, YIPF3 and YIPF4 interact with ATG8 proteins through LIR motifs and are mobilized into autophagosomes that traffic to lysosomes in a process that requires the canonical autophagic machinery. Cells lacking YIPF3 or YIPF4 are selectively defective in elimination of a specific cohort of Golgi membrane proteins during nutrient stress. Moreover, YIPF3 and YIPF4 play an analogous role in Golgi remodelling during programmed conversion of stem cells to the neuronal lineage in vitro. Collectively, the findings of this study reveal prioritization of membrane protein cargo during nutrient-stress-dependent proteome remodelling and identify a Golgi remodelling pathway that requires membrane-embedded receptors.

PARK15/FBXO7 is dispensable for PINK1/Parkin mitophagy in iNeurons and HeLa cell systems.

The protein kinase PINK1 and ubiquitin ligase Parkin promote removal of damaged mitochondria via a feed-forward mechanism involving ubiquitin (Ub) phosphorylation (pUb), Parkin activation, and ubiquitylation of mitochondrial outer membrane proteins to support the recruitment of mitophagy receptors. The ubiquitin ligase substrate receptor FBXO7/PARK15 is mutated in an early-onset parkinsonian-pyramidal syndrome. Previous studies have proposed a role for FBXO7 in promoting Parkin-dependent mitophagy. Here, we systematically examine the involvement of FBXO7 in depolarization and mt UPR-dependent mitophagy in the well-established HeLa and induced-neurons cell systems. We find that FBXO7-/- cells have no demonstrable defect in: (i) kinetics of pUb accumulation, (ii) pUb puncta on mitochondria by super-resolution imaging, (iii) recruitment of Parkin and autophagy machinery to damaged mitochondria, (iv) mitophagic flux, and (v) mitochondrial clearance as quantified by global proteomics. Moreover, global proteomics of neurogenesis in the absence of FBXO7 reveals no obvious alterations in mitochondria or other organelles. These results argue against a general role for FBXO7 in Parkin-dependent mitophagy and point to the need for additional studies to define how FBXO7 mutations promote parkinsonian-pyramidal syndrome.

IsobaricQuant enables cross-platform quantification, visualization, and filtering of isobarically-labeled peptides.

In mass spectrometry (MS)-based quantitative proteomics, labeling with isobaric mass tags such as iTRAQ and TMT can substantially improve sample throughput and reduce peptide missing values. Nonetheless, the quantification of labeled peptides tends to suffer from reduced accuracy due to the co-isolation of co-eluting precursors of similar mass-to-charge. Acquisition approaches such as multistage MS3 or ion mobility separation address this problem, yet are difficult to audit and limited to expensive instrumentation. Here we introduce IsobaricQuant, an open-source software tool for quantification, visualization, and filtering of peptides labeled with isobaric mass tags, with specific focus on precursor interference. IsobaricQuant is compatible with MS2 and MS3 acquisition strategies, has a viewer that allows assessing interference, and provides several scores to aid the filtering of scans with compression. We demonstrate that IsobaricQuant quantifications are accurate by comparing it with commonly used software. We further show that its QC scores can successfully filter out scans with reduced quantitative accuracy at MS2 and MS3 levels, removing inaccurate peptide quantifications and decreasing protein CVs. Finally, we apply IsobaricQuant to a PISA dataset and show that QC scores improve the sensitivity of the identification of protein targets of a kinase inhibitor. IsobaricQuant is available at https://github.com/Villen-Lab/isobaricquant.

Coisolation of Peptide Pairs for Peptide Identification and MS/MS-Based Quantification.

Stable-isotope labeling with amino acids in cell culture (SILAC)-based metabolic labeling is a widely adopted proteomics approach that enables quantitative comparisons among a variety of experimental conditions. Despite its quantitative capacity, SILAC experiments analyzed with data-dependent acquisition (DDA) do not fully leverage peptide pair information for identification and suffer from undersampling compared to label-free proteomic experiments. Herein, we developed a DDA strategy that coisolates and fragments SILAC peptide pairs and uses y-ions for their relative quantification. To facilitate the analysis of this type of data, we adapted the Comet sequence database search engine to make use of SILAC peptide paired fragments and developed a tool to annotate and quantify MS/MS spectra of coisolated SILAC pairs. This peptide pair coisolation approach generally improved expectation scores compared to the traditional DDA approach. Fragment ion quantification performed similarly well to precursor quantification in the MS1 and achieved more quantifications. Lastly, our method enables reliable MS/MS quantification of SILAC proteome mixtures with overlapping isotopic distributions. This study shows the feasibility of the coisolation approach. Coupling this approach with intelligent acquisition strategies has the potential to improve SILAC peptide sampling and quantification.

Long-Term Results Following Repair for Degenerative Mitral Regurgitation - Analysis of Factors Influencing Durability.

BACKGROUND: The majority of patients with degenerative mitral regurgitation (DMR) are amenable to reconstructive procedures. There is debate regarding factors that influence long-term durability with respect to repair technique, valve remodelling and progressive myxomatous change. METHODS: A total of 685 patients with DMR underwent mitral valve repair by a single surgeon between 1991 and 2011 with follow-up completed at 31 December 2016. Repair rate for patients undergoing surgery for DMR was over 90%. Mean age was 64 years (18-89) with 66.2% male, 47% NYHA class III-IV, and 20% had permanent atrial fibrillation (PAF). Major associated procedures were performed in 28% of patients (189); including coronary artery bypass graft (CABG) (127), aortic valve replacement (15), aortic root surgery (3) and tricuspid valve annuloplasty (61). RESULTS: Operative mortality (≤30 days) occurred in four patients (0.58%). At 20 years, survival was 58%, freedom from reoperation was 90% and freedom from reoperation and non-operated recurrent MR >2+ (relapse) was 78%. Factors influencing survival were advancing age, left ventricular (LV) dysfunction (ejection fraction <60% or end systolic dimension >40mm), New York Heart Association (NYHA) III-IV and PAF. Predictors of relapse were the degree of residual intraoperative mitral regurgitation (p<0.001), anterior leaflet prolapse (p<0.001) and the addition of a sliding annuloplasty in isolated posterior leaflet repair (p=0.023). The majority of reoperations were for technical issues related to the original repair. A competent valve at 6 months to 3 years postoperatively predicted an excellent long-term result. CONCLUSION: The great majority of degenerative mitral valves are repairable regardless of age with excellent long-term results achievable following surgery. Survival is reduced by significant symptoms, LV dysfunction and preoperative PAF. Repair is best performed before these features develop. Durability is largely dependent on the technical performance of the repair and degree of residual MR on the post-pump transoesophageal echocardiogram. We recommend surgery should be performed by surgeons specialising and skilled in mitral valve repair.

Improved surfaceome coverage with a label-free nonaffinity-purified workflow.

The proteins of the cellular plasma membrane (PM) perform important functions relating to homeostasis and intercellular communication. Due to its overall low cellular abundance, amphipathic character, and low membrane-to-cytoplasm ratio, the PM proteome has been challenging to isolate and characterize, and is poorly represented in standard LC-MS/MS analyses. In this study, we employ sucrose gradient ultracentrifugation for the enrichment of the PM proteome, without chemical labeling and affinity purification, together with GeLCMS and use subsequent bioinformatics tools to select proteins associated with the PM/cell surface, herein referred to as the surfaceome. Using this methodology, we identify over 1900 cell surface associated proteins in a human acute myeloid leukemia cell line. These surface proteins comprise almost 50% of all detected cellular proteins, a number that substantially exceeds the depth of coverage in previously published studies describing the leukemia surfaceome.

Radiation risk reduction in cardiac electrophysiology through use of a gridless imaging technique.

AIMS: It has been previously demonstrated that use of appropriate frame rates coupled with minimal use of high-dose digital acquisition can limit radiation risk to patients undergoing diagnostic and therapeutic electrophysiology (EP). Imaging without the anti-scatter grid has been proposed as a means of achieving further radiation reduction. We evaluate application of a gridless imaging technique to deliver further reductions in radiation risk to both patients and personnel. METHODS AND RESULTS: Radiation and clinical data for EP procedures performed for 16 months from March 2012 were monitored. The period was divided into three phases: Phase 1 (March 2012-June 2012) provided a performance baseline (radiation output modelling and procedural risk adjustment calibration), Phase 2 (July 2012-September 2012) confirmation of performance with the grid, and Phase 3 (September 2012-June 2013) gridless imaging period. Statistical process control (SPC) charts were used to monitor for changes in radiation use and clinical outcomes (procedural success). Imaging without the grid halved the levels of radiation delivered in undertaking EP procedures. Although there was a perceptible impact on image quality with the grid removed. Review of the SPC chart monitoring procedural outcomes did not identify any discernable adverse impact on success rates. Selected use of the gridless technique is recommended with re-introduction of the grid in larger patients or during aspects of the procedure where image quality is important (e.g. transeptal punctures). CONCLUSION: Use of a gridless imaging technique can contribute to a significant reduction in radiation risk to both patients and operators during cardiac EP procedures.

How to bring the NHS back from the permanent brink of collapse.

Three sets of coordinated actions are required if the health service is to pull back from the edge of the cliff - and stay there.

Defining a kinetic mechanism for l-DOPA 2,3 dioxygenase, a single-domain type I extradiol dioxygenase from Streptomyces lincolnensis.

l-DOPA-2,3-dioxygenase from Streptomyces lincolnensis is a single domain type I extradiol dioxygenase of the vicinal oxygen chelate superfamily and catalyzes the second step in the metabolism of the propylhygric acid moiety of the antibiotic, lincomycin. In this report, the kinetic mechanism of l-DOPA dioxygenase is interrogated using stopped-flow in order to determine microscopic rate constants. Pre-steady state, progress curve and steady-state data were combined in a global kinetic analysis using KinTek Explorer in order to define and constrain a kinetic model for the type I l-DOPA dioxygenase. The data are best described by a four step mechanism, in which the cyclization of the enzymatic product is not enzyme catalyzed.

Combinatorial selective ER-phagy remodels the ER during neurogenesis.

The endoplasmic reticulum (ER) employs a diverse proteome landscape to orchestrate many cellular functions, ranging from protein and lipid synthesis to calcium ion flux and inter-organelle communication. A case in point concerns the process of neurogenesis, where a refined tubular ER network is assembled via ER shaping proteins into the newly formed neuronal projections to create highly polarized dendrites and axons. Previous studies have suggested a role for autophagy in ER remodelling, as autophagy-deficient neurons in vivo display axonal ER accumulation within synaptic boutons, and the membrane-embedded ER-phagy receptor FAM134B has been genetically linked with human sensory and autonomic neuropathy. However, our understanding of the mechanisms underlying selective removal of the ER and the role of individual ER-phagy receptors is limited. Here we combine a genetically tractable induced neuron (iNeuron) system for monitoring ER remodelling during in vitro differentiation with proteomic and computational tools to create a quantitative landscape of ER proteome remodelling via selective autophagy. Through analysis of single and combinatorial ER-phagy receptor mutants, we delineate the extent to which each receptor contributes to both the magnitude and selectivity of ER protein clearance. We define specific subsets of ER membrane or lumenal proteins as preferred clients for distinct receptors. Using spatial sensors and flux reporters, we demonstrate receptor-specific autophagic capture of ER in axons, and directly visualize tubular ER membranes within autophagosomes in neuronal projections by cryo-electron tomography. This molecular inventory of ER proteome remodelling and versatile genetic toolkit provide a quantitative framework for understanding the contributions of individual ER-phagy receptors for reshaping ER during cell state transitions.

Lysosomal storage disease proteo/lipidomic profiling using nMOST links ferritinophagy with mitochondrial iron deficiencies in cells lacking NPC2.

Lysosomal storage diseases (LSDs) comprised ~50 monogenic diseases characterized by the accumulation of cellular material in lysosomes and associated defects in lysosomal function, but systematic molecular phenotyping is lacking. Here, we develop a nanoflow-based multi-omic single-shot technology (nMOST) workflow allowing simultaneously quantify HeLa cell proteomes and lipidomes from more than two dozen LSD mutants, revealing diverse molecular phenotypes. Defects in delivery of ferritin and its autophagic receptor NCOA4 to lysosomes (ferritinophagy) were pronounced in NPC2-/- cells, which correlated with increased lyso-phosphatidylcholine species and multi-lamellar membrane structures visualized by cryo-electron-tomography. Ferritinophagy defects correlated with loss of mitochondrial cristae, MICOS-complex components, and electron transport chain complexes rich in iron-sulfur cluster proteins. Strikingly, mitochondrial defects were alleviated when iron was provided through the transferrin system. This resource reveals how defects in lysosomal function can impact mitochondrial homeostasis in trans and highlights nMOST as a discovery tool for illuminating molecular phenotypes across LSDs.

Risk modelling in quality clinical registries: monitoring lesion treatment failure rate in percutaneous coronary interventions.

AIMS: This paper describes the development of a risk adjustment (RA) model predictive of individual lesion treatment failure in percutaneous coronary interventions (PCI) for use in a quality monitoring and improvement program. METHODS AND RESULTS: Prospectively collected data for 3972 consecutive revascularisation procedures (5601 lesions) performed between January 2003 and September 2011 were studied. Data on procedures to September 2009 (n=3100) were used to identify factors predictive of lesion treatment failure. Factors identified included lesion risk class (p<0.001), occlusion type (p<0.001), patient age (p=0.001), vessel system (p<0.04), vessel diameter (p<0.001), unstable angina (p=0.003) and presence of major cardiac risk factors (p=0.01). A Bayesian RA model was built using these factors with predictive performance of the model tested on the remaining procedures (area under the receiver operating curve: 0.765, Hosmer-Lemeshow p value: 0.11). Cumulative sum, exponentially weighted moving average and funnel plots were constructed using the RA model and subjectively evaluated. CONCLUSION: A RA model was developed and applied to SPC monitoring for lesion failure in a PCI database. If linked to appropriate quality improvement governance response protocols, SPC using this RA tool might improve quality control and risk management by identifying variation in performance based on a comparison of observed and expected outcomes.

Performance monitoring in cardiac surgery: application of statistical process control to a single-site database.

BACKGROUND: Graphical Statistical Process Control (SPC) tools have been shown to promptly identify significant variations in clinical outcomes in a range of health care settings. We explored the application of these techniques to quantitatively inform the routine cardiac surgical (CAS) morbidity and mortality (M&M) review processes at a single site. METHODS: Baseline clinical and procedural data relating to 5265 consecutive cardiac surgical procedures, performed at St Andrew's War Memorial Hospital (SAWMH) between the 1st January 2003 and the 30th April 2012, were retrospectively evaluated. A range of appropriate clinical outcome indicators (COIs) were developed and evaluated using a combination of Cumulative Sum charts, Exponentially Weighted Moving Average charts and Funnel Plots. Charts were updated regularly and discussed at the cardiac surgery unit's bi-monthly M&M meetings. Risk adjustment (RA) for the COIs was developed and validated for incorporation into the charts to improve monitoring performance. RESULTS: Discrete and aggregated measures, including blood product/reoperation, major acute post-procedural complications, cardiopulmonary bypass duration and Length of Stay/Readmission < 28 days have proved to be valuable measures for monitoring outcomes. Instances of variation in performance identified using the charts were examined thoroughly and could be related to changes in clinical practice (e.g. antifibrinolytic use) as well as differences in individual operator performance (in some instances, driven by case mix). CONCLUSIONS: SPC tools can promptly detect meaningful changes in clinical outcome thereby allowing early intervention to address altered performance. Careful interpretation of charts for group and individual operators has proven helpful in detecting and differentiating systemic versus individual variation.

Use of graphical statistical process control tools to monitor and improve outcomes in cardiac surgery.

BACKGROUND: Graphical Statistical Process Control (SPC) tools have been shown to promptly identify significant variations in clinical outcomes in a range of health care settings. We explored the application of these techniques to qualitatively inform the routine cardiac surgical morbidity and mortality (M&M) review process at a single site. METHODS: Baseline clinical and procedural data relating to 4774 consecutive cardiac surgical procedures, performed between the 1st January 2003 and the 30th April 2011, were retrospectively evaluated. A range of appropriate performance measures and benchmarks were developed and evaluated using a combination of CUmulative SUM (CUSUM) charts, Exponentially Weighted Moving Average (EWMA) charts and Funnel Plots. Charts have been discussed at the unit's routine M&M meetings. Risk adjustment (RA) based on EuroSCORE has been incorporated into the charts to improve performance. RESULTS: Discrete and aggregated measures, including Blood Product/Reoperation, major acute post-procedural complications and Length of Stay/Readmission<28 days have proved to be usable measures for monitoring outcomes. Monitoring trends in minor morbidities provides a valuable warning of impending changes in significant events. Instances of variation in performance have been examined and could be related to differences in individual operator performance via individual operator curves. CONCLUSION: SPC tools facilitate near "real-time" performance monitoring allowing early detection and intervention in altered performance. Careful interpretation of charts for group and individual operators has proven helpful in detecting and differentiating systemic vs. individual variation.

Combinatorial selective ER-phagy remodels the ER during neurogenesis.

The endoplasmic reticulum (ER) employs a diverse proteome landscape to orchestrate many cellular functions ranging from protein and lipid synthesis to calcium ion flux and inter-organelle communication. A case in point concerns the process of neurogenesis: a refined tubular ER network is assembled via ER shaping proteins into the newly formed neuronal projections to create highly polarized dendrites and axons. Previous studies have suggested a role for autophagy in ER remodeling, as autophagy-deficient neurons in vivo display axonal ER accumulation within synaptic boutons, and the membrane-embedded ER-phagy receptor FAM134B has been genetically linked with human sensory and autonomic neuropathy. However, our understanding of the mechanisms underlying selective removal of ER and the role of individual ER-phagy receptors is limited. Here, we combine a genetically tractable induced neuron (iNeuron) system for monitoring ER remodeling during in vitro differentiation with proteomic and computational tools to create a quantitative landscape of ER proteome remodeling via selective autophagy. Through analysis of single and combinatorial ER-phagy receptor mutants, we delineate the extent to which each receptor contributes to both magnitude and selectivity of ER protein clearance. We define specific subsets of ER membrane or lumenal proteins as preferred clients for distinct receptors. Using spatial sensors and flux reporters, we demonstrate receptor-specific autophagic capture of ER in axons, and directly visualize tubular ER membranes within autophagosomes in neuronal projections by cryo-electron tomography. This molecular inventory of ER proteome remodeling and versatile genetic toolkit provides a quantitative framework for understanding contributions of individual ER-phagy receptors for reshaping ER during cell state transitions.

Monitoring radiation use in cardiac fluoroscopy imaging procedures.

PURPOSE: Timely identification of systematic changes in radiation delivery of an imaging system can lead to a reduction in risk for the patients involved. However, existing quality assurance programs involving the routine testing of equipment performance using phantoms are limited in their ability to effectively carry out this task. To address this issue, the authors propose the implementation of an ongoing monitoring process that utilizes procedural data to identify unexpected large or small radiation exposures for individual patients, as well as to detect persistent changes in the radiation output of imaging platforms. METHODS: Data used in this study were obtained from records routinely collected during procedures performed in the cardiac catheterization imaging facility at St. Andrew's War Memorial Hospital, Brisbane, Australia, over the period January 2008-March 2010. A two stage monitoring process employing individual and exponentially weighted moving average (EWMA) control charts was developed and used to identify unexpectedly high or low radiation exposure levels for individual patients, as well as detect persistent changes in the radiation output delivered by the imaging systems. To increase sensitivity of the charts, we account for variation in dose area product (DAP) values due to other measured factors (patient weight, fluoroscopy time, and digital acquisition frame count) using multiple linear regression. Control charts are then constructed using the residual values from this linear regression. The proposed monitoring process was evaluated using simulation to model the performance of the process under known conditions. RESULTS: Retrospective application of this technique to actual clinical data identified a number of cases in which the DAP result could be considered unexpected. Most of these, upon review, were attributed to data entry errors. The charts monitoring the overall system radiation output trends demonstrated changes in equipment performance associated with relocation of the equipment to a new department. When tested under simulated conditions, the EWMA chart was capable of detecting a sustained 15% increase in average radiation output within 60 cases (<1 month of operation), while a 33% increase would be signaled within 20 cases. CONCLUSIONS: This technique offers a valuable enhancement to existing quality assurance programs in radiology that rely upon the testing of equipment radiation output at discrete time frames to ensure performance security.

The role of quantitative feedback in coronary angiography radiation reduction.

OBJECTIVE: To evaluate the benefits of radiation education with and without feedback reporting in altering clinician radiation use behaviour in performing coronary angiography (CA). DESIGN: A retrospective review of radiation use (fluoroscopy time) in coronary angiograms performed between July 1996 and December 2005 by 10 cardiologists to assess the impact of various interventions aimed at minimizing radiation risk. The impact of interventions such as education and audit/feedback was correlated against radiation use using cumulative sum and cumulative expected minus observed charts. SETTING: Private Hospital in Brisbane, Australia. PARTICIPANTS: Ten cardiologists. INTERVENTION: Education and audit/feedback. RESULTS: Baseline radiation use subject to standard guidelines was stable. Group performance charts show a modest transient improvement in radiation use associated with an education intervention alone. However, regular detailed personalized feedback comparing an individual's radiation use to group and external benchmarks was successful in achieving sustained reduction in overall radiation use. For individual participants, significant improvement was noted in 7 of 10 cardiologists. CONCLUSION: Although an improved theoretical understanding of effective radiation hygiene strategies might contribute to reduced radiation use, this study suggests that regular detailed quantitative feedback supporting education is an effective tool in altering radiation use in CA. Understanding triggers that stimulate change in clinician behaviour is critical to the design of systems to optimize clinical performance. Confidentially reported benchmarking systems may be a useful tool to alter clinician behaviour.

Reassessment of radiation risks from electrophysiology procedures compared to coronary angiography.

BACKGROUND: Electrophysiology (EP) procedures have been reported to carry a significantly greater radiation risk than that of coronary angiography (CA). This is largely due to numerous reports linking severe deterministic radiation effects to long procedure and fluoroscopy times (FTs). This study documents low radiation doses achieved by strategies involving operator training and education as well as equipment and technique optimisation to reduce radiation risks. METHODS: Records relating to 732 diagnostic EP and 1744 therapeutic EP procedures performed between January 2002 and December 2007 were analysed. Data from 1458 diagnostic only CA procedures performed in 2006 was used for comparison. For each procedure type, FT, number of digital frames acquired and estimated effective dose (E) were compared. RESULTS: Although the FT for CA procedures is significantly less than for therapeutic EP procedures (FT for diagnostic EP being similar), EP procedures generally are associated with lower E, the exception being procedures for atrial fibrillation (AF). CONCLUSION: Through the application of a comprehensive exposure minimisation strategy, the radiation risk to patients undergoing diagnostic and, therapeutic EP procedures (except AF ablation procedures) is significantly less than that faced by patients undergoing CA. E, however, is heavily dependent on procedure type and as such care must be taken in undertaking generalised comparisons for audit and benchmarking purposes.

Measures of radiation exposure in cardiac imaging and the impact of case complexity.

BACKGROUND: Although it is important for a patient's radiation related risks to be considered when recommending a cardiac imaging procedure, few clinicians appreciate the level of exposure involved. This paper provides a comprehensive set of radiation exposure metrics for common angiography procedures highlighting links to factors that influence radiation dose. METHODS: Radiation use metrics and various clinical findings for diagnostic procedures and number of lesions treated, vessels treated and stents deployed for interventional cases were analysed. RESULTS: Data relating to 1088 coronary angiography (CA), 256 angioplasty and 167 CA/angioplasty procedures were examined. The median effective dose (E) (mSv) for these procedures (including inter-quartile range) were, respectively, 3.3 (2.1-5.1), 7.5 (4.5-14.1) and 11.6 (6.9-16.1). For CA, E varied with the number of vessels (p<0.01) while for angioplasty E was linked to the number of vessels (p<0.01), lesions (p<0.01) and stents (p<0.01). CONCLUSION: Radiation exposure metrics for common cardiac angiography imaging procedures have been documented and linked to procedure complexity. This has implications for performance monitoring when comparing radiation usage between users, facilities, times and technologies.