Assessing lactate concentration as a predictor of 28-day in-hospital mortality in the presence of ethanol: A retrospective study of emergency department patients.

BACKGROUND: Presence of ethanol (EtOH) may alter the relationship between blood lactate concentrations and mortality. This study compares lactate-associated mortality risk in the presence and absence of EtOH. METHODS: We performed a retrospective cohort study including all patients, age >17 years, presenting from January 2012-December 2018, to an urban, academic emergency department, with a clinically measured lactate. Data were electronically abstracted from the medical record. The primary outcome was 28-day in-hospital mortality. Patients were grouped by EtOH test results as follows: 1) present (any EtOH detected), 2) absent (EtOH concentration measured and not detected), or 3) not ordered. Marginal analysis was used to calculated probability of mortality for fixed values of lactate and model covariates. RESULTS: Of 40,956 adult emergency department patients with measured lactate, we excluded 768 (1.89%) for lactate >10.0 mmol/L, leaving 40,240 for analysis: 4,066 (10.1%) EtOH present, 10,819 (26.9%) EtOH absent, 25,355 (63%) EtOH not ordered. Of these, 1790 (4.4%) had 28-day in-hospital mortality. Marginal probability of mortality calculated for specific lactate values found less risk for EtOH Present patients versus EtOH absent patients at lactate 0.0 mmol/L (0.8% [95%CI: 0.5-1.2%] vs 3.2% [2.8-3.6%]), 2.0 mmol/L (1.5% [1.1-1.9%] vs 4.0% [3.7-4.3%]), 4.0 mmol/L (2.6% [2.2-3.1%] vs 5.0% [4.6-5.4%]), until 6.0 mmol/L (4.5% [3.7-5.4%] vs 6.2% [5.4-7.0%]). CONCLUSION: EtOH presence significantly alters lactate-associated mortality risk when lactate <6.0 mmol/L. Emergency department clinicians should interpret these lactate values with caution and consider other data for risk stratification when EtOH is present.

Squamous epithelial cell presence reduces accuracy of urinalysis for prediction of positive urine cultures.

BACKGROUND: Diagnostic value of urinalysis specimens contaminated with squamous epithelial cells (SEC) from the genital surfaces is assumed to be limited compared to clean-catch samples. However, no studies have quantified the change in predictive value in the presence of SECs for individual urinalysis markers. METHODS: In a retrospective, single center cohort study, we analyzed all urine cultures sent from the ED over a 26-month period with corresponding urinalysis results. Cultures were classified as positive with growth of >104 colony forming units of pathogenic bacteria, negative if no growth, or contaminated for all other results. UA specimens were classified as contaminated or clean based on SEC presence. Accuracy of urinalysis markers for prediction of positive cultures was calculated as an area under the curve (AUC) and was compared between contaminated and clean UA specimens. RESULTS: 6490 paired UA and urine cultures were analyzed, consisting of 3949 clean and 2541 contaminated samples. SEC presence was less common with male gender, older age, and smaller BMI. Urine cultures were 19.2% positive overall, and SECs were more common in contaminated cultures. AUCs for individual markers ranged from 0.557 to 0.796, with pyuria, bacteriuria, and leukocyte esterase having higher AUC in clean samples over contaminated. CONCLUSION: Analysis of AUC for individual urinalysis markers showed reduced diagnostic accuracy in the presence of SECs. SEC presence also reflected much higher rates of contaminated cultures. These results support the reduced reliance on contaminated UA specimens for ruling in UTI in ED patients.

Comparison of bedside screening methods for frailty assessment in older adult trauma patients in the emergency department.

BACKGROUND: Frailty is linked to poor outcomes in older patients. We prospectively compared the utility of the picture-based Clinical Frailty Scale (CFS9), clinical assessments, and ultrasound muscle measurements against the reference FRAIL scale in older adult trauma patients in the emergency department (ED). METHODS: We recruited a convenience sample of adults 65 yrs. or older with blunt trauma and injury severity scores <9. We queried subjects (or surrogates) on the FRAIL scale, and compared this to: physician-based and subject/surrogate-based CFS9; mid-upper arm circumference (MUAC) and grip strength; and ultrasound (US) measures of muscle thickness (limbs and abdominal wall). We derived optimal diagnostic thresholds and calculated performance metrics for each comparison using sensitivity, specificity, predictive values, and area under receiver operating characteristic curves (AUROC). RESULTS: Fifteen of 65 patients were frail by FRAIL scale (23%). CFS9 performed well when assessed by subject/surrogate (AUROC 0.91 [95% CI 0.84-0.98] or physician (AUROC 0.77 [95% CI 0.63-0.91]. Optimal thresholds for both physician and subject/surrogate were CFS9 of 4 or greater. If both physician and subject/surrogate provided scores <4, sensitivity and negative predictive value were 90.0% (54.1-99.5%) and 95.0% (73.1-99.7%). Grip strength and MUAC were not predictors. US measures that combined biceps and quadriceps thickness showed an AUROC of 0.75 compared to the reference standard. CONCLUSION: The ED needs rapid, validated tools to screen for frailty. The CFS9 has excellent negative predictive value in ruling out frailty. Ultrasound of combined biceps and quadriceps has modest concordance as an alternative in trauma patients who cannot provide a history.

A Sepsis-related Diagnosis Impacts Interventions and Predicts Outcomes for Emergency Patients with Severe Sepsis.

INTRODUCTION: Many patients meeting criteria for severe sepsis are not given a sepsis-related diagnosis by emergency physicians (EP). This study 1) compares emergency department (ED) interventions and in-hospital outcomes among patients with severe sepsis, based on the presence or absence of sepsis-related diagnosis, and 2) assesses how adverse outcomes relate to three-hour sepsis bundle completion among patients fulfilling severe sepsis criteria but not given a sepsis-related diagnosis. METHODS: We performed a retrospective cohort study using patients meeting criteria for severe sepsis at two urban, academic tertiary care centers from March 2015 through May 2015. We included all ED patients with the following: 1) the 1992 Consensus definition of severe sepsis, including two or more systemic inflammatory response syndrome criteria and evidence of organ dysfunction; or 2) physician diagnosis of severe sepsis or septic shock. We excluded patients transferred to or from another hospital and those <18 years old. Patients with an EP-assigned sepsis diagnosis created the "Physician Diagnosis" group; the remaining patients composed the "Consensus Criteria" group. The primary outcome was in-hospital mortality. Secondary outcomes included completed elements of the current three-hour sepsis bundle; non-elective intubation; vasopressor administration; intensive care unit (ICU) admission from the ED; and transfer to the ICU in < 24 hours. We compared proportions of each outcome between groups using the chi-square test, and we also performed a stratified analysis using chi square to assess the association between failure to complete the three-hour bundle and adverse outcomes in each group. RESULTS: Of 418 patients identified with severe sepsis we excluded 54, leaving 364 patients for analysis: 121 "Physician Diagnosis" and 243 "Consensus Criteria." The "Physician Diagnosis" group had a higher in-hospital mortality (12.4% vs 3.3%, P < 0.01) and compliance with the three-hour sepsis bundle (52.1% vs 20.2%, P < 0.01) compared with the "Consensus Criteria" group. An incomplete three-hour sepsis bundle was not associated with a higher incidence of death, intubation, vasopressor use, ICU admission or transfer to the ICU in <24 hours in patients without a sepsis diagnosis. CONCLUSION: "Physician Diagnosis" patients more frequently received sepsis-specific interventions and had a higher incidence of mortality. "Consensus Criteria" patients had infrequent adverse outcomes regardless of three-hour bundle compliance. EPs' sepsis diagnoses reflect risk-stratification beyond the severe sepsis criteria.

Hidden Markov Models for Protein Domain Homology Identification and Analysis.

Protein domain identification and analysis are cornerstones of modern proteomics. The tools available to protein domain researchers avail a variety of approaches to understanding large protein domain families. Hidden Markov Models (HMM) form the basis for identifying and categorizing evolutionarily linked protein domains. Here I describe the use of HMM models for predicting and identifying Src Homology 2 (SH2) domains within the proteome.

Proteomic Clustering Analysis of SH2 Domain Datasets.

Proteomic clustering analysis provides a means of identifying relationships and visualizing those relationships in an extremely complex field of study with many interacting parts. With recent high-throughput studies of Src Homology 2 (SH2) domains, many and varied datasets are being amassed. A strategy for analyzing patterns between these large datasets is required to transform the information into knowledge. The methods for creating neighbor-joining phylogenetic trees, pairs scatter plots, and two-dimensional hierarchical clustering heatmaps are just a few of the diverse methods available to a proteomic researcher. This chapter examines selecting objects to be analyzed, selecting comparison functions to apply to those objects, and pseudo-code for processing data and preparing it for various types of analyses. Here I apply clustering analysis to previous collections of SH2 domains datasets to bring insight into new binding or specificity patterns between the different SH2 domains.

SRC Homology 2 Domain Binding Sites in Insulin, IGF-1 and FGF receptor mediated signaling networks reveal an extensive potential interactome.

Specific peptide ligand recognition by modular interaction domains is essential for the fidelity of information flow through the signal transduction networks that control cell behavior in response to extrinsic and intrinsic stimuli. Src homology 2 (SH2) domains recognize distinct phosphotyrosine peptide motifs, but the specific sites that are phosphorylated and the complement of available SH2 domains varies considerably in individual cell types. Such differences are the basis for a wide range of available protein interaction microstates from which signaling can evolve in highly divergent ways. This underlying complexity suggests the need to broadly map the signaling potential of systems as a prerequisite for understanding signaling in specific cell types as well as various pathologies that involve signal transduction such as cancer, developmental defects and metabolic disorders. This report describes interactions between SH2 domains and potential binding partners that comprise initial signaling downstream of activated fibroblast growth factor (FGF), insulin (Ins), and insulin-like growth factor-1 (IGF-1) receptors. A panel of 50 SH2 domains screened against a set of 192 phosphotyrosine peptides defines an extensive potential interactome while demonstrating the selectivity of individual SH2 domains. The interactions described confirm virtually all previously reported associations while describing a large set of potential novel interactions that imply additional complexity in the signaling networks initiated from activated receptors. This study of pTyr ligand binding by SH2 domains provides valuable insight into the selectivity that underpins complex signaling networks that are assembled using modular protein interaction domains.

The SH2 domain-containing proteins in 21 species establish the provenance and scope of phosphotyrosine signaling in eukaryotes.

The Src homology 2 (SH2) domains are participants in metazoan signal transduction, acting as primary mediators for regulated protein-protein interactions with tyrosine-phosphorylated substrates. Here, we describe the origin and evolution of SH2 domain proteins by means of sequence analysis from 21 eukaryotic organisms from the basal unicellular eukaryotes, where SH2 domains first appeared, through the multicellular animals and increasingly complex metazoans. On the basis of our results, SH2 domains and phosphotyrosine signaling emerged in the early Unikonta, and the numbers of SH2 domains expanded in the choanoflagellate and metazoan lineages with the development of tyrosine kinases, leading to rapid elaboration of phosphotyrosine signaling in early multicellular animals. Our results also indicated that SH2 domains coevolved and the number of the domains expanded alongside protein tyrosine kinases and tyrosine phosphatases, thereby coupling phosphotyrosine signaling to downstream signaling networks. Gene duplication combined with domain gain or loss produced novel SH2-containing proteins that function within phosphotyrosine signaling, which likely have contributed to diversity and complexity in metazoans. We found that intra- and intermolecular interactions within and between SH2 domain proteins increased in prevalence along with organismal complexity and may function to generate more highly connected and robust phosphotyrosine signaling networks.

SH2 domains recognize contextual peptide sequence information to determine selectivity.

Selective ligand recognition by modular protein interaction domains is a primary determinant of specificity in signaling pathways. Src homology 2 (SH2) domains fulfill this capacity immediately downstream of tyrosine kinases, acting to recruit their host polypeptides to ligand proteins harboring phosphorylated tyrosine residues. The degree to which SH2 domains are selective and the mechanisms underlying selectivity are fundamental to understanding phosphotyrosine signaling networks. An examination of interactions between 50 SH2 domains and a set of 192 phosphotyrosine peptides corresponding to physiological motifs within FGF, insulin, and IGF-1 receptor pathways indicates that individual SH2 domains have distinct recognition properties and exhibit a remarkable degree of selectivity beyond that predicted by previously described binding motifs. The underlying basis for such selectivity is the ability of SH2 domains to recognize both permissive amino acid residues that enhance binding and non-permissive amino acid residues that oppose binding in the vicinity of the essential phosphotyrosine. Neighboring positions affect one another so local sequence context matters to SH2 domains. This complex linguistics allows SH2 domains to distinguish subtle differences in peptide ligands. This newly appreciated contextual dependence substantially increases the accessible information content embedded in the peptide ligands that can be effectively integrated to determine binding. This concept may serve more broadly as a paradigm for subtle recognition of physiological ligands by protein interaction domains.

High-throughput phosphotyrosine profiling using SH2 domains.

Protein tyrosine phosphorylation controls many aspects of signaling in multicellular organisms. One of the major consequences of tyrosine phosphorylation is the creation of binding sites for proteins containing Src homology 2 (SH2) domains. To profile the global tyrosine phosphorylation state of the cell, we have developed proteomic binding assays encompassing nearly the full complement of human SH2 domains. Here we provide a global view of SH2 domain binding to cellular proteins based on large-scale far-western analyses. We also use reverse-phase protein arrays to generate comprehensive, quantitative SH2 binding profiles for phosphopeptides, recombinant proteins, and entire proteomes. As an example, we profiled the adhesion-dependent SH2 binding interactions in fibroblasts and identified specific focal adhesion complex proteins whose tyrosine phosphorylation and binding to SH2 domains are modulated by adhesion. These results demonstrate that high-throughput comprehensive SH2 profiling provides valuable mechanistic insights into tyrosine kinase signaling pathways.

The human and mouse complement of SH2 domain proteins-establishing the boundaries of phosphotyrosine signaling.

SH2 domains are interaction modules uniquely dedicated to the recognition of phosphotyrosine sites and are embedded in proteins that couple protein-tyrosine kinases to intracellular signaling pathways. Here, we report a comprehensive bioinformatics, structural, and functional view of the human and mouse complement of SH2 domain proteins. This information delimits the set of SH2-containing effectors available for PTK signaling and will facilitate the systems-level analysis of pTyr-dependent protein-protein interactions and PTK-mediated signal transduction. The domain-based architecture of SH2-containing proteins is of more general relevance for understanding the large family of protein interaction domains and the modular organization of the majority of human proteins.