A Review of Calcineurin Biophysics with Implications for Cardiac Physiology.

Calcineurin, also known as protein phosphatase 2B, is a heterodimeric serine threonine phosphatase involved in numerous signaling pathways. During the past 50 years, calcineurin has been the subject of extensive investigation. Many of its cellular and physiological functions have been described, and the underlying biophysical mechanisms are the subject of active investigation. With the abundance of techniques and experimental designs utilized to study calcineurin and its numerous substrates, it is difficult to reconcile the available information. There have been a plethora of reports describing the role of calcineurin in cardiac disease. However, a physiological role of calcineurin in healthy cardiomyocyte function requires clarification. Here, we review the seminal biophysical and structural details that are responsible for the molecular function and inhibition of calcineurin. We then focus on literature describing the roles of calcineurin in cardiomyocyte physiology and disease.

Neuronal phase consistency tracks dynamic changes in acoustic spectral regularity.

The brain parses the auditory environment into distinct sounds by identifying those acoustic features in the environment that have common relationships (e.g., spectral regularities) with one another and then grouping together the neuronal representations of these features. Although there is a large literature that tests how the brain tracks spectral regularities that are predictable, it is not known how the auditory system tracks spectral regularities that are not predictable and that change dynamically over time. Furthermore, the contribution of brain regions downstream of the auditory cortex to the coding of spectral regularity is unknown. Here, we addressed these two issues by recording electrocorticographic activity, while human patients listened to tone-burst sequences with dynamically varying spectral regularities, and identified potential neuronal mechanisms of the analysis of spectral regularities throughout the brain. We found that the degree of oscillatory stimulus phase consistency (PC) in multiple neuronal-frequency bands tracked spectral regularity. In particular, PC in the delta-frequency band seemed to be the best indicator of spectral regularity. We also found that these regularity representations existed in multiple regions throughout cortex. This widespread reliable modulation in PC - both in neuronal-frequency space and in cortical space - suggests that phase-based modulations may be a general mechanism for tracking regularity in the auditory system specifically and other sensory systems more generally. Our findings also support a general role for the delta-frequency band in processing the regularity of auditory stimuli.

Synchronous and asynchronous theta and gamma activity during episodic memory formation.

To test the hypothesis that neural oscillations synchronize to mediate memory encoding, we analyzed electrocorticographic recordings taken as 68 human neurosurgical patients studied and subsequently recalled lists of common words. To the extent that changes in spectral power reflect synchronous oscillations, we would expect those power changes to be accompanied by increases in phase synchrony between the region of interest and neighboring brain areas. Contrary to the hypothesized role of synchronous gamma oscillations in memory formation, we found that many key regions that showed power increases during successful memory encoding also exhibited decreases in global synchrony. Similarly, cortical theta activity that decreases during memory encoding exhibits both increased and decreased global synchrony depending on region and stage of encoding. We suggest that network synchrony analyses, as used here, can help to distinguish between two major types of spectral modulations: (1) those that reflect synchronous engagement of regional neurons with neighboring brain areas, and (2) those that reflect either asynchronous modulations of neural activity or local synchrony accompanied by global disengagement from neighboring regions. We show that these two kinds of spectral modulations have distinct spatiotemporal profiles during memory encoding.

Predictive Modeling of Phenotypic Antimicrobial Susceptibility of Selected Beta-Lactam Antimicrobials from Beta-Lactamase Resistance Genes.

The outcome of bacterial infection management relies on prompt diagnosis and effective treatment, but conventional antimicrobial susceptibility testing can be slow and labor-intensive. Therefore, this study aims to predict phenotypic antimicrobial susceptibility of selected beta-lactam antimicrobials in the bacteria of the family Enterobacteriaceae from different beta-lactamase resistance genotypes. Using human datasets extracted from the Antimicrobial Testing Leadership and Surveillance (ATLAS) program conducted by Pfizer and retail meat datasets from the National Antimicrobial Resistance Monitoring System for Enteric Bacteria (NARMS), we used a robust or weighted least square multivariable linear regression modeling framework to explore the relationship between antimicrobial susceptibility data of beta-lactam antimicrobials and different types of beta-lactamase resistance genes. In humans, in the presence of the blaCTX-M-1, blaCTX-M-2, blaCTX-M-8/25, and blaCTX-M-9 groups, MICs of cephalosporins significantly increased by values between 0.34-3.07 µg/mL, however, the MICs of carbapenem significantly decreased by values between 0.81-0.87 µg/mL. In the presence of carbapenemase genes (blaKPC, blaNDM, blaIMP, and blaVIM), the MICs of cephalosporin antimicrobials significantly increased by values between 1.06-5.77 µg/mL, while the MICs of carbapenem antimicrobials significantly increased by values between 5.39-67.38 µg/mL. In retail meat, MIC of ceftriaxone increased significantly in the presence of blaCMY-2, blaCTX-M-1, blaCTX-M-55, blaCTX-M-65, and blaSHV-2 by 55.16 µg/mL, 222.70 µg/mL, 250.81 µg/mL, 204.89 µg/mL, and 31.51 µg/mL respectively. MIC of cefoxitin increased significantly in the presence of blaCTX-M-65 and blaTEM-1 by 1.57 µg/mL and 1.04 µg/mL respectively. In the presence of blaCMY-2, MIC of cefoxitin increased by an average of 8.66 µg/mL over 17 years. Compared to E. coli isolates, MIC of cefoxitin in Salmonella enterica isolates decreased significantly by 0.67 µg/mL. On the other hand, MIC of ceftiofur increased in the presence of blaCTX-M-1, blaCTX-M-65, blaSHV-2, and blaTEM-1 by 8.82 µg/mL, 9.11 µg/mL, 8.18 µg/mL, and 1.04 µg/mL respectively. In the presence of blaCMY-2, MIC of ceftiofur increased by an average of 10.20 µg/mL over 14 years. The ability to predict antimicrobial susceptibility of beta-lactam antimicrobials directly from beta-lactamase resistance genes may help reduce the reliance on routine phenotypic testing with higher turnaround times in diagnostic, therapeutic, and surveillance of antimicrobial-resistant bacteria of the family Enterobacteriaceae.

Comparing individual antimicrobial resistant and multi-drug resistant Salmonella enterica across serotypes, sampling sources, sampling periods, and food animal types in the United States (2014-2018).

This study aimed to compare the antimicrobial-resistant Salmonella enterica profiles from three sampling sources cecal contents, HACCP (during processing), and retail meat using phenotypic antibiotic susceptibility and serotype data gathered from 2014 and 2018. Antimicrobial resistance data for 29 major Salmonella serotypes from three sampling sources and associated food animal types (cattle, swine, chicken, and turkey) were obtained from the database of the United States National Antimicrobial Resistance Monitoring System. Using multivariable logistic regression models, we compared individual and multi-drug resistance (MDR) in Salmonella enterica between the three sampling sources, food animal types, sampling period, and Salmonella serotypes. Across the three sources and throughout the sampling period, the recovery of antimicrobial-resistant Salmonella enterica - including MDR, MDR-AmpC, and ACSSuT - among food animal types were dependent on the sampling period and, in some cases, sampling sources and period for the selected antimicrobials. The predicted probability of antimicrobial resistance was greater in Salmonella serotypes from turkey compared to other food animal types, conditional on sampling sources. Ceftriaxone-resistant (OR=0.83, 95% CI: 0.69-0.99), and Sulfisoxazole-resistant (OR=0.84, 95% CI: 0.72-0.98) Salmonella serotypes were less likely to be recovered from the Hazard Analysis and Critical Control Point (HACCP) sources than with the cecal sources. Except for Salmonella serotypes Dublin and Newport, most of the Salmonella serotypes were less likely to be resistant to the selected antimicrobials, or found as MDR, compared to serotype Typhimurium. This study offers an integrated view on the predicted probability of MDR Salmonella serotypes, as well as insights into which serotypes are persistent, emerging or declining across sampling sources and food animal types in the United States.

Human disease-associated calmodulin mutations alter calcineurin function through multiple mechanisms.

Calmodulin (CaM) is a ubiquitous, calcium-sensing protein that regulates a multitude of processes throughout the body. In response to changes in [Ca2+], CaM modifies, activates, and deactivates enzymes and ion channels, as well as many other cellular processes. The importance of CaM is highlighted by the conservation of an identical amino acid sequence in all mammals. Alterations to CaM amino acid sequence were once thought to be incompatible with life. During the last decade modifications to the CaM protein sequence have been observed in patients suffering from life-threatening heart disease (calmodulinopathy). Thus far, inadequate or untimely interaction between mutant CaM and several proteins (LTCC, RyR2, and CaMKII) have been identified as mechanisms underlying calmodulinopathy. Given the extensive number of CaM interactions in the body, there are likely many consequences for altering CaM protein sequence. Here, we demonstrate that disease-associated CaM mutations alter the sensitivity and activity of the Ca2+-CaM-enhanced serine/threonine phosphatase calcineurin (CaN). Biophysical characterization by circular dichroism, solution NMR spectroscopy, stopped-flow kinetic measurements, and MD simulations provide mechanistic insight into mutation dysfunction as well as highlight important aspects of CaM Ca2+ signal transduction. We find that individual CaM point mutations (N53I, F89L, D129G, and F141L) impair CaN function, however, the mechanisms are not the same. Specifically, individual point mutations can influence or modify the following properties: CaM binding, Ca2+ binding, and/or Ca2+kinetics. Moreover, structural aspects of the CaNCaM complex can be altered in manners that indicate changes to allosteric transmission of CaM binding to the enzyme active site. Given that loss of CaN function can be fatal, as well as evidence that CaN modifies ion channels already associated with calmodulinopathy, our results raise the possibility that altered CaN function contributes to calmodulinopathy.

Resistance to extended-spectrum cephalosporins in Escherichia coli and Salmonella enterica isolated from food-producing animals: Ecological study from selected national surveillance programs.

Extended-spectrum cephalosporins (ESC) are categorized by World Health Organization as critically important antimicrobials with limited therapeutic alternatives for the treatment of severe bacterial infections in humans. Preserving the effectiveness of ESC requires continuous monitoring of resistance and comparison of associated data across national surveillance programs in the face of globalization. In this ecological study, we compared ESC resistance in Escherichia coli and Salmonella enterica isolated from food-producing animals from 2003 to 2019 between nine countries (Canada, Denmark, Finland, Japan, Netherlands, Norway, Sweden, United Kingdom, and the United States). Using the beta-regression model, compared to Canada, non-selective ESC-R Salmonella enterica was less likely isolated from food producing animals in other eight countries (Odds ratio range: 0.07-0.76). We observed an interaction between the country and the year with a significantly decreased proportion (P < 0.05) of non-selective ESC-R Escherichia coli from the Netherlands, the United Kingdom, and the United States compared to Canada over the years. There was a linear correlation between non-selective ESC-R Escherichia coli and ESC use from Netherlands (Spearman's ρ = 0.91, P < 0.0001). For the six European countries, the interaction between the country and year showed a significant decrease in the proportion of selective ESC-R Escherichia coli over the years for the Netherlands compared to Denmark (P = 0.002). While there were variations in the proportion of beta-lactamase genes reported over the years, blaCTX-M and blaCMY-2 genes were commonly detected among the selective ESC-R Escherichia coli. This study reveals variability in the recovery of ESC-resistant bacteria among the countries that seems likely influenced by the individual country policy on the use of critically important antimicrobials and resistance surveillance programs. However, there is a need for harmonization and consistency in food animal sources of bacterial isolates used in surveillance programs within and between the countries for easy comparability.

Suicide risk in older adults: evaluating models of risk and predicting excess zeros in a primary care sample.

Research is needed that examines theory-based risk factors for suicide in older adults. The interpersonal theory of suicide (Joiner, 2005; Van Orden et al., 2010) provides specific hypotheses regarding variables that contribute to the development and variability in death ideation and suicide ideation; however, data suggest that older adults may not report suicide ideation in research settings or to treatment providers even when they experience it (Heisel et al., 2006). The purpose of this study was to test theory-based predictions regarding variables that contribute to death ideation (i.e., a passive wish to die) and suicide ideation in older adults. This study introduces the application of zero-inflated negative binomial regression (ZINB) to the study of suicidal behavior. ZINB was used to test theory-based predictions, while also testing a hypothesis regarding variables associated with denial of suicide ideation among participants who endorsed risk factors associated with suicide risk. Participants included 239 adults aged 60 and older recruited from primary care clinics who completed a variety of self-report instruments. The results of this study indicated that perceived burdensomeness and hopelessness were significantly associated with variability in death ideation. Additional results indicated that elevated scores on thwarted belonging, the interaction between perceived burdensomeness and hopelessness, and the interaction between thwarted belonging and perceived burdensomeness were associated with a significant reduction in the probability of a participant being a suicide ideator. These results offer substantial support for the interpersonal theory of suicide. The implications of these findings are discussed.

Myoglobin-CO conformational substate dynamics: 2D vibrational echoes and MD simulations.

Two-dimensional (2D) infrared vibrational echoes were performed on horse heart carbonmonoxymyoglobin (MbCO) in water over a range of temperatures. The A(1) and A(3) conformational substates of MbCO are found to have different dephasing rates with different temperature dependences. A frequency-frequency correlation function derived from molecular dynamics simulations on MbCO at 298 K is used to calculate the vibrational echo decay. The calculated decay shows substantial agreement with the experimentally measured decays. The 2D vibrational echo probes protein dynamics and provides an observable that can be used to test structural assignments for the MbCO conformational substates.