Faculty Bibliography

PURPOSE OF REVIEW/OBJECTIVE:Hypernatremia is a relatively frequent electrolyte disorder seen in critically ill patients. As many as 27% of patients in intensive care units (ICUs) develop hypernatremia of variable severity during an ICU stay. Debate among specialists often ensues as to whether to correct hypernatremia or not. Some practitioners, particularly intensivists, believe that correction of hypernatremia with fluids may cause expansion of the extracellular fluid volume (ECFV) thereby worsening ventilation and impeding extubation. Other practitioners, including many nephrologists, do not expect correction of hypernatremia to lead to clinically apparent ECFV expansion, and fear other deleterious effects of hypernatremia. In this review we address the controversy regarding appropriate practice. FINDINGS/RESULTS:There are no randomized, clinical trials (RCTs) to guide the administration of electrolyte-free fluid administration in hypernatremic patients. However, there are associations, demonstrated in the literature, suggesting that hypernatremia of any severity will increase the mortality and length of stay in these patients. These associations generally support the practice of correction of hypernatremia. In addition, our knowledge of the distribution of total body water influences us towards correcting hypernatremia as an appropriate therapy. We do not expect that adequate RCTs addressing this question will be performed. SUMMARY/CONCLUSIONS:Allowing persistence of any degree of hypernatremia is associated with increased mortality, length of stay (LOS) and postdischarge mortality. We expect that proper use of electrolyte-free water intake will avoid adverse outcomes.

The engineering of autologous patient T cells for adoptive cell therapies has revolutionized the treatment of several types of cancer¹. However, further improvements are needed to increase response and cure rates. CRISPR-based loss-of-function screens have been limited to negative regulators of T cell functions^2-4 and raise safety concerns owing to the permanent modification of the genome. Here we identify positive regulators of T cell functions through overexpression of around 12,000 barcoded human open reading frames (ORFs). The top-ranked genes increased the proliferation and activation of primary human CD4⁺ and CD8⁺ T cells and their secretion of key cytokines such as interleukin-2 and interferon-γ. In addition, we developed the single-cell genomics method OverCITE-seq for high-throughput quantification of the transcriptome and surface antigens in ORF-engineered T cells. The top-ranked ORF-lymphotoxin-β receptor (LTBR)-is typically expressed in myeloid cells but absent in lymphocytes. When overexpressed in T cells, LTBR induced profound transcriptional and epigenomic remodelling, leading to increased T cell effector functions and resistance to exhaustion in chronic stimulation settings through constitutive activation of the canonical NF-κB pathway. LTBR and other highly ranked genes improved the antigen-specific responses of chimeric antigen receptor T cells and γδ T cells, highlighting their potential for future cancer-agnostic therapies⁵. Our results provide several strategies for improving next-generation T cell therapies by the induction of synthetic cell programmes.

BACKGROUND:With the explosion of COVID-19 globally, it was unclear if people with Parkinson's disease (PD) were at increased risk for severe manifestations or negative outcomes. OBJECTIVES/OBJECTIVE:To report on people with PD who had suspected or confirmed COVID-19 to understand how COVID-19 manifested in PD patients. METHODS:We surveyed PD patients who reported COVID-19 to their Movement Disorders specialists at Columbia University Irving Medical Center and respondents from an online survey administered by the Parkinson's Foundation that assessed COVID-19 symptoms, general clinical outcomes and changes in motor and non-motor PD symptoms. RESULTS:Forty-six participants with PD and COVID-19 were enrolled. Similar to the general population, the manifestations of COVID-19 among people with PD were heterogeneous ranging from asymptomatic carriers (1/46) to death (6/46). The most commonly reported COVID-19 symptoms were fever/chills, fatigue, cough, weight loss, and muscle pain. Worsening and new onset of motor and non-motor PD symptoms during COVID-19 illness were also reported, including dyskinesia, rigidity, balance disturbances, anxiety, depression, and insomnia. CONCLUSION/CONCLUSIONS:We did not find sufficient evidence that PD is an independent risk factor for severe COVID-19 and death. Larger studies with controls are required to understand this further. Longitudinal follow-up of these participants will allow for observation of possible long-term effects of COVID-19 in PD patients.

OBJECTIVES/OBJECTIVE:Salivary hypofunction and xerostomia, are common side effects of radiotherapy, negatively impacting quality of life. The OraRad study presents results on the longitudinal impact of radiotherapy on salivary flow and patient-reported outcomes. PATIENTS AND METHODS/METHODS:Prospective, multicenter cohort study of 572 patients receiving curative-intent head and neck radiotherapy (RT). Stimulated salivary flow (SSF) rate and patient-reported outcomes were measured prior to RT and at 6- and 18-months post-RT. Linear mixed effects models examined the relationship between RT dose and change in salivary flow, and change in patient-reported outcomes. RESULTS:544 patients had baseline salivary flow measurement, with median (IQR) stimulated flow rate of 0.975 (0.648, 1.417) g/min. Average RT dose to parotid glands was associated with change in salivary flow post-RT (p < 0.001). Diminished flow to 37% of pre-RT level was observed at 6 months (median: 0.358, IQR: 0.188 to 0.640 g/min, n = 481) with partial recovery to 59% of pre-RT at 18 months (median: 0.575, IQR: 0.338 to 0.884 g/min, n = 422). Significant improvement in patient-reported swallowing, senses (taste and smell), mouth opening, dry mouth, and sticky saliva (p-values < 0.03) were observed between 6 and 18 months post-RT. Changes in swallowing, mouth opening, dry mouth, and sticky saliva were significantly associated with changes in salivary flow from baseline (p-values < 0.04). CONCLUSION/CONCLUSIONS:Salivary flow and patient-reported outcomes decreased as a result of RT, but demonstrated partial recovery during follow-up. Continued efforts are needed to improve post-RT salivary function to support quality of life.

Ambiguous images elicit bistable perception, wherein periods of momentary perceptual stability are interrupted by sudden perceptual switches. When intermittently presented, ambiguous images trigger a perceptual memory trace in the intervening blank periods. Understanding the neural bases of perceptual stability and perceptual memory during bistable perception may hold clues for explaining the apparent stability of visual experience in the natural world, where ambiguous and fleeting images are prevalent. Motivated by recent work showing the involvement of the right inferior frontal gyrus (rIFG) in bistable perception, we conducted a transcranial direct-current stimulation (tDCS) study with a double-blind, within-subject cross-over design to test a potential causal role of rIFG in these processes. Subjects viewed ambiguous images presented continuously or intermittently while under EEG recording. We did not find any significant tDCS effect on perceptual behavior. However, the fluctuations of oscillatory power in the alpha and beta bands predicted perceptual stability, with higher power corresponding to longer percept durations. In addition, higher alpha and beta power predicted enhanced perceptual memory during intermittent viewing. These results reveal a unified neurophysiological mechanism sustaining perceptual stability and perceptual memory when the visual system is faced with ambiguous input.

In understanding circuit operations, a key problem is the extent to which neuronal spiking reflects local computation or responses to upstream inputs. We addressed this issue in the hippocampus by performing combined optogenetic and pharmacogenetic local and upstream inactivation. Silencing the medial entorhinal cortex (mEC) largely abolished extracellular theta and gamma currents in CA1 while only moderately affecting firing rates. In contrast, CA3 and local CA1 silencing strongly decreased firing of CA1 neurons without affecting theta currents. Each perturbation reconfigured the CA1 spatial map. However, the ability of the CA1 circuit to support place field activity persisted, maintaining the same fraction of spatially tuned place fields and reliable assembly expression as in the intact mouse. Thus, the CA1 network can induce and maintain coordinated cell assemblies with minimal reliance on its inputs, but these inputs can effectively reconfigure and assist in maintaining stability of the CA1 map.

G protein-coupled receptors (GPCRs) regulate many pathophysiological processes and are major therapeutic targets. The impact of disease on the subcellular distribution and function of GPCRs is poorly understood. We investigated trafficking and signaling of protease-activated receptor 2 (PAR₂) in colitis. To localize PAR₂ and assess redistribution during disease, we generated knockin mice expressing PAR₂ fused to monomeric ultrastable green fluorescent protein (muGFP). PAR₂-muGFP signaled and trafficked normally. PAR₂ messenger RNA was detected at similar levels in Par₂-mugfp and wild-type mice. Immunostaining with a GFP antibody and RNAScope in situ hybridization using F2rl1 (PAR₂) and Gfp probes revealed that PAR₂-muGFP was expressed in epithelial cells of the small and large intestine and in subsets of enteric and dorsal root ganglia neurons. In healthy mice, PAR₂-muGFP was prominently localized to the basolateral membrane of colonocytes. In mice with colitis, PAR₂-muGFP was depleted from the plasma membrane of colonocytes and redistributed to early endosomes, consistent with generation of proinflammatory proteases that activate PAR₂ PAR₂ agonists stimulated endocytosis of PAR₂ and recruitment of Gα_q, Gα_i, and β-arrestin to early endosomes of T84 colon carcinoma cells. PAR₂ agonists increased paracellular permeability of colonic epithelial cells, induced colonic inflammation and hyperalgesia in mice, and stimulated proinflammatory cytokine release from segments of human colon. Knockdown of dynamin-2 (Dnm2), the major colonocyte isoform, and Dnm inhibition attenuated PAR₂ endocytosis, signaling complex assembly and colonic inflammation and hyperalgesia. Thus, PAR₂ endocytosis sustains protease-evoked inflammation and nociception and PAR₂ in endosomes is a potential therapeutic target for colitis.

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an arrhythmia syndrome due to gene mutations that render RYR2 calcium release channels hyperactive, causing spontaneous calcium release and delayed afterdepolarizations (DADs). What remains unknown is the cellular source of ventricular arrhythmia triggered by DADs - Purkinje cells in the conduction system or ventricular cardiomyocytes in the working myocardium. To answer this question, we used a genetic approach in mice to knock out cardiac calsequestrin either in Purkinje cells or in ventricular cardiomyocytes. Total loss of calsequestrin in the heart causes a severe CPVT phenotype in mice and humans. We found that loss of calsequestrin only in ventricular myocytes produced a full-blown CPVT phenotype, whereas mice with loss of calsequestrin only in Purkinje cells were comparable to wild-type mice. Subendocardial chemical ablation or restoration of calsequestrin expression in subendocardial cardiomyocytes neighboring Purkinje cells was sufficient to protect against catecholamine-induced arrhythmias. In silico modeling demonstrated that DADs in ventricular myocardium can trigger full action potentials in the Purkinje fiber, but not vice versa. Hence, ectopic beats in CPVT are likely generated at the Purkinje-myocardial junction via a heretofore unrecognized tissue mechanism, whereby DADs in the ventricular myocardium trigger full action potentials in adjacent Purkinje cells.

Understanding how excitatory (E) and inhibitory (I) inputs are integrated by neurons requires monitoring their subthreshold behavior. We probed the subthreshold dynamics using optogenetic depolarizing pulses in hippocampal neuronal assemblies in freely moving mice. Excitability decreased during sharp-wave ripples coupled with increased I. In contrast to this "negative gain," optogenetic probing showed increased within-field excitability in place cells by weakening I and unmasked stable place fields in initially non-place cells. Neuronal assemblies active during sharp-wave ripples in the home cage predicted spatial overlap and sequences of place fields of both place cells and unmasked preexisting place fields of non-place cells during track running. Thus, indirect probing of subthreshold dynamics in neuronal populations permits the disclosing of preexisting assemblies and modes of neuronal operations.