Formulation Development of a COVID-19 Recombinant Spike Protein-Based Vaccine.

The purpose of this study was to develop a formulation for a recombinant prefusion spike protein vaccine against SARS-CoV-2. It was found that the spike protein was susceptible to aggregation due to mechanical stress. Therefore, formulation studies were initiated focused on screening pharmaceutical excipients capable of preventing this. The screening of a panel of potential stabilizing conditions found that Tween 20 could inhibit mechanically induced aggregation. A concentration-dependent study indicated that a higher concentration of Tween 20 (0.2% v/v) was required to prevent conformational changes in the trimer. The conformational changes induced by mechanical stress were characterized by size exclusion chromatography (SEC) and hydrogen-deuterium exchange mass spectrometry (HDX-MS), indicating the formation of an extended trimeric conformation that was also unable to bind to antibodies directed to the S2 domain. Long-term stability modeling, using advanced kinetic analysis, indicated that the formulation containing 0.2% (v/v) Tween 20 at a neutral pH was predicted to be stable for at least two years at 2 °C to 8 °C. Additional stabilizer screening conducted by thermal shift assay indicated that sucrose and glycerol were able to significantly increase the spike protein melting temperature (Tm) and improve the overall thermostability of the spike protein in a short-term stability study. Thus, while 0.2% (v/v) Tween 20 was sufficient to prevent aggregation and to maintain spike protein stability under refrigeration, the addition of sucrose further improved vaccine thermostability. Altogether, our study provides a systematic approach to the formulation of protein-based COVID-19 vaccine and highlights the impact of mechanical stress on the conformation of the spike protein and the significance of surfactants and stabilizers in maintaining the structural and functional integrity of the spike protein.

Perioperative and Long-Term Outcomes in Infants Undergoing a Tracheostomy from a Neonatal Intensive Care Unit.

OBJECTIVE: The purpose of this study was to identify risk factors for perioperative complications and long-term morbidity in infants from the neonatal intensive care unit (NICU) presenting for a tracheostomy. METHODS: This single-center retrospective cohort study included infants in the NICU presenting for a tracheostomy from August 2011 to December 2019. Primary outcomes were categorized as either a perioperative complication or long-term morbidity. A severe perioperative complication was defined as having either (1) an intraoperative cardiopulmonary arrest, (2) an intraoperative death, (3) a postoperative cardiopulmonary arrest within 30 days of the procedure, or (4) a postoperative death within 30 days of the procedure. Long-term morbidities included (1) the need for gastrostomy tube placement within the tracheostomy hospitalization and (2) the need for diuretic therapy, pulmonary hypertensive therapy, oxygen, or mechanical ventilation at 12 and 24 months following the tracheostomy. RESULTS: One-hundred eighty-three children underwent a tracheostomy. The mean age at tracheostomy was 16.9 weeks while the mean post-conceptual age at tracheostomy was 49.7 weeks. The incidence of severe perioperative complications was 4.4% (n = 8) with the number of pulmonary hypertension medication classes preoperatively (OR: 3.64, 95% CI: (1.44-8.94), p = 0.005) as a significant risk factor. Approximately 81% of children additionally had a gastrostomy tube placed at the time of the tracheostomy, and 62% were ventilator-dependent 2 years following their tracheostomy. CONCLUSION: Our study provides critical perioperative complications and long-term morbidity data to neonatologists, pediatricians, surgeons, anesthesiologists, and families in the expected course of infants from the NICU presenting for a tracheostomy. LEVEL OF EVIDENCE: 3 Laryngoscope, 134:1945-1954, 2024.

Screening for ADHD in Adult Patients With Epilepsy: Prevalence of Symptoms and Challenges to Diagnosis.

OBJECTIVE: Given the complex nature of seizure disorders and their treatments, ADHD may be underdiagnosed in this population. We hypothesized that a higher percentage of patients presenting to a seizure clinic would endorse ADHD symptoms compared to rates reported in the general population and that formal screening for ADHD symptoms would identify patients with previously undiagnosed comorbid ADHD. METHODS: In this study, we surveyed 312 adults in a seizure clinic using the Adult ADHD Self-Report Scale (ASRS-v1.1). RESULTS: We found that 90 patients (28.8%) screened positive with the ASRS-v1.1, but only nine of these patients were able to complete neuropsychological testing,. Out of these patients, only one was diagnosed with possible ADHD. CONCLUSION: Through this process, we identified many challenges to making a new ADHD diagnosis in this population, including attention deficits due to other medical or psychiatric diagnoses, a positive urine drug screen, lack of collateral report/information about developmental history, and barriers to neuropsychological evaluation.

Associations of Incomplete SARS-CoV-2 Vaccination among Patients with Unstable Housing in Houston.

Vaccination is a safe and effective way to protect against SARS-CoV-2. Two of the three authorized SARS-CoV-2 vaccines require two doses, presenting logistical challenges. Those with unstable housing face barriers that amplify these challenges. In this study, we utilized a database maintained by Healthcare for the Homeless-Houston to determine the rates of partial vaccination among those with unstable housing in Houston (n=294). We then performed post-hoc analyses to identify predictors of partial vaccination. Our key finding was that 30% of those with unstable housing missed their second dose, a proportion far higher than the national average. Those with permanent supportive housing and those who had a Harris County Gold Card (financial assistance for health care costs) were more likely to return for dose two, while those who were younger, living on the streets, or staying in a temporary homeless shelter were more likely to miss the second dose.

Impact of Palliative Care on Interhospital Transfers to the Intensive Care Unit.

Community hospitals will often transfer their most complex, critically ill patients to intensive care units (ICUs) of tertiary care centers for specialized, comprehensive care. This population of patients has high rates of morbidity and mortality. Palliative care involvement in critically ill patients has been demonstrated to reduce over-utilization of resources and hospital length of stays. We hypothesized that transfers from community hospitals had low rates of palliative care involvement and high utilization of ICU resources. In this single-center retrospective cohort study, 848 patients transferred from local community hospitals to the medical ICU (MICU) and cardiac care unit (CCU) at a tertiary care center between 2016-2018 were analyzed for patient disposition, length of stay, hospitalization cost, and time to palliative care consultation. Of the 848 patients, 484 (57.1%) expired, with 117 (13.8%) having expired within 48 hours of transfer. Palliative care consult was placed for 201 (23.7%) patients. Patients with palliative care consult were statistically more likely to be referred to hospice (p<0.001). Over two-thirds of palliative care consults were placed later than 5 days after transfer. Time to palliative care consult was positively correlated with length of hospitalization among MICU patients (r=0.79) and CCU patients (r=0.90). Time to palliative consult was also positively correlated with hospitalization cost among MICU patients (r=0.75) and CCU patients (r=0.86). These results indicate early palliative care consultation in this population may result in timely goals of care discussions and optimization of resources.

The development of the Supplemental Nutrition Assistance Program enrollment accessibility (SNAP-access) score.

BACKGROUND: The Supplemental Nutrition Assistance Program (SNAP) is a federal public benefit providing food assistance to millions of Americans. However, it is typically administered by states, creating potential variation in accessibility and transparency of information about enrollment for people with disabilities. OBJECTIVE: To develop and demonstrate the use of a method to assess the accessibility and transparency of information about the disability-inclusive process and practices of SNAP enrollment. METHODS: Cross-sectional data was collected from SNAP landing and enrollment webpages from all 50 U.S. states, the District of Columbia, and New York City from June-August 2021. Based on principles of universal design and accessibility, scores were determined for each SNAP program across three areas: flexibility in the enrollment process (6 points), efficiency of finding information about enrollment on SNAP websites (6 points), and the accessibility of SNAP webpages (6 points). Total scores were the sum of these sub-categories (18 points maximum). RESULTS: Of the 52 SNAP programs assessed, mean scores were 10.66 (SD = 2.51) for the total score, 2.67 (SD = 0.91) for flexibility in the enrollment process, 3.32 (SD = 1.19) for efficiency of finding information about enrollment on SNAP websites, and 4.67 (SD = 1.72) for the accessibility of SNAP webpages. No programs received the maximum flexibility score (6 points) on flexibility, 2 programs received the maximum on efficiency, and 31 programs the maximum on accessibility. CONCLUSIONS: We found differences in the accessibility, flexibility, and efficiency of SNAP program enrollment information available on SNAP websites and outline room for improvement across all three of these areas.

Connecting DCX, COMT and FMR1 in social behavior and cognitive impairment.

Genetic variants of DCX, COMT and FMR1 have been linked to neurodevelopmental disorders related to intellectual disability and social behavior. In this systematic review we examine the roles of the DCX, COMT and FMR1 genes in the context of hippocampal neurogenesis with respect to these disorders with the aim of identifying important hubs and signaling pathways that may bridge these conditions. Taken together our findings indicate that factors connecting DCX, COMT, and FMR1 in intellectual disability and social behavior may converge at Wnt signaling, neuron migration, and axon and dendrite morphogenesis. Data derived from genomic research has identified a multitude of genes that are linked to brain disorders and developmental differences. Information about where and how these genes function and cooperate is lagging behind. The approach used here may help to shed light on the biological underpinnings in which key genes interface and may prove useful for the testing of specific hypotheses.

Sterol Sponge Mechanism Is Conserved for Glycosylated Polyene Macrolides.

Amphotericin-like glycosylated polyene macrolides (GPMs) are a clinically and industrially important family of natural products, but the mechanisms by which they exert their extraordinary biological activities have remained unclear for more than half a century. Amphotericin B exerts fungicidal action primarily via self-assembly into an extramembranous sponge that rapidly extracts ergosterol from fungal membranes, but it has remained unclear whether this mechanism is applicable to other GPMs. Using a highly conserved polyene-hemiketal region of GPMs that we hypothesized to represent a conserved ergosterol-binding domain, we bioinformatically mapped the entirety of the GPM sequence-function space and expanded the number of GPM biosynthetic gene clusters (BGCs) by 10-fold. We further leveraged bioinformatic predictions and tetrazine-based reactivity screening targeting the electron-rich polyene region of GPMs to discover a first-in-class methyltetraene- and diepoxide-containing GPM, kineosporicin, and to assign BGCs to many new producers of previously reported members. Leveraging a range of structurally diverse known and newly discovered GPMs, we found that the sterol sponge mechanism of fungicidal action is conserved.

PAT solutions to monitor adsorption of Tetanus Toxoid with aluminum adjuvants.

The focus of this study was to examine the small-scale adsorption process of Tetanus Toxoid (TT) as a model protein antigen to aluminum phosphate (AlPO4) and aluminum oxyhydroxide (AlOOH) adjuvants with real-time monitoring by in-line ReactIR™, ParticleTrack™ based on Focused Beam Reflectance Measurement (FBRM) and EasyViewer™ probes. The adsorption process of AlPO4 and AlOOH with TT using was monitored in the small-scale reactors. Conformational changes in TT were monitored using in-line infrared probe ReactIR, whereas particle formation associated with protein adsorption were measured by particle size, count, and imaging tools, such as ParticleTrack with FBRM and EasyViewer probes. ParticleTrack distribution results and kinetic measurements were also supported by observations made using EasyViewer. In addition to EasyMax, BioBLU reactor was also used for the adsorption experiments. ReactIR with ATR-Fiber probe was effectively able to monitor adsorption progress of TT to AlOOH and to AlPO4. ReactIR, EasyViewer, and ParticleTrack provided detailed mechanistic and kinetic information for reaction of TT with AlPO4 and AlOOH. These in-situ measurements revealed a possible multi-step process for TT to AlPO4 which may be an indication of antigen adsorption.

Absence of electrocardiographic left ventricular hypertrophy in patients undergoing Transcatheter aortic valve replacement is associated with increased mortality.

BACKGROUND: We examined the association between the absence ECG LVH and all-cause mortality in patients with severe AS undergoing TAVR. METHODS: We conducted a retrospective single center study on 399 TAVR patients from 2012 to 2016. ECGs were reviewed for LVH diagnosed by Sokolow-Lyon's voltage criteria. All patients met echocardiographic criteria for LVH. Logistic regression was used to examine the association between ECG LVH and covariates. Survival analysis was performed using Cox regression analysis and Kaplan Meier curves. RESULTS: Patients without ECG LVH were younger (81.0 ± 8.4 vs. 84.0 ± 7.7 years, p = 0.001) with a higher BMI (29.3 ± 7.0 vs. 27.1 ± 5.6 kg/m2, p = 0.006) and lower FEV1 (65.6 ± 22.8 vs. 74.1 ± 21.6%, p = 0.002). In multivariable analysis, increased BMI and decreased FEV1 remained predictive of the absence of ECG LVH. Over a mean follow-up time of 32 (± 17.0) months, the 5-year cumulative survival was 79% in the ECG LVH group and 58% in the group without ECG LVH (p = 0.039). Absence of ECG LVH remained predictive of all-cause mortality (HR 1.56, 95% CI 1.01-2.59, p = 0.045) in multivariable Cox regression analysis. When patients were grouped by comorbidities, patients with the highest mortality were those with increased BMI or decreased FEV1. CONCLUSIONS: Absence of LVH by ECG criteria in patients with severe AS undergoing TAVR was associated with increased all-cause mortality. Routinely performed, noninvasive and inexpensive ECG may aid in identification of high-risk patients that may not benefit from TAVR and warrant further evaluation of underlying comorbidities.

Endocrine regulation of MFS2 by branchless controls phosphate excretion and stone formation in Drosophila renal tubules.

How inorganic phosphate (Pi) homeostasis is regulated in Drosophila is currently unknown. We here identify MFS2 as a key Pi transporter in fly renal (Malpighian) tubules. Consistent with its role in Pi excretion, we found that dietary Pi induces MFS2 expression. This results in the formation of Malpighian calcium-Pi stones, while RNAi-mediated knockdown of MFS2 increases blood (hemolymph) Pi and decreases formation of Malpighian tubule stones in flies cultured on high Pi medium. Conversely, microinjection of adults with the phosphaturic human hormone fibroblast growth factor 23 (FGF23) induces tubule expression of MFS2 and decreases blood Pi. This action of FGF23 is blocked by genetic ablation of MFS2. Furthermore, genetic overexpression of the fly FGF branchless (bnl) in the tubules induces expression of MFS2 and increases Malpighian tubule stones suggesting that bnl is the endogenous phosphaturic hormone in adult flies. Finally, genetic ablation of MFS2 increased fly life span, suggesting that Malpighian tubule stones are a key element whereby high Pi diet reduces fly longevity previously reported by us. In conclusion, MFS2 mediates excretion of Pi in Drosophila, which is as in higher species under the hormonal control of FGF-signaling.

Oxygen and lack of oxygen in fetal and placental development, feto-placental coupling, and congenital heart defects.

Low oxygen concentration (hypoxia) is part of normal embryonic development, yet the situation is complex. Oxygen (O2 ) is a janus gas with low levels signaling through hypoxia-inducible transcription factor (HIF) that are required for development of fetal and placental vasculature and fetal red blood cells. This results in coupling of fetus and mother around midgestation as a functional feto-placental unit (FPU) for O2 transport, which is required for continued growth and development of the fetus. Defects in these processes may leave the developing fetus vulnerable to O2 deprivation or other stressors during this critical midgestational transition when common septal and conotruncal heart defects (CHDs) are likely to arise. Recent human epidemiological and case-control studies support an association between placental dysfunction, manifest as early onset pre-eclampsia (PE) and increased serum bio-markers, and CHD. Animal studies support this association, in particular those using gene inactivation in the mouse. Sophisticated methods for gene inactivation, cell fate mapping, and a quantitative bio-reporter of O2 concentration support the premise that hypoxic stress at critical stages of development leads to CHD. The secondary heart field contributing to the cardiac outlet is a key target, with activation of the un-folded protein response and abrogation of FGF signaling or precocious activation of a cardiomyocyte transcriptional program for differentiation, suggested as mechanisms. These studies provide a strong foundation for further study of feto-placental coupling and hypoxic stress in the genesis of human CHD.