The EPHA3 protein tyrosine kinase, a member of the ephrin receptor family, regulates cell fate, cell motility, and cell–cell interaction. These cellular events are critical for tissue development, immunological responses, and the processes of tumorigenesis. Earlier studies revealed that signaling via the STK4-encoded MST1 serine-threonine protein kinase, a core component of the Hippo pathway, attenuated EPHA3 expression. Here, we investigated the mechanism by which MST1 regulates EPHA3. Our findings have revealed that the transcriptional regulators YAP1 and TEAD1 are crucial activators of EPHA3 transcription. Silencing YAP1 and TEAD1 suppressed the EPHA3 protein and mRNA levels. In addition, we identified putative TEAD enhancers in the distal EPHA3 promoter, where YAP1 and TEAD1 bind and promote EPHA3 expression. Furthermore, EPHA3 knockout by CRISPR/Cas9 technology reduced cell–cell interaction and cell motility. These findings demonstrate that EPHA3 is transcriptionally regulated by YAP1/TEAD1 of the Hippo pathway, suggesting that it is sensitive to cell contact-dependent interactions.
This study investigated the influences of neighborhood factors (residential stability and neighborhood disadvantage) and variants of the serotonin transporter linked polymorphic region (5-HTTLPR) genotype on the development of substance use among African American children aged 10-24 years. To accomplish this, a harmonized data set of five longitudinal studies was created via pooling overlapping age cohorts to establish a database with 2,689 children and 12,474 data points to span ages 10-24 years. A description of steps used in the development of the harmonized data set is provided, including how issues such as the measurement equivalence of constructs were addressed. A sequence of multilevel models was specified to evaluate Gene × Environment effects on growth of substance use across time. Findings indicated that residential instability was associated with higher levels and a steeper gradient of growth in substance use across time. The inclusion of the 5-HTTLPR genotype provided greater precision to the relationships in that higher residential instability, in conjunction with the risk variant of 5-HTTLPR (i.e., the short allele), was associated with the highest level and steepest gradient of growth in substance use across ages 10-24 years. The findings demonstrated how the creation of a harmonized data set increased statistical power to test Gene × Environment interactions for an under studied sample.
Collective sex venues such as sex clubs are strategic sites to promote sexual health among sexual and gender minority individuals. We present qualitative findings from a multiple-method study on the acceptability of sexual-health services at collective sex venues in New York City (NYC) among attendees who identified as men, transgender, or gender non-conforming. In a survey used for sample selection (n = 342), most respondents (82.7%) agreed that “having outreach workers at sex venues is a good thing.” Interviewees (n = 30) appreciated how on-site services could promote sexual health in their community. They felt peer workers should be familiar with collective sex venues and share demographic characteristics with attendees. Some participants felt workers should keep some boundaries from attendees, while others felt they could be fully integrated in the environment, suggesting that either peer outreach or popular-opinion leader types of interventions could be feasible.
The kidney is a main site of erythropoietin production in the body. We developed a new method for the detection of Epo protein by deglycosylation-coupled Western blotting. Detection of deglycosylated Epo enables the examination of small changes in Epo production. Using this method, we investigated the effects of angiotensin II (ATII) on Epo production in the kidney. ATII stimulated the plasma Epo concentration; Epo, HIF2α, and PHD2 mRNA expression in nephron segments in the renal cortex and outer medulla; and Epo protein expression in the renal cortex. In situ hybridization and immunohistochemistry revealed that ATII stimulates Epo mRNA and protein expression not only in proximal tubules but also in collecting ducts, especially in intercalated cells. These data support the regulation of Epo production in the kidney by the renin–angiotensin– aldosterone system (RAS).
We evaluated the relationship between estimated 24-hour urinary sodium excretion from the Kawasaki, Tanaka, and INTERSALT (International Study of Sodium, Potassium, and Blood Pressure) formulas and blood pressure (BP). We pooled 10 034 person-visit data from 3 cohort studies in Bangladesh that had measured 24-hour urine sodium (m-24hUNa), potassium, creatinine excretion, and BP. We used m-24hUNa, potassium, and creatinine where necessary, rather than spot urine values in the formulas. Bland-Altman plots were used to determine the bias associated with formula-estimated sodium relative to m-24hUNa. We compared the sodium excretion and BP relationships from m-24hUNa versus formula-estimated sodium excretions, using restricted cubic spline plots for adjusted multilevel linear models. All formulas overestimated 24-hour sodium at lower levels but underestimated 24-hour sodium at higher levels. There was a linear relationship between m-24hUNa excretion and systolic BP, while estimated sodium excretion from all 3 formulas had a J-shaped relationship with systolic BP. The relationships between urine sodium excretion and diastolic BP were more complex but were also altered by using formulas. All formulas had associations with BP when a sex-specific constant sodium concentration was inserted in place of measured sodium. Since we used the m-24hUNa, potassium, and creatinine concentrations in formulas, the J-shaped relationships are due to intrinsic problems in the formulas, not due to spot urine sampling. Formula-estimated 24-hour urine sodium excretion should not be used to examine the relationship between sodium excretion and BP since they alter the real associations.
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Jennifer Foster;
S Castillo-Parra;
M Espinoza;
AM Rojas-Serey;
J Diego Maya;
V Sabaj Diez;
V Aliaga Castillo;
M Castillo Nino;
L Romero;
G Hawes Barrios
Multiple interprofessional integrated modules (MIIM) 1 and 2 are two required, cross-curricular courses developed by a team of health professions faculty, as well as experts in education, within the Faculty of Medicine of the University of Chile. MIIM 1 focused on virtual cases requiring team decision-making in real time. MIIM 2 focused on a team-based community project. The evaluation of MIIM included student, teacher, and coordinator perspectives. To explore the perceptions of this interprofessional experience quantitative data in the form of standardised course evaluations regarding teaching methodology, interpersonal relations and the course organisation and logistics were gathered. In addition, qualitative perceptions were collected from student focus groups and meetings with tutors and coordinators. Between 2010 and 2014, 881 students enrolled in MIIM. Their evaluation scores rated interpersonal relations most highly, followed by organisation and logistics, and then teaching methodology. A key result was the learning related to interprofessional team work by the teaching coordinators, as well as the participating faculty. The strengths of this experience included student integration and construction of new knowledge, skill development in making decisions, and collective self-learning. Challenges included additional time management and tutors’ role. This work requires valuation of an alternative way of learning, which is critical for the performance of future health professionals.
Sexual and gender minority individuals who attend collective sex venues (CSVs; establishments where people can have sex in groups or the presence of others) are at elevated risk for HIV and STIs. On-site sexual health interventions have been attempted at CSVs, but attendees’ interest in receiving such services is under-investigated. This paper presents results from a 2020 online cross-sectional survey completed by 342 sexual and gender minority individuals who attended CSVs in New York City. Interest in services such as on-site testing for STIs, testing vans near CSVs, and informational referrals was overall high, particularly among younger participants. Among participants who reported being HIV negative, those of younger age and those who were not using PrEP reported being more likely to take an HIV test if it would be offered at CSVs. In open-text survey responses, participants expressed interest in CSVs providing free prevention services such as HIV/STI testing, PEP, PrEP, and STI medications or vaccination, as well as in ways to improve norms surrounding condom use and consent at these venues. Some participants expressed barriers to on-site services such as privacy concerns, preexisting access to health services, an emphasis on personal responsibility, and negative reactions to the presence of service providers. However, some participants also felt that these services could be delivered in a positive, acceptable, and non-judgmental way, especially by involving CSV organizers and attendees in their implementation. Findings from this study can inform future initiatives to develop sexual health interventions at CSVs.
Efforts to reduce the health and ecological burdens of household biomass combustion are underway in Ghana, principally by promoting clean cookstoves and fuels. Recent studies have focused on the sustained use of clean cookstoves, but sometimes household adopt a new cookstove and then end use of that stove. In this study, we introduce a novel framework for understanding and encouraging household transitions to cleaner cooking: clean fuel discontinuance. We leveraged data from the Ghana Randomized Air Pollution and Health Study (GRAPHS)(N = 1412) where pregnant women received either improved biomass (BioLite) or dual burner LPG stoves for free. LPG users were given free LPG refills during GRAPHS. Weekly questionnaires were administered. Stove use monitors tracked a sub-cohort (n = 220) 6 months before and after the fuel subsidy. We examined social and ecological determinants of stove use and discontinuance. Overall intervention stove use adherence was high throughout GRAPHS, with self-reported use at 69% and 86% of participant-weeks for BioLite and LPG arms respectively. Participants used intervention stoves less for meals requiring vigorous stirring. Burns from intervention stoves decreased use among BioLite (RR: 0.96, p = 0.009), but not LPG users. Device breakage was mentioned as an impediment in 18% of free-text responses for LPG users and 1% for BioLite. Tree canopy within a spatial buffer—a plausible proxy for biomass fuels access—was the only variable explaining LPG discontinued stove use in adjusted Cox time-to-event analyses (HR = −0.56, p < 0.001). Future studies should consider the stove use discontinuance framework.
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Zobedia Cruz-Monserrate;
Kristyn Gumpper;
Sabrina Kaul;
Niharika Badi;
Samantha Terhorst;
Kelly Dubay;
Gregory Lesinski;
William Fisher;
Amy McElhany;
Luis F Lara;
Somashekar Krishna;
Thomas Mace;
Natalia Higuita-Castro;
Lilibeth Ortega-Pineda;
Michael A Freitas;
Alice Hinton;
Dhiraj Yadav;
Phil A Hart;
Stephen J Pandol;
Saima Ahmed;
Benoit Fatou;
Hanno Steen;
Darwin L Conwell
Objectives Endoscopic pancreatic function tests are used to diagnose pancreatic diseases and are a viable source for the discovery of biomarkers to better characterize pancreatic disorders. However, pancreatic fluid (PF) contains active enzymes that degrade biomolecules. Therefore, we tested how preservation methods and time to storage influence the integrity and quality of proteins and nucleic acids. Methods We obtained PF from 9 subjects who underwent an endoscopic pancreatic function test. Samples were snap frozen at the time of collection; after 1, 2, and 4 hours on ice; or after storage overnight at 4°C with or without RNase or protease inhibitors (PIs). Electrophoresis and mass spectrometry analysis determined protein abundance and quality, whereas nucleic acid integrity values determined DNA and RNA degradation. Results Protein degradation increased after 4 hours on ice and DNA degradation after 2 hours on ice. Adding PIs delayed degradation. RNA was significantly degraded under all conditions compared with the snap frozen samples. Isolated RNA from PF-derived exosomes exhibited similar poor quality as RNA isolated from matched PF samples. Conclusions Adding PIs immediately after collecting PF and processing the fluid within 4 hours of collection maintains the protein and nucleic acid integrity for use in downstream molecular analyses.
Spheroid cultures derived from explanted cancer specimens are an increasingly utilized resource for studying complex biological processes like tumor cell invasion and metastasis, representing an important bridge between the simplicity and practicality of 2-dimensional monolayer cultures and the complexity and realism of in vivo animal models. Temporal imaging of spheroids can capture the dynamics of cell behaviors and microenvironments, and when combined with quantitative image analysis methods, enables deep interrogation of biological mechanisms. This paper presents a comprehensive open-source software framework for Temporal Analysis of Spheroid Imaging (TASI) that allows investigators to objectively characterize spheroid growth and invasion dynamics. TASI performs spatiotemporal segmentation of spheroid cultures, extraction of features describing spheroid morpho-phenotypes, mathematical modeling of spheroid dynamics, and statistical comparisons of experimental conditions. We demonstrate the utility of this tool in an analysis of non-small cell lung cancer spheroids that exhibit variability in metastatic and proliferative behaviors.