Search

Importance: Federally qualified health centers (FQHCs) are federally funded community health clinics that provide comprehensive care to underserved populations, making them potential opportunities to offer eye care and address unmet health care needs. Evaluating the presence of eye care services at FQHCs in Florida is important in understanding and addressing possible gaps in care for the state's large uninsured and underserved populations., Objective: To determine whether FQHCs in Florida are currently offering eye care services, where they are available, what services are being offered, and who provides them., Design, Setting, and Participants: This study used a cross-sectional design conducted within 1 year (from November 2023 to February 2024). FQHCs listed in the US Health Resources and Services Administration database were contacted by telephone to inquire about the presence of eye care services. The FQHCs were located in both urban and rural areas in Florida to assess accessibility of eye care services in the state. School-based health centers and nonophthalmic specialty care health centers were excluded. A total of 437 FQHCs were included., Main Outcomes and Measures: Primary outcomes included the presence of eye care services, types of services offered, clinician type (optometrists or ophthalmologists), frequency of services, and availability of pediatric services., Results: Among 437 FQHCs contacted, only 39 (8.9%) reported offering eye care services. These services primarily included vision examinations, glasses prescriptions, and dilated eye examinations. Optometrists were the primary providers of services at all clinics, with no clinics reporting care by ophthalmologists. The frequency of services varied considerably, ranging from daily to bimonthly. Thirty-seven (94.9%) of the 39 clinics offered pediatric eye care services., Conclusions and Relevance: The low prevalence of FQHCs with eye care services and the absence of ophthalmologist-provided care highlight a gap in access to eye care for underserved populations in Florida. These findings support investigations into implementing eye care services and interventions at FQHCs that might enhance access and equity in eye care.

Polycystic kidney disease (PKD) is a genetic disorder characterized by bilateral cyst formation. We showed that PKD cells and kidneys display metabolic alterations, including the Warburg effect and glutaminolysis, sustained in vitro by the enzyme asparagine synthetase (ASNS). Here, we used antisense oligonucleotides (ASO) against Asns in orthologous and slowly progressive PKD murine models and show that treatment leads to a drastic reduction of total kidney volume (measured by MRI) and a prominent rescue of renal function in the mouse. Mechanistically, the upregulation of an ATF4-ASNS axis in PKD is driven by the amino acid response (AAR) branch of the integrated stress response (ISR). Metabolic profiling of PKD or control kidneys treated with Asns-ASO or Scr-ASO revealed major changes in the mutants, several of which are rescued by Asns silencing in vivo. Indeed, ASNS drives glutamine-dependent de novo pyrimidine synthesis and proliferation in cystic epithelia. Notably, while several metabolic pathways were completely corrected by Asns-ASO, glycolysis was only partially restored. Accordingly, combining the glycolytic inhibitor 2DG with Asns-ASO further improved efficacy. Our studies identify a new therapeutic target and novel metabolic vulnerabilities in PKD., (© 2024. The Author(s).)

Transcriptional and translational control are key determinants of gene expression, however, to what extent these two processes can be collectively coordinated is still poorly understood. Here, we use Nanopore long-read sequencing and cap analysis of gene expression (CAGE-seq) to document the landscape of 5' and 3' untranslated region (UTR) isoforms and transcription start sites of epidermal stem cells, wild-type keratinocytes and squamous cell carcinomas. Focusing on squamous cell carcinomas, we show that a small cohort of genes with alternative 5'UTR isoforms exhibit overall increased translational efficiencies and are enriched in ribosomal proteins and splicing factors. By combining polysome fractionations and CAGE-seq, we further characterize two of these UTR isoform genes with identical coding sequences and demonstrate that the underlying transcription start site heterogeneity frequently results in 5' terminal oligopyrimidine (TOP) and pyrimidine-rich translational element (PRTE) motif switches to drive mTORC1-dependent translation of the mRNA. Genome-wide, we show that highly translated squamous cell carcinoma transcripts switch towards increased use of 5'TOP and PRTE motifs, have generally shorter 5'UTRs and expose decreased RNA secondary structures. Notably, we found that the two 5'TOP motif-containing, but not the TOP-less, RPL21 transcript isoforms strongly correlated with overall survival in human head and neck squamous cell carcinoma patients. Our findings warrant isoform-specific analyses in human cancer datasets and suggest that switching between 5'UTR isoforms is an elegant and simple way to alter protein synthesis rates, set their sensitivity to the mTORC1-dependent nutrient-sensing pathway and direct the translational potential of an mRNA by the precise 5'UTR sequence., (© 2022. The Author(s).)

MicroRNA (miRNA) loaded Argonaute (AGO) complexes regulate gene expression via direct base pairing with their mRNA targets. Previous works suggest that up to 60% of mammalian transcripts might be subject to miRNA-mediated regulation, but it remains largely unknown which fraction of these interactions are functional in a specific cellular context. Here, we integrate transcriptome data from a set of miRNA-depleted mouse embryonic stem cell (mESC) lines with published miRNA interaction predictions and AGO-binding profiles. Using this integrative approach, combined with molecular validation data, we present evidence that < 10% of expressed genes are functionally and directly regulated by miRNAs in mESCs. In addition, analyses of the stem cell-specific miR-290-295 cluster target genes identify TFAP4 as an important transcription factor for early development. The extensive datasets developed in this study will support the development of improved predictive models for miRNA-mRNA functional interactions., (© 2022 The Authors. Published under the terms of the CC BY NC ND 4.0 license.)

The Argonaute proteins (AGOs) are well known for their role in post-transcriptional gene silencing in the microRNA (miRNA) pathway. Here we show that in mouse embryonic stem cells, AGO1&2 serve additional functions that go beyond the miRNA pathway. Through the combined deletion of both Agos, we identified a specific set of genes that are uniquely regulated by AGOs but not by the other miRNA biogenesis factors. Deletion of Ago2&1 caused a global reduction of the repressive histone mark H3K27me3 due to downregulation at protein levels of Polycomb repressive complex 2 components. By integrating chromatin accessibility, prediction of transcription factor binding sites, and chromatin immunoprecipitation sequencing data, we identified the pluripotency factor KLF4 as a key modulator of AGO1&2-regulated genes. Our findings revealed a novel axis of gene regulation that is mediated by noncanonical functions of AGO proteins that affect chromatin states and gene expression using mechanisms outside the miRNA pathway., (Copyright © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

There is a paucity of data regarding racial disparities in the survival of patients with indolent non-Hodgkin lymphomas (iNHL) in the contemporary time-period. Hence, we sought to determine whether racial disparities exist in the survival of patients with iNHLs in the US. We included 68 059 adult patients with follicular lymphoma (FL, n = 41 943), marginal zone lymphoma (MZL, n = 22 485), and lymphoplasmacytic lymphoma/Waldenström macroglobulinemia (LPL/WM, n = 3631) who were diagnosed in the US between 2000 and 2017. Race was categorized as White, Black, Asian/Pacific Islander, or American Indian/Alaska Native (API/AI). The primary outcome was relative survival (RS), which was estimated using flexible parametric survival models. The RS estimates varied according to race and disease histology but were consistently lower for racial minorities, including those diagnosed during the most recent 5-year time-period of 2012-2017. On multivariable analysis for RS, Black patients with FL had a 32% higher excess mortality rate compared to White patients [adjusted excess hazard ratio (aEHR), 1.32; 95% CI, 1.15-1.51; p < .001], corresponding to a difference of 55 (95% CI, 24-86) excess deaths per 10 000 person-years. For MZL, Black patients had a 40% higher excess mortality rate compared to White patients (aEHR 1.40; 95% CI, 1.18-1.66; p < .001), corresponding to a difference of 62 (95% CI, 26-98) excess deaths per 10 000 person-years. No significant racial differences were detected for patients with WM. The greatest disparity was seen for younger Black patients with FL. Our findings highlight the need for interventions to improve the outcomes of Black patients with iNHLs, particularly younger Black patients with FL., (© 2021 Wiley Periodicals LLC.)

The fibroblast growth factor (FGF) and the transforming growth factor-β (TGF-β) pathways are both involved in the maintenance of human embryonic stem cells (hESCs) and regulate the onset of their differentiation. Their converging functions have suggested that these pathways might share a wide range of overlapping targets. Published studies have focused on the long-term effects (24-48 h) of FGF and TGF-β inhibition in hESCs, identifying direct and indirect target genes. In this study, we focused on the earliest transcriptome changes occurring between 3 and 9 h after FGF and TGF-β inhibition to identify direct target genes only. Our analysis clearly shows that only a handful of target transcripts are common to both pathways. This is surprising in light of the previous literature, and has implications for models of cell signaling in human pluripotent cells. In addition, we identified STOX2 as a novel primary target of the TGF-β signaling pathway. We show that STOX2 might act as a novel SMAD2/4 cofactor. Taken together, our results provide insights into the effect of cell signaling on the transcription profile of human pluripotent cells.

Aim of the Study: For out-of-hospital-cardiac-arrest (OHCA) due to ventricular fibrillation (VF) guidelines recommend early defibrillation followed by chest compressions for two minutes before analyzing shock success. If rhythm analysis reveals VF again, it is obscure whether VF persisted or reoccurred within the two-minutes-cycle of chest compressions after successful defibrillation. We investigated the time of VF-recurrence in OHCA., Methods: We examined all cases of OHCA presenting with initial VF rhythm at arrival of ALS-ambulance (Marburg-Biedenkopf-County, 246.648 inhabitants) from January 2014 to March 2018. Three independent investigators analyzed corpuls3® ECG-recordings. We included ECG-data from CPR-beginning until four minutes after the third shock. VF termination was defined as the absence of a VF-waveform within 5 s of shock delivery. VF recurrence was defined as the presence of a VF-waveform in the interval 5 s post shock delivery., Results: We included 185 shocks in 82 patients. 74.1% (n = 137) of all shocks terminated VF, but VF recurred in 81% (n = 111). The median (IQR) time of VF-recurrences was 27 s (13.5 s/80.5 s) after shock. 51.4% (n = 57) of VF-recurrence occurred 5-30 s after shock, 13.5% (n = 15) VF-recurrence occurred 31-60 s after shock, 21.6% (n = 24) of VF-recurrence occurred 61-120 s after shock, 13.5% (n = 15) of VF-recurrence occurred 121-240 s after shock., Conclusions: Although VF was terminated by defibrillation in 74.1%, VF recurred in 81% subsequent to the chest compression interval. Thus, VF reappears frequently and early. It is unclear to which extend chest compressions influence VF-relapse. Further studies need to re-evaluate the algorithm, timing of antiarrhythmic therapy or novel defibrillation strategies to minimize refibrillation during shockable OHCA., (Copyright © 2020 Elsevier B.V. All rights reserved.)

RNA sequencing (RNA-seq) has become a standard procedure to investigate transcriptional changes between conditions and is routinely used in research and clinics. While standard differential expression (DE) analysis between two conditions has been extensively studied, and improved over the past decades, RNA-seq time course (TC) DE analysis algorithms are still in their early stages. In this study, we compare, for the first time, existing TC RNA-seq tools on an extensive simulation data set and validated the best performing tools on published data. Surprisingly, TC tools were outperformed by the classical pairwise comparison approach on short time series (<8 time points) in terms of overall performance and robustness to noise, mostly because of high number of false positives, with the exception of ImpulseDE2. Overlapping of candidate lists between tools improved this shortcoming, as the majority of false-positive, but not true-positive, candidates were unique for each method. On longer time series, pairwise approach was less efficient on the overall performance compared with splineTC and maSigPro, which did not identify any false-positive candidate.

Stratification of patients with pancreatic ductal adenocarcinoma (PDAC) remains a key challenge in the field of clinical oncology. No predictive biomarkers have yet been found for any available treatment options. Previously, we identified SERPINB7 as a putative biomarker for PDAC and thus, herein, we aimed to validate our previous findings and assessed the predictive value of SERPINB7. Patients who underwent surgery and received gemcitabine (gem) or gemcitabine plus nab-paclitaxel (gem/nab) as adjuvant therapy, between 2011 and 2017, were included in this study (n = 57). Expression level of SERPINB7 was assessed in tumor tissue by immunohistochemistry (IHC) and RNA in situ hybridization (RNA ISH). Its association with disease-free survival (DFS) and overall survival (OS) was investigated. While IHC did not show any correlation between survival and the protein level of SERPINB7, RNA ISH revealed that expression of SERPINB7 was associated with a poor DFS (P = .01) and OS (P = .002) in the gem group but not in the gem/nab. Adjusted Cox-regression analysis confirmed the independent predictive value of SERPINB7 on OS (P = .006, HR: 3.47; 95% CI: 1.49-8.09) in the gem group. In conclusion, SERPINB7 was identified as the first predictive RNA biomarker for PDAC. This study suggests that patients who expressed SERPINB7 might receive another treatment than gem alone., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

The purpose of this project was to explore (a) how nursing students regarded debriefing sessions at the end of a laboratory skills class and (b) the cognitive, affective, and social learning outcomes of debriefing sessions. Survey questionnaires were administered to 378 prelicensure nursing students after their nursing skills laboratory class that included a debriefing session. Most students either agreed or strongly agreed that debriefing facilitated the discussion about their experience and reflection on learning.

Pancreatic cancer is a dismal disease with a mortality rate almost similar to its incidence rate. To date, there are neither validated predictive nor prognostic biomarkers for this lethal disease. Thus, the aim of the present study was to retrospectively investigate the capability of biochemical parameters and molecular profiles to predict survival of patients with metastatic pancreatic ductal adenocarcinoma (mPDAC) who participated in a phase II clinical trial to test the safety and efficacy of the combination treatment of capecitabine plus nab-paclitaxel. Herein, we investigated the association of 18 biochemical parameters obtained from routine diagnosis and the clinical outcome of the 30 patients enrolled in the clinical trial. Furthermore, we analysed formalin-fixed paraffin-embedded (FFPE) tumour tissue to identify molecular biomarkers via RNA seq and the Illumina TruSeq Amplicon Cancer panel which covers 48 hotspot genes. Our analysis identified SERPINB7 as a novel transcript and a DNA mutation signature that might predict a poor outcome of disease. Moreover, we identified the bilirubin basal level as an independent predictive factor for overall survival in our study cohort.

Analysis of gene expression has contributed to a plethora of biological and medical research studies. Microarrays have been intensively used for the profiling of gene expression during diverse developmental processes, treatments and diseases. New massively parallel sequencing methods, often named as RNA-sequencing (RNA-seq) are extensively improving our understanding of gene regulation and signaling networks. Computational methods developed originally for microarrays analysis can now be optimized and applied to genome-wide studies in order to have access to a better comprehension of the whole transcriptome. This review addresses current challenges on RNA-seq analysis and specifically focuses on new bioinformatics tools developed for time series experiments. Furthermore, possible improvements in analysis, data integration as well as future applications of differential expression analysis are discussed.

Phenotypic differences between closely related species are thought to arise primarily from changes in gene expression due to mutations in cis-regulatory sequences (enhancers). However, it has remained unclear how frequently mutations alter enhancer activity or create functional enhancers de novo. Here we use STARR-seq, a recently developed quantitative enhancer assay, to determine genome-wide enhancer activity profiles for five Drosophila species in the constant trans-regulatory environment of Drosophila melanogaster S2 cells. We find that the functions of a large fraction of D. melanogaster enhancers are conserved for their orthologous sequences owing to selection and stabilizing turnover of transcription factor motifs. Moreover, hundreds of enhancers have been gained since the D. melanogaster-Drosophila yakuba split about 11 million years ago without apparent adaptive selection and can contribute to changes in gene expression in vivo. Our finding that enhancer activity is often deeply conserved and frequently gained provides functional insights into regulatory evolution.

Neuronal L-type voltage-gated calcium channels (LTCCs) are involved in several physiological functions, but increased activity of LTCCs has been linked to pathology. Due to the coupling of LTCC-mediated Ca(2+) influx to Ca(2+)-dependent conductances, such as KCa or non-specific cation channels, LTCCs act as important regulators of neuronal excitability. Augmentation of after-hyperpolarizations may be one mechanism that shows how elevated LTCC activity can lead to neurological malfunctions. However, little is known about other impacts on electrical discharge activity. We used pharmacological up-regulation of LTCCs to address this issue on primary rat hippocampal neurons. Potentiation of LTCCs with Bay K8644 enhanced excitatory postsynaptic potentials to various degrees and eventually resulted in paroxysmal depolarization shifts (PDS). Under conditions of disturbed Ca(2+) homeostasis, PDS were evoked frequently upon LTCC potentiation. Exposing the neurons to oxidative stress using hydrogen peroxide also induced LTCC-dependent PDS. Hence, raising LTCC activity had unidirectional effects on brief electrical signals and increased the likeliness of epileptiform events. However, long-lasting seizure-like activity induced by various pharmacological means was affected by Bay K8644 in a bimodal manner, with increases in one group of neurons and decreases in another group. In each group, isradipine exerted the opposite effect. This suggests that therapeutic reduction in LTCC activity may have little beneficial or even adverse effects on long-lasting abnormal discharge activities. However, our data identify enhanced activity of LTCCs as one precipitating cause of PDS. Because evidence is continuously accumulating that PDS represent important elements in neuropathogenesis, LTCCs may provide valuable targets for neuroprophylactic therapy.

Feline infectious peritonitis (FIP) is a lethal immunopathological disease caused by feline coronaviruses (FCoVs). Here, we describe a reverse genetics approach to study FIP by assessing the pathogenicity of recombinant type I and type II and chimeric type I/type II FCoVs. All recombinant FCoVs established productive infection in cats, and recombinant type II FCoV (strain 79-1146) induced FIP. Virus sequence analyses from FIP-diseased cats revealed that the 3c gene stop codon of strain 79-1146 has changed to restore a full-length open reading frame (ORF).

Physiological polyamines are required in various biological processes. In the current study, we used norspermidine, a structural analog of the natural polyamine spermidine, to investigate polyamine uptake in the model organism Caenorhabditis elegans. Norspermidine was found to have two remarkable effects: it is toxic for the nematode, without affecting its food, Escherichia coli; and it hampers RNA interference. By characterizing a norspermidine-resistant C. elegans mutant strain that has been isolated in a genetic screen, we demonstrate that both effects, as well as the uptake of a fluorescent polyamine-conjugate, depend on the transporter protein CATP-5, a novel P(5B)-type ATPase. To our knowledge, CATP-5 represents the first P(5)-type ATPase that is associated with the plasma membrane, being expressed in the apical membrane of intestinal cells and the excretory cell. Moreover, genetic interaction studies using C. elegans polyamine synthesis mutants indicate that CATP-5 has a function redundant to polyamine synthesis and link reduced polyamine levels to retarded postembryonic development, reduced brood size, shortened life span, and small body size. We suggest that CATP-5 represents a crucial component of the pharmacologically important polyamine transport system, the molecular nature of which has not been identified so far in metazoa.

The effectiveness of a 20% solution of mannitol-bisacodyl for oral use in preparing patients for barium enemas was investigated, in an investigator-blind control situation. The results were evaluated and compared with other methods currently in use. No comparative study where mannitol had been used could be found in the world literature. The mannitol-bisacodyl regimen was found to be an acceptable and reliable method for colon preparation in a high percentage of cases. Side-effects and changes in electrolyte balance of body fluids were recorded. The regimen was found to be safe for use on any patient irrespective of race, age or clinical condition and is not contraindicated in cases of cardiac and renal failure.