A NOVEL ROLE FOR NEUROTENSIN IN REGULATION OF STEM CELL FUNCTION IN THE SMALL INTESTINE

Neurotensin (NT) is a nutrient-regulated gut hormone that plays important roles in lipid absorption, obesity, metabolic disorders, and normal and neoplastic growth in the intestine. In this study, we 1) elucidate the mechanisms regulating NT release from endocrine cells, 2) examine the role of NT on proliferation and stem cell function in the small intestine and 3) define the effects of NT on colorectal cancer stem cells. We report that NT release from endocrine cells is enhanced by the MAPK scaffold protein Kinase Suppressor of Ras 1 (KSR1) and the exocyst complex component 70 (Exo70). Moreover, free fatty acid stimulated release of NT is attenuated by inhibition of KSR1, MEK, ERK1/2, and Exo70, indicating these proteins as possible therapeutic targets for modulating the negative effects of NT on lipid absorption, obesity, and metabolic dysfunction. Functionally, we show that NT plays an evolutionarily conserved role in maintenance of intestinal stem cells during nutritional stress. NT is required for the induction of WNT/β-catenin signaling and ISC-specific gene expression during fasting and promotes intestinal stem cell self-renewal. In the murine small intestine, loss of NT impairs crypt progenitor cell proliferation via downregulation of ERK1/2 signaling, cyclin D1 expression, and cell cycle related gene expression programs. Loss of NT impairs intestinal stem cell-specific gene expression and intestinal stem cell function after fasting, whereas NT overexpression prevents intestinal stem cell depletion in the midgut of Drosophila fed a nutrient-reduced diet. We extend these findings to reveal a similar role for NT in regulation of stem cell function in colorectal cancer. NT enhances WNT/β-catenin signaling and cancer stem cell self-renewal in the human colon carcinoma cell line HCT116 and in intestinal tumor organoids derived from APC mutant models of colorectal cancer. Mechanistically, we show that NT induces phosphorylation of LRP6 in an ERK1/2 dependent manner in colorectal cancer cells and intestinal tumor organoids, indicating that NT promotes WNT/β-catenin signaling via activation of the ERK1/2 signaling pathway. In contrast, loss of NT impairs cancer stem cell function in tumor organoids derived from APCmin/NT-/-mice, and our preliminary findings indicate that NT-deficiency significantly increases lifespan in APC mutant mice. Collectively, the studies within this dissertation identify NT as a positive regulator of normal and neoplastic growth within the small intestine through regulation of stem cell function and activation of the WNT/β-catenin pathway and identify potential therapeutic targets for modulating the effects of NT in the small intestine

LeguTec – mechanische Beikrautregulierung im Sojaanbau in Luxemburg

Fünf mechanische Beikrautregulierungsmethoden im Sojaanbau werden unter praxisnahen Bedingungen auf drei Bio-Betrieben in Luxemburg seit dem Frühjahr 2018 getestet. Die Feldversuche werden in vier Wiederholungen einschließlich Kontrollparzellen durchgeführt. Verschiedene Boniturparameter werden vor und nach jeder Regulierung sowie zur Blüte und zur Ernte erhoben, um die Effizienz der verwendeten Techniken in Bezug auf den Ertrag zu bewerten. Erste Resultate zeigen höhere Erträge und eine geringere Beikrautdeckung zur Blüte in den Hackvarianten im Vergleich zu den Striegelvarianten

Kinase Suppressor of Ras 1 and Exo70 Promote Fatty Acid-Stimulated Neurotensin Secretion Through ERK1/2 Signaling

Neurotensin is a peptide hormone released from enteroendocrine cells in the small intestine in response to fat ingestion. Although the mechanisms regulating neurotensin secretion are still incompletely understood, our recent findings implicate a role for extracellular signal–regulated kinase 1 and 2 as positive regulators of free fatty acid-stimulated neurotensin secretion. Previous studies have shown that kinase suppressor of Ras 1 acts as a molecular scaffold of the Raf/MEK/extracellular signal–regulated kinase 1 and 2 kinase cascade and regulates intensity and duration of extracellular signal–regulated kinase 1 and 2 signaling. Here, we demonstrate that inhibition of kinase suppressor of Ras 1 attenuates neurotensin secretion and extracellular signal–regulated kinase 1 and 2 signaling in human endocrine cells. Conversely, we show that overexpression of kinase suppressor of Ras 1 enhances neurotensin secretion and extracellular signal–regulated kinase 1 and 2 signaling. We also show that inhibition of extracellular signal–regulated kinase 2 and exocyst complex component 70, a substrate of extracellular signal–regulated kinase 2 and mediator of secretory vesicle exocytosis, potently inhibits basal and docosahexaenoic acid-stimulated neurotensin secretion, whereas overexpression of exocyst complex component 70 enhances basal and docosahexaenoic acid-stimulated neurotensin secretion. Together, our findings demonstrate a role for kinase suppressor of Ras 1 as a positive regulator of neurotensin secretion from human endocrine cells and indicate that this effect is mediated by the extracellular signal–regulated kinase 1 and 2 signaling pathway. Moreover, we reveal a novel role for exocyst complex component 70 in regulation of neurotensin vesicle exocytosis through its interaction with the extracellular signal–regulated kinase 1 and 2 signaling pathway

Neurotensin Regulates Proliferation and Stem Cell Function in the Small Intestine in a Nutrient-Dependent Manner

BACKGROUND & AIMS: Intestinal stem cells (ISCs) are sensitive to dietary alterations and nutrient availability. Neurotensin (NT), a gut peptide localized predominantly to the small bowel and released by fat ingestion, stimulates the growth of intestinal mucosa under basal conditions and during periods of nutrient deprivation, suggesting a possible role for NT on ISC function. METHODS: Leucine-rich repeat-containing G-protein coupled receptor 5-Enhanced Green Fluorescent Protein (Lgr5-EGFP) NT wild type (Nt+/+) and Lgr5-EGFP NT knockout (Nt-/-) mice were fed ad libitum or fasted for 48 hours. Small intestine tissue and crypts were examined by gene expression analyses, fluorescence-activated cell sorting, Western blot, immunohistochemistry, and crypt-derived organoid culture. Drosophila expressing NT in midgut enteroendocrine cells were fed a standard diet or low-energy diet and esg-green fluorescent protein+ ISCs were quantified via immunofluorescence. RESULTS: Loss of NT impaired crypt cell proliferation and ISC function in a manner dependent on nutrient status. Under nutrient-rich conditions, NT stimulated extracellular signal-regulated kinases 1 and 2 signaling and the expression of genes that promote cell-cycle progression, leading to crypt cell proliferation. Under conditions of nutrient depletion, NT stimulated WNT/β-catenin signaling and promoted an ISC gene signature, leading to enhanced ISC function. NT was required for the induction of WNT/β-catenin signaling and ISC-specific gene expression during nutrient depletion, and loss of NT reduced crypt cell proliferation and impaired ISC function and Lgr5 expression in the intestine during fasting. Conversely, the expression of NT in midgut enteroendocrine cells of Drosophila prevented loss of ISCs during nutrient depletion. CONCLUSIONS: Collectively, our findings establish an evolutionarily conserved role for NT in ISC maintenance during nutritional stress. GSE182828

Hearing Screening in North Carolina\u27s NICU and Well-Baby Nurseries: Impact of JCIH 2019 and COVID-19

Purpose: Over an 18-month period in 2020–2021, the North Carolina Early Hearing Detection and Intervention (EHDI) program in collaboration with the North Carolina Leadership Education in Neurodevelopmental and Related Disabilities (LEND) program conducted a statewide examination of newborn hearing screening practices in North Carolina’s 24 Neonatal Intensive Care Units (NICU) and 86 well-baby nurseries to determine how current protocols and procedures conform to those recommended by the Joint Committee on Infant Hearing (JCIH) in its Year 2019 Position Statement: Principles and Guidelines for Early Hearing Detection and Intervention Programs. The COVID-19 pandemic emerged during the study period and motivated a second aim, to examine the impact of the pandemic on infant hearing screening. Results: Our findings revealed that the hospitals in North Carolina are fully committed to their hearing screening programs as demonstrated by a 100% response rate and numerous strengths in both the NICU and well-baby nurseries. Even so, for many hospitals we identified opportunities for program development or improvement based on JCIH 2019 recommendations, especially those concerning oversight of the screening program by a pediatric audiologist, direct referral to an audiologist for NICU babies who fail the in-hospital screening, and audiology referral for well babies who fail the outpatient rescreen. Following the investigation, the NC-EHDI program has worked in partnership with hospitals to provide information, technical assistance, and resources based on our findings and recommendations. The authors would be happy to share the survey instruments and other resources developed for this project with EHDI programs in other states interested in conducting a similar study

GABA(A) receptors containing (alpha)5 subunits in the CA1 and CA3 hippocampal fields regulate ethanol-motivated behaviors: an extended ethanol reward circuitry

GABA receptors within the mesolimbic circuitry have been proposed to play a role in regulating alcohol-seeking behaviors in the alcohol-preferring (P) rat. However, the precise GABA(A) receptor subunit(s) mediating the reinforcing properties of EtOH remains unknown. We examined the capacity of intrahippocampal infusions of an alpha5 subunit-selective ( approximately 75-fold) benzodiazepine (BDZ) inverse agonist [i.e., RY 023 (RY) (tert-butyl 8-(trimethylsilyl) acetylene-5,6-dihydro-5-methyl-6-oxo-4H-imidazo [1,5a] [1,4] benzodiazepine-3-carboxylate)] to alter lever pressing maintained by concurrent presentation of EtOH (10% v/v) and a saccharin solution (0.05% w/v). Bilateral (1.5-20 microgram) and unilateral (0.01-40 microgram) RY dose-dependently reduced EtOH-maintained responding, with saccharin-maintained responding being reduced only with the highest doses (e.g., 20 and 40 microgram). The competitive BDZ antagonist ZK 93426 (ZK) (7 microgram) reversed the RY-induced suppression on EtOH-maintained responding, confirming that the effect was mediated via the BDZ site on the GABA(A) receptor complex. Intrahippocampal modulation of the EtOH-maintained responding was site-specific; no antagonism by RY after intra-accumbens [nucleus accumbens (NACC)] and intraventral tegmental [ventral tegmental area (VTA)] infusions was observed. Because the VTA and NACC contain very high densities of alpha1 and alpha2 subunits, respectively, we determined whether RY exhibited a "negative" or "neutral" pharmacological profile at recombinant alpha1beta3gamma2, alpha2beta3gamma2, and alpha5beta3gamma2 receptors expressed in Xenopus oocytes. RY produced "classic" inverse agonism at all alpha receptor subtypes; thus, a neutral efficacy was not sufficient to explain the failure of RY to alter EtOH responding in the NACC or VTA. The results provide the first demonstration that the alpha5-containing GABA(A) receptors in the hippocampus play an important role in regulating EtOH-seeking behaviors

FINCH: A Blueprint for Accessible and Scientifically Valuable Remote Sensing Satellite Missions

Satellite remote sensing missions have grown in popularity over the past fifteen years due to their ability to cover large swaths of land at regular time intervals, making them suitable for monitoring environmental trends such as greenhouse gas emissions and agricultural practices. As environmental monitoring becomes central in global efforts to combat climate change, accessible platforms for contributing to this research are critical. Many remote sensing missions demand high performance of payloads, restricting research and development to organizations with sufficient resources to address these challenges. Atmospheric remote sensing missions, for example, require extremely high spatial and spectral resolutions to generate scientifically useful results. As an undergraduate-led design team, the University of Toronto Aerospace Team’s Space Systems Division has performed an extensive mission selection process to find a feasible and impactful mission focusing on crop residue mapping. This mission profile provides the data needed to improve crop residue retention practices and reduce greenhouse gas emissions from soil, while relaxing performance requirements relative to many active atmospheric sensing missions. This is accompanied by the design of FINCH, a 3U CubeSat with a hyperspectral camera composed of custom and commercial off-the-shelf components. The team’s custom composite payload, the FINCH Eye, strives to advance performance achieved at this form factor by leveraging novel technologies while keeping design feasibility for a student team a priority. Optical and mechanical design decisions and performance are detailed, as well as assembly, integration, and testing considerations. Beyond its design, the FINCH Eye is examined from operational, timeline, and financial perspectives, and a discussion of the supporting firmware, data processing, and attitude control systems is included. Insight is provided into open-source tools that the team has developed to aid in the design process, including a linear error analysis tool for assessing scientific performance, an optical system tradeoff analysis tool, and data processing algorithms. Ultimately, the team presents a comprehensive case study of an accessible and impactful satellite optical payload design process, in hopes of serving as a blueprint for future design teams seeking to contribute to remote sensing research

Detailed Analysis of a Contiguous 22-Mb Region of the Maize Genome

Most of our understanding of plant genome structure and evolution has come from the careful annotation of small (e.g., 100 kb) sequenced genomic regions or from automated annotation of complete genome sequences. Here, we sequenced and carefully annotated a contiguous 22 Mb region of maize chromosome 4 using an improved pseudomolecule for annotation. The sequence segment was comprehensively ordered, oriented, and confirmed using the maize optical map. Nearly 84% of the sequence is composed of transposable elements (TEs) that are mostly nested within each other, of which most families are low-copy. We identified 544 gene models using multiple levels of evidence, as well as five miRNA genes. Gene fragments, many captured by TEs, are prevalent within this region. Elimination of gene redundancy from a tetraploid maize ancestor that originated a few million years ago is responsible in this region for most disruptions of synteny with sorghum and rice. Consistent with other sub-genomic analyses in maize, small RNA mapping showed that many small RNAs match TEs and that most TEs match small RNAs. These results, performed on ∼1% of the maize genome, demonstrate the feasibility of refining the B73 RefGen_v1 genome assembly by incorporating optical map, high-resolution genetic map, and comparative genomic data sets. Such improvements, along with those of gene and repeat annotation, will serve to promote future functional genomic and phylogenomic research in maize and other grasses

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead