bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification of a High‐Risk Cardiovascular Disease Population: Combining Computational Modeling and Bioluminescence Imaging
Authors: Head Trajen , Dau Peter , Andreev Victor , Deo Sapna , Daftarian Pirouz , Goldschmidt‐Clermont Pascal , Daunert Sylvia 
Published: November 2023

Cardiovascular disease (CVD) is the leading cause of mortality globally, responsible for over 17.3 million deaths annually (~31% of all deaths). Over the next decade, this value is projected to increase, surpassing 23.6 million annual deaths by 2030. The vast majority of these deaths are caused by complications resulting from progressive thickening of the arterial wall, and the formation and growth of arterial plaques within the vessel lumen. Subsequent plaque rupture may potentially lead to blood clot (thrombus) formation. The combined effect of these processes is the reduction of vessel diameter (stenosis) and subsequent reduction in blood flow (ischemia), while complete occlusion of a vessel via thrombus formation may precipitate a heart attack or stroke. Utilizing discrete event simulation (DES) modeling, we have examined the various processes of lesion formation and growth, and demonstrated that an increased number of raised lesions in an individual at an early age represents a reliable biomarker for increased risk of subsequent sudden cardiac death (SCD). As a result of this finding, we examined the levels of known CVD risk factors for a group of ~3,000 young people (ages 15–34). Using generalized linear regression modeling (GLM) we investigated how these factors contribute to lesion formation and growth. We identified a subpopulation (~13%) exhibiting an increased number of raised lesions. However, we found no correlation with the accelerated atherosclerosis and measured levels of risk factors in this group, suggesting one or more “hidden” risk factors. Thus, we began work to develop a biosensor capable of identifying this “high‐risk” group presenting a significantly increased number of lesions. Atherosclerotic plaques possess a large number of apoptotic and necrotic cells. We designed and expressed a fusion protein capable of targeting and binding to these apoptotic cells (Annexin V), as well as generating a detectable bioluminescent signal (RLuc8). This Annexin‐Renilla Fusion Protein (ArFP) was subsequently characterized and applied both in vitro and in vivo, demonstrating that apoptosis detection is possible in a number of samples including vascular tissue. Plaque identification through apoptosis detection with this unique sensor provides another step towards the early identification and possible intervention of those at the highest risk of CVD‐related death.Support or Funding InformationThis work was supported in part by grants from the National Institutes of Health [1R01GM114321 and R01GM047915]. Dr. Sylvia Daunert is grateful to the Miller School of Medicine of the University of Miami for the Lucille P. Markey Chair in the Department of Biochemistry and Molecular Biology.
Altered Brain Hemoglobin Gene Expression in the Frontal Cortex of Patients with Alzheimer's and Acquired Creutzfeldt‐Jakob's Disease
Authors: Vanni Silvia , Zattoni Marco , Moda Fabio , Tagliavini Fabrizio , Haik Stephane , Deslys Jean‐Philippe , Zanusso Gianluigi , Ironside James W , Ferrer Isidre , Kovacs Gabor , Legname Giuseppe 
Published: November 2023

ObjectiveThe aim of this study was to shed light on the molecular mechanism involved in the onset and the progression of human prion diseases. In particular, we investigated specific alterations that occur in variant and iatrogenic forms of Creutzfeldt‐Jakob disease (vCJD and iCJD) infected brains in comparison to other sporadic (sCJD) or genetic (gCJD) prion and non‐prion neurodegenerative diseases.MethodsWe collected 132 samples from frontal cortex of frozen brain tissue, from prion‐infected patients (vCJD n=20, iCJD n=11, sCJD n=25, gCJD = 28), patients with Alzheimer's disease (AD n=18) and age‐matched controls (n=30). RNA was extracted and only samples with a RIN≥5 were included in the study RT‐qPCR was performed for the selected transcripts, with ACTB, RPL19, GAPDH and B2M as reference genes. In addition, specific erythrocyte marker (ALAS2) expression analysis was performed in order to account for the blood contamination.ResultsPreviously (Barbisin et al. 2014), we reported a significant down‐regulation in the gene expression of both neuronal/glial hemoglobin chains (nHBB and nHBA2) in a macaque model of intracranial BSE‐infection. By contrast, we observed a significant up‐regulation of nHBA1/2 in vCJD brains together with a significant down‐regulation of nHBB in iCJD patients. In addition, both chains display a strong down‐regulation in AD brains, in accordance with earlier results reported in literature (Ferrer et al. 2011).ConclusionsBesides an involvement of oxygen‐ and iron‐related pathways unbalance as pathological events shared among different neurodegenerative diseases, these data point towards distinct transcriptional responses depending on the neuropathological alterations in human prion diseases. In particular, the initial site of misfolding event (CNS vs peripheral tissue) and the related difference in disease timing, together with specific molecular and conformational features of the pathological agent of the disease (such as Bovine Spongiform Encephalopathy (BSE)‐derived vCJD, various sCJD and iCJD strains or other misfolded protein events such Aβ accumulation in AD) seem to determine the peculiar hemoglobin dysregulation in different neurodegenerative diseases. In conclusion, these results suggest that gene expression of nHBB and nHBA2 in the CNS is differentially affected by different prion strains.Support or Funding InformationThis work was supported by a Fondo per gli Investimenti della Ricerca di Base (FIRB) program project [grant number RBAP11FRE9 to G.L.] from Ministero dell'Istruzione, dell'Università e della Ricerca (MIUR) Italy.
Probing the Structural and Biochemical Basis for the Development of Cushing's Syndrome caused by a Somatic Mutant of Protein Kinase A
Authors: Walker Caitlin , Karamafrooz Adak , Wang Yingjie , Taylor Susan , Veglia Gianluigi 
Published: November 2023

Protein phosphorylation is a ubiquitously conserved signaling mechanism playing a crucial role in activating or deactivating vital cellular signaling pathways. Dysregulation of these pathways lead to human diseases, such as heart disease and cancer. Recently somatic mutations in the gene PRKACA coding for the catalytic domain of cAMP‐dependent protein kinase A (PKA‐C) have been identified as frequent genetic alterations in adrenocortical adenomas responsible for adrenal Cushing's syndrome. One such mutation, L205R is located at the interface between the catalytic and regulatory subunits of PKA. Crystal structures of L205R complexed with ATP and the short‐length of the heat stable protein kinase A inhibitor (PKI5‐24) are mostly superimposable to the wild‐type PKA‐C structure, with the exception of the orientation of the R205 side chain, that is reflected in a slightly different positioning of the pseudo‐substrate.To understand how this mutation affects both the regulation and activity of PKA‐C, we performed thermostability, kinetics, and binding analyses using biochemical and biophysical techniques as well as NMR spectroscopy. We discovered that the mutation causes a significant reduction in binding affinity for substrates and regulatory domains. In addition, preliminary NMR dynamic studies show that PKA‐C L205R exhibits increased conformational dynamics, suggesting that this single‐site mutation might influence the allosteric communication within the enzyme. Based on these results, we propose that this somatic mutation might alter the binding interface as well as the conformational dynamics of the enzyme causing aberrant phosphorylation and contributing to the development of the disease.Support or Funding InformationNational Institutes of Health, GM100301
Suppressor of IKK epsilon (SIKE) acts as a bridge between innate immune signalling pathway and cytoskeletal rearrangements
Authors: Wittenberg Karli A , Slykas Frank A , Sonnenschein Halie A , Bell Jessica K. 
Published: November 2023

Innate immunity coordinates the first line of defense against invading pathogens. This involves the recruitment and subsequent migration of nearby immune cells to the site of infection, allowing the pathogen to be consumed via phagocytosis to sequester the infection. These processes require the rapid disassembly, rearrangement, and repolymerization of the network of structural proteins known as the cytoskeleton. Migration and phagocytosis are one of the functional consequences of innate immune receptor triggered signaling pathways. Signals coalesce at catalytic hubs, such as kinases, which serve to amplify and regulate the signal. In the anti‐viral response, TANK binding kinase 1 (TBK1) functions as a catalytic hub. Suppressor of IKK epsilon (SIKE) acts as a high affinity, alternate TBK1 substrate, although SIKE's downstream function is not yet defined. The primary goal of this work was to assess the effect of SIKE on cytoskeletal rearrangements in migration and phagocytosis. For these studies, a CRISPR knockout cell line was derived, SIKE‐CR, from chronic myelogenous leukemia (CML) cell line KBM‐7 named HAP1. Wound healing assays of HAP1 parental cells and SIKE‐CR illustrated that the absence of SIKE significantly decreased cell migration. In the mouse macrophage RAW 264.7 cell line, SIKE expression was knocked down via lentiviral delivery of shRNA targeting SIKE. Phagocytosis was assessed in WT and SIKE knockdown cells via uptake of latex beads labeled with IgG‐FITC. Reduced SIKE expression increased phagocytic uptake. To determine SIKE interactions with cytoskeletal proteins, co‐immunoprecipitations (cIP), in vitro precipitation (IVP) reactions, and enzyme‐linked immunosorbent assays (ELISA) were completed and demonstrated SIKE:tubulin and SIKE:α‐actinin complexes. Together, these studies establish an interaction between SIKE and cytoskeletal proteins and suggest that SIKE functions in cytoskeletal rearrangement associated with migration and phagocytosis.Support or Funding InformationWork supported by NIH grant R21AI107447 and USD SURE.
Breast Tumor Cell‐derived FGF‐5 Induces MEKK1‐dependent Chemokine Expression in Mammary Fibroblasts; Implications for Tumor Microenvironment
Authors: Cuevas Bruce Daniel 
Published: November 2023

Breast tumors contain both transformed epithelial cells and non‐transformed stroma cells, and accumulating experimental evidence strongly suggests that this heterogeneous tumor microenvironment influences tumor growth and metastasis. Previously, we demonstrated that MAPK/ERK kinase kinase 1 (MEKK1) regulates gene expression in fibroblasts and breast tumor cells, and that transgene‐induced breast tumor metastasis is markedly inhibited in MEKK1‐deficient mice. In this work, we utilized expression array analysis of heterotypic cell cultures to determine the role of MEKK1 in stroma gene expression induced by breast tumor cells. We discovered that the breast tumor cell line MDA‐MB 231 constitutively expresses fibroblast growth factor 5 (FGF‐5), and that exposure to either tumor cell‐conditioned media or recombinant FGF‐5 induces chemokine expression in mammary fibroblasts. We demonstrate that FGF‐5‐induced expression of C‐C Motif Chemokine Ligand 5 (CCL5/RANTES) is markedly reduced in MEKK1‐deficient mammary fibroblasts, and that CCL5 expression requires MEKK1 kinase activity but is independent of MEKK1 ubiquitin ligase activity. Furthermore, we report that FGF‐5 initiates MEKK1 kinase activation and signaling in mammary fibroblasts. Finally, we show that breast tumor cell chemotaxis induced by fibroblasts is markedly inhibited by exposure to the CCL5 receptor antagonist Maraviroc, and that MEKK1‐deficient mammary fibroblasts are significantly less effective at promoting tumor cell chemotaxis than cells that express MEKK1. Overall, our findings suggest that MEKK1 is a key regulator of the stroma cell response to tumor cell‐derived FGF‐5. We conclude that MEKK1 is part of the signaling mechanism regulating a tumor/stroma paracrine loop that influences function of both transformed cells and stroma cells within breast tumors. Future studies in our lab will investigate the utility of the FGF‐5/CCL5 signaling axis as a set of targets that can be therapeutically inhibited to reduce tumor growth and progression.Support or Funding InformationAmerican Cancer Society, Illinois Div.
Mir‐16 and Mir‐34a Suppress Growth of a Variety of Human Cancer Cells
Authors: Jones Eyone , Mazirka Pavel , McNurlan Margaret A , Brink Peter , Caso Giuseppe 
Published: November 2023

MicroRNAs (miRs) can have a pivotal role in the regulation of tumor growth and progression and could represent important therapeutic agents. MiR‐16 and miR34a levels are reduced in many human cancers and have potential as tumor suppressors.The aim of this study was to test the direct effect of miR‐16 and miR34a on the growth of different types of human cancer cells. Human prostate (PC3) and breast cancer (MBA‐MD‐231) and melanoma (UACC62) cells were transfected with miR‐16 or miR34a. Cell proliferation was assessed over a 120 h period using an MTT based assay. MiR‐induced apoptosis was also evaluated in transfected PC3 cells by flow cytometry using Annexin V and PI staining.Increasing intracellular levels of miR‐16 had a profound suppressive effect on the growth of all tumor types. After 120 h cell number was decreased by 86% in PC3, by 56% in MDA‐MD‐231, and by 80% in UACC62 cells transfected with miR‐16 compared to controls (P<0.001). MiR‐34a significantly depressed growth in prostate and breast cancer cells. 120 h after transfection with mir‐34a cell number was 56% in PC3 and 66% lower in MBA‐MD‐231 cells than controls (P<0.001). No significant effect of miR‐34a was detected on melanoma cell growth. Both miRs increased apoptosis in PC3 prostate cancer cells, with miR‐16 showing a larger effect. The number of apoptotic cells increased from 6.4 % (control) to 20.5 % and 16.2% after treatment respectively with miR‐16 and miR34a.In summary, miR‐16 showed a profound suppressive effect on growth of human prostate and breast cancer, and melanoma cells. MiR34 depressed the growth of breast and prostate cancer cells but did not show a significant effect on UACC62 melanoma cells. The negative effect on growth of miR‐16 and miR‐34a on PC3 prostate cancer cells results from both suppression of cell proliferation and enhancement of apoptosis.Support or Funding InformationSupported by the Stony Brook Foundation TRO‐Catacosinos Cancer Translational Researcher Award, and by the Dept. of Surgery, Stony Brook University Medical Center (Small Grant Program)
Subpopulations of Neurotensin Neurons in the Lateral Hypothalamic Area Respond to Distinct Cues and Contribute to Energy Balance by Discrete Mechanisms and Projections
Authors: Brown Juliette Anne , Wright Anna , Bugescu Raluca , Woodworth Hillary L , Kurt Gizem , Leinninger Gina M 
Published: November 2023

In the lateral hypothalamic area (LHA), some neurons, express the neuropeptide neurotensin (Nts) and regulate feeding, drinking and physical activity. Many LHA Nts neurons also contain the inhibitory neurotransmitter GABA, but some modulate glutamate (glut) signaling. This suggests that there are different subtypes of LHA Nts neurons. LHA Nts neuronal terminals have been seen within both the Ventra Tegmental Area (VTA) and Substantia Nigra Compacta (SNc) implying that subpopulations of LHA Nts neurons have distinct midbrain projection targets and mechanisms of action. Furthermore, some LHA Nts neurons co‐express the long form of the leptin receptor (LepRb) and are activated by the anorectic hormone leptin (NtsLepRb neurons) while others are activated by dehydration (NtsDehy neurons). I examined the hypothesis that NtsLepRb and NtsDehy neurons are distinguishable from each other based on their expression of GABA or glut and whether they project preferentially to the VTA or SNc. Until now, the lack of reagents to simultaneously identify Nts, GABA and glutamate prevented this analysis. To overcome this, we designed a dual genetic recombinase approach in order to simultaneously label Nts and GABA or glutamate‐containing neurons. We first generated mice that express FlpO recombinase specifically in Nts neurons (NtsFlpO mice) which we crossed with mice that express Cre recombinase in GABA or glutamate neurons. The resulting offspring expressed FlpO recombinase in Nts neurons and Cre in GABA/Glutamate Neurons; NtsFlpO/vGatCre or NtsFlpO/vGlutCre mice respectively. I injected vectors for FlpO‐inducible green fluorescent protein (GFP) and Cre‐inducible red fluorescent protein (RFP) in the LHA these mice with causes GFP expression only within LHA Nts neurons and RFP expression only in GABA/Glut neurons enabling the simultaneous detection of Nts and GABA and/or glutamate containing neurons via fluorescent microscopy. To examine the midbrain projections of each subtype of LHA Nts neurons, Fluorogold (FG) retrograde tract tracer was injected into the VTA or SNc of mice that express GFP only in Nts neurons (NtsGFP mice) and followed with leptin, or dehydration treatment. Brains were analyzed via immunostaining to label NtsLepRb, or NtsDehy neurons and FG. Our data shows that many LHA Nts neurons contain GABA and some express glutamate. Additionally, both LHA NtsLepRb neurons project to the VTA to the SNc but LHA NtsDehy neurons do not project to either midbrain region. The neurotransmitter content of these cue‐distinct subpopulations will be determined using Leptin and Dehydration treatment too.Support or Funding InformationThis work is supported by NIH awards to JAB (1F31 DK107081‐01A1, 5 T32 ES007255‐27) and GML (RC105025)
Elucidating Peptide Pheromone Signaling in Pathogen Conversations to Understand Advantages of Social Cooperation
Authors: Federle Michael J 
Published: November 2023

Cell‐to‐cell communication (quorum sensing) of Gram‐positive bacteria commonly employs peptide pheromones that transmit information between bacterial cells to facilitate coordinated gene expression across a population. In bacterial pathogens, quorum‐sensing pathways are found to contribute to virulence, biofilm development, antibiotic resistance and species evolution. In Firmicutes, pheromone receptors that are located within the cytosol encompasses a large protein family, designated RNPP (Rap/Npr/PlcR/PrgX), that contains a common structural domain responsible for binding pheromones. Though related by structure, primary amino acid sequences of RNPP proteins and their corresponding peptide ligands are disparate. Likewise, the interactions between receptors and ligands, and the mechanisms of activation resulting from these interactions are empirically unique individually for the few prototypical systems studied and thus, overall, the family remains underexplored. It is our goal to elucidate the sensory‐transduction mechanisms of these molecular switches to facilitate methodologies that can be used to modulate communication pathways to treat or prevent disease. Using genetic, biochemical, structural and chemical‐biology approaches, we have begun to elucidate signaling pathways among streptococcal species, have characterized receptors and their interactions with ligands, and have identified modulatory compounds that interfere with normal signaling. For the Rgg2/3 signaling pathway in Streptococcus pyogenes, pheromones induce phenotypes that are consistent with an ability for bacteria to colonize the host asymptomatically, and we hypothesize that social coordination of this behavior reinforces conformity to avoid host innate‐immunity stimulation.Support or Funding InformationNIH R01AI091779 and R01AI425452, and the Burroughs Wellcome Fund Investigators in the Pathogenesis of Infectious Diseases
Genotypically distinct Helicobacter pylori causes Gastric and Peptic Ulcer diseases in Kashmir Indian patients
Authors: Rehman Shabina , Bharti Alok C , Das Bhudev C , Talukder Jamil 
Published: November 2023

BackgroundAlthough 50% of the world population is infected, only a small fraction of people infected with H. pylori suffers from active gastrointestinal (GI) diseases such as gastric ulcer, peptic ulcer and gastric cancer. Genotypically varying virulent H. pylori such as cytotoxin associated gene (Cag A) and vaculating assoctiated cytotoxin (VacA) have been shown associated with gastric and duodenal ulcer patients living in different countries, respectively. Significant Kashmir population has been identified to have GI diseases. However, it is not known whether Kashmir patients with gastric and peptic ulcer diseases are infected with different or same virulent H. pylori.AimThis study was initiated to identify whether patients with gastric and peptic ulcers, and gastric cancer are infected with same or different H. pylori genotypes.Methods263 patients scheduled for upper endoscopy and 50 healthy volunteers were recruited for this study. Endoscopy examination divided the patients into non‐ulcer Dyspepsia (NUD), Peptic ulcer Disease (PUD) and Gastric cancer (GC). Biopsy specimens were taken from stomach (antrum and fundus) and duodenum from each patient. H. Pylori infection was confirmed by Rapid urease test (RUT), glm ureC and culture. H. pylori genomic DNA was isolated from H. pylori culture and biopsy specimen using phenol chloroform method. CagA and VacA genotypes were confirmed by polymerase chain reaction (PCR). H. pylori antigen was detected by immunohistochemistry.ResultsEndoscopic examination grouped the 213 patients into NUD (65), PUD (50) and GC (98). glm ureC analyses established 69%, 80% and 65% of NUD, PUD and GC patients were H. pylori positive, respectively. PCR analysis revealed that: 1) 78%, 80% and 75% of the NUD, PUD and GC patients were infected with CagA positive strains of H. Pylori, respectively; and 2) 76%, 85% and 80% of the NUD, PUD and GC patients were infected with VacA positive strains of H. pylori, respectively. Of the 50 normal healthy volunteers 15 (30%) were also identified H. pylori positive.ConclusionThe most virulent genotype VacA is significantly over‐expressed in all the disease groups indicating a strong association of this genotype with H. pylori ‐ associated disease pathogenesis.
Age distribution of different gastro‐duodenal disease groups in JK and Delhi region.

Region
NUD
PUD
GC
Normal
p value

JK
38.8±8.7 (n=65)
44.1±12.6 (n=50)
52.9±11.95 (n=98)
46.94±9.42 (n=50)
0.07a<0.001b<0.001b

Delhi
38.1±9.6 (n=55)
43.7±11.5 (n=62)
48.3±12.9 (n=50)
36.1±11.7 (n=50)
0.05a<0.00 lb<0.22c

NUD‐ non ulcer dyspepsia; PUD‐peptic ulcer disease; GC‐gastric cancer; JK – Jammu and Kashmir.
p value for NUD vs. PUD,
NUD vs. GC; and
PUP vs. GC