bluuebunny/crossref_metadata_2025_split huggingface dataset:
PDE4 Inhibitor Reverses Alcohol Impairment of CFTR Function and Protects Against Bacterial Pneumonia
Authors: Stanford Denise , Rasmussen Lawrence , Antony Linto , Allen Antonio , Raju S. V. 
Published: May 2022

INTRODUCTIONAlcohol is a significant risk factor for respiratory infections. Excessive alcohol users suffer from more frequent lung infections, severe symptoms, and poor antibiotic response. Mucociliary clearance (MCC) is a primary lung defense mechanism against inhaled/aspirated pathogens and is impaired by alcohol use. Previously, we found diminished MCC associates with reduced CFTR‐mediated ion transport in alcohol‐fed rats. In addition, CFTR dysfunction correlated with increased cAMP degradation by phosphodiesterase‐4 (PDE4) enzymes and reduced channel gating. Here, we test whether roflumilast, a clinically used PDE4 inhibitor, can preserve cAMP levels in alcohol‐fed rats and promote CFTR function and clearance of Klebsiella pneumoniae (KP, ATCC 43816), to which alcohol users are disproportionately susceptible.METHODSAge and gender‐matched rats (CFTR+/+) were pair‐fed with Lieber‐DeCarli alcohol diet or isocaloric control diet for eight weeks. CFTR activity was assayed by Ussing chamber electrophysiology of tracheal explants. Rats were randomly treated with vehicle/roflumilast (5mg/kg/day) by oral gavage starting 7 days before the end of 8‐week alcohol diet. Airway surface liquid (ASL) depth, periciliary layer (PCL) height, mucus viscosity, and transport (MCT) on excised trachea were quantified with micro‐optical coherence tomography (µOCT) imaging. In addition, lung function was assessed by flexivent oscillometry.RESULTSAlcohol administration reduced CFTR‐mediated ion transport in rat airways. µOCT image analysis of excised trachea from these rats indicated decreased PCL depth and increased mucus viscosity. Despite the normal ciliary beating, there was a noticeable delay in MCT in alcohol‐fed rats. Roflumilast treatment significantly increased CFTR activity in both control and alcohol‐administered rats. Increased CFTR activation by roflumilast resulted in augmented PCL depth, reduced mucus viscosity, and significantly accelerated MCT rate. While roflumilast partially improved clearance of KP, there was pronounced protection against increased airway resistance and lung injury. Histopathological examination of lung sections indicates roflumilast reduced the severity of multifocal pneumonia with edema, fibrin deposition, and numerous bacteria in alveolar spaces. These improvements were more evident in control rats over those administered with alcohol. Ongoing studies are exploring how roflumilast altered immune cell recruitment and inflammatory signaling to reduce infectious injury.CONCLUSIONSAcquired (non‐genetic) CFTR dysfunction may be a suitable target to improve mucus clearance impaired by alcohol use. Administration of PDE4 inhibitors such as roflumilast may boost lung host defense against bacterial pathogens and improve pneumonia outcomes in individuals affected by excessive alcohol use.
Nicotine Treatment Resulted in Decreased Protein Expression of Prorenin and Soluble Prorenin Receptor in 3T3‐L1 cells
Authors: Collins Katelyn B. , Nichols Kellea B. , Poupeau Audrey , Taylor Alexis , Yiannikouris Frederique 
Published: May 2022

Over 36.5% of the United States population is affected by obesity, which is a major contributing factor to the rise of hypertension and cardiovascular disease. Furthermore, the CDC reports that over 16 million Americans suffer from nicotine‐related diseases. Nicotine consumption causes an increase in blood pressure and a decrease in adipogenesis. The prorenin receptor (PRR) and its soluble form (sPRR) are newly discovered components of a blood pressure regulatory pathway, the renin angiotensin system (RAS), and are upregulated during times of obesity. Currently, there is no research to understand the contrasting relationship between nicotine and PRR. Therefore, in this present study, we aimed to determine whether nicotine regulates blood pressure through an adipocyte PRR‐dependent.3T3‐L1 cells were grown to confluence and differentiated over 8 days. On day 8, cells were treated with nicotine dose response treatments: vehicle (water), 0.01 µM nicotine, 1 µM nicotine, or 10 µM nicotine. After 24‐hour treatment, cells were collected to measure protein levels of PRR and it soluble form, sPRR. Additionally, gene expression levels of PRR, renin, 18S, furin, and S1P were measured. Media from the cells was collected after 2 hours, 4 hours, and 24 hours of incubation to measure the secretion levels of sPRR into the media.Nicotine significantly decreased PRR protein expression at 1µM (n=2) compared to vehicle treatment (n=6) (vehicle: 1.00 0.37au; 1 µM: 0.46 0.09au; p <0.05). Nicotine did not affect the gene expression of PRR but significantly decreased PRR and sPRR protein levels in a dose response manner compared to vehicle treatment (vehicle: 1.00 0.42au, n=6; 0.01 µM: 0.47 0.18au, n=4, 1uM: 0.32 0.12au, n=2; p <0.05). Nicotine did not change the expression of S1P, and furin, two enzymes involved in PRR cleavage. Finally, renin and angiotensinogen (AGT) increased significantly in differentiated adipocytes treated with nicotine compared to those treated with vehicle. Together our data indicated that nicotine influences local renin angiotensin system by post‐transcriptionally down‐regulating PRR and sPRR protein and by upregulating renin and AGT. Together, these data suggest that nicotine could assert its effect on blood pressure and adipogenesis via an adipocyte‐PRR dependent mechanism.
Global DNA Methylation Status in Relation to Resistance Training with High vs Low Loads to Failure
Authors: Sexton Casey , Godwin Joshua , Ruple Bradley , Mcintosh Mason , Osburn Shelby , Hollingsworth Blake , Agostinelli Philip , Kavazis Andreas , Zeigenfuss Timothy , Lopez Hector , Smith Ryan , Young Kaelin , Dwaraka Varun , Mobley Christopher , Sharples Adam , Roberts Michael 
Published: May 2022

ObjectiveOur objective was to determine how global DNA methylation status of skeletal muscle differs with resistance training (RT) to failure in trained participants using either 80% of their estimated one‐repetition maximum (Est. 1‐RM) (80FAIL) or 30% of their Est. 1‐RM (30FAIL).HypothesesWe hypothesize there will be a significant decrease in global DNA methylation with both RT conditions, but that 30FAIL RT will elicit a greater magnitude of hypomethylation than 80FAIL RT.MethodsPreviously trained college‐aged males (n= 11, age 23 ± 4 years, percent fat 11.4 ± 6.4%, training experience 4 ± 3 years, squat strength relative to body weight 1.7 ± 0.3) voluntarily underwent two bouts of RT. Participants completed both the 80FAIL and 30FAIL conditions (separated by one week). For each bout of RT, participants completed 4 sets of back squats and 4 sets leg extensions with either the 80FAIL or 30FAIL training conditions. Muscle biopsies were collected from the vastus lateralis before (PRE), 3 hours (3hPOST), and 6 hours (6hrPOST) after each RT bout. DNA was then batch‐isolated from muscle tissue and submitted for analysis using the Illumina MethylationEPIC array.ResultsTotal number of repetitions performed were significantly higher for 30FAIL training vs 80FAIL training (p< .001), however total training volume (sets x reps x load) was not significantly different between conditions (p= 0.571). Global methylation changes between PRE, 3hPOST, and 6hPOST are presented herein. In addition, significant differences between conditions are emphasized for each post‐exercise time point.ConclusionsWith this study we expand our understanding of how the manipulation of RT variables affect epigenetic modifications in skeletal muscle.
Loss of Myosin Vb leads to dysregulation of colonic goblet cell structure and function
Authors: Digrazia Jessica , Engevik Melinda , Engevik Amy C. 
Published: May 2022

BackgroundIntestinal goblet cells secrete a protective mucus layer preventing bacteria and harmful ingested substances from contacting the columnar epithelial cells that line the intestine. Single cell RNAseq data has revealed that goblet cells have high expression of Myosin 5b (Myo5b), a molecular motor that regulates vesicle trafficking in epithelial cells. Myo5b is most widely studied in the small intestine, but it’s role in the colonic epithelium and particularly goblet cells is unknown. A recent study found that Myo5b expression was reduced in inflammatory bowel disease (IBD), a condition that is characterized by dysfunctional mucus; potentially providing a link between Myo5b and goblet cell function. We hypothesized that loss of Myo5b would perturb goblet cell mucus production.MethodsAdult control and intestinal specific tamoxifen inducible VillinCreERT2;Myo5bflox/flox(Myo5b KO) mice were injected intraperitoneally with 2 mg of tamoxifen to deplete intestinal Myo5b. After four days colonic tissue was collected and analyzed.ResultsWe found that loss of Myo5b in vivo results in reduced number of goblet cells in the colon by H&E staining. Muc2 immunostaining and alcian blue staining similarly demonstrated a reduction of mucin within goblet cells. Mucin proteins are heavily o‐glycosylated which makes up 80% of the molecular weight. Normally, mucin structures are terminated by sialic acid which provides a negative charge that protects the mucus protein from bacterial degradation. Staining for sialic acid with the lectin SNA revealed a decrease in sialic acid levels in the mucus of Myo5b KO mice compared to controls. Once sialic acid is removed from mucins, the underlying glycan structures are exposed. Using the lectin WGA to identify the internal glycan oligosaccharide n‐acetyl glucosamine, we observed increased exposure of n‐acetyl glucosamine in Myo5b KO mice compared to control mice; which was consistent with our sialic acid findings. Intestinal goblet cells also secrete wound healing factors including the peptide Trefoil factor 3 (TFF3). Similar to the pattern we observed for Muc2 and sialic acid content, we found decreased levels of TFF3 in the mucosa of Myo5b KO mice.ConclusionsOur data shows that loss of functional Myo5b in the colonic epithelium results in decreased goblet cell numbers and alterations in mucin composition. These data suggest that Myo5b may play an important role in goblet cell mucus production and that mutations affecting Myo5b function may contribute to IBD.
CYP1A1 and CYP1A2 Form Distinct Protein Complexes with NADPH‐cytochrome P450 Reductase, Cytochrome b5, and Heme Oxygenase‐1
Authors: Connick Patrick , Reed James R. , Backes Wayne L. 
Published: May 2022

The cytosolic face of the endoplasmic reticulum (ER) membrane contains a variety of enzymes involved in oxidative metabolism. Among these, NADPH‐cytochrome P450 reductase (POR) and cytochrome b5 (CYB5) provide reducing electrons necessary for cytochrome P450 (P450) and heme oxygenase‐1 (HO‐1) activity. HO‐1 catalyzes the first and rate‐limiting step in free heme catabolism while the P450s are responsible for the metabolism of endogenous and exogenous compounds, including a majority of prescribed drugs. P450s have been shown to form heteromeric and homomeric complexes with other P450s and these complexes have been associated with alterations in P450 activity. Additionally, the P450 catalytic cycle requires 2 electrons—while POR is able to provide both, CYB5 can provide the second electron in some cases. However, binding of POR and CYB5 to the same P450 has been reported to be mutually exclusive, raising the question of how CYB5 can stimulate P450 activity if POR must dissociate prior to CYB5 binding.In order to shed light on this question, we set out to collect more information on the protein‐protein interaction network that the ER‐membrane proteins participate in. Using bioluminescence resonance energy transfer (BRET), we are able to detect interactions between a pair of proteins and, further, to measure the stability of that pair in the presence of a third protein. To detect protein‐protein interactions, HEK293T cells were co‐transfected with vectors encoding two potentially interacting proteins tagged with either Renilla luciferase (Rluc) or GFP2.While human CYP1A2 showed evidence of a specific complex, with analysis of BRET data showing the presence of homodimers, human CYP1A1 did not form a specific homomeric complex. CYP1A1 and CYP1A2 also differed in how they interacted with other ER membrane proteins. Both proteins showed evidence of heteromeric complex formation with HO‐1, but CYP1A1 only did so in the presence of excess POR. POR and CYB5 do not appear to interact with each other and in the presence of CYP1A2, HO‐1, and several other P450s, a complex between POR and CYB5 was not detected. However, a complex including the two proteins was detected in the presence of CYP1A1 which indicated the presence of a ternary CYB5‐POR‐CYP1A1 complex. The presence of a ternary complex could explain the stimulation of P450 activity by CYB5, though the specific mechanism remains to be determined.
Gut Dysbiosis: A Review of the Effects of a High‐Fat Diet on Iron Regulation and Brain‐Derived Neurotrophic Factor (BDNF) Expression in the Brain‐Gut Axis
Authors: Ty Darren , Kania Barbara , Sotelo Alexis , Wisco Jonathan 
Published: May 2022

IntroductionThe human microbiome in relation to the ‘gut‐brain’ bi‐directional axis is becoming increasingly relevant in our study to understand the effects of gastro‐intestinal microbiota on brain development and neurological diseases such as Alzheimer’s disease. One particular protein of interest is called brain‐derived neurotrophic factor (BDNF), which is a protein found in the brain that is important for learning and memory through stabilization of long‐term potentiation (LTP). Through past research, its expression levels have been found to be greatly affected by diet and exercise. Furthermore, in events such as aging, decreased levels of BDNF is associated with cognitive decline and increased susceptibility to neurodegenerative diseases such as Alzheimer’s disease (AD). Our study aims to look at how the human microbiome, particularly that of a Western diet, affects BDNF expression levels and modulation in the ‘gut‐brain’ bi‐directional axis.MethodsIn order to explore our research topic further, we conducted a literature review of about 80 papers using keywords: Small intestines, brain, liver, hepcidin, TMAO, BDNF, iron, astrocytes, Western Diet, microbiota, gut microbiome, gut dysbiosis, iron, inflammation, oxidative stress, IL‐6, vagus nerve.ResultsAfter reviewing papers, it was concluded that a Western diet decreases brain BDNF expression. However, the exact mechanism of how gut microbiota modulates this expression and function in the brain is unclear. Some papers mentioned BDNF in the gut and the periphery, but not much else other than that they existed.ConclusionsThis study sparked further questions: Does gut microbiota affect brain BDNF directly through the gut‐brain axis? Or is it indirect through an intermediate such as gut BDNF? Is there a relationship between gut BDNF and brain BDNF? It is known that brain BDNF is associated with learning and memory and is important for preserving cognitive abilities, however little is known about how gut microbiota affects it through the gut‐brain axis or even its relation to other peripheral BDNF. Looking forward, we plan on measuring different protein levels, including that of BDNF, in the brains and guts across different cohorts of mice. The cohorts will differ in diet, age, and having the APP gene. From this analysis we hope to find more insight on the unknown mechanism that bridges gut microbiota to BDNF in the brain, which may ultimately help in finding ways to prevent cognitive decline seen in many neurological diseases.
Effects of Sustained Hypobaric Hypoxia on Amplitude of Forced Hemodynamic Oscillations During Central Hypovolemia
Authors: Anderson Garen K. , Rosenberg Alexander , Barnes Haley , Bird Jordan , Pentz Brandon , Byman Britta R. , Jendzjowsky Nicholas , Wilson Richard , Day Trevor , Rickards Caroline 
Published: May 2022

IntroductionForcing oscillations in arterial pressure and cerebral blood flow at 0.1 Hz during simulated hemorrhage protects cerebral oxygenation at both low and high altitude. Arterial pressure oscillations at 0.1 Hz are endogenously driven by rhythmic fluctuations in sympathetic nerve activity. As hypobaric hypoxia increases basal sympathetic activity, we hypothesize that the amplitude of forced oscillations in arterial pressure and cerebral blood flow during simulated hemorrhage will be greater at high altitude compared to low altitude.Methods8 healthy human participants (4 M, 24.7 ± 4.1 y; 4 F, 34.3 ± 8.3 y) underwent a hypovolemic oscillatory lower body negative pressure (OLBNP) protocol, where chamber pressure reduced to ‐60 mmHg then oscillated every 5‐s between ‐30 mmHg and ‐90 mmHg over 10‐min (0.1 Hz). This protocol was performed at both low altitude (LA; Calgary, Alberta, Canada; 1045 m) and high altitude (HA; White Mountain, California, USA; 3800 m). Mean arterial pressure (MAP), mean middle cerebral artery velocity (MCAv), and cerebral tissue oxygenation (ScO2) were recorded continuously. Frequency analysis (via continuous wavelet transform) was used to quantify oscillations in MAP and mean MCAv at ~0.1 Hz. Data were analyzed with linear mixed‐models and paired t‐tests. All data are represented as mean ± SD.ResultsBaseline amplitude of oscillations were similar between HA and LA for MAP (1.9 ± 0.6 mmHg vs. 1.2 ± 0.5 mmHg; P = 0.47) and mean MCAv (0.9 ± 0.4 cm/s vs. 1.1 ± 0.3 cm/s; P = 0.91). Oscillatory amplitudes increased with 0.1 Hz OLBNP and altitude for MAP (ANOVA main effect, OLBNP: P < 0.001, Altitude: P = 0.007) and mean MCAv (ANOVA main effect, OLBNP: P = 0.002, Altitude: P = 0.008). Amplitude of oscillations during OLBNP were greater at HA for both MAP (4.0 ± 2.1 mmHgvs. 2.6 ± 1.4 mmHg, P = 0.05) and mean MCAv (2.4 ± 1.1 cm/svs. 0.9 ± 0.4 cm/s; P = 0.01). The relative (%∆) decrease in ScO2 was not different between HA and LA (‐0.63 ± 0.92 % vs. ‐2.56 ± 2.61 %, P = 0.11).ConclusionsOscillatory amplitudes at 0.1 Hz in both MAP and mean MCAv increased during OLBNP at high altitude. This effect may be due, in part, to the sympathoexcitatory stimulus of hypobaric hypoxia, and does not alter the protection of cerebral tissue oxygenation in this environment.
Serum and Glucocorticoid‐Inducible Kinase 1 (SGK1): An Important Contributor to Diarrhea and Malabsorption in Microvillus Inclusion Disease (MVID)
Authors: Ahsan Md Kaimul , dos Reis Diego Carlos , Barbieri Andrea , Sumigray Kaelyn , Nottoli Timothy , Salas Pedro J. , Ameen Nadia A. 
Published: May 2022

Microvillus Inclusion Disease (MVID) is a lethal congenital diarrheal disease resulting from loss of function mutations in the actin motor myosin VB (MYO5B) that regulates apical traffic. MVID remains without treatment to reverse the severe diarrhea that leads to death. MVID diarrhea results from both increased fluid secretion and malabsorption of ions and carbohydrates in the small intestine. Serum and Glucocorticoid‐inducible kinase 1 (SGK1), is a potent regulator of ion transporters including cystic fibrosis transmembrane conductance regulator (CFTR) in the intestine. Since SGK1 upregulates CFTR function in the intestine, we hypothesized that loss of SGK1 could potentially reduce MVID diarrhea by decreasing CFTR fluid secretion.Using CRISPR‐Cas 9 approaches, we first generated MYO5B‐floxed mice and to generate conditional MYO5B‐KO (R26creER;MYO5Bf/f) mice, we cross bred with R26creER mice. Tamoxifen‐treated R26creER;MYO5Bf/fmice resulted in characteristic features of human MVID including severe diarrhea, Microvillus Inclusions (MIs), defective apical traffic, depolarization of proteins, and ion transporters including CFTR, NHE3 and DRA. MYO5B‐KO mice also showed increased phosphorylation of SGK1 and PDK1 in the intestine. SGK1‐floxed (SGK1f/f)mice were crossed with R26creER;MYO5Bf/fmice to generate conditional MYO5B/SGK1 DcKO (R26creER;MYO5Bf/f;SGK1f/f) upon Tamoxifen induction. Surprisingly, tamoxifen treatment (3 day) of MYO5B/SGK1 DcKO mice resulted in more severe diarrhea compared to MYO5B (R26creER;MYO5Bf/f) mice. Immunoblots of intestinal lysates revealed decreased CFTR, increased alpha‐ and beta‐ENaC, increased phosphorylation of PDK1, Nedd4‐2 and pPKCιThr563 in MYO5B/SGK1 DcKO vs. MYO5B ScKO mice. Finally, fecal glucose loss was higher with reduced SGLT1 and GLUT2 by immunoblot in MYO5B/SGK1 DcKO vs MYO5B ScKO mice. We conclude that activation of SGK1 pathway and CFTR contribute to worsening of diarrhea and carbohydrate malabsorption in MVID.
miR‐133a Mitigates Ferroptosis by Attenuating Fatty Acid Metabolism in the Diabetic Heart
Authors: Kambis Tyler N. , Kar Sumit , Mishra Paras K. 
Published: May 2022

Background/hypothesis of the studyMyocardial lipid accumulation occurs prior to the development of cardiac dysfunction in patients with diabetes mellitus (DM). miR‐133a prevents DM‐induced myocardial lipid deposition and is decreased in the DM heart, however its cardioprotective mechanisms remain unknown. Ferroptotic cell death which results from an increase in lipid peroxidation combined with a failure of lipid‐antioxidant mechanisms in the presence of iron, is upregulated in the DM heart. Thus, we hypothesize that overexpression of miR‐133a protects against ferroptotic cell death via inhibition of lipid accumulation, leading to decreased lipid peroxidation in the DM heart. We developed a novel Akita/miR‐133aTg (Ak/Tg) mouse model by crossbreeding DM Akita (Ak) with cardiac specific miR‐133a Tg (Tg) to elucidate how miR‐133a regulates ferroptotic cell death in the DM heart.Methods and ResultsDeep sequencing analysis revealed that miR‐133a activated fatty acid oxidation in the DM heart (p‐value=4.59E‐11). Protein levels of transcription factor Peroxisome proliferator‐activated receptor gamma (PPARG) (WT: 0.195±0.012, Ak: 0.23±0.012, Ak/Tg: 0.173±0.009, Tg: 0.16±0.012) and its coactivator Peroxisome proliferator‐activated receptor‐gamma coactivator (PGC‐1α) (WT: 0.162±0.006, Ak: 0.224±0.012, Ak/Tg: 0.149±0.011, Tg: 0.145±0.010). Furthermore, citrate synthase activity was increased in the presence of miR‐133a (WT: 0.793±0.043, Ak: 0.735±0.026, Ak/Tg: 0.994±0.055, Tg: 0.929±0.018), implying an increase of lipid clearance. Protein levels of glutathione peroxidase (GPX4), an antioxidant that inhibits the production of peroxidized lipids, were increased by the presence of miR‐133a (WT: 0.038±0.003, Ak: 0.024±0.002, Ak/Tg: 0.040±0.004, Tg: 0.030±0.006). miR‐133a also decreased levels of DM‐induced peroxidized lipids (WT: 203.9±12.69, Ak: 231.9±7.252, Ak/Tg: 209.8±8.885, Tg: 251.9±14.26).ConclusionsThese data support that miR‐133a decreases myocardial lipid deposition via increased lipid metabolism while simultaneously increasing levels of lipid antioxidant GPX4, leading to lower levels of peroxidized lipids resulting in the mitigation of ferroptotic cell death in the DM heart.