bluuebunny/crossref_metadata_2025 huggingface dataset:
The superior laryngeal nerve is involved in a brief apneic response to intralaryngeal, but not in the lethal ventilatory arrest to intravenous, administration of fentanyl (FNT) in rats
Authors: Zhuang Jianguo , Shi Shan , Gao Xiuping , Xu Fadi
Published: May 2022

Rapid intravenous (IV) injection of FNT induces a sudden ventilatory arrest within a few minutes or even seconds, which is the major cause of overdose FNT‐induced death. Our previous studies have shown that right atrial or IV bolus injection of FNT in anesthetized rats produced a vagal‐mediated apnea or a lethal ventilatory arrest in a dose‐dependent manner (Zhuang et al, AJP 2012 and EB Meeting 2022). The superior laryngeal nerve (SLN) containing the laryngeal sensory afferents also travels in the vagus nerve and is capable of triggering the apneic response associated with hypertension and bradycardia upon activation. Here, we asked whether local administration of FNT into the larynx affected cardiorespiratory activities via acting on the SLN and to what extent the sudden ventilatory arrest induced by IV injection of FNT depended on the SLN integrity. FNT at 15 µg was either sprayed (5‐10 µl) into the larynx or bolus IV‐injected (50 µl) in the anesthetized rats with and without bilateral SLN transection. We found that FNT sprayed into the larynx induced an immediate brief apnea (~3.5‐fold of baseline TE) and laryngeal constriction (but not closure) associated with hypertension and bradycardia. The cardiorespiratory response disappeared after transection of SLN. Interestingly, the same dose of FNT given by IV injection induced a ventilatory arrest (apneic duration >8‐fold longer than that produced by the local administration) associated with laryngeal closure, hypotension and bradycardia. These responses to IV injection of FNT were not significantly affected by SLN transection. Our data suggest that the SLN is responsible for the brief apneic response to laryngeal application of FNT, but not for the lethal ventilatory arrest induced by rapid IV injection of FNT.
When is the Same Not the Same? Issues of Question Equivalence in Online Exam Pools
Authors: Leupen Sarah , Hodges Linda C. , Bass Sarah , Carpenter Tara , Goolsby‐Cole Cody , Stanwyck Liz
Published: May 2022

During the pandemic, the use of question pools for online testing was recommended to mitigate cheating, exposing multitudes of STEM students across the globe to this practice. Yet little systematic analysis of the practice apparently exists. In this study, we undertook an investigation of student performance on our questions in online exam pools across several STEM courses: upper‐level physiology, general chemistry, and introductory physics. We found that the difficulty of creating analogous questions in a pool varied by question type, with quantitative problems being the easiest to vary without altering average student performance. However, when instructors created pools by rearranging aspects of a question, posing opposite counterparts of concepts, or formulating questions assessing the same learning objective, we sometimes discovered student learning differences between seemingly closely‐related ideas, illustrating the challenge of our own expert blind spot. We provide suggestions for instructors on steps to take to improve the equity of question pools, such as being cautious in how many variables one changes in a specific pool and “test driving” proposed questions in lower stakes assessments.
Effects of a Bonded Arginine Silicate Inositol combination (nooLVL®) on Cognitive function in esports Gamers
Authors: Emerson Katie , Perez Ojalvo Sara , Greenberg Danielle
Published: May 2022

Objectives

The purpose of this open‐label study was to assess the ability of a bonded arginine silicate inositol combination (nooLVL) to affect reaction time and responses to cognitive tasks in esports gamers. nooLVL is a non‐stimulant complex of bonded arginine silicate with inositol. Previous studies have evaluated nooLVL in esports gamers showing improvements in accuracy, decision making, and reaction time during gaming. This study looks to further detect cognitive changes in esports gamers on an array of cognitive tests developed by Cambridge Brain Sciences.

Methods

Assessments were evaluated on six Cambridge Brain Sciences (CBS) cognitive tasks: Feature Match, Double Trouble, Mental Rotations, Polygons, Odd One Out, and Spatial Planning. These tests measure visuospatial processing, deductive reasoning, intelligent behavior, concentration, and attention. The subjects were healthy adult males and females (age >18; n=9) with minimum previous gaming experience of 3‐5 hours of video games per week for at least 6 months. All study participants consumed 1600mg per day of nooLVL (600mg arginine, 105mg silicon, 385mg inositol and 60mg potassium) for seven days. They completed the CBS tests at baseline, on day 1 after a single dose of nooLVL, and again on day 7.

Results

Mean changes within group at baseline to day 1, day 1 to day 7 and baseline to day 7 were analyzed with paired t‐tests. Significant differences were found in Double Trouble (baseline vs day 1 [p=0.0019], day 1 vs day 7 [p=0.0369] and baseline vs day 7 [p=0.0005]), Mental Rotations (baseline to day 1 [p=0.0506] and baseline to day 7 [p=0.0192]), and Spatial Planning (baseline to day 7 was observed [p=0.0266]). The Double Trouble assessment measures concentration, Mental Rotations examination measures the function of visual representation in the brain, and Spatial Planning assessment measures intelligent behavior through forethought and sequencing performance.

Outcomes of the Feature Match, Polygons, and Odd One Out tests did not show significance.

Conclusions

These data show that nooLVL® a complex of bonded arginine silicate with inositol, when used in esport gamers, improves outcomes on certain functioning tests such as visual processing, concentration, and spatial learning as early as after a single dose and lasting up to seven days.
Correlation between Consumption of Fast‐food, Sugary Food, and High Caffeine Intake with Dental‐Related Anxieties
Authors: Ahmed Eeshah , Malik Sana , Begdache Lina
Published: May 2022

Existing literature has investigated the relationship between dental anxieties and oral health, yet the relationship between diet and oral health status has yet to be examined. The purpose of this research study was to determine the correlational relationships that exist between oral health, dental anxiety, and nutrition. Data was collected through an anonymous Google Forms survey which was distributed to patients at various dental offices in the Binghamton‐Vestal, NY area as well among Binghamton University students. The distributed survey was composed of the validated Food‐Mood Questionnaire with questions pertaining to demographics, dental health and dental health anxieties. A total of 506 responses were collected and analyzed using Pearson’s Correlation Coefficient in SPSS version 25.0. Our results suggest that females are more likely to experience dental anxiety than men. An increase in the consumption of low‐quality‐energy dense foods, including pre‐packaged food, fast food, and sugary foods, a pattern similar to the Western diet, as well as caffeine consumption of 4 times or more per week, were positively correlated (P < 0.01) with increased nervousness and anxiety at the dental office. Frequent consumption of low‐quality‐energy dense foods was also positively correlated with increased feelings of general mental health distress. Our findings provide a proof of concept that dietary patterns are potentially associated with dental anxiety, and modulation of these patterns may diminish dental‐related mental distress.
Teaching Collaboration Skills to Undergraduate Biochemistry and Chemistry Students
Authors: Mertz Pamela S. , Sherrer Shanen M. , Bowers Geoffrey M.
Published: May 2022

We want our students to develop effective collaboration skills, but how do we intentionally teach teamwork and assess it? The Department of Chemistry and Biochemistry at St. Mary’s College of Maryland has scaffolded collaboration skills throughout the curriculum for the chemistry and biochemistry programs at three levels, foundational, developing, and capstone. The skills are tied to signature assignments at each level. In Biochemistry I and II, we are using a team contract at the beginning of semester‐long team projects (a bioinformatics project and a grant proposal project) where students identify their strengths, review expectations, and make plans for group communication. At the end of the semester, each student assesses their own contributions and those of their team members for various parts of the project. In addition, this academic year we started using a common collaboration rubric in multiple courses for students to evaluate self and team members using the following subcategories: quality of work, commitment, leadership, communication, and analysis. In Biochemistry I and II, we used this rubric for multiple assignments that are part of the semester long projects. In the General Chemistry lab, we modified this rubric to fit into a form that reflects these same collaboration attributes after each lab experience, so students can assess and report privately on their experiences as part of their collaboration grade for the course. A similar collaboration rubric is completed by students for each of five team‐based laboratories in Physical Chemistry I, enabling us to obtain honest feedback and assign a collaboration grade. We will discuss how the use of these forms has affected collaborative projects in our courses.
Arteria Lusoria and its relation to Dysphagia Lusoria. A systematic review
Authors: Syed Qareen H. , Valenzuela Juan J. , Orellana Mathias , Farfan Emilio , Gold Marjorie , Beas Antonia
Published: May 2022

Introduction

Anatomically, the aberrant subclavian artery is the last branch of the aortic arch. On its way to the right upper limb it runs posterior to the esophagus. During this passage it can compress the esophagus, producing Lusoria Dysphagia.

Materials and methods

Different databases were subjected to a systematized literature search, using the following criteria: articles published between 2000 and 2020, and articles that established a clinical correlation between an aberrant subclavian artery and Lusoria Dysphagia.

Results

After full text analysis of the articles that met the criteria, 48 articles were found in which three types of aberrant arteries were identified. These were associated with symptoms of Dysphagia Lusoria in the subjects of the analyzed studies. The most common type of variation in the origin of the aberrant artery was Type I, an aberrant right subclavian artery that arises as the last branch of the aortic arch, maintaining the origins of the other branches of this arch. The most common course was retroesophageal, which was most commonly associated with the symptoms of patients with Dysphagia Lusoria.

Conclusions

The knowledge of the variations and courses of these types of aberrant arteries and their clinical associations is relevant for specialist professionals who treat patients diagnosed with Dysphagia Lusoria.
The sciatic nerve and and its relation to the piriformis muscle syndrome regarding daily clinical practice. A review
Authors: Beas Antonia , Salas Catalina , Valenzuela Juan José , Orellana Mathias , Syed Qareen , Santana Andrés
Published: May 2022

Introduction

The sciatic nerve innervates most of the muscles of the lower limb, so any alteration in it'sits function or clinical presentation could directly affect the functioning of the lower limb. Therefore, the normal anatomy, anatomical variants and how these can influence clinical implications in the gluteal region are of importance in daily clinical practice.

Methodsodology

A systematic review was carried out in the scientific literature to answer our research question. We searched in the databases MEDLINE, SCOPUS, WOS, Scielo, GOOGLE SCHOLAR and other scientific sources, using search terms in relation to our stated objectives and connected through Boolean terms.

Results

The results of this study show that the variations of the sciatic nerve can be ample, which associated to the variability in the geographic regions of the studies, sex of the sample and incidence of the variation, will make it necessary to consider different multifactorial components for the anatomical analysis of sciatic nerve variations and for clinical considerations in patients with and without these anatomical variations.

Conclusion

With the results of our review, we can infer that the sciatic nerve presents a variability of distribution patterns, which may be presented in the classical anatomical literature as a variability that is found differently in the population. In addition to inferring that the variation of the sciatic nerve can be little attributed to the symptoms of the piriformis muscle syndrome.
A Novel Murine Model for Studying Alcohol‐associated Biliary Dysfunction
Authors: Yan Shengmin , Ma Michelle , Yin Xiao‐Ming
Published: May 2022

Alcoholic hepatitis (AH) is a severe, potentially life‐threatening form of alcohol‐associated liver disease (ALD) with limited therapeutic options. Existing evidence indicates that cholestasis and biliary dysfunction are common in AH patients and can worsen the prognosis. However, most ALD murine models used today can only recapitulate hepatic steatosis with limited features of AH, which significantly curbs research related to ALD pathogenesis. To develop a murine model that is feasible and less time‐consuming, we included a subtoxic dose of α‐naphthylisothiocyanate (ANIT), a well‐studied intrahepatic cholestasis inducer, in an alcohol feeding model, as a second hit. Mice were fed with an ethanol diet (EtOH) for 10 days or 4 weeks, followed by ANIT administration 48 hours before sample collection. Our results showed that ANIT did not cause overt liver injury in pair‐fed mice, but an observable increase in serum levels of AST in mice fed EtOH for 10 days. Remarkably, in the 4‐week ALD model, serum levels of ALT, AST, total bile acids (TBA), and hepatic levels of TBA were all significantly increased in mice given EtOH and ANIT, indicating a synergistic effect of EtOH and ANIT on liver injury that is dependent on EtOH feeding. In the liver, 4‐week EtOH‐fed mice displayed cholangitis, lytic cell death, and neutrophil infiltration around the periportal area following ANIT treatment as shown by H&E and LY6G staining. In contrast, pair‐fed mice did not show abnormal liver histology following ANIT treatment. Expression of several inflammation‐related genes that are increased in livers from AH patients were analyzed. Consistent with histological observations, hepatic expression of genes like Cxcl1, Ccl2, Ccl20, and Lcn2 were significantly increased in 4‐week EtOH‐fed mice following ANIT treatment, suggesting that ANIT synergistically enhanced EtOH‐induced immune response in the liver. Taken together, our results suggest that EtOH‐dependent changes in the liver can cause higher susceptibility to biliary injury and that a subtoxic dose of ANIT pushes alcohol‐associated liver damage towards a more severe phenotype in mice. Overall, our results support a treatment scheme for alcohol‐associated cholestasis that is feasible, less time‐consuming, cost‐effective, and with modulable severity of phenotypes.
SDS‐induced amyloid fibrillation of acidic fibroblast growth factor
Authors: Alraawi Zeina , Maity Sanhita
Published: May 2022

Fibroblast growth factors and their receptors regulate a broad range of biological functions from developmental processes during embryogenesis to various physiological roles in the adult state, including regulation of angiogenesis, wound healing, and metabolism. hFGF‐1 also plays a key role in neuroprotection against neurotoxicity in Alzheimer’s disease (AD). The anionic surfactant sodium dodecyl sulfate (SDS) is widely used as a model for induction of amyloid‐like fibrils. In this context, we examined the effects of SDS on FGF1 using a variety of biophysical techniques including, Transmission Electron Microscopy (TEM). The secondary structure of hFGF1 was monitored by incubating hFGF1 with different concentrations of SDS utilizing the far‐UV CD (190‐250 nm) spectroscopy. We found that from 0.08‐0.15mM concentration of SDS, the negative peak at ~206 shifted gradually towards ~212nm while at 0.2mM concentration, far‐UV CD spectra of hFGF‐1 exhibit significant changes with striking shifting of negative peak ~212 nm to ~218 nm and disappearance of positive band at ~228nm which signifies substantial loss of native β‐barrel structure. The aggregation of hFGF‐1 is detected systematically by observing the absorbance at 350nm. The results indicate that at 0.2mM SDS, hFGF1 highly aggregates. In addition to far‐UV CD spectra and turbidity measurement, the influence of SDS on the tertiary structural changes of hFGF1 was studied by mentoring the intrinsic fluorescence of hFGF1 incubated with various concentrations of SDS. The study reveals that 0.2mM SDS is the concentration that affect mostly on the tertiary structure of hFGF1 because of the highest ratio of 305 to 350 at this concentration. Thioflavin‐T (ThT) assay for quantification of Amyloid‐like fibril formation was used to investigate the concentration of SDS that induce maximum hfGF1 fibrils. This study showed that 0.2mM SDS is the concentration that induced the optimum formation of fibrils hFGF1. Based on the results of these experiments, we wanted to do more investigation to achieve more evidence about the effect of SDS on the fibrils hFGF1. Therefore, morphological study by TEM was employed to examine the effect of 0.2mM SDS and 0.6mM SDS on the formation of fibrils hFGF1. At incubation for 1‐hour fibrillar aggregates of hFGF‐1 with average diameter at about 20‐25nm is observed in presence of 0.2mM SDS. All the previous results using biophysical techniques indicate that SDS induces formation of fibrils hFGF1 concentration which assist to better understanding about the causative of the fibrils hFGF1 which lead to find a remedy for the disease related with it.
Proline reduces half‐life of homocysteine thiolactone
Authors: Bhattacharya Reshmee , Singh Laishram R.
Published: May 2022

Homocysteine (Hcy) is a sulfur containing toxic amino acid synthesized from sulfur metabolism pathway. Elevated level of Hcy is associated with various cardiovascular complications including atherosclerosis. One of the likely mechanisms underlying harmful effects of Hcy is the chemical modification of proteins by homocysteine thiolactone (HTL), a highly reactive cyclic thioester of Hcy. HTL preferentially forms amide bonds with ɛ‐ amino group of protein lysine, arginine or histidine residues in a non‐enzymatic mechanism; a process referred to as “protein N‐homocysteinylation resulting in the impairment of protein functions. Therefore, novel approaches that could inhibit or delay protein N‐homocysteinylation would be therapeutically important to curb hyperhomocysteinemia. In the present communication, we screened for different cellular molecules produced under stressful insults for their ability to suppress covalent modification of proteins by HTL. We found that out of several molecules, proline has the potential to inhibit protein N‐homocysteinylation in a non‐enzymatic fashion. We also discovered that proline treatment increases Hcy levels in HeLa cells and induces cell cycle arrest. Our study highlights the importance of proline treatment in reducing toxicity load in hyperhomocysteinemic/homocystinuric patients.
Cryo‐EM structures of the Human GATOR1‐Rag‐Ragulator Complex Reveal a Spatial‐Constraint Regulated GAP Mechanism
Authors: Egri Shawn , Ouch Christna , Chou Hui‐Ting , Yu Zhiheng , Song Kangkang , Xu Chen , Shen Kuang
Published: May 2022

mTORC1 controls cellular processes in response to nutrient availability. Amino acid signals are transmitted to mTORC1 through the Rag GTPases, which are localized on the lysosomal surface by Ragulator. The Rag GTPases receive amino acid signals from upstream regulators. One negative regulator, GATOR1, is a GTPase activating protein (GAP) for RagA. GATOR1 binding to the Rag GTPases occurs via either of two modes: an inhibitory mode that has low enzymatic activity but high affinity, and a GAP mode that has high enzymatic activity but low affinity. How these two binding interactions coordinate to process amino acid signals is unknown. Here, we resolved three cryo‐EM structural models of the GATOR1‐Rag‐Ragulator complex, with the Rag‐Ragulator subcomplex occupying the inhibitory site, the GAP site, and both sites simultaneously. These structural models, together with the spatial constraints from the lysosomal membrane, reveal how GATOR1 coordinates the nucleotide loading states of both Rag subunits to transmit amino acid signals.
LncRNAs imported into mitochondria possess distinct features stratified by machine learning that promote interaction with the mitochondrial import protein PNPase
Authors: Taylor Andrew , Hathaway Quincy , Robart Aaron , Cook Chris , Kunovac Amina , Durr Andrya , Rizwan Saira , Cramer Evan , Starcovic Sarah , Hollander John
Published: May 2022

Understanding the localization and regulatory activity of extra‐nuclear long non‐coding RNAs (lncRNAs) is especially critical in the context of the mitochondrion, which possesses a genome separate from the nucleus that dictates bioenergetic function and is influenced by a broad range of pathologies. Prior research including efforts from our own laboratory have identified Polynucleotide Phosphorylase (PNPase) to be critical for selective RNA passage through the mitochondrial membrane, including microRNAs and other ncRNAs. However, the dynamics of lncRNA binding with PNPase have not yet been determined. Understanding how lncRNAs interact with PNPase is a crucial initial step for predicting their ability to be imported into mitochondria and how the process can be manipulated to regulate mitochondrial genome expression.

The objective of this study was to evaluate lncRNAs present in mitochondria and classify sequence‐based features that might permit interaction with PNPase. Sequencing performed on mitochondrial and cytoplasmic isolates from human and mouse cardiac tissue identified over 500 mitochondrially‐localized nuclear genome‐encoded lncRNAs. Crosslinked immunoprecipitation (CLIP) of mitochondrial isolates using antibodies for PNPase pulled down predominantly lncRNAs in both human and mouse. The most highly bound lncRNA sequences were run though supervised machine learning using 10‐fold cross validation through Support Vector Machines (SVM) and Classification and Regression Trees (CART) machine learning algorithms, which identified stratification of primary and secondary sequence features as compared to random RNA and low binding lncRNA sequences, with an accuracy of 82% for SVM and an area under curve value of 0.89 for CART. Primary sequence features such as dissimilarities from coding sequences, various k‐mer frequencies, and overall GC content correlated with increased lncRNA interaction with PNPase (p<0.001), as did secondary features such as minimum free energy and electron‐ion interaction pseudopotential (p<0.001). These features were integrated into an artificial RNA fragment sequence that was gel shifted with purified PNPase protein and showed higher binding affinity than a random RNA fragment of equal length.

LncRNAs are a unique species of ncRNA with a diverse range of regulatory functions. Understanding the mechanism behind lncRNA import into the mitochondrion could lead to novel therapeutic interventions to preserve bioenergetic activity. This study indicates that PNPase has affinity for rankable primary sequence and secondary structural features within lncRNAs, and application of the most critical features can be used to improve interaction with PNPase and potentially import into mitochondria.
Claudin‐23 Strengthens the Colonic Epithelial Barrier by Regulating Claudin‐3 and ‐4 proteins in the Tight Junction Plasma Membrane
Authors: Raya‐Sandino Arturo , Lozada‐Soto Kristen M. , Garcia‐Hernandez Vicky , Luissint Anny‐Claude , Koval Michael , Parkos Charles A. , Nusrat Asma
Published: May 2022

Colonic epithelial barrier function is controlled by differential expression of Claudin (CLDN) proteins in the crypt‐luminal axis. We recently identified the expression of a CLDN family member, CLDN‐23, in intestinal epithelial cells (IECs). However, the role of CLDN‐23 in regulating epithelial barrier function has not been identified. Analysis of the human and murine intestinal crypt‐luminal axis revealed increased CLDN‐23 expression in differentiated IECs facing the lumen compared to the proliferative crypt‐base IECs. Similarly, CLDN‐23 expression was higher in differentiated human and murine IECs cultured as a monolayer on permeable supports compared to proliferative IECs cultured in a 3D matrigel matrix. To investigate the contribution of CLDN‐23 in regulating IEC barrier function in vivo, we generated inducible intestinal‐epithelia specific (Villin‐CreERT2) Cldn23 knockout mice (Cldn23ERΔIEC). Functional analysis displayed increased intestinal mucosal permeability to 4kDa FITC dextran, indicating compromised epithelial barrier function. To confirm this observation, we created human IECs in which CLDN‐23 expression was either silenced or enhanced in vitro. CLDN‐23 loss increased paracellular permeability to 4kDa FITC dextran and reduced transepithelial electrical resistance (TEER) consistent with a leaky barrier. Conversely, CLDN‐23 overexpression resulted in improved barrier function. Altogether, these data suggest that CLDN‐23 controls the IEC barrier function.

Evaluation of CLDN protein interactions employing co‐culture of HeLa cells expressing CLDN‐23 with those expressing either CLDN‐2, CLDN‐3, or CLDN‐4 showed that CLDN‐23 interacts in trans with barrier‐forming CLDN‐3 and CLDN‐4, but not with pore‐forming CLDN‐2. Using in situ proximity ligation assay, we observed close association in cis between CLDN‐23 and barrier‐forming CLDN‐3 and CLDN‐4 in the TJ plasma membrane, but not with CLDN‐2. These findings suggest that CLDN‐23 interacts in both cis and trans with barrier‐forming CLDN‐3 and CLDN‐4 in differentiated IECs. Interestingly, the overexpression of CLDN‐23 in human IECs revealed up‐regulation of barrier‐forming CLDN‐3 protein expression in the TJ while decreasing the pore‐forming CLDN‐2 protein at this site. In conclusion, we identified that CLDN‐23 strengthens the colonic barrier properties by orchestrating the composition of CLDN protein interactions in epithelial TJs.
The Role of Scleraxis in Inducing Vascular Stiffness
Authors: Al‐Hattab Danah S. , Moffat Teri , Ledingham Allison , Czubryt Michael P.
Published: May 2022

Vascular fibrosis (VF) is one of the most significant pathologies related to blood vessel dysfunction. VF results in an impairment of vascular tone due to an increase in extracellular matrix proteins (ECM) and vascular smooth muscle synthesis within the vessel wall, thus increasing vascular stiffness and reducing lumen diameter. VF is both caused by and exacerbates numerous vascular diseases including diabetes, atherosclerosis and in particular, hypertension, which accounts for 13% of deaths globally (WHO World Health Report, 2002). Multiple studies have investigated various therapeutic approaches to interfere with VF, but to date this condition remains untreated, thus novel approaches to target fibrosis development and progression are urgently required.

Our lab identified the transcription factor scleraxis as a novel master regulator of cellular phenotype conversion. We demonstrated that scleraxis is sufficient to induce fibroblast to myofibroblast phenotype conversion, a critical step in the development of fibrosis in many tissue types. Scleraxis also induces Epithelial to Mesenchymal Transition (EMT) and cell migration as part of development and cell differentiation. Some studies have shown that mechanical stretch in tendons increases expression of scleraxis, while additional studies reported that stretch in rabbit aorta induces the vascular smooth muscle proliferative phenotype. Our preliminary data has revealed that scleraxis is expressed in VSMCs in the arterial wall, and we have found elevated scleraxis expression in high pressure versus low pressure regions of vessels. We thus hypothesize that scleraxis is sufficient and necessary to induce vascular fibrosis.

Our data shows that scleraxis overexpression increases thickness of the aortic vascular wall in scleraxis overexpression mice, via histological sections, thus contributing to the progression of vascular fibrosis. In vitro studies show that scleraxis overexpression in Human Aortic Smooth Muscle Cells (HAOSMC) induces their proliferation and migration using different assays (CCk8/Edu+ proliferation assay, Transwell migration assay, gene expression assays). Scleraxis overexpression also induces hypertrophy of HAOSMC shown via immunostaining. This suggests that scleraxis overexpression induces VSMC plasticity from contractile to synthetic‐proliferative phenotype, thus inducing VSMC hyperplasia.

New & Noteworthy: Our findings indicate that scleraxis plays a role in VSMC plasticity via scleraxis‐ mediated transcriptional regulation of VSMC proliferation and migration. This work has potential relevance to many related diseases that feature VSMC activation, including atherosclerosis and restenosis.
Does Cell Proliferation Affect Embryonic Flexure and Torsion?
Authors: Wang Dereck , Ramasubramanian Ashok
Published: May 2022

Two vital and simultaneous morphogenetic events that occur during early embryo development are the flexure and torsion (F&T) of the embryonic body axis. Both contribute to development of the fetal position, which can be observed in avian and mammalian embryos. Correct progression of F&T is crucial for the proper development of the heart and other organs. Embryonic flexure can be divided into three segments: cranial, cervical, and thoracic. In this study, we focus on cervical flexure and torsion of the embryonic body axis. Differential cell proliferation, the difference in the rate of division of cells between regions in the developing embryo, is one mechanism that could influence F&T. Goodrum & Jacobson (1981) observed thickening of the dorsal midbrain relative to the ventral midbrain during cranial flexure. Miller & White (1998) have shown using mouse embryos that differential growth could cause torsion of an axis confined to an attached sphere.

We performed studies inhibiting cell proliferation. Fertile white leghorn chick eggs were incubated to approximately Hamburger and Hamilton (HH) Stage 12 or 13. At these stages, F&T have just begun. We then added aphidicolin, a mitotic inhibitor, to whole embryo culture. Embryos were observed at 0, 5, and 10 hours to see if F&T were repressed. We hypothesized that the addition of aphidicolin would inhibit progression of F&T. We developed quantitative schemes to score the extent of F&T progression from HH 12 to HH 15. Embryos were scored from 0 to 4 based on the degree of cervical flexure and were also scored from 0 to 4 based on the degree of torsion. Afterward, the difference in F&T scores of same embryos at time t=0 h and t=10 h was calculated to determine the extent of both. These scores were then averaged for multiple embryos.

Compared with controls (n=46), embryos incubated in 400 µM aphidicolin (n=18) flexed and rotated at a significantly lower rate. For flexion, the mean difference in scores for control and treated embryos were 1.85 and 1.22, respectively. For torsion, the mean difference in scores for control and treated embryos were 1.62 and 1.25, respectively. These differences were statistically significant. It is noted that global application of aphidicolin did impact the overall health of embryos. However, our preliminary results indicate that differential cell proliferation could play a role in rotation and flexure progression in the developing chicken embryo.

References:

Goodrum, G. R., & Jacobson, A. G. (1981). Cephalic flexure formation in the chick embryo. Journal of Experimental Zoology, 216(3), 399‐408. doi:10.1002/jez.1402160308

Miller, S. A., & White, R. D. (1998). Right‐Left Asymmetry of Cell Proliferation Predominates in Mouse Embryos Undergoing Clockwise Axial Rotation. The Anatomical Record, 250(1), 103‐108. doi:10.1002/(sici)1097‐0185(199801)250:13.0.co;2‐s
Uncovering the Ciliogenesis Regulatory Network
Authors: Wesselman Hannah M. , Wingert Rebecca
Published: May 2022

Ciliopathies comprise disorders associated with aberrant function of essential organelles called cilia. Cilia are found in almost all vertebrate cells, with a subset of this population being motile and/or multiciliated. Over 1,000 proteins have been associated with the ciliary proteome, yet the regulatory circuit controlling the formation of cilia remains lesser known. Our lab has identified transcription factors and coactivators that contribute to the ciliogenesis circuit. Such factors include orphan nuclear receptor Esrrγ and coactivator Ppargc1a/PGC1a. Independently, these factors regulate both mono‐ and multi‐ciliated cells in the early signaling node, ear, and embryonic kidney (pronephros). Absence of these factors results in decreased multiciliated cells, decreased cilia length, and increased unciliated basal bodies. Considering these phenotypic similarities, we hypothesized a cooperative relationship between Esrrγ and Pppargc1a in the context of ciliated cell development. By modelling dual heterozygosity with genetic interaction studies, we found that ppargc1aworks synergistically with esrrγato regulate prostaglandin signaling to support ciliary outgrowth and multiciliated cell fate choice. While previous studies have alluded to the interaction of Esrrγ and Ppargc1a/PGC1a in metabolism, this is the first report of a cooperative effect that is essential for ciliogenesis or lineage decisions in development. This study has far‐reaching implications, as cilia are present in various tissues, and these factors may serve as therapeutic targets for ciliopathic conditions and birth defects.
Phosphorylation‐specific recruitment of human SCAF6 to RNA polymerase II during transcription regulation
Authors: Moreno Rosamaria Y. , Zhang Y. Jessie
Published: May 2022

The C‐terminal domain (CTD) of human RNA polymerase II (RNAPII) acts as a regulatory hub for RNA processing and progression of eukaryotic transcription. Transcriptional control is fine‐tuned through dynamic phosphorylation of the 52 repetitive heptads on the CTD to generate post‐translational modification patterns that precisely recruit cellular proteins according to transcriptional stages. However, it remains unclear what transcription regulators are differentially recruited by specific phospho‐marks. We assembled an enzymatic toolbox to recapitulate the dynamic phosphorylation patterns of CTD during transcription. We mapped the individual modification sites in high precision with advanced mass spectrometry strategies such as ultraviolet photodissociation. Utilizing a label‐free quantitative proteomics approach, we identified proteins that interact with different phosphoisoforms of Pol II CTD. Specifically, we identified RNA‐binding protein SCAF6 that are recruited for transcription by Ser2 and Thr4 phosphorylation. We characterized the interaction of SCAF6 with differentially phosphorylated CTD. The timely recruitment of SCAF6 is then investigated in cells using genome‐wide sequencing. In conclusion, we have expanded the repertoire of RNA‐binding proteins that directly interact with the CTD of RNA Pol II and illuminate the concerted interplay between splicing and transcription.
Sucrose reinforcement effects following inhibition of AMPA and NMDA‐type glutamate receptors in the ventromedial prefrontal cortex
Authors: Huerta Sanchez Laura L. , Lanz Mariana , Doan Hoa , Li Taylor , Szumlinski Karen
Published: May 2022

The glutamatergic system is purported to play an important role in the self‐administration of both drug and non‐drug reinforcers, such as sucrose. Sucrose‐experienced rats exhibit blunted glutamate release within the ventromedial prefrontal cortex (vmPFC) while cocaine‐experienced rats exhibit increased cue‐induced glutamate release within the vmPFC. While the behavioral phenomenon seems to be similar, there may be different mechanisms of glutamate in cocaine‐ and sucrose‐seeking. To address this question, this study investigated the role of vmPFC AMPA‐ and NMDA ‐type glutamate receptors in sucrose‐taking. For this, male and female Sprague‐Dawley rats were implanted with bilateral guide cannulae within the vmPFC and were trained to self‐administer sucrose pellets (6hr/day) for 10 days. Using a within‐subjects design, rats were microinjected (0.5 μl per side) with the GluA2‐lacking selective AMPA antagonist Naspm (40 μg), the non‐selective AMPA receptor antagonist NBQX (1 μg), the GluN2A‐selective NMDA antagonist PPPA (0.43 μg), the GluN2B‐selective NMDA antagonist TCN‐237 (0.002 μg) and the non‐selective NMDA receptor antagonist D‐AP5 (2.5 μg). Following each microinjection, rats underwent a 2‐hr test for sucrose self‐administration. In males, microinjection of D‐AP5 increased responding for sucrose reinforcement, while no significant effects were observed for the other compounds tested. While in females, there were no significant effects observed for any of the compounds tested. These results indicate that activation of NMDA receptors within the vmPFC suppress sucrose self‐administration in male rats, whereas AMPA receptors do not appear to be involved.
Activation of Endothelial Potassium Channels Lowers Pulmonary Arterial Pressure in Pulmonary Hypertension
Authors: Daneva Zdravka , Chen Yen‐Lin , Sonkusare Swapnil K.
Published: May 2022

Endothelium‐dependent vasodilation of small pulmonary arteries (PAs) lowers the resting pulmonary arterial pressure (PAP). Moreover, a loss of endothelium‐dependent vasodilation is a major contributor to elevated PAP in pulmonary hypertension (PH). Therefore, therapeutic strategies specifically targeting endothelial vasodilatory mechanisms may have beneficial effects on PAP in PH. In this regard, endothelial Ca2+ signaling mechanisms promote PA dilation and may present potential therapeutic targets for lowering PAP in PH. Under homeostatic conditions, Ca2+ signals activate endothelial nitric oxide synthase (eNOS) and Ca2+‐sensitive intermediate and small conductance potassium (IK and SK) channels to dilate PAs. eNOS activity and NO signaling are impaired in PH. However, the activity of IK/SK channels and its effect on PAP in PH have not been investigated. We hypothesized that IK/SK channels can be targeted for lowering PAP in PH. A mouse model of PH (chronic hypoxia + Sugen 5416, hereafter CH+Su, for 3 weeks) showed elevated right ventricular systolic pressure (RVSP, an indirect indicator of PAP) and development of pulmonary arterial lesions. We recently showed that adenosine triphosphate (ATP) dilates PAs through Ca2+‐dependent activation of eNOS in normal mice (Daneva et al., PNAS, 2021). PA pressure myography experiments demonstrated that ATP was unable to dilate PAs from CH+Su mice, confirming a loss of eNOS‐dependent vasodilation in these mice. However, PA dilation in response to IK/SK channel agonist NS309 (1 μM) was not different between PAs from control and CH+Su mice. Whole‐cell patch‐clamp studies in freshly isolated endothelial cells showed that IK/SK channel activity is not impaired in the endothelium from CH+Su mice. Finally, intraperitoneal administration of the IK/SK channel activator better suited for in vivo treatment, SKA‐31 (30 mg/kg), lowered the RVSP in CH+Su mice. Cardiac functional magnetic resonance imaging studies revealed that acute SKA‐31 administration does not alter the heart rate, stroke volume, ejection fraction, and cardiac output. Importantly, the RVSP‐lowering effect of SKA‐31 was not observed in normal mice. Collectively, our data reveal unaltered IK/SK channel activity in PH and provide proof of principle that IK/SK activation can be used as a strategy for lowering PAP in PH.
Distinct Wnt Signaling Thresholds Required for Polyposis in the Proximal and Distal Stomach
Authors: Delgado Elizabeth , Keeley Theresa M. , Hibdon Elise S. , McGowan Kevin P. , Gillilland Merritt G. , Colacino Justin A. , Stoffel Elena , Samuelson Linda C.
Published: May 2022

Objective

The Wnt pathway is critical for gastrointestinal stem cell proliferation. Patients with Familial Adenomatous Polyposis (FAP) have germline mutations in the adenomatous polyposis coli (APC)gene, resulting in dysregulated Wnt signaling and a polyposis syndrome with stark regional differences in disease severity. In the stomach, the proximal corpus contains numerous polyps, while few polyps are located in the distal antrum. Here, we tested the hypothesis that there are regional differences in sensitivity to Wnt signaling along the gastric proximal‐distal axis, and that a ‘just‐right’ level of Wnt signaling is required for gastric polyposis.

Methods

To model gastric polyp formation in FAP patients, we used mice with an inducible Apcloss‐of‐function mutation. Gastric tissue responses were studied one month after maximal Wnt activation in homozygous Sox2‐CreERT2; Apc580flox/flox(APC‐/‐) and submaximal Wnt activation in heterozygous Sox2‐CreERT2; Apc580flox/+ (APC+/‐) mice. To gain insight into regional differences in Wnt response, RNA sequencing was performed on corpus and antral organoids treated with the Wnt inhibitor XAV939.

Results

In the distal stomach, we observed hyperproliferation and tissue expansion in homozygous APC‐/‐ mice with no change in heterozygous APC+/‐mice. In contrast, the proximal stomach did not show an effect in homozygous APC‐/‐ mice but showed a modest increase in proliferation in heterozygous mice. PCR analysis of APC‐/‐ tissue confirmed APC gene deletion in the antrum, but not corpus, suggesting regional differences in maintenance of homozygous mutant stem cells. Following acute Wnt inhibition, corpus and antral organoids shared many Wnt‐regulated genes. However, Gene Ontology Enrichment analysis showed genes associated with cell differentiation, cell population proliferation, and cell cycle regulation were upregulated in antral organoids but downregulated in corpus organoids. KEGG analysis indicated downregulation of gastric cancer genes in both tissue organoids; however, only antral organoids showed downregulation of signaling pathways regulating pluripotency.

Conclusion

The corpus and antrum exhibit differences in the Wnt signaling threshold required to initiate polyposis. Whereas submaximal levels of Wnt signaling are sufficient to initiate growth in the corpus, maximal levels are needed for antral hyperproliferation. Notably, ‘high‐Wnt’ stem cells are retained in the antrum and lost from the corpus, suggesting regional differences in Wnt regulation of gastric stem cells. Our findings in mouse highlight how heterozygous APC mutations may be sufficient for proximal stomach polyposis—a clinically relevant observation, as the vast majority of FAP patient gastric polyps are benign sessile growths located in the proximal stomach, with less frequent antral adenomatous polyps more closely associated with gastric cancer.
Caffeine Augments the Lactate and Interleukin‐6 Response to Moderate‐Intensity Exercise in Men But Not Women
Authors: Abbotts Kieran , Ewell Taylor , Bomar Matthew , Butterklee Hannah , Rebik Jordan , Bell Christopher
Published: May 2022

The release of interleukin 6 (IL‐6) from contracting skeletal muscle is thought to contribute to some of the health benefits bestowed by exercise. This IL‐6 response appears proportional to exercise volume. Unfortunately, high volumes of exercise are not feasible for all people. Caffeine augments the magnitude of increase in circulating concentration of IL‐6 in response to high‐intensity and long‐duration exercise, in men. Caffeine is also known to increase circulating concentrations of lactate during exercise. One of the mechanisms thought to contribute to IL‐6 release from exercising skeletal muscle is lactate production. We hypothesized that caffeine, ingested prior to moderate‐intensity exercise, would lead to greater circulating concentrations of lactate and IL‐6 in a study population comprising both men and women. 15 healthy adults (9 males and 6 females, aged 26±7 years, (mean ± SD)) completed 30‐minutes of moderate‐intensity cycle ergometer exercise, equivalent to the ventilatory threshold, after ingesting either caffeine (6 mg/kg) or placebo. Arterialized‐venous blood was collected throughout each of the exercise sessions. Compared with placebo, caffeine increased end‐exercise circulating concentrations of lactate (5.72±3.95 vs. 7.14±4.66 mmol/L, P<0.001) but not end‐exercise IL‐6 (1.84±0.97 vs. 2.37±1.04 pg/mL, P=0.139). However, when women were excluded from the analysis, caffeine augmented (P=0.04) the magnitude of increase of end‐exercise IL‐6 concentration (1.80±0.86 vs. 2.57±1.21 pg/mL); this effect was further exaggerated after 30‐minutes of inactive recovery (3.81±2.32 vs. 5.06±3.22 pg/mL). Noteworthy, caffeine evoked greater end‐exercise lactate concentrations in data sets containing only men (P=0.02) and only women (P=0.002) but did not influence the IL‐6 response in women (P=0.94). Our preliminary data imply that in men unable/unwilling to perform high‐intensity and/or long duration exercise, caffeine may potentially enhance the IL‐6 mediated health benefits of relatively short, moderate intensity exercise.
Effects of T‐2 Toxin on the Intestinal Cecum Microbiome of Broiler Chickens
Authors: Grozina Alena A. , Ovchinnikova Natalya V. , Yildirim Elena A. , Ilina Larisa A. , Filippova Valentina A. , Laptev Georgiy Y. , Brazhnik Eugeniy A. , Dubrovin Andrei V. , Tyurina Daria G.
Published: May 2022

Many of the World's studies are aimed at identifying specific microbial markers of chicken intestines, including those markers that are associated with diseases. While mycotoxins can lead to bacterial disorder, gut microbiota could play a useful role in the prevention of mycotoxicosis. For this reason, it is important to characterize the bacterial populations that are involved in the dysbacteriosis caused by mycotoxins. This study was aimed the influence of diet contaminated with T‐2 toxin on caecum microbiome of Smena‐8 broilers. The experiment was carried out on 33‐day‐old broiler chickens for 14 days according to the Basic Principles of the Declaration of Helsinki. 4 experimental groups (5 chickens in each) were formed based on the concentration of the added T‐2 toxin: control 1, that received a diet without the contamination of T‐2 toxin, trial 2 ‐ received a diet with the addition of T‐2 toxin at a concentration of 100 μg/kg, trial 3 ‐ 200 μg/kg, trial 3 ‐ 400 μg/kg. Broiler chicken caecal content's genomic DNA was extracted and amplified of 16S rRNA NGS sequencing by the Miseq sequencer (Illumina, USA). The results showed that some microorganisms, in comparison with the control group, completely disappeared, and some, on the contrary, appeared when the feed was contaminated with T‐2 toxin. Members of the Mycoplasmataceae family and the phylum Candidatus Saccharibacteria have appeared in the intestines of chickens in trials 2,3 and 4. The average number of representatives of the Mycoplasmataceaefamily, among which pathogens are often found, increased (P≤0.05) under the influence of the toxin. The number of the Mycoplasmataceae family in groups in trial 2,3,4 was 0.0063 ± 0.0003, 0.0093 ± 0.0005 and 0.0130 ± 0.0007% respectively. On the contrary, representatives of other families (including Anaerolineaceae, Rickettsiaceae, Neisseriaceae), which are found in the control, completely disappeared from the intestines of chickens when contaminated with T‐2 toxin (trials 2,3 and 4). Thus, it can be concluded that T‐2 toxin provokes dysbiosis in the intestines of chickens. The number of the Lactobacillaceaefamily increased depending on the increase in the dose of mycotoxin – in trials 2,3 and 4 it was higher than in controls by 31.6, 45.6 and 46.3%, respectively. In all likelihood, lactobacilli are more resistant to mycotoxins than other members of the microbiota. This may be due to the fact that Lactobacillaceaemay play a role in the mycotoxin detoxification process. We believe that Lactobacillaceae, Mycoplasmataceae, Candidatus Saccharibacteria, Anaerolineaceae, Rickettsiaceae, Neisseriaceae are indicator groups of microorganisms that could be relevant biomarkers of toxicity of T‐2 toxin in chickens.