bluuebunny/crossref_metadata_2025_split huggingface dataset:
Long Non‐coding RNAs Mediate Heart Failure with Preserved Ejection Fraction Associated with Thyroid Dysfunction
Authors: Bounds Curren , Chakraborty Sankalpa , Dickerson Bryce , Shethwala Ruhi , Rajagopalan Viswanathan 
Published: May 2022

IntroductionHeart failure with preserved ejection fraction (HEpEF) carries significantly high morbidity and mortality. HFpEF is associated with preserved contractile function and can present with diastolic dysfunction, cardiac fibrosis and hypothyroidism (low levels of thyroid hormones, THs). We previously demonstrated safe, cardioprotective effects with treatment of active form of THs in multiple rodent models of cardiovascular disorders. Long‐noncoding ribonucleic acids (lncRNAs; >200 nucleotides long) account for large majority of the genome with significant transcriptional potential. However, their role in HFpEF associated with TH dysfunction is unclear. We hypothesized that lncRNAs act as mediators in development of hypothyroid HFpEF.MethodsHearts were isolated from obese ZSF1 rats that presented with HFpEF phenotype. Wild type and ZSF1 lean rats served as controls. Cardiac ultrasound and morphometric analyses were performed. RNA was isolated from left ventricular (LV) tissues and studied using Agilent LncRNA Microarray platform and analyzed using Feature Extraction and GeneSpring. Fibroblasts were isolated from wild type adult rat hearts and treated with TH receptor inhibitor. Real‐time quantitative polymerase chain reaction was used to study lncRNA expression.ResultsStudies revealed that total thyroxine TH levels significantly decreased in five‐six‐month old (early; p<0.05) ZSF1 obese rats compared to ZSF1 lean controls (45% decrease) and wild type controls (42% decrease), and this remained low at 13 months (late; p<0.05). Total tri‐iodothyronine TH levels also followed similar pattern in the early time‐point (37%‐42% decrease). ZSF1 obese rats showed increased normalized heart weight, LV weight and liver weight indicating cardiac hypertrophy and development of HF. In early obese rats, echocardiography showed preserved LV EF and significantly elevated E/E’ indicating diastolic dysfunction (p<0.05). LncRNA Microarray analysis including unsupervised hierarchical clustering showed that several unique lncRNAs were significantly altered (p<0.05; >2‐fold change) in obese LVs compared to the lean or wild type control counterparts and select lncRNA expression was validated with qPCR. Furthermore, isolated wild type adult rat cardiac fibroblasts subjected to TH receptor inhibition (mimicking hypothyroidism) in vitro showed significant increase in lncRNA expression that was also increased in obese ZSF1 hearts presenting with hypothyroidism (p<0.05).ConclusionsThese results indicate that lncRNAs may play important roles in in HFpEF pathogenesis associated with hypothyroidism. These studies will serve as significant step in identification of key lncRNAs in cardiac‐thyroid axis offering potentially valuable translational opportunities.
α1B‐Adrenoceptor is Required for Chemokine (C‐X‐C Motif) Receptor 4 Mediated Chemotaxis in THP1 Cells
Authors: Enten Garrett , Gao Xianlong , Majetschak Matthias 
Published: May 2022

IntroductionChemokine (C‐X‐C motif) receptor 4 (CXCR4) is a key regulator of leukocyte trafficking under physiological conditions and in numerous inflammatory disease processes. Previously, we demonstrated that CXCR4 hetero‐oligomerizes with α1B‐adrenoceptors (ARs), through which the receptors cross‐talk. The existence and function of these hetero‐oligomers in leukocytes, however, is unknown. Thus, we tested whether such heteromeric complexes are detectable in the human monocytic cell line THP1 and in freshly isolated human monocytes, and evaluated how α1B‐AR knockdown by CRISPR/Cas9 gene editing in THP1 cells affects CXCR4‐mediated chemotaxis.MethodsPeripheral blood mononuclear cells were isolated by density gradient centrifugation from human whole blood; monocytes were then isolated via negative selection using MACS LS, an indirect magnetic labeling system. Proximity ligation assays (PLA) were used to visualize individual receptors and receptor‐receptor interactions in THP1 cells and freshly isolated monocytes. CRISPR/Cas9 gene editing was utilized to create a α1B‐AR knockout strain (ADRA1BKO); wild type THP1 cells were used as a negative control (ADRA1Bwt). Cell migration toward the cognate CXCR4 agonist chemokine (C‐X‐C motif) ligand 12 (CXCL12) was tested employing a 96 well Boyden chamber. Transmigrated cells were counted automatically utilizing high contrast bright field and post‐imaging particle analyses. The chemotactic index (ChI) was calculated as the ratio of cells that transmigrated in the presence versus the absence of CXCL12. Data were analyzed with ANOVA/Tukey’s post‐hoc. Data are mean ± SE from n=4 experiments performed on different days. A 2‐tailed p<0.05 was considered significant.ResultsPLA confirmed the presence of endogenously and constitutively expressed CXCR4:α1B‐AR heteromers in freshly isolated monocytes and THP1 cells. PLA signals for α1B‐AR and CXCR4:α1B‐AR heteromers were not detectable in ADRA1BKO cells. CXCR4 signals were not affected by CRISPR/Cas9 gene knockout. ADRA1Bwtcells showed a bell‐shaped chemotactic dose response to CXCL12 with a maximal ChI of 9.1±1.1 at 100 nM. CRISPR/Cas9 knockout of α1B‐AR reduced the chemotactic response to CXCL12 by 89±52% (p<0.05). ADRA1BKO results were replicated in an additional α1B‐AR knockout strain to rule out clonal effects.ConclusionOur observations suggest that CXCR4 heteromerizes with α1B‐AR in human monocytes and that CXCL12‐mediated chemotaxis depends on the formation of CXCR4:α1B‐AR heteromers. These data provide evidence for a novel molecular mechanism by which CXCR4‐mediated chemotaxis in monocytes/leukocytes is controlled.
Mitochondrial Telomerase Prevents Chemotherapy‐Induced Cardiovascular Toxicity
Authors: Brandt Lukas , Hader Shelby N. , Norwood Toro Laura E. , Gutterman David D. , Beyer Andreas M. 
Published: May 2022

Anti‐cancer therapy (including doxorubicin (Dox)) is frequently used to combat growth of tumors. Chemotherapy (CTx) induced cardiotoxicity manifests as cardiomyopathy and can ultimately progress to heart failure and significantly contributes to morbidity and mortality among cancer survivors. Limited evidence demonstrates the impact of Dox on vascular endothelial function and possible contribution to progression of heart failure. TERT, the catalytic subunit of telomerase, has been increasingly recognized for its benificial effects on vascular quiescence via suppression of mitochondrial ROS to maintain mitochondrial integrity. Anthracycline treatment (e.g., Dox) correlates with both, suppressed TERT expression and activity as well as increased mitochondrial DNA damage. These combined effects of inadequate TERT levels and loss of mitochondrial integrity may contribute to CTx induced cardiotoxicity.The goal of this study was to define the contribution of TERT to CTx‐induced endothelial dysfunction and to cardiac dysfunction. We hypothesize that TERT gain of function prevents, while TERT loss of function augments CTx‐induced microvascular and cardiac dysfunction. We modulated TERT activation with a known telomerase activator cycloastragenol (CAG) via dietary supplement.In TERT WT animals, treatment with Dox significantly reduced endothelial dependent dilator function as early as two weeks after first dose. Dietary supplementation with CAG prevented Dox induced decrease in endothelial function (A). In contrast, in TERT‐specific KO animals Dox induced vascular damage was increased and CAG treatment failed to prevent endothelial dysfunction (B). Cardiac function followed similar patterns. In WT rats Dox caused significant reductions in left ventricular ejection fraction (LVEF) six weeks after the first dose of Dox, which was prevented by dietary supplementation with CAG. The reduction in LVEF was augmented in TERT KO animals and not rescued by CAG (C). Collectively, our data showed that Dox induced endothelial dysfunction was prevented by the pharmacological activation of TERT via CAG and furthermore suggests, that Dox induced endothelial dysfunction manifests before or independent of a cardiac damage. Likewise, CAG administration was sufficient to prevent reductions in LVEF relative to baseline in Dox treated WT animals. Specificity of a TERT mediated phenotype via CAG was established by failure of rescue in TERT KO rats. Ultimately, our results indicate that CAG has the potential as therapeutic to counteract the detrimental effects of CTx in the cardiovascular system via activation of TERT.
Variation in ORF3a Protein of SARS‐CoV‐2 Decreases The Severity of Host Cell Damage
Authors: Landherr Maria , Cypress Michael W. , Chaput Isabel , Jhun Bong Sook , O‐Uchi Jin , Polina Iuliia 
Published: May 2022

IntroductionSevere acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) primarily targets the respiratory system. However, direct SARS‐CoV‐2 infection and viral protein expression have also been reported in other organs, which potentially contributes to multi‐organ dysfunction and increased mortality in COVID‐19. Since the original Wuhan‐type genome was sequenced, several genetic variants of SARS‐CoV‐2 have emerged with differing pathophysiological properties such as the levels of transmissibility, disease severity, and mortality. We previously reported that the protein encoded by open reading frame 3a (ORF3a), a critical protein for SARS‐CoV‐2 replication and release, is found in the mitochondria of host cells and could increase oxidative stress and apoptotic signaling. ORF3a‐Q57H is a highly recurrent variation and the most commonly found variant of ORF3a. Interestingly, the Q57H variant is associated with increased transmissibility, but lower mortality.AimTo investigate the impact of the ORF3a‐Q57H variant on host cell damage.MethodsPlasmids carrying ORF3a‐Q57H were generated by PCR‐based site mutagenesis using Wuhan‐type ORF3a (ORF3a‐WT) as a template. Whole cell lysates were prepared from HEK293T cells and H9c2 cardiac myoblasts expressing ORF3a‐WT or the mutant ORF3a‐Q57H and used for biochemical assays. Live cell imaging for assessing subcellular localization of ORF3a/ORF3a‐Q57H, mitochondrial reactive oxygen species (mROS), and caspase 3 activity in H9c2 cells was performed by confocal microscopy.ResultsIn whole cell lysates, we found that ORF3a‐Q57H exhibits significantly higher protein expression compared to ORF3a‐WT. However, there is no significant difference in the ability of mitochondrial trafficking between ORF3a‐WT and ORF3a‐Q57H assessed by live cell imaging using GFP‐tagged ORF3a/ORF3a‐Q57H with mitochondria‐targeted DsRed. Next, we investigated the effects of ORF3a expression on apoptotic and mitophagy signaling by quantifying the caspase 3 activity and LC3A/B ratio. We found that ORF3a‐Q57H has significantly lower apoptosis and mitophagy signaling compared to WT despite its higher protein expression levels. ER‐stress signaling was not activated in either ORF3a‐WT or ORF3a‐Q57H, as assessed by markers including Glucose‐regulated protein 78/94 and C/EBP‐homologous protein. Lastly, live cell imaging using the mitochondrial superoxide‐sensitive dye Mitosox Red revealed that ORF3a‐WT significantly increases mROS levels, but ORF3a‐Q57H expression does not.ConclusionSARS‐CoV‐2‐ORF3a‐Q57H causes lower oxidative stress and cell damage compared to the Wuhan‐type variant, which could help explain the decreased mortality associated with the ORF3a‐Q57H variant. These results provide novel insights on how genetic variations of SARS‐CoV‐2 influence the pathophysiology and clinical severity of COVID‐19.
Ablation of PDE4A Exacerbates Pseudomonas aeruginosa‐Induced Acute Lung Injury in Mice
Authors: Boyd Abigail , Abou Saleh Lina , Aragon Ileana , Fiedler Edward , Irelan Daniel , Spadafora Domenico , Barrington Robert , King Judy , Richter Wito 
Published: May 2022

Type 4 cyclic nucleotide phosphodiesterases (PDE4s) comprise a family of four isoenzymes, PDE4A to D, that hydrolyze the second messenger cAMP. Three of these enzymes, PDE4A, PDE4B and PDE4D, are widely expressed throughout pulmonary and immune cell types, and their concurrent inhibition with non/PAN‐selective PDE4 inhibitors is well‐known to exert potent anti‐inflammatory benefits. To determine if PDE4 inhibition may represent a therapeutic approach for acute lung injury resulting from severe pneumonia, we explored the effect of genetic ablation of individual PDE4 subtypes, or their concurrent inhibition with PAN‐PDE4 inhibitors, in a model of acute intra‐tracheal Pseudomonas aeruginosa (PA) infection in mice. We have shown recently that selective ablation of PDE4B is sufficient to protect mice from lung injury in this model by reducing pulmonary and systemic levels of pro‐inflammatory cytokines, as well as pulmonary vascular leakage and mortality. With the present study we explored whether selective inactivation of PDE4A or treatment with PAN‐PDE4 inhibitors may provide additional therapeutic benefits.MethodsMice deficient in PDE4A (PDE4A‐KO) and their wildtype (WT) littermate controls were infected intratracheally with the PA lab strain PA01 and subjected 16 h later to bronchoalveolar lavage (BAL), cardiac blood draws, and tissue extractions. Alternatively, vascular/alveolar leakage was assessed by measuring Evans Blue Dye extravasation. To test the effect of PAN‐PDE4 inhibition, mice were treated with the PDE4 inhibitor Piclamilast (5 mg/kg, i.p.) at 2 h prior to PA01 infection.ResultsCompared to WT controls, PDE4A‐KO mice displayed a hyper‐inflammatory phenotype characterized by increased levels of pro‐inflammatory cytokines, immune cells (neutrophils) in BAL fluid, and vascular leakage. Despite these hyperinflammatory responses, the bacterial load in lungs was also increased in PDE4A‐KO mice compared to WT controls. Treatment with a PAN‐PDE4 inhibitor produced incongruent effects, with an increased bacterial load mirroring the effect of PDE4A ablation, whereas reduced levels of inflammatory cytokines such as TNFα mimicked the effect of PDE4B ablation.ConclusionsAblation of PDE4A produces detrimental effects in this model of acute PA‐lung infection. Current studies aim to determine whether an impaired bacterial killing, and the resulting increase in bacterial load, are responsible for the hyperinflammation, or whether these are independent effects of PDE4A ablation. PAN‐PDE4 inhibition mimics some of the beneficial effects of selective PDE4B ablation, but also some of the detrimental effects of PDE4A ablation. These findings suggest that selective inactivation of PDE4B may be a more effective therapeutic approach, compared to PAN‐PDE4 inhibition, in settings of acute PA‐lung infection.
Exploring Sex Differences in Renal Sodium Transporters with Four Core Genotype (FCG) Model
Authors: McDonough Alicia A. , Ralph Donna L. , Soong Joanne , Carrisoza‐Gaytan Rolando , Kleyman Thomas R. , Satlin Lisa M. 
Published: May 2022

In the kidney, sex‐specific differences in Na+ transporter profiles along the nephron are now evident (“transporters” = co‐transporters, channels, claudins and pumps, their phosphorylation (p)): females (F) exhibit variable differences vs males (M) along the proximal tubule (PT) and higher distal (DT) and collecting duct (CD) Na+ transporter abundance and activity vs. M. Circulating levels of gonadal hormones determine sex differences in many physiologic traits, however, recent studies indicate that sex differences may also be due to the sex chromosome complement (SCC; XX vs.XY). The novel FCG C57BL/6 mouse model dissociates gonadal sex (ovaries (F) or testes (M)) from sex chromosomal complement (SCC; XX, XY). That is, traits influenced by gonadal hormones are similar between MXX and MXY (testes) vs FXX and FXY (ovaries); whereas traits influenced by SCC are similar between MXX and FXX vs. MXY and FXY (independent of gonads).MethodWe implemented the 4CG model with transporter profiling (semi‐quantitative immunoblotting; n = 4‐5 mice/genotype) to reveal the contributions of gonadal hormones vs. SCC on the sexual dimorphisms in transporter protein abundance along the nephron.Results(all P<0.05): Along the PT: OAT1 was lower and AQP1 and SGLT2 higher in abundance in gonadal F (FXX) vs M (MXY); SCC also contributed to differences: NHE3 was lower by 15‐30% in MXX vs. MXY and SGLT2 was 15% lower in FXY vs FXX. Inthe medullary thick ascending limb (TAL): alpha and beta NKATPase are 60% higher in FXX vs MXY and 25‐30% lower in FXY vs FXX; alpha was 15% lower in MXX vs. MXY. Along the cortical TAL and DT: NKCC2, NKCC2p, NCC, NCCp, alpha and beta NKATPase, claudin 7, SPAK and SPAKp kinase abundance were 1.4 ‐ 2.6 ‐ fold higher in FXX vs MXY; SCC differences were limited to 15%, 30%, and 20% lower NCC, and NCCp, and claudin 7 in MXX vs MXY, respectively. Along the CD: ENaC subunits were 30‐50% higher, claudin‐8 2‐fold higher, pendrin and UMOD 20% higher in FXX vs. MXY; SCC differences included 15‐20% lower ROMK and Kir 4.1 and ENaC beta and gamma subunits in MXX vs MXY. Results provide evidence for impact of both gonadal hormones and SCC on kidney transporter abundance, in some cases, offsetting the effects of the other. Further progress will come from defining whether X vs Y genes contribute to the differences. These findings have potential impact on electrolyte and blood pressure (patho)physiology in F vs. M.
Effect of Prolonged High‐Altitude Exposure on Heart Rate Response to Central Sleep Apnea
Authors: Berthelsen Lindsey , Steele Andrew , Vanden Berg Emily , Bird Jordan , Thrall Scott , Wilson Richard , Jendzjowsky Nicholas , Day Trevor , Steinback Craig 
Published: May 2022

Central sleep apnea (CSA) is common at high‐altitude (>2500m) and is characterized by a cyclical pattern of hyperventilation and transient arterial hypocapnia, triggering apnea. We have demonstrated that voluntary wakeful apnea at altitude promotes a greater degree of bradycardia and cardiac arrhythmias than at sea level. However, whether bradyarrhythmias are observed during CSA at altitude has not been investigated. We investigated the duration dependent effect of altitude on heart rate changes during periods of apnea overnight. We hypothesized that overnight apnea would result in a greater magnitude of bradycardia with prolonged hypoxic exposure.Central sleep apnea (Apnea‐hypopnea index; AHI) was assessed overnight at low altitude (1045m), early (Night 2‐3) and late (Night 9‐10) at high altitude (N=14; 6 female). Heart rate (HR) and rhythm (electrocardiogram; ECG lead II) was collected alongside sleep data (RESMED Apnealink) to quantify sleep apnea events and desaturations. HR data were averaged over 10s prior to and 10s following apneas. Delta HR during apnea (BL HR – Nadir HR apnea) was determined to characterize bradycardia. Differences in heart rate prior to, during, and post apnea were compared using a repeated‐measures ANOVA design, with a Holm‐Sidak post‐hoc analysis where main effect of condition was significant. The magnitude of bradycardia during apneas between early and late time points were compared using a paired t‐test. Additional analyses of sex differences were performed using a two‐way mixed model ANOVA.AHI was augmented at high altitude (main effect p=0.012) but not different between early and late time points (p=0.193). The incidence of central apneic events was greater late at high‐altitude compared to low altitude (p=0.0045). Overnight oxygen saturation was decreased at high altitude (main effect of time, p<0.01) but was elevated late (84 ± 3%) compared to early (82 ± 4%) at altitude (p=0.038). Average HR overnight was similar between early (74 ± 11bpm) and late (69 ± 10bpm) time points (p=0.134). HR dropped during apneas at early (‐6.2±4.6 bpm; p=0.018) and late (‐6.4±2.8 bpm; p<0.001) timepoints. Female participants had a lower AHI at high altitude (p=0.006) but tended to have a greater bradycardia (‐7.7± 5.6bpm early, ‐7.6±2.9bpm late) compared to men (‐3.9±0.68bpm early; ‐4.5±1.4bpm late; main effect of sex, p=0.052).We demonstrate that CSA at altitude is associated with bradycardia that does not appear to become greater with prolonged exposure. It appears females may be less susceptible to CSA but have a greater bradycardic response during apneas compared to males.
Mimicking a Model: A Novel Approach to Pelvis and Perineum Dissection
Authors: McManus Lauren , Reynolds Amberly 
Published: May 2022

Pelvic dissections are difficult for students to visualize and complete given the standard dissection approach and limited time provided in standard anatomy curricula at U.S. medical schools. The purpose of this project is to propose a novel approach to dissecting the pelvis with aims of increasing student understanding of pelvic anatomy. The proposed novel technique allows for ease of viewing by making the donor more mobile and more model‐like, like what is seen with common anatomical models used at medical schools.The technique begins with transverse sectioning of the body into, an upper, mid, and lower section with the mid‐section containing the pelvic region. This section allows the dissector to utilize the pelvic bowl in a standing position, show cross‐sectional anatomy of the thigh muscles, and provide focus on three different dissection pathways. The pathways created include, neurovascular, organ, and muscle specific.On a preliminary dissection, this method provided ease at dissecting this region, a structurally similar view to live bodies and models, and additional pathways for learning. This dissection was done by a first‐year medical student six months following typical anatomical dissection. The technique is beginning to redefine in situ study of the pelvis and perineum by providing an alternative, more focused way to utilize donors for this region. Models tend to provide the “perfect” anatomy, which can be useful in understanding the typical relationships, however, it is important for students to understand this region and the relationships found with each structure to help better prepare them for future patients.Future studies to enhance this technique will determine how the pathways can be better utilized. In having students perform this technique alongside the typical techniques, a comparison can be made with a larger population. By having subjective feedback, it can show how students interacted with the dissection, as well as how it was utilized throughout this region of study.