bluuebunny/crossref_metadata_2025_split huggingface dataset:
Chronic kidney disease‐induced insulin resistance is mediated by increased phosphorylation of SGK1 and enhanced SGLT2 expression in NTS
Authors: Chen Hsin-Hung , Cheng Pei-Wen , Tseng Ching-Jiunn 
Published: April 2020

BackgroundThe serum and glucocorticoid inducible‐kinase‐1 (SGK1) expression is regulated during discrete developmental stages and pathological conditions, such as hypertension and diabetic neuropathy. In the present study, chronic kidney disease (CKD) in rats enhanced SGK1 function and regulation within the nucleus tractus solitarii (NTS). This study aimed to assess the regulatory mechanism of SGK1 in the NTS in rats presenting CKD‐induced insulin resistance.MethodsEight‐week‐ and nine‐week‐old male Wistar–Kyoto (WKY) rats underwent nephrectomy of the left and right kidneys, respectively. After subjecting the rats to CKD‐induced hypertension for 2 weeks, they were treated with SGK1 inhibitor (GSK650394, 100 mM) for another 2 weeks. Furthermore, the rats underwent 5/6 nephrectomy for 4 weeks; thereafter, they were tested for blood pressure, blood and urine chemistry, renal function, and protein expression. Mann–Whitney U test was used to compare the mean rank differences in scales of the general parameters between the CKD (sham, CKD, and CKD + GSK650394) groups, and the values were presented as mean ± SEM.ResultsRats with CKD revealed proteinuria, lower urine sodium and chloride levels and hyperglycemia, while treatment with SGK1 inhibitor significantly decreased these effects. Immunoblotting analysis further revealed that phospho‐SGK1 (Ser78), phosphoinositide‐dependent kinase‐1 (PDK1), and sodium‐glucose co‐transporter 2 (SGLT2) expressions increased in the NTS of CKD rats. Furthermore, inhibition of phospho‐SGK1 (Ser78) by GSK650394 significantly decreased the expression of PDK1 and SGLT2 in the NTS of CKD rats.ConclusionOur results suggest that inhibition of SGK1 expression in the NTS by intracerebroventricularly injected SGK1 inhibitor decreases the fasting serum glucose by negatively regulating CKD‐induced SGK1 signaling, reducing SGK1 (Ser78) phosphorylation, and by inhibiting the PDK1 and SGLT2 expressions in the NTS. These results indicate that SGK1 regulates the cardiometabolic syndrome in the NTS of CKD rats.Support or Funding Information107‐2320‐B‐075B‐002‐MY2、107‐2320‐B‐075B‐001
Sensitizing Aptamer INSR‐A43 Enhances Insulin Sensitivity and Sensitizes Insulin Receptor Signaling Pathways
Authors: Park Mangeun , Yunn Na-OH , Ryu Sung Ho 
Published: April 2020

Aptamers are single‐stranded oligonucleotides (DNA or RNA) and form a unique three‐dimensional (3D) structure based on its sequence. These aptamers have high affinity and specificity to target molecule and recognize 3D structure of target molecule surface. For this reason, aptamers have been in the spotlight as a universal capture. Especially, aptamers can modulate target protein as allosteric agonist or allosteric antagonist. However, aptamer‐based allosteric enhancer or sensitizer (hereafter ‐ sensitizer) has not been elucidated. Here, for the first time, we show the INSR‐A43, which is aptamer‐based sensitizer for the insulin receptor. Because insulin receptor plays a key role in regulation of glucose metabolism, it is important to specifically modulate the insulin receptor. INSR‐A43 binds to allosteric site of the insulin receptor and then increases insulin binding to the insulin receptor. Taken together, INSR‐A43 effectively enhances insulin sensitivity to the insulin receptor and sensitizes insulin receptor and insulin receptor‐related downstream signaling pathways. We expect that aptamers can be applied as an attractive tool for receptor sensitization.Support or Funding InformationThis research was supported by Grobal Research Laboratory (GRL) Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (No. NRF‐2016K1A1A2912722)
Parental obesity exacerbates cardiovascular responses to acute stress in male, but not female, offspring
Authors: do Carmo Jussara M. , Moak Sydney P. , Dai Xuemei , Hall John E. , Silva Alexandre A. D. A. 
Published: April 2020

The proportion of women and men of reproductive age who are overweight and obese has been increasing considerably, and maternal obesity and excessive gestational weight gain may predispose their offspring to health‐related consequences during childhood and adulthood. Obesity may also be associated with increased cardiovascular responses to mental stress. However, there is no available evidence on the effects of intergenerational obesity on cardiovascular responses to acute stress in adult offspring. In this study, we examined whether the pressor response to acute stress is exacerbated in obese offspring from obese parents. Offspring fed normal diet (ND) born from ND parents (N‐F1‐N, n=5 per sex) used as control, offspring fed high fat diet (HFD) born from ND parents (N‐F1‐H, n=5 per sex) and offspring fed HFD born from HFD parents (H‐F1‐H, n=5 per sex) were used. Parents were fed HFD or ND after they were weaned, while the offspring were placed on HFD from weaning until the end of experiments. Mice were implanted with telemetry probes for measurement of mean arterial pressure (MAP) and heart rate (HR). After a 10 day recovery period, baseline MAP and HR were continuously measured for 30 minutes before, during 5‐minute air‐jet stress consisting of pulses of compressed air directed at the animal’s forehead, and 30 minutes after the stress test. Male and female H‐F1‐H mice were heavier than N‐F1‐H and N‐F1‐N control mice (47.3±0.8 vs. 42.5±1.9 and 33.1±0.2 g and 41.1±0.1 vs. 37.2±0.1 and 24.1±0.1 g, respectively). Male H‐F1‐H mice had higher MAP (110±4 mmHg) compared to N‐F1‐H and control mice (100±6 and 99±3 mmHg). Acute air‐jet stress significantly increased MAP by 36±2 mmHg in male H‐F1‐H mice compared to only 26±3 and 28±3 mmHg in male N‐F1‐H and control mice, respectively. MAP remained significantly higher post stress in H‐F1‐H mice (133±2 vs. 114±6 and 110±3 mmHg). Baseline HR and HR responses to air‐jet were similar among the male groups. In females, however, baseline MAP and HR as well as MAP and HR responses to air‐jet stress test were similar among groups. These results indicate that male, but not female, offspring fed a HFD from obese parents have higher baseline blood pressure (BP) and greater BP responses during acute stress, and their BP remains elevated for a longer period post stress. These observations also suggest that parental obesity may have greater effect on male offspring cardiovascular system compared to female offspring.Support or Funding InformationNIDDK 1RO1121411, NHLBI‐PO1HL51971, NIGMS P20GM104357 and U54GM115428
N‐ and C‐Terminal Alzheimer’s Aβ Heterogeneity Modulates the Balance Between Brain Clearance and Amyloid Formation
Authors: Ghiso Jorge , Cabrera Erwin , Neubert Thomas A. , Rostagno Agueda 
Published: April 2020

Impaired brain clearance and abnormal Aβ degradation are today considered key events for the formation and progressive accumulation of soluble neurotoxic oligomers and the development of synaptic pathology, one of the strongest correlates to cognitive impairment in Alzheimer’s disease (AD). Clearance studies, mostly centered in monomeric Aβ40, have provided a basic assessment of the participating mechanisms, but the complex molecular and structural heterogeneity of the brain Aβ peptidome –going far beyond the classic Aβ40/Aβ42 dichotomy–has been largely overlooked. The most common heterogeneity resides in the multiple truncated fragments that consistently populate the Aβ peptidome and of which little is known in regard to their homeostatic regulation and potential contribution to disease pathogenesis. Tissue specimens from AD cases and transgenic models subjected to differential solubility extractions followed by immunoprecipitation coupled to mass spectrometry revealed the extent and characteristics of the multiple N‐ and C‐terminal truncated Aβ species. The data indicate that C‐terminal truncations increase Aβ solubility and abrogate oligomerization and fibrillogenesis, features clearly associated with clearance mechanisms, whereas N‐terminal truncations, particularly abundant components of parenchymal and vascular amyloid deposits in AD, non‐human primates, and multiple APP transgenic models drive enhanced pro‐amyloidogenic properties. Biophysical studies using synthetic homologues confirmed the differences in solubility further illustrating the contrasting oligomerization/fibrillization characteristics of the various Aβ truncated derivatives, and novel antibodies recognizing specific N‐ and C‐terminal truncations differentially immunolabeled amyloid deposits in AD brains and transgenic models. Intracerebral injections of monomeric and oligomeric radiolabeled homologues revealed striking differences in their brain clearance characteristics, showing that while soluble derivatives exhibited a fast brain removal, oligomerization largely increased brain retention, a characteristic particularly evident in fragments truncated at Phe4, topographically abundant in fibrillar, thioflavin positive plaque cores. Overall, our data indicate that degradation at the C‐terminus generates highly soluble Aβ fragments that are common components of interstitial and cerebrospinal fluids, and likely associated to catabolic/clearance mechanisms whereas truncations at the N‐terminus favor oligomerization and brain retention, with the potential to exacerbate the process of amyloid formation and self‐perpetuate the amyloidogenic loop.Support or Funding InformationNIH grants AG059595 and AG051266, and the BrightFocus Foundation grant A2015275S
A High‐throughput CRISPR Interference Screen to Dissect Functional Regulators of GPCR/cAMP Signaling
Authors: Semesta Khairunnisa Mentari , Tian Ruilin , Kampmann Martin , von Zastrow Mark , Tsvetanova Nikoleta G. 
Published: April 2020

The beta‐adrenergic (β2‐AR) receptor is a prototypical GPCR that controls important functions in the nervous, pulmonary, and cardiovascular systems. β2‐AR signaling involves multiple factors in both G‐protein dependent and independent signaling, resulting in diverse signaling outcomes. To understand the complexity of this pathway, we have performed the first unbiased, high‐throughput functional genomics screen of β2‐AR signaling using cAMP reporter and CRISPR interference (CRISPRi) silencing. We generated a reporter cell line using a construct encoding cAMP‐response element (CREB) fused to a ProteoTuner destabilizing domain and green fluorescent protein and another construct encoding catalytically‐dead Cas9 (dCas9) fused to the transcriptional repressor KRAB. Reporter cells were transduced with genome‐wide lentiviral sgRNA library to silence individual genes. After induction of β2‐AR signaling, we sorted the edited cells based on reporter expression and subjected them to deep sequencing and statistical analysis of sgRNA enrichment. We obtained 32 positive and 14 negative β2‐AR/cAMP signaling modulators enriched in factors involved in transcription initiation, RNA splicing, and RNA metabolic processes. We validated TP53 and NUP93 as novel positive modulators of β2‐AR signaling using dual luciferase cAMP assay and RT‐qPCR of cAMP‐dependent target gene, consistent with our cAMP reporter result. We found that the TP53‐dependent modulation of this pathway can partially be explained by lower transcription and surface expression of the β2‐AR receptors. These findings demonstrate a successful application of our reporter in a high‐throughput approach to identify previously uncharacterized modulators of the β2‐AR/cAMP pathway. This study reveals broader potential applications of our cAMP reporter as a platform to understand other aspects of GPCR signal transduction such as biased signaling, receptor‐specific molecular mechanisms, and drug discovery.Support or Funding InformationThis work is supported by the National Institute of Mental Health R00 award (MH109633) awarded to Nikoleta Tsvetanova.
Pleiotropic Activation of Endothelial Function by Angiotensin II Receptor Blockers is Crucial to Their Protective Anti‐vascular Remodeling Effects
Authors: Tehrani Arash Y. , Fameli Nicola , White Zoe , Esfandiarei Mitra , van Breemen Casey , Bernatchez Pascal 
Published: April 2020

RationaleEndothelial function is strongly associated with vascular health and homeostasis. However, therapeutically relevant approaches capable of stimulating the protective properties of the endothelium remain elusive.ObjectiveTo determine whether modulation of the Angiotensin II (AngII) system with Angiotensin receptor blockers (ARBs) can improve endothelial function and promote downstream vascular homeostasis.Methods and ResultsWe show that all 4 ARBs tested (losartan, telmisartan, olmesartan, and valsartan) can promote robust chronic activation of endothelial nitric oxide (NO) in mice. With telmisartan, a well‐known pleiotropic ARB, in vivo treatment lead to a 58% inhibition of vascular contractility after two weeks of treatment and up to 77% at 6 months, in a fully L‐NAME sensitive fashion. Endothelial NO synthase (eNOS) knockout (KO) mice showed complete resistance against the endothelial function effects of telmisartan, whereas the Angiotensin converting enzyme inhibitor (ACEi) captopril showed little to no endothelial function activity at a similar blood pressure (BP) lowering dose. In a myograph chamber, direct stimulation of aortic vessels with telmisartan in the absence of AngII stimulated endothelial function, confirming pleiotropism. Moreover, in vitro stimulation of cultured endothelial cells caused rapid intracellular Calcium ([Ca2+]i) mobilization, which is a known upstream event of eNOS activation. In vivo, telmisartan completely abolished aortic wall remodeling in aging animals as well as a well‐established mouse model of Marfan syndrome (MFS), which causes accelerated and pre‐mature vascular aging and remodeling of the aortic root. In addition, in vivo inhibition of NOS activity with L‐NAME rendered telmisartan inactive in a BP‐independent manner.ConclusionARBs have unique and robust endothelial function activation properties independent of their inhibitory effects on the AngII system. Direct stimulation of the endothelial NO system with ARBs results in vascular protection and homeostasis independent of BP lowering effects, which supports a broader prophylactic use of this class of medication.Support or Funding InformationSupported by CIHR, HSFC, BCKDF, Marfan Foundation, SPH Foundation and Rare Disease Foundation.
Heart Rate Response of Chicken Embryos to Room Temperature at Different Ages
Authors: Sycip Oliver C. , Brundage Cord M. 
Published: April 2020

Chicken (Gallus gallus domesticus) embryos are used as a research model. Embryotic heart rate (HR) can be monitored non‐invasively for health as well as a physiological response to experimental stimuli. References ranges for embryotic HR vary with cardiovascular and embryonic development. These may be influenced by the environment in which they are measured. A decline in heart rate (HR) has been documented in embryos following removal from incubation. This was hypothesized to be a response to the drop in ambient temperature. The magnitude and duration of this response may vary with embryonic development. To evaluate the effect of ambient temperature on chicken embryo HR, 10‐day, 15‐day, and 18‐day old chicken embryos (n = 20, 15 and 15 respectively) were placed on HR monitors for 10 min at both incubation (37.7 ± 0.1 °C) and room (25.6 ± 0.4 °C) temperatures. The pattern of HR change was similar in both temperature groups. HR predictably increased with embryonic age (p < 0.05); HR across all ages decreased similarly (p > 0.10). After a decrease in HR after 3 min (−30 ± 39 bpm, p < 0.01), HR did not continue to decline (p > 0.05). Differences in both HR between embryotic stages and changes in HR over the 10 minute trials were temperature independent (p > 0.05; p > 0.10). This would suggest that chicken embryo research can be conducted at room temperature within 10 min (in 10 – 18‐day embryos) without temperature significantly impacting HR. Initial decreases in HR may not be due to temperature, but rather other factors associated with movement and manipulation. When looking at changes in HR, egg handling procedures and embryo age should be considered and controlled.Support or Funding InformationResearch reported in this publication was supported by the MENTORES (Mentoring, Educating, Networking, and Thematic Opportunities for Research in Engineering and Science) project, funded by a Title V grant, Promoting Post‐baccalaureate Opportunities for Hispanic Americans (PPOHA) | U.S. Department of Education, Washington, D.C. PR/Award Number: P031M140025. The content is solely the responsibility of the authors and does not necessarily represent the official views of the Department of Education.
A Glial Cell Inhibitor Blocks Epithelial Barrier Repair in a Pig Model of Intestinal Ischemia
Authors: Sheridan Ana , Pridgen Tiffany , Odle Jack , Van Landeghem Laurianne , Blikslager Anthony , Ziegler Amanda 
Published: April 2020

The gut epithelium functions as a barrier against toxic luminal contents which can repair efficiently after injury to prevent systemic illness and death. However, we have shown that repair is severely hindered in neonates as compared to juveniles in our translational pig model of intestinal ischemia. The subepithelial enteric glial cell (EGC) network is known to promote epithelial repair by paracrine signaling mechanisms. This EGC network develops postnatally; therefore, we believe this repair defect in neonates is due to an underdeveloped EGC network. In support of this, we hypothesize that EGC inhibition in juveniles will block epithelial barrier repair after ischemic injury mimicking the neonatal phenotype. Ischemia‐injured jejunum of juvenile pigs was recovered ex vivo with and without fluoroacetate (FA), a glial inhibitor. Transepithelial electrical resistance (TEER) was monitored as a measure of barrier function, and tissues were collected for imaging analysis and primary EGC culture. FA inhibited TEER recovery at all tested concentrations (P<0.0001). Histology showed 500μM FA optimally inhibits epithelial repair (P=0.0196) without directly damaging the epithelium (P=0.5509). Cellular metabolism marker c‐fos has been optimized for immunofluorescence in control tissues and ongoing imaging work will validate selective inhibition of EGC metabolism by 500μM FA. In addition, pig EGC isolation and culture has been optimized, so that future work will confirm inhibitory effects of FA on EGC signaling functions in vitro. Understanding the development of glial‐epithelial crosstalk in barrier repair will ultimately guide novel clinical solutions to improve outcomes in neonatal patients affected by intestinal injury.Support or Funding InformationUNC CGIBD Large Animal Models Core (P30 DK034987), UNC CGIBD Basic Science Research Training Fellowship (NIH T32 5T32DK007737), UNC CGIBD Pilot Feasibility Grant (P30 DK034987), USDA National Institute of Food and Agriculture (Projects 1007263 and 07985)
Dynamic Regulation of TET2 by Phosphorylation and Dephosphorylation at Ser99
Authors: Kundu Anirban , Shelar Sandeep , Ghosh Arindam P. , Ballestas Marry P. , Kirkman Richard P. , Nam Hye-Young , Brinkley Garrett J. , Karki Suman , Mobley James A. , Varambally Sooryanarayana , Sudarshan Sunil 
Published: April 2020

The ten‐eleven translocation enzyme was discovered when an 8‐year‐old boy diagnosed with acute myeloid leukemia (AML) containing the t(10;11)(q22;q23) in 2003. Three members, TET1, TET2 and TET3, belong to this dioxygenase family of enzymes. TET2 is widely implicated in cancer and in aging due to its role as a global epigenetic modifier catalyzing oxidation of 5‐methylcytosine (5mC) to 5‐hydroxymethylcytosine (5hmC), 5‐formylcytosine (5fC) and then 5‐carboxylcytosine (5caC) in 2‐oxogluterate (2‐OG) dependent manner. TET2 mutations are prevalent in malignancies including acute AML. Notably, most TET2 mutations in AML are monoallelic. Given the presence of a remaining wild type allele in these malignancies, characterizing the mechanism that regulate TET2 protein could have therapeutic significance.Human TET2 is 2002 amino acids in length. The catalytic domain is located in the C‐terminal portion of the protein between amino acids 1129–1936. The C‐terminal catalytic domain comprises a cysteine rich domain (1129–1312 amino acids) followed by a double stranded β‐helix domain (DSBH) which make a compact fold to execute the catalytic activity. The C‐terminal part of TET2 has been reported capable of oxidizing 5‐mC to 5‐hmC independent of the N‐terminus. As such, the function of the N‐terminus of this large protein remains poorly characterized.Here, we identify that several isoforms of the 14‐3‐3 family of proteins bind TET2. 14‐3‐3s bind TET2 when phosphorylated at serine 99 (S99). AMPK‐mediated phosphorylation at S99 promotes TET2 stability and increases global DNA 5‐hydroxymethylcytosine. 14‐3‐3s’ interaction with TET2 serves to protect S99 phosphorylation. Disruption of this interaction leads to both reduced TET2 phosphorylation and decreased protein stability. Furthermore, we identify that the protein phosphatase 2A (PP2A) can interact with TET2 and dephosphorylates S99. siRNA knockdown of PP2A (B‐alpha subunit) or pharmacologic inhibition of PP2A was able to significantly increase TET2 phosphorylation and protein level as well. Collectively, our study provides novel insights into the role of the N‐terminal domain in TET2 regulation. Moreover, they demonstrate the dynamic nature of TET2 protein regulation that could have therapeutic implications for disease states resulting from reduced TET2 levels and/or activity.Support or Funding InformationThe research reported in this article was supported by R01CA200653, I01BX002930, and a Mike Slive Foundation Grant (to S.S.) and in part by the UAB Comprehensive Cancer Center (P30CA013148).