bluuebunny/crossref_metadata_2025_split huggingface dataset:
Loss of CDK4/6 Activity Is Synthetic Lethal with VHL Inactivation in Clear Cell Renal Cell Carcinoma
Authors: Nicholson Hilary E , Housden Benjamin , Perrimon Norbert , Kaelin William G 
Published: June 2021

Clear cell renal cell carcinoma (ccRCC) is the leading form of kidney cancer, constituting >70% of all cases.1 Inactivation of the von Hippel‐Lindau (VHL) tumor suppressor gene and its protein product, pVHL, occurs in ~90% of ccRCCs.2 pVHL targets the HIF2α transcription factor for degradation. Without functional pVHL, HIF2α accumulates and induces inappropriate transcription of angiogenic, invasive, and growth‐promoting genes. Drugs that inhibit HIF2α or its downstream target VEGF are active against ccRCC, but not all patients respond to these agents and none are cured by them. I hypothesized that pVHL loss might create specific dependencies that could then be targeted therapeutically. To look for such synthetic lethal targets, I used RNAi in isogenic Drosophila cells that have or lack wild‐type pVHL and used CRISPR/Cas9 and chemical compound libraries in isogenic human ccRCC cell lines that have or lack wild‐type pVHL. Analysis of the overlap amongst the hits from these screens identified an increased requirement for CDK4/6 activity in pVHL‐defective ccRCC cells compared to their pVHL‐proficient counterparts. In secondary assays I confirmed that pharmacologic inhibition of CDK4/6 by either Abemaciclib or Palbociclib preferentially reduced viability of VHL−/− cells as compared to VHL+/+ cells across a variety of ccRCC cell lines. Importantly, inhibition of VHL−/− ccRCC cells by Palbociclib was abrogated by expressing a Palbociclib‐resistant CDK6 cDNA or by knockout of the canonical CDK4/6 target pRB. Sensitivity to Palbociclib could be reversed by expressing exogenous pVHL, but not a pVHL mutant lacking its known substrate docking site. HIF2α knockout in VHL−/− cells did not eliminate the effect of CDK4/6 inhibition, and pharmacological stabilization of HIF2α in VHL+/+ cells did not sensitize them to CDK4/6 inhibition. Therefore HIF2α is neither necessary nor sufficient for the synthetic lethal relationship between VHL and CDK4/6. Moreover, the combination of the HIF2α inhibitor PT2399 and the CDK4/6 inhibitor Palbociclib synergistically suppressed proliferation of VHL−/− ccRCC in vitro. I have now confirmed that both Palbociclib and Abemaciclib suppress VHL−/− ccRCC tumor growth in nude mice, including tumors that are PT2399‐resistant. I am currently asking whether these drugs can enhance the activity of PT2399 in vivo.Support or Funding InformationThis work was supported by the Dana‐Farber Cancer Institute and the National Cancer Institute (F32 CA220849‐02) (HN), and the Howard Hughes Medical Institute (WGK). We would like to thank Nicky Persky in the Johannessen Lab at Broad Institute for the entry clone containing CDK6‐D104S, as well as Rebecca Jennings in the Signoretti Lab at DFCI for immunohistochemistry.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Cardiomyocyte‐Specific EGFR Deletion Differentially Alters Cardiac Contractility and Remodeling
Authors: Guo Shuchi , Song Jianliang , Carter Rhonda , Lucchese Ana Maria , Gao Erhe , Cheung Joseph , Tilley Douglas 
Published: June 2021

Epidermal Growth Factor Receptor (EGFR), a member of the Erbb receptor tyrosine kinase family, is essential to the development of multiple tissues and organs. Due to the embryonic lethality of global EGFR deletion, the impact of EGFR signaling in the adult heart, normally or in response to pathological stimuli, has not been well‐explored. Using mice with floxed EGFR alleles crossed with αMHC‐Cre mice, we have generated a cardiomyocyte‐specific constitutive EGFR knockout mouse model (CM‐EGFR‐KO) to address its role in the heart. Compared to their wild‐type (WT) littermate controls, CM‐EGFR‐KO mice displayed age‐related development of cardiac contractile dysfunction, marked by impaired diastolic relaxation and Ca2+ handling at the individual myocyte level, as monitored via echocardiographic, hemodynamic, myocyte contractility and Ca2+ transient analyses. RNASeq analysis of CM‐EGFR‐KO versus WT littermate left ventricular samples revealed temporally‐dependent alterations in the expression of a number of myofilament, Ca2+ handling and fetal genes. The contractile defect initially occurs between 7 and 9 weeks of age and is maintained over time without overt cardiac hypertrophic or fibrotic remodeling until 10 months of age, by which time the mice ultimately succumb to severe heart failure and reduced lifespan. Chronic administration of the β‐adrenergic receptor (βAR) agonist isoproterenol effectively compensated for the contractile dysfunction in CM‐EGFR‐KO mice, an effect that was reversible upon cessation of infusion. Notably, βAR‐mediated cardiomyocyte hypertrophy was attenuated in CM‐EGFR‐KO mice compared to WT littermate controls. Treatment of 16 week‐old CM‐EGFR‐KO mice with AAV9‐cTnT‐EGFR permanently rescued cardiomyocyte‐specific EGFR expression and contractile function, suggesting that the contractile dysfunction phenotype observed in these mice is not secondary to aberrant cardiac development, but is specific to EGFR‐dependent regulation of contractile signaling.Support or Funding InformationNIH grant HL105414 to DGTThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
The CYP4A/20‐HETE Axis Regulates Ischemia‐induced Neovascularization via Its Actions on Endothelial Progenitor and Preexisting Endothelial Cells
Authors: Tang Samantha , Chen Li , Zhang Frank F , Gotlinger Katherine H , Falck John R , Schwartzman Michal L , Arbab Ali S , Guo Austin M 
Published: June 2021

20‐hydroxyeicosatetraenoic acid (20‐HETE) was recently identified as a novel contributor of ischemia‐induced neovascularization based on the key observation that pharmacological interferences of the CYP4A/20‐HETE axis decrease ischemic neovascularization. The objective of the present study is to determine whether the underlying cellular mechanisms involve endothelial progenitor cells (EPC) and pre‐existing endothelial cells (EC).We found that ischemia leads to a time‐dependent increase in 20‐HETE that is endothelial in origin using LC/MS/MS analysis and immunofluorescent (IF) microscopy. This is accompanied by increases in SDF‐1a expressions, EPC mobilization from bone marrow, and subsequent homing to ischemic tissues. Pharmacological interferences of the CYP4A/20‐HETE axis with a 20‐HETE synthesis inhibitor Dibromo‐dodecenyl‐methylsulfimide(DDMS) or a 20‐HETE antagonist N‐(20‐hydroxyeicosa‐6(Z), 15(Z)‐dienoyl) glycine(6, 15‐20‐HEDGE) significantly attenuated these increases. Importantly, we also determined that 20‐HETE plays a novel role in maintaining EPC functions and increases in the expression of Oct4, Sox2, and Nanog expression that are indicative of increased progenitor cell stemness. Flow cytometric analysis revealed that pharmacological interferences of the CYP4A/20‐HETE axis decrease the EPC population in culture, whereas 20‐HETE increases the cultured EPC population. Furthermore, ischemia also markedly increased the proliferation, oxidative stress, and ICAM‐1 expression in preexisting EC in the hindlimb gracilis muscles. We found that these increases were markedly negated by DDMS and 6, 15‐20‐HEDGE.Taken together, the CYP4A/20‐HETE axis regulates ischemia‐induced compensatory neovascularization via its combined actions on promoting EPC and local preexisting EC responses that are associated with increased neovascularization.Support or Funding InformationAmerican Heart Association [Grant 11SDG6870004]; National Institutes of Health [Grants HL34300], [Grant DK38226]; Robert A. Welch Foundation [GL625910]This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
High Salt Up‐Regulates Ca2+‐Sensing Receptor and Na+/Ca2+ Exchanger Expression in Kidneys of Dahl Salt‐Sensitive Rats
Authors: Odutola Samuel O , Awumey Emmanuel M 
Published: June 2021

The G protein‐coupled Ca2+‐sensing receptor (CaSR) provides the primary mechanism for Ca2+e‐mediated regulation of the release of parathyroid hormone (PTH) as a result of parathyroid cells sensing small changes in the [Ca2+]e. However, the CaSR is also involved in the fine regulation of serum Ca2+ levels and renal Ca2+ excretion, independent of PTH secretion. The mechanisms underlying these latter effects are not clear. We studied the effect of high salt on urinary ion excretion, blood pressure and expression of the CaSR and NCX1in kidneys of Dahl SR and SS rats. Our results show that four weeks of feeding a high salt diet increased BP in SS, increased urinary Na+/Ca2+ excretion in SR and SS but reduced K+ excretion. The high salt diet caused minimal structural changes in the kidneys, as determined by haematoxylin/eosin (H&E) and periodic acid‐Schiff (PAS) staining. Immunohistochemical analysis of tissue sections showed localization of the CaSR and NCX1 in the thick ascending limb of the loop of Henle, collecting ducts, cortical distal tubules, which was up‐regulated by the high salt in SS animals. NCX1 expression and localization in this study tracks closely with the CaSR. Thus, expression levels and localization patterns of these protein suggest that CaSR modulation of ion transporter and channel proteins could account for the difference in ion excretion observed between SS and SR rats as well as their responses to the high salt diet.Support or Funding InformationNIH Grant SC1 HL136278This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Apocynin Alters Redox Signaling in Blood Vessels of Spontaneously Hypertensive Rats
Authors: Graton Murilo Eduardo , Potje Simone Regina , Troiano Jéssica Antonini , do Vale Gabriel Tavares , Benevides Priscila Scarpim , Gonçalves Emily Thalia , de Melo Stevanato Nakamune Ana Cláudia , Tirapelli Carlos Renato , Bendhack Lusiane Maria , Antoniali Cristina 
Published: June 2021

Apocynin reduces blood pressure and prevents development of endothelial dysfunction in SHR. Mechanisms underlying the effects of apocynin in SHR remain unclear. Angiotensin (Ang) II regulates NOX, the major source of ROS in cardiovascular system. We hypothesized that apocynin, acting as NOX inhibitor, may alter pressor and vasoconstrictor effects of Ang II in SHR. We analyzed the effect of Ang II on blood pressure, reactivity on aorta and mesenteric resistance arteries, plasmatic antioxidant capacity, NOX and NOS expression and activity, AT1 and AT2 receptors expression in blood vessels of SHR treated with apocynin SHR treated with apocynin (30 mg/Kg/day, p.o.). In SHR, apocynin reduced pressor effect of Ang II, increased plasmatic antioxidant capacity, ablated aortic and mesenteric NOX‐dependent ROS production, NOX2 and p47phox overexpression demonstrating that apocynin acts as a NOX inhibitor in blood vessels of SHR. Moreover, apocynin increased aortic and plasmatic nitrate/nitrite levels, but did not alter mesenteric NOS activity or eNOS expression, diminished AT1 expression in aortas and mesenteric bed, and increased AT2 expression in aortas from SHR. Apocynin increased endothelial modulation on Ang II vasoconstriction in resistance arteries from SHR. All these results demonstrated that in vivo treatment with apocynin induces important alterations of several mechanisms that lead to the reduction of the pressor and vasoconstrictor effects of Ang II in SHR. Apocynin effect involves further mechanisms besides the modulation of vascular ROS, which improve NO availability in vascular cells of SHR. These integrated data could help us to understand the promising effect of apocynin as an antihypertensive drug different from those currently used clinically.Support or Funding InformationApproved by local ethics committee CEUA FOA 450/2015.Financial support: CNPq, CAPES and FAPESP (2016/22180‐9).This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Inhibition of Marfan‐associated Aortic Root Dilation by Angiotensin II Receptor Blockers May Be Independent of Blood Pressure Lowering
Authors: Tehrani Arash Y , White Zoe , Milad Nadia , van Breemen Casey , Esfandiarei Mitra , Bernatchez Pascal 
Published: June 2021

BackgroundMarfan syndrome (MFS) is a connective tissue disorder resulting in accelerated vascular aging, aortic root widening and aneurysm if unmanaged. Despite encouraging pre‐clinical data, recent clinical trials have shown underwhelming efficacy of anti‐hypertensive angiotensin II receptor type 1 (ATR1) blocker (ARB) losartan in MFS patients. We have recently reported that losartan mediates its anti‐aortic root remodeling effects in blood pressure (BP)‐ and ATR1‐independent fashions, and instead acts by improving endothelial function and protective endothelial nitric oxide (NO) synthase‐mediated NO release. Considering the unique mechanism of action of losartan in MFS, whether other ARBs can provide greater protection against MFS aortic root widening is unknown.ObjectiveHere we compare the efficacy of other ARBs previously linked to endothelial function activity with losartan at blocking aortic root widening in MFS mice.Methods and ResultsAortic root dilation, aortic vessel remodeling, BP, and aortic contractility were compared between MFS and wild‐type (WT) mice after an 18‐week treatment with losartan, telmisartan, valsartan, or vehicle. Losartan and telmisartan doses were titrated to give similar hemodynamic effects in vivo, resulting in a 20–30% decrease in BP in both groups, while valsartan was chosen at a non‐BP lowering dose. Untreated MFS mice showed increased aortic root widening compared to WT controls (1.84 ± 0.05mm vs 1.52 ± 0.03mm; p<0.05) as assessed by high frequency ultrasound imaging. Despite differences in BP lowering, all ARBs significantly reduced aortic root widening in the MFS group by 10–16% (p<0.05). Histological assessment revealed a 26–37% (p<0.05) reduction in MFS aortic root medial thickening, and a 50% reduction in elastic fiber fragmentation (p<0.0001) in all treatment groups. Using ex vivo small chamber myography, we showed that all ARBs reduced phenylephrine‐induced aortic contractility by 60–80% (p<0.05) in both MFS and WT aortas in a NO‐dependent fashion.ConclusionThis is the first animal study to compare the effects of ARBs other than losartan in MFS. Despite differential BP effects, telmisartan and valsartan were effective at blocking MFS aortic root widening and activating endothelial function. This suggests BP lowering to be of minor therapeutic value in MFS management and thus aortic root protection could be achieved with lesser side effects for MFS patients. Future clinical trials should focus on other ARBs than losartan for the management of MFS.Support or Funding InformationSupported by CIHR, HSFC, BCKDF, Marfan Foundation, SPH Foundation and Rare Disease Foundation.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Bidirectional Effects of EZH2 Inhibitor in Endometriosis
Authors: Brunty Sarah , Ray‐Wright Kristeena , Santanam Nalini 
Published: June 2021

Endometriosis is a complex disorder that afflicts many women with chronic pelvic pain and infertility. It is believed that polycomb group of proteins, such as enhancer of zeste homolog 2 (EZH2), which plays a major role in cancer, also might contribute to the etiology of endometriosis. We and others have shown upregulation of EZH2 in human endometriotic tissues. However, the factors responsible for this upregulation is unknown. This study hypothesizes that the components of peritoneal fluid (PF) plays a dynamic role in epigenetic pathways in endometriosis. To test this hypothesis we measured EZH2 expression and activity (activation of H3K27me3) in endometrial cells. Ishikawa (human endometrial) cells were treated with 1% PF from IRB‐approved and consented women with and without endometriosis (n=6–8/group age and cycle matched). A subset of cells was also treated with various concentrations of GSK126 (5 nM to 10 μM) either before or after PF treatment. Real‐time PCR was used to analyze EZH2 expression in these treated cells. Our results showed bidirectional effects of GSK126 on EZH2 and H3k27me3 expression, depending on the order of the drug treatment (before or after 24 hours of PF treatment) and the source of the PF (control vs endo). When cells were treated with endo PF, there was a 2–3 fold induction in EZH2 expression when cells were treated with endo PF compared to the media control. Surprisingly, there was a dose dependent increase in EZH2 mRNA expression with GSK126 pre‐treatment followed by the PF treatments. Almost a 9‐fold increase in EZH2 expression was observed when cells were pre‐treated with 2.5 μM GSK126 followed by control PF (*p<0.05), whereas pretreatment of GSK followed by endo PF had no such effects on EZH2 expression. However, when cells were pre‐treated with PF followed by the drug the EZH2 expression levels went down. Western blot of EZH2 and H3K27me3 showed similar findings. When MTT assay was performed to examine cell viability under the above experimental conditions, no significant changes in viability was observed at the 48 or 96 hour time points. These results suggest for an intermediate pathway modulated by the PF components that might be at play. The current studies are investigating the possible role for ARID1A mutation in the bidirectional effect of the EZH2 inhibitors in endometriosis.Support or Funding InformationPhRMA Foundation Pre Doctoral Fellowship in Pharmacology/ToxicologyThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.
Resting Cerebral Blood Flow Immediately Post Concussion is correlated to Cognitive Performance
Authors: Iring Stephanie , Favre Michelle , Brewer Kelly , De La Cruz Leslie , Liu Jie , Breen Paul P , Falvo Michael J. , Serrador Jorge J. 
Published: June 2021

Sports related concussion is estimated to affect more than 3.8 million Americans a year. Despite the large number of concussions, our understanding of the mechanisms underlying concussion are limited. We also lack an accurate diagnostic tool to provide an indication of concussion severity. Currently, most concussion assessments use cognitive testing to assess severity. Our lab has been examining use of Doppler ultrasound in the field as a possible novel objective measures of concussion severity. We recruited players from recreational rugby tournaments. Players who were seen at the medical tent and determined not to need further treatment were offered the opportunity to participate. A control group of players who had played a rugby match but did not receive a head hit were also recruited. During the tournaments, we used a portable tent based lab to obtain measures of internal carotid cerebral blood flow by imaging the artery to obtain diameter and velocity. Additionally, we measured both beat‐by‐beat blood pressure (Finometer) as well as end tidal CO2 using a nasal cannula during rest, both in the seated and supine position. In a subgroup of 47 players (28 controls, 19 concussed, Mean age= 30, range= 19–66 yrs) out of our large study (125 players, 49 concussed), we had players perform a Simple Reaction Time Test (SRTT), Trails A & B, Digit Symbol Substitution Test (DSST), Paced Auditory Serial Addition Test (PASAT) with 3 sec and 2 sec delay and a Digit Span Memory Test. Concussed players had significantly higher SCAT total number of symptoms (15 ± 11 vs 5 ± 7, P=0.029) as well as Symptom Severity Score (33 ± 18 vs 7 ± 9, P=0.002). Concussed players only performed significantly worse on the SRTT (353 ± 128 ms vs 278 ± 44 ms, P=0.046) while trending on DSST (54 ± 15 ms vs 63± 4 ms, P=0.085). Concussed players demonstrated significantly greater cerebral blood flow when seated (646 ± 180 mL/min vs 415 ± 85 mL/min, P=0.001). Cerebral blood flow was also significantly correlated to performance on SRTT (r=0.84, P=0.035), Trails B Time (r=0.94, P=0.006), DSST (r=−0.88, P=0.021) and Digit Span (r=−0.89, P=0.019). Surprisingly increased seated cerebral blood flow was found in the concussed players and associated with worse cognitive performance. This initial study suggests that cerebral blood flow immediately post‐concussion may be an objective indicator of severity. Future work is needed to examine how immediate measures of cerebral blood flow are predictive of cognitive recovery and post‐concussive symptoms.Support or Funding InformationThis work was supported by Dept. of Pharmacology, Physiology, and Neuroscience, Rutgers and the War Related Illness and Injury Study Center.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.