bluuebunny/crossref_metadata_2025_split huggingface dataset:
Structural and Functional Characterization of the Metastatic RhoGEF P‐Rex1 and its Regulation by PtdIns(3,4,5)P3: Towards Inhibitory Small Molecule Development
Authors: Cash Jennifer N. , Davis Ellen M. , Tesmer John J.G. 
Published: November 2023

Phosphatidylinositol 3,4,5‐trisphosphate (PIP3)‐dependent Rac exchanger 1 (P‐Rex1) is a Rho guanine‐nucleotide exchange factor (RhoGEF) that regulates cell motility and is strongly associated with cancer metastasis. P‐Rex1 is synergistically recruited to the cell membrane and activated by PIP3 and heterotrimeric G protein βγ (Gβγ) subunits, positioning the enzyme downstream of multiple classes of cell surface receptors that control processes such as cytoskeleton rearrangement and cell migration. P‐Rex1 has thus become an attractive therapeutic target for the suppression of cancer metastasis. However, development of inhibitors against P‐Rex1 is hindered by the fact that its regulatory mechanisms are poorly understood. My primary goal is to define the molecular basis for regulation of P‐Rex1 by PIP3 and Gβγ using X‐ray crystallography and biochemical and cell‐based assays, with the aim of identifying the surfaces of P‐Rex1 that are important for interaction with these molecules as well as their mechanisms of activation. Thus far, I have determined crystal structures of the tandem Dbl homology (DH)/pleckstrin homology (PH) domain catalytic core of P‐Rex1 in complexes with its substrate small GTPases Cdc42 and Rac1, as well as of the independent PH domain in complex with Ins(1,3,4,5)P4, a soluble analog of PIP3. Using site‐directed mutagenesis, we have shown that the PH domain is necessary and sufficient for PIP3‐dependent activation. Interestingly, mutation of residues within the PIP3 binding site does not abrogate membrane binding, suggesting that PIP3 activates P‐Rex1 through an allosteric mechanism as opposed to simple recruitment to the cell membrane. These studies have also opened the door to medium‐throughput screens anticipated to identify small molecule probes that compete with physiological regulators of P‐Rex1. These compounds could potentially be developed into molecules that block PIP3 binding, thereby inhibiting P‐Rex1 activation in cells and, ultimately, blocking cancer metastasis.Support or Funding InformationThis work was supported by an American Cancer Society – Michigan Cancer Research Fund Postdoctoral Fellowship (PF‐14‐224‐01‐DMC) to J.C. and an American Heart Association Undergraduate Student Research Program Award (14UFEL20510027) to E.D.
Comparative Analysis of Organophosphate Developmental Neurotoxicity in a Freshwater Planarian
Authors: Hagstrom Danielle , Zhang Siqi , Hirokawa Hideto , Taylor Palmer , Collins Eva‐Maria S. 
Published: November 2023

Organophosphates (OPs) are among the most commonly used pesticides in the U.S. and act by irreversibly inhibiting acetylcholinesterase (AChE), leading to cholinergic overstimulation due to increased synaptic levels of acetylcholine, paralysis and death. Their environmental abundance, although below acutely toxic levels to humans, has been suggested to be at least partially responsible for the observed increase in childhood neurodevelopmental disorders. However, it is unknown through which mechanisms chronic, low‐dose prenatal and infant OP exposure causes neurotoxicity in the developing brain. In this study, we analyzed the toxicological profiles of five common OPs (chlorpyrifos, dichlorvos, diazinon, malathion and parathion) using a multi‐endpoint planarian screening platform. The freshwater planarian, Dugesia japonica, is an excellent in vivo model for developmental neurotoxicology studies because its remarkable regenerative capabilities allows development to be induced at will by amputation. Moreover, because of their short development time (12 days) and similar size, regenerating and adult animals can be tested simultaneously using the same assays to identify development‐specific toxicity. Importantly, despite its simplicity, features of the planarian brain are conserved in the mammalian brain on the molecular and structural levels making mechanistic analysis of neurotoxicity in this animal directly relevant to human health. With this platform, we characterized effects on lethality, regeneration and a range of stimulated and unstimulated behaviors. We found that the different OPs varied widely in toxic dose, affected endpoints and developmental sensitivity. Lastly, through biochemical analysis of AChE activity in OP‐treated animals, we found that, depending on the OP, prolonged inhibition of AChE was not necessary for or necessarily indicative of toxic effects. This indicates that toxicity may be induced by alternative mechanisms, necessitating further in depth mechanistic studies.Support or Funding InformationDH supported by the NIH Cellular and Molecular Genetics Training Grant. EMSC supported by the Hellman foundation, the Burroughs Wellcome Fund CASI award and the Alfred P. Sloan Fellowship
Glutathione Transferase P1 (GSTP1) is Modified by Palmitate
Authors: Marensi Vanessa , Yap Megan , Ji Yuhuan , Lin Cheng , Berthiaume Luc , Leslie Elaine M 
Published: November 2023

Glutathione transferase P1 (GSTP1) protects cells from carcinogens by catalyzing their conjugation with the tripeptide glutathione (γ‐Glu‐Cys‐Gly). GSTP1 is also involved in cell signaling, proliferation and apoptosis. Overexpression of GSTP1 in tumors and single nucleotide polymorphic variants are associated with anti‐cancer drug resistance and poor prognosis. In contrast, inactivation of GSTP1 due to epigenetic promoter silencing increases susceptibility to certain cancer types, with prostate cancer as the best studied example. GSTP1 is classically described as a cytosolic enzyme; however, we have reported that it is strongly associated with the plasma membrane and the strength is comparable to the integral membrane protein Na+/K+‐ATPase. We hypothesize that the addition of a hydrophobic component is required to allow its strong interaction with membranes. Palmitoylation is the reversible post‐translational addition of a 16‐C saturated fatty acid to proteins, most commonly on Cys residues through a thioester bond. We found that GSTP1 is modified by palmitate. However, Cys‐less (Cys to Ser or Cys to Ala) mutants expressed in MCF7 cells surprisingly retained palmitoylation. In addition, treatment of palmitoylated GSTP1 with NaOH, which cleaves thioester bonds, did not remove palmitate. These data suggested that GSTP1 is modified by palmitate on at least one non‐Cys residue. We also demonstrated that GSTP1 can be non‐catalytically palmitoylated (or autoplamitoylated). Peptide sequencing by ESI‐MS/MS of the autopalmitoylated GSTP1 revealed that Cys48 and Cys102 undergo S‐palmitoylation and Lys103 undergoes N‐palmitoylation. N‐palmitoylation of an internal Lys residue is a rare observation and provides an explanation for the resistance of GSTP1 palmitoylation to NaOH treatment. In conclusion, we have identified palmitoylation as a novel post‐translational modification of GSTP1. This research lays the foundation for understanding the fundamental biology of GSTP1 and how palmitoylation may influence its structure, function and membrane association.Support or Funding InformationCanadian Institute of Health Research (CIHR), Alberta Innovates Health Solutions (AIHS) and Alberta Cancer Foundation (ACF)
Anti‐vascular ageing effects of endothelial SIRT1 ‐ communicating with smooth muscle cells of the arteries
Authors: Wang Yu , Xu Aimin , Vanhoutte Paul M 
Published: November 2023

Arterial remodeling is not only a physiological process during the formation of blood vessels, but also plays an important role in the development of cardiovascular diseases. Crosstalks between the endothelium and the surrounding vascular smooth muscle cells control the arterial remodeling process in response to changes in blood flow and nutrient supply. Endothelial disruption and dysfunction facilitate the adverse arterial remodeling and alter the mechanical responses of the blood vessel wall. Despite this information, little is known about the molecular pathways by which arterial remodeling is controlled and regulated. SIRT1 is a longevity regulator possessing potent anti‐cardiometabolic ageing properties. The present study identifies endothelial SIRT1 as a key modulator of arterial remodeling. SIRT1 facilitates the ubiquitination and proteasomal degradation of LKB1, a senescence‐inducing factor in endothelial cells. LKB1 binds to the promoter sequences of a gene encoding transforming growth factor beta (TGFbeta). By promoting the protein complex formation with HERC2, a giant scaffold protein possessing an E3 ligase activity, SIRT1 mediates the selective degradation of acetylated LKB1, thus preventing the trans‐activation of TGFbeta. In mice, overexpression of SIRT1 in endothelial cells abolishes the hypertrophic arterial remodeling caused by eNOS deficiency. In conclusion, SIRT1 prevents endothelial senescence‐induced pathological vascular remodeling and represents a promising anti‐vascular ageing drug target for the prevention of diseases such as restenosis, hypertension and atherosclerosis.Support or Funding InformationThis work was supported by the grants from Hong Kong Research Grant Council [General Research (HKU780613M and 17121714 ) and Collaborative Research Fund (C7055‐14G)].
AMELIORATIVE EFFECT OF METHANOLIC EXTRACT OF AZADIRACHTA INDICA LEAF ON LEAD ACETATE‐INDUCED CARDIOTOXOCITY, NEPHROTOXICITY AND GENOTOXICITY IN MALE WISTAR RATS
Authors: Adeyeoluwa Temitayo Esther , Adejumobi Abiola Olumuyiwa , Ola‐Davies Olufunke Eunice , Adedapo Adeolu Alex , Oyagbemi Ademola Adetokunbo , Omobowale Temidayo Olutayo 
Published: November 2023

The persistent existence of lead in the environment is associated with severe health issues due to its cumulative effect and its ability to affect virtually every organ in the body. Recent discoveries have attributed the tissue damaging and inflammatory effects of lead toxicity to oxidative stress. Natural antioxidants are now being employed in the alleviation of lead‐induced toxicity and oxidative stress as they have been observed to be effective with little or no side effects associated.This study is designed to investigate the ameliorative and antioxidative effects of Azadirachta indica on lead induced cardiotoxicity and nephrotoxicity. Cardiotoxicity and nephrotoxicity was established following oral administration of lead acetate (1mg/ml and 2mg/ml) for six weeks. However, rats were treated with 100mg/kg and 200mg/kg body weight of methanolic extract of Azadiracta indica (MEAI) after oral administration of lead acetate.Lead acetate administration led to a significant increase in heart rate, blood pressure and markers of oxidative stress in cardiac and renal tissues. Enzymic and non‐enzymic antioxidant defense systems were increased by lead acetate toxicity, there was however a persistent increase in glutathione‐S‐transferase (GST) activity. Renal nitric oxide (NO) content was significant reduced whereas cardiac NO content increased significantly. However, treatment of rats with 100mg/kg and 200mg/kg body weight of methanolic extract of Azadiracta indica (MEAI) after oral administration of lead acetate led to improvement in antioxidant defense systems, reduction in markers of oxidative stress, restoration of blood pressure and electrocardiographic (ECG) changes respectively. Lead acetate also precipitated genotoxicity which was significantly attenuated by MEAI. The pericardial inflammation and necrosis as well as the peritubular inflammation observed in the histology of the heart and kidney tissues. This architectural anarchy was ameliorated by MEAI.In conclusion, the results of this study show that Azadirachta indica ameliorated lead acetate‐induced cardiotoxicity and nephrotoxicity through free radical scavenging activity and antioxidant defense system. Furthermore, heart rate, systolic and diastolic pressure and other haemodynamic parameters were restored to near normal by MEAI. Hence, Azadirachta indica is a potential drug candidate against environmental pollutant such as lead acetate.
Increased Coronary Artery Dissection Rate and Severity in Murine Model of Marfan Syndrome
Authors: Milad Nadia , Sellers Stephanie L , Seidman Michael , Bernatchez Pascal N 
Published: November 2023

PurposeMarfan syndrome (MFS) is an autosomal dominant genetic disorder that leads to a variety of clinical presentations ‐ most seriously, aortic root dissection and aneurysm. Spontaneous coronary artery dissection, although rare in the general population, is associated with high rate of mortality and occurs more frequently in MFS patients due to their characteristic arterial wall weakness. MFS patients are often prescribed anti‐hypertensives such as Losartan, an angiotensin II type 1 receptor (ATR1) antagonist, but recent clinical trials have cast doubt on their effectiveness in preventing clinical endpoints. We investigated the rate and severity of coronary artery dissection in a murine model of MFS compared to wild‐type mice.MethodsExperimental mice were bred with littermate controls using the common Fbn1C1039G/+ model of MFS giving rise to wild‐type and MFS groups. At 6 or 22 months of age, hearts were fixed, sectioned and stained with hematoxylin and eosin, Movat's pentachrome and iron stains in order to visualize coronary artery dissection. Dissection was scored using a binary system (dissection or no dissection) and a severity‐weighted system (0 represents no dissection, 1 ‐ slight dissection, 2 ‐ clear dissection, 3 ‐ severe dissection).ResultsAt 6 months of age, we saw an unexpectedly high rate of mild coronary artery dissection in wild‐type mice compared to MFS alone (100% in wild‐type vs. 50% in MFS) but when weighted for severity there was no difference between groups (1.0 for wild‐type vs. 0.88 in MFS). However at 22 months of age, the MFS group displayed a higher rate of dissection compared to wild‐type (80% vs. 33%) as well as an increase in dissection severity (1.6 vs. 0.33, respectively).ConclusionsTo our knowledge, this is the first genetic mouse model of coronary artery dissection. The unexpectedly high rate of mild dissection in wild‐type mice at 6 months of age (all were scored 1 for slight dissection) may be due to low N values at this time‐point or may be part of normal vessel maturation, unfortunately little study on coronary artery dissection in mice has been conducted. However, the observed increase in both rate and severity in MFS mice at 22 months compared to wild‐type is likely due to vessel wall fragility and elastic fibre fragmentation characteristic of MFS which left coronary arteries susceptible to further damage. Future research into the effects of Losartan treatment on rate and severity of coronary artery dissection should be done.Support or Funding InformationThis research was funded by CIHR grant awarded to Dr. Pascal N Bernatchez.
Decreased Interleukin‐6 Signaling in Female Mice Early Post‐Myocardial Infarction Attenuates Neutrophil Infiltration and Limits Cardiac Dilation and Rupture
Authors: DeLeon‐Pennell Kristine , Iyer Rugmani Padmanabhan , Ma Yonggang , Yabluchanskiy Andriy , Lindsey Merry L 
Published: November 2023

Survival after myocardial infarction (MI) is improved in female compared to male mice, yet the mechanisms remain undefined. We hypothesized lower acute inflammation in female mice leads to improved survival and cardiac function post‐MI. We used our Mouse Heart Attack Research Tool (mHART) 1.0 database that has 888,000 data points. We queried the database and found >350 3–7 month C57BL/6J mice that qualified for this study. In addition to improved survival, rupture of the left ventricle (LV; rupture/total deaths) was also reduced in females (11%; 1/9) versus males (46%; 16/35; p<0.05). Out of 52 plasma analytes measured at day (D)0‐D7 post‐MI (n≥10/sex/day), interleukin (IL)‐6 positively correlated with end diastolic dimension (EDD) in males only (R2=0.47; p<0.05). Female IL‐6 levels were significantly lower at D1 and D3 compared to males (all p<0.05). LV dilation was decreased in females (EDD/LV mass index=1318 ± 67 mm/mg) versus males (1487 ± 48 mm/mg; p<0.05; n≥10/sex) at D7 post‐MI. Bioinformatic analysis of 168 genes at D1 post‐MI identified IL‐6ra, the IL‐6 receptor, as the top feature linked to sex differences (p<0.05; n≥8/sex). IL‐6ra was decreased 2‐fold in females versus males. Since IL‐6 regulates neutrophil infiltration, we assessed neutrophil numbers; there was a 2‐fold reduction in females compared to males at D1 and D3 post‐MI (both p<0.05; n=6/sex/day). In conclusion, utilizing mHART 1.0, females had decreased neutrophil infiltration, as well as LV dilation and rupture, possibly due to reduced IL‐6 signaling.Support or Funding InformationThis work was supported by AHA [14POST18770012 to RPI and 15SDG22930009 to YM], and by NIH [HHSN 268201000036C (N01‐HV‐00244) for the San Antonio Cardiovascular Proteomics Center, U54GM114833 for the UCLA Center of Excellence for Big Data, and R01HL075360 to MLL] and from the Biomedical Laboratory Research and Development Service of the Veterans Affairs Office of Research and Development Award [5I01BX000505 to MLL], and by HL051971 and GM104357.
Non‐Resolving Chemokine Response Dysregulates Cardiosplenic And Cardiorenal Network Following Myocardial Infarction In Aging
Authors: Halade Ganesh V , Kain Vasundhara , Ingle Kevin , Black Laurence M 
Published: November 2023

Chemokine signal drives the innate inflammatory response following myocardial infarction (MI), though how it provokes non‐resolving chronic inflammation in aging is unclear. To answer this, we hypothesized that resolution reprograms, specifically proresolving lipid mediators formed in innate immune response, may be dysregulated in aging post‐MI, influencing cardiorenal and cardiosplenic network. Young (6 months) and aging (18 months) mice were fed standard lab chow (LC) and safflower oil (SO)‐enriched diet for 2 months and were then subjected to coronary artery ligation in order to induce MI. Despite similar infarct areas and reduced fractional shortening post‐MI, splenic mass spectroscopy lipid profiling revealed higher levels arachidonic acid (AA) derived pro‐inflammatory metabolites, but minimal formation of eicosanoids, docosanoids D‐ and E‐ series resolvins in SO‐fed group compared with LC group. SO fed aging mice displayed higher levels of pro‐inflammatory thromboxanes and tetranor‐12‐HETEs metabolites (p<.0.5) compared to adult‐SO fed mice. Data suggests that aging decreases the expression levels of 12‐lipoxyegenase (LOX) and 15‐LOX and 5‐LOX (all p<0.05) in infarcted left ventricle compared to young LC fed mice, but increased in kidney and spleen post‐MI. Excess intake of SO decreased formyl peptide receptor 2 (FPR2) compared to young SO‐fed mice, indicating non‐resolving response Increased SO intake also caused higher NGAL expression, suggesting kidney injury with LC group. The decreased expression of Arg‐1 in response to MI suggested the delayed healing in aging mice compared to young mice. In summary, aging magnifies the post‐MI chemokine‐driven innate response and impairs the healing response by stimulating prolonged neutrophil trafficking and proinflammatory lipid mediators.Support or Funding InformationThe authors acknowledge support from NIH‐NCCAM K99/R00 AT006704 and UAB start‐up funds, awarded to GVH.
Pericytes Improve Motor Recovery After Spinal Cord Injury in Both Sexes
Authors: Mayo Jamie N. , Bearden Shawn E. 
Published: November 2023

ObjectiveNew capillary growth can improve recovery after spinal cord injury (SCI). Pericytes, microvascular cells, can be stimulated in vitro to induce new capillary growth. The hypothesis was tested that stimulated pericytes would improve motor recovery in both sexes in a pediatric model of SCI.MethodsHuman placental pericytes and pericytes stimulated with cobalt chloride were injected into the site of a thoracic hemi‐section of the spinal cord in rat pups on postnatal day three (P3). Hindlimb motor recovery (assessed using the Basso, Beattie, and Bresnahan motor score scale) and Doppler blood flow perfusion at the site of transection were measured on P10. Immunohistochemistry was used to quantify vessel and neurofilament density on P10.ResultsPericyte, but not stimulated pericyte, treatment improved motor recovery in both sexes (males, 2.722 ± 0.31 fold score improvement; females, 3.824 ± 0.58 fold score improvement, p < 0.05). Pericyte treatment supported a stable, flow regulated vascular bed in both sexes and greater neurofilament density (males, 1.9 ± 0.09 fold increase; females, 2.031 ± 0.1553 fold increase, p < 0.05). There were significant effects of treatment and interactions between treatment and sex in vessel density after SCI (p < 0.05). In the vehicle control group, females had a higher vessel density seven days after injury than males (p < 0.05). This pattern persisted following pericyte injection (p < 0.05), and the average vessel density was not different than their sex‐matched controls (p > 0.05). With stimulated pericyte injection, males had a greater vessel density then their sex‐matched controls (p < 0.05) and no longer had lower vessel density then females. Sex was a significant source of variation (p < 0.05) for endothelial proliferation. Within the vehicle control group there was less endothelial proliferation in female tissue than there was in male tissue (p < 0.05).ConclusionsPericytes and stimulated pericytes interact differently with the host environment after SCI. Injected directly after injury, naïve pericytes as opposed to stimulated pericytes have more potential to improve motor recovery. It appears that either the timing of or capacity for the initiation of an angiogenic program following CNS injury might differ between males and females, even before sexual maturity. Additionally, because female rats had greater vessel density than males but lesser endothelial proliferation it is possible that females may be better able to maintain vessel viability after initial injury. The results from this study highlight the importance of appreciating that even when males and females reach the same gross functional end measurements, they might do so by different structural and molecular mechanisms.Support or Funding InformationHL106548, P20 GM103408