bluuebunny/crossref_metadata_2025_split huggingface dataset:
Exacerbated Headache‐Related Pain in the Single Prolonged Stress Model of Post‐traumatic Stress Disorder
Authors: Standifer Kelly M. , Simpson‐Durand Cindy , Zhang Yong 
Published: November 2023

Post‐traumatic stress disorder (PTSD) frequently shares co‐morbidity with chronic pain and negatively impacts the course of treatment for both disorders. Single‐prolonged stress (SPS), an established animal model for PTSD, models many of the symptoms of PTSD reported in humans including exaggerated negative feedback of the HPA axis, hypocortisolism, enhanced fear and anxiety responses and cognitive impairment. We and others reported that SPS induces long‐lasting mechanical and thermal allodynia and visceral and inflammatory hypersensitivity. Chronic headache pain is one of the most commonly reported co‐morbid pain conditions by PTSD patients and one of the most resistant to effective treatment, yet this has never been modeled in an animal model of PTSD. We modified the chronic headache pain model recently developed by Pradhan and colleagues (2014) to investigate the contribution of SPS to sodium nitroprusside (SNP)‐induced hyperalgesia. SPS and control Sprague Dawley rats were randomly divided into SNP or vehicle‐treated groups. SNP (2 mg/kg, i.p.) was injected every second day from day 7 to 15 after initiation of SPS. Rat paw withdrawal threshold (PWT) to von Frey stimuli and tail flick latencies (TFL) dramatically decreased as early as 7 days after SPS and lasted until at least day 21. Basal PWT and TFL, as assessed prior to each SNP administration, also significantly decreased during the SNP treatment period. The lower pain thresholds recovered 7 days following the final SNP injection in SNP group, but not in SPS/SNP group. Nociceptin/orphanin FQ (N/OFQ) plays an important role in maintenance of hyperalgesia and allodynia following SPS. Elevated N/OFQ levels observed in cerebrospinal fluid (CSF) of SPS rats were even higher in CSF of the SPS/SNP group. Real‐time PCR revealed that glial fibrillary acidic protein (GFAP) and N/OFQ peptide (NOP) receptor mRNA expression increased in dorsal root ganglia (DRG) 21 days after SPS exposure; mRNA increases in the SPS/SNP group was more pronounced than SPS or SNP alone. PreproN/OFQ mRNA expression decreased in both SPS and SPS/SNP rats compared to control, but no difference was observed between the two groups. SNP treatment alone did not alter GFAP, NOP receptor or N/OFQ mRNA levels in DRG. Significant interaction between SNP and SPS was noted following immunoblot analysis of TNF alpha protein expression in DRG and NOP receptor expression in trigeminal ganglia. Our results indicate that chronic SNP‐induced nociceptive hypersensitivity (headache pain) is exaggerated in rats exposed to PTSD, and that N/OFQ may play an important role in the interaction between SPS and SNP.The animal protocol was approved by the OUHSC IACUC committee; studies conformed to the FASEB Statement of Principles for the use of animals in research and education.Support or Funding InformationUniversity of Oklahoma Health Sciences Center Vice President for Research and Presbyterian Health Foundation Bridge Grant Programs.
Spatial encoding of cyclic AMP signaling specificity by GPCR endocytosis
Authors: Tsvetanova Nikoleta , von Zastrow Mark 
Published: November 2023

G protein‐coupled receptors (GPCRs) comprise the largest family of signaling receptors and a very important class of therapeutic targets. GPCRs are well known to signal via cyclic AMP (cAMP) production at the plasma membrane, but it is now clear that various receptors also signal after internalization. This endosome‐derived signal is already known to modify the temporal profile of downstream responsiveness but it remains unknown, particularly for signals transduced by diffusible second messengers, if there is any functional significance to the spatial segregation of initiation sites. Using the beta2‐adrenoceptor (β2‐AR) as a model, we show that endocytosis is required for the full repertoire of downstream cAMP‐dependent transcriptional control. Next, we describe an orthogonal optogenetic approach to definitively establish that the location of cAMP production is indeed the critical variable determining the transcriptional response. Further, we demonstrate that the endocytosis‐dependent transcriptional signal is highly sensitive and saturable, effectively limiting the downstream cellular response to a uniform level. These properties allow the endosome to function as a ‘noise filter’, reducing variability of the downstream response and thus enhancing reliability of cognate ligand detection. In summary, our findings reveal a discrete principle for achieving cellular signaling specificity, based on endosome‐mediated spatial encoding of intracellular second messenger production and ‘location aware’ downstream transcriptional control.Support or Funding InformationNational Institute on Drug Abuse of the NIH; American Heart Association Postdoctoral Fellowship
Methamphetamine‐Induced Ubiquitination and Trafficking of Dopamine Transporters
Authors: Hinshaw Tyler , Sun Haiguo , Leussen Deborah , Chen Rong 
Published: November 2023

IntroductionThe dopamine transporter (DAT) is the primary terminator of synaptic dopamine transmission by reuptake of dopamine, and is a major target for psychostimulants including cocaine and methamphetamine. DAT undergoes constitutive and stimulated endocytosis and postendocytic trafficking. It has been shown that psychostimulants cause aberrant trafficking of DAT although the mechanisms remain elusive. Posttranslational modification of transporters by ubiquitin conjugation has recently emerged as a major regulatory mechanism of internalization and intracellular sorting of transporters and receptors. However, it is unknown whether psychostimulants alter ubiquitination of DAT. Moreover, recent evidence indicates that membrane rafts play an important role in DAT trafficking and activity. Thus, the purpose of this study is to investigate the consequence of neurotoxic treatment of methamphetamine on DAT ubiquitination and localization in lipid rafts.MethodsMale Sprague Dawley rats were treated with methamphetamine (7.5 mg/kg, IP, every 2 hrs for a total of 4 injections) or saline. This treatment regimen is known to induce hyperthermia which produces neurotoxicity. To prevent the effect of hyperthermia on DAT function, rats were cooled following each methamphetamine injection. Animals were sacrificed two or ten days after the treatment. Striatal tissues were immunoprecipitated with specific DAT antibody and immunoblotted with ubiquitin antibody. The localizations of striatal DAT in lipid rafts were examined using discontinuous sucrose gradient.ResultsOur data showed that neurotoxic doses of methamphetamine significantly enhanced DAT ubiquitination only two days after the treatment, suggesting a transient effect of methamphetamine on DAT posttranslational modification. The heightened DAT ubiquitination was not observed after 10 days, which indicate DAT ubiquitination is reversible. We also examined the changes of DAT localization in lipid rafts and non‐lipid rafts. Methamphetamine‐induced alterations in DAT ubiquitination and translocation in lipid rafts may be associated with impaired DAT activity.Support or Funding InformationThis project is supported by the ASPET Zannoni Summer Undergraduate Research Fellow (SURF) Individual Awards and NIH DA006634.
EGF Receptor Transactivation by α1‐Adrenoceptors on Prostatic Hyperplastic Cells
Authors: Silva Claudia Lucia M , Nascimento‐Viana Jessica B , Alcántara‐Hernández Rocío , García‐Sáinz J Adolfo , Noël François 
Published: November 2023

Benign prostatic hyperplasia (BPH) is a progressive urological disorder related to an imbalance between cell proliferation and apoptosis. BPH is characterized by an increased number of epithelial, smooth muscle and stromal cells. Prostatic enlargement contributes to the lower urinary tract symptoms (LUTS/BPH). The participation of epidermal growth factor (EGF) receptor in G protein‐coupled receptor signaling is a process of transactivation critical for mitogenic activity in some models. Here we focused on the putative role of EGF receptor transactivation mediated by α1D‐adrenoceptors in stromal cell growth. Human BPH cells were treated with phenylephrine (3 μM) or vehicle (control) for either 15 min or 48h, and cell growth and protein expression were examined by Trypan Blue exclusion/MTT and Western blotting assays, respectively. Cell treatment with 3 μM phenylephrine (48h) induced a cell growth effect comparable to 0.1 μM EGF (166.3 ± 13.7 and 157.4 ± 14%, n = 5 experiments performed in quadruplicate, for phenylephrine and EGF, respectively, P < 0.001). The effect of phenylephrine was blocked by the α1D‐adrenoceptor antagonists BMY7378 (50 nM; 92.8 ± 4.7%, n = 5, P < 0.001), LDT3 (50 nM; 94.2 ± 8.2%, n = 5, P < 0.001) and LDT5 (50 nM; 92.6 ± 9.1%, n = 5, P < 0.001) suggesting that these receptors are involved in cell growth. Then we evaluated the downstream α1D‐adrenoceptor signaling. Phenylephrine (3 μM, 15 min) induced p‐ERK expression and the α1D‐adrenoceptor antagonists also blocked this effect (n = 4). Furthermore, the phenylephrine‐induced cell growth was blocked by the metalloproteinase inhibitor GM6001 (10 μM), the selective EGF receptor inhibitor AG1478 (5 μM) and MEK inhibitor PD98059 (1 μM). In conclusion, these results suggest that EGF receptor transactivation by α1D‐adrenoceptor is relevant for human stromal cell proliferation and may contribute to prostatic enlargement and LUTS/BPH.Support or Funding InformationCNPq, Brazil [455436/2014‐2]
Development of a System for Realtime Assessment of β1‐Adrenergic Receptor Association with Epidermal Growth Factor Receptor
Authors: Guo Shuchi , Carter Rhonda , Tilley Douglas 
Published: November 2023

β‐adrenergic receptors (βAR) are G‐protein coupled receptors (GPCR) highly expressed in the heart and essential for coordinating responses to neurohormonal stimulation. βAR stimulation leads to the activation of a number of important proximal signaling pathways. Gs protein‐, β‐arrestin‐ and epidermal growth factor receptor (EGFR)‐dependent signaling pathways each contribute to acute and chronic regulation of cardiac function and remodeling. In heart failure, βAR‐mediated transactivation of EGFR plays a protective role via enhanced cardiomyocyte survival, thus is an attractive pathway to target for potential therapeutic development. We have previously shown that β1AR and EGFR associate in a complex, even at endogenous levels of expression in the heart, and that disruption of this interaction leads to alterations in downstream signaling that may prevent pro‐survival effects. Indirect evidence has suggested that this association is differentially regulated by G protein‐dependent and ‐independent mechanisms, and we hypothesize that GPCR kinases (GRKs) are the proximal regulatory signal for this receptor complex. To begin test the differential roles of G protein‐versus GRK‐dependent regulation of β1AR‐EGFR association, we have developed a series of FRET assays with which to quantify in realtime their impact on cAMP generation, β‐arrestin recruitment, β1AR‐EGFR affinity and EGFR activation. We have applied U2S cells as our initial testing model with adenoviral infection of FRET reporters for each readout, and using isoproterenol (ISO) as a full agonist standard for activation of each pathway, we have successfully established each assay. Using this platform we continue to test the relative impact of G protein‐dependent versus –independent signaling on β1AR‐EGFR association.Support or Funding InformationNIH HL105414
Glucagon‐Like Peptide‐1 (GLP‐1) Receptor Expression on Human Umbilical Vein Endothelial Cells (HUVEC) in Response to Cellular Activation
Authors: Noormohamed Nadia , Kiel Dan 
Published: November 2023

PurposeThe Glucagon‐Like Peptide‐1 (GLP‐1) receptor is the target for certain anti‐diabetic medications. The GLP‐1 receptor is a cell‐surface receptor that elicits a number of responses upon activation leading to reduced blood glucose and various other effects. A number of cell‐surface proteins have been shown to change expression level in response to various stimuli, such as nitric oxide and cytokines. Alterations in the level of cell‐surface receptors can increase or decrease responses through that receptor. The purpose of this study is to measure GLP‐1 receptor expression on human umbilical vein endothelial cells (HUVEC) in response to cellular activation by various compounds.MethodsNitric oxide and tumor necrosis factor alpha (TNF‐alpha) have been shown to induce changes in levels of cell‐surface receptors. Puerarin is an isoflavone found in a number of plants and herbs that stimulates endothelial nitric oxide synthase, leading to the generation of intracellular nitric oxide. Nitric oxide activates intracellular enzymes, such as guanylate cyclase, leading to the generation of second messengers. TNF‐alpha is a cytokine that increases transcription of various genes including those for cell surface adhesion molecules involved in the inflammatory process. HUVEC monolayers were treated with puerarin, TNF‐alpha, or vehicle. After 24 hours of treatment, cells were harvested. Equivalent amounts of cellular homogenates were separated via electrophoresis, and proteins transferred to a membrane. The membrane was probed with an antibody directed against the GLP‐1 receptor. Visualization of the band corresponding to the GLP‐1 receptor was achieved using chemiluminescence, and quantified using densitometry. Neither of the treatments produced large changes in GLP‐1 receptor expression. GLP‐1 receptor expression on HUVEC may be less responsive to cellular stimulation compared to other proteins on HUVEC.
LCT Gene Variant ‐13910 C>T, Dairy Intake, and 25‐Hydroxyvitamin D Plasma Levels: A Mendelian Randomization Study
Authors: Alharbi Ohood , El‐Sohemy Ahmed 
Published: November 2023

This abstract is from the Experimental Biology 2016 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.BackgroundThe LCT ‐13910 C>T gene variant is associated with lactose intolerance (LI) in different ethnic groups. Individuals with LI often limit or avoid dairy consumption, a main dietary source of vitamin D in North America, which may lead to inadequate vitamin D intake. However, the association between LCT genotype and circulating 25(OH)‐vitamin D levels is not clear. The objective of this study was to determine the prevalence of genotypes predictive of LI in different ethnic groups living in Canada and to determine whether LCT genotype is associated with dairy intake and 25(OH)D plasma levels.MethodsFasting blood samples were drawn from a total of 1,400 participants aged 20–29 years from the Toronto Nutrigenomics and Health Study for genotyping and plasma 25(OH)D analysis. Dairy intake was assessed using a one‐month, 196‐item Toronto‐modified Willett food frequency questionnaire. The prevalence of the LI gene variant LCT ‐13910C>T was compared using. Using the Mendelian randomization (MR) approach, we used analysis of covariance to determine the association between LCT genotypes and dairy intake, as well as 25(OH)D levels.ResultsApproximately 33% of Caucasians, 99% of East Asians, 74% of South Asians, and 59% of those with other or mixed ethnicities had the CC genotype predictive of LI. The CC genotype was significantly associated with lower total dairy intake (p=0.001), lower skim milk intake (p=0.002), and lower plasma 25(OH)D (p=0.0004). The CC and the CT genotypes were associated with a two‐fold, and 50% greater risk, respectively, of suboptimal plasma 25(OH)D.ConclusionThe CC genotype predictive of LI is associated with lower plasma 25(OH)D, which is due, at least in part, to lower intake of dairy products, particularly skim milk. Increased risk of suboptimal levels of vitamin D was also observed among those with the CT genotype suggesting an intermediate effect of heterozygotes.Support or Funding Information: Research support from the Advanced Foods and Materials Network
Association Between LCT ‐13910 C>T Genotype and Plasma 25‐Hydroxyvitamin D with Premenstrual Symptoms
Authors: Jarosz Alicia Caroline , Alharbi Ohood , El‐Sohemy Ahmed 
Published: November 2023

BackgroundVitamin D has been associated with premenstrual symptoms, but the findings have been inconclusive. Dairy is a good dietary source of vitamin D, but individuals with lactose intolerance consume less dairy and have lower plasma 25(OH)‐vitamin D levels. The LCT ‐ 13910 C>T gene variant is a strong predictor of lactose intolerance where those who have the CC genotype limit their intake of dairy. The objective of this study was to determine the association between 25(OH)‐vitamin D status and LCT genotype on premenstrual symptoms.Methods964 women aged 20–29 years participating in the Toronto Nutrigenomics and Health Study provided data on their premenstrual symptoms and fasting blood samples were taken for genotyping and plasma 25(OH) D analysis. Optimal, sufficient, insufficient, and deficient vitamin D status was defined as ≥75, 50–74.9, 30–49.9, and <30 nmol/L, respectively. Logistic regressions were used to calculate the odds ratio (OR) and 95% confidence interval (CI) for the associations between vitamin D status and premenstrual symptoms. A Mendelian Randomization approach was used in the analysis of covariance to determine the association between LCT genotype and premenstrual symptoms.ResultsThe odds ratio (95% CI) for experiencing clumsiness was 2.63 (1.32–5.21) for subjects with deficient plasma 25(OH)‐vitamin D status compared to those with optimal status. The corresponding odds ratios (95% CIs) for those experiencing confusion and with deficient or insufficient plasma 25(OH)‐vitamin D were 2.57 (1.42–4.67) and 1.82 (1.09–3.05), respectively, compared to those with optimal status. Plasma 25(OH)‐vitamin D status was not associated with experiencing any of the other premenstrual symptoms (acne, bloating, cramps, mood swings, increased appetite, fatigue, headache, anxiety, sexual desire insomnia, nausea, depression, or desire to be alone). The odds ratios (95% CIs) for experiencing clumsiness and cramps were 2.12 (1.08–4.17) and 0.55 (0.33–0.92), respectively, for individuals with the LCT CC genotype compared to those with the TT genotype. LCT genotype was not significantly associated with any of the other symptoms.ConclusionOur findings indicate that plasma 25‐Hydroxyvitamin D status and LCT genotype are associated with certain premenstrual symptoms, suggesting that dairy might be associated with both an increased and decreased risk of certain premenstrual symptoms.Support or Funding InformationResearch support: A.C.J. is a recipient of a Natural Sciences and Engineering Research Council Graduate Scholarship
Systems Pharmacology Approach to Identify Potential Therapeutic Small‐Molecules for Treatment of Diabetic Peripheral Neuropathy
Authors: McGregor Brett A. , Porter James E. , Feldman Eva L. , Hur Junguk 
Published: November 2023

Diabetic peripheral neuropathy (DPN) is one of the most common complications of diabetes. Despite extensive research the underlying mechanisms leading to this complication are not fully understood, and there are no specific treatments or means to predict DPN onset or progression. In this study, we employed a systems pharmacology approach to identify DPN‐related transcriptional networks and pathways conserved across various animal models and species, to identify drug‐candidates with therapeutic potentials.From our Diabetic Neuropathy Microarray Knowledge‐Base (DNMKB), we selected DPN‐related microarray data performed in sciatic nerve samples from type 1 diabetes (streptozotocin‐treated) and type 2 diabetes (db/db, ob/ob, high‐fat diet fed) murine models and human subjects with non‐progressive and progressive DPN. Differentially expressed genes (DEGs) were identified between non‐diabetic and diabetic samples in murine models, and between non‐progressive and progressive samples in human. Our in‐house literature mining program, SciMiner, was used to construct a transcriptional network for each DEG set, based on the sentence‐level gene‐gene co‐citation information in the complete PubMed abstracts. These networks were then compared using a network‐comparison program TALE (tool for approximate large graph matching) to identify overlapping relations between networks. These comparisons indicated that genes involved in the reelin signaling pathway, matrix metalloproteinases inhibition, and proliferation regulation to be differentially expressed and conserved between datasets.The genes that demonstrate the most dramatic fold change while overlapping across species and various DPN models were defined as the representative gene expression signature of DPN. This DPN signature was searched against the drug‐perturbed gene expression profile data from the Library of Integrated Network‐based Cellular Signatures (LINCS) database. LINCS, aiming to create a network‐based understanding of gene expression when cells are exposed to different perturbing agents, have extensively profiled the transcriptomic perturbations of over 20,000 small‐molecules in various cell lines and primary cells. Our expression profile‐based comparison analysis identified over 50 potential small‐molecule compounds with potential of reversing the direction of expression changes identified in the central pathways involved in our DPN networks. The top candidates included methyltransferase inhibitors and metabotropic glutamate inhibitors.A systems pharmacology approach, integrating of multiple bio/chem‐informatics analyses, identified DPN‐associated pathways that are highly conserved across various murine models and across species, as well as potential small‐molecules for DPN.Support or Funding InformationThe study was supported by the Diabetic Complications Consortium (DiaComp) Pilot & Feasiblity Grant (to J.H.), and the American Diabetes Association (to E.L.F).