bluuebunny/crossref_metadata_2025_split huggingface dataset:
New Strategies for Probing and Inhibiting CDK4/cyclin D Hyperactivity in Melanoma
Authors: Prével Camille , Pellerano Morgan , Bihel Frederic , Meunier Laurent , Morris May 
Published: June 2021

Uncontrolled proliferation of melanocytes results in melanoma, one of the most aggressive forms of skin cancer, which further metastasizes into a deadly form of cancer. The p16‐cyclin D‐CDK4/6‐retinoblastoma protein pathway is dysregulated in 90% of melanomas and activation of this pathway potently cooperates with mutant BRAF or NRAS in transformation of melanocytes. CDK4/cyclin D kinase hyperactivation, associated with mutation of CDK4, amplification of cyclin D or loss of p16INK4a leads to increased risk of developing melanoma. This kinase therefore constitutes a key biomarker and an emerging pharmacological target.The limited efficacy of current therapeutics for treatment of melanoma and the lack of diagnostic assays to detect the early onset of this disease call for new generations of targeted therapeutics and novel approaches to probe melanoma‐specific biomarkers. To this aim, we engineered a fluorescent biosensor which reports on CDK4 kinase hyperactivity in vitro and in living cells, which we employed to develop a diagnostic assay using human skin biopsies. We further designed biosensors for drug discovery purposes, which discriminate compounds that target the interface between CDK4 and cyclin D, or that interfere with conformational dynamics of CDK4 activation loop, which were successfully applied to screen for novel and potent classes of CDK4 inhibitorsOur work highlights the potency of fluorescent biosensors for developing diagnostic strategies to monitor cancer‐associated alterations in CDK4 activity as well as drug discovery programmes.
The Effects of Lying, Sitting and Standing on Energy Expenditure in Young, Adult Women
Authors: Popp Collin , Bridges William , Najm Nadia , Traylor Daniel , Jesch Elliot 
Published: June 2021

BackgroundSedentary behavior and the increased risk for metabolic disease are well established. Standing at work or at home to decrease time sedentary can increase daily energy expenditure, however, little research has focused on the energy expenditure differences between various sedentary positions (i.e., lying, sitting).ObjectiveThe purpose of this study was to investigate the energy expenditure of lying, sitting and standing.MethodsTwenty‐eight young, healthy females reported to the metabolic laboratory where each completed all positions (lying, sitting and standing) in random order. Inspired and expired gases were collected for 45‐minutes (15 minutes for each position), and all three positions were measured during one continuous test. Oxygen consumption (VO2) and carbon dioxide production (VCO2) were measured to estimate resting energy expenditure (REE) and respiratory quotient (RQ). Statistical analyses used repeat measures ANOVA to analyze all variables and post hoc t‐tests.ResultsThere were positional differences for most variables. Standing VO2 (L/min) was 9.5% and 7.7% greater than lying VO2 (L/min) and sitting VO2 (L/min), respectively. Standing RQ was lower than lying RQ, but not different from sitting RQ. Standing REE (kcal/min) was 9.0 % greater than lying REE (kcal/min), and 7.1% greater than sitting REE (kcal/min), however, lying REE and sitting REE were not different from each other. Relative standing REE (kcal/min/kg) was 8.8% greater than lying REE (kcal/min/kg) and 6.4% greater than sitting REE (kcal/min/kg).ConclusionOur results indicate that standing has small, but significant effect on energy expenditure when compared to lying and sitting.
Validation of Photography for Determining Food Intakes of High School Students
Authors: Barrett Sheila , Kyle Ashley , Skaradzinski Veronica , Miu Leong Calvin 
Published: June 2021

PurposeTo determine if photography is a reliable data collection tool for measuring the food intakes of high school students who participate in the National School Lunch Program (NSLP). Secondly, to compare the energy and macronutrient content of the NSLP meals with meals eaten when not at school.MethodResearchers visited one high school in Illinois and measured the amounts of food consumed by the students using photographs of before and after food trays and weighed food wastes for comparisons. Three researchers not involved with the weighing of the food wastes, estimated the percentage wastes from the photographs. Estimated percentage wastes were converted into gram amounts by using the actual gram weight of the food items served. Actual wastes were weighed and compared with the estimated gram wastes. Students completed a demographic questionnaire and a three day food record which was used to determine the contribution of calories and macronutrients consumed when not at school. Twenty‐two students from grades 9‐12 participated in this pilot study, mean age was 16.41±1.30.Results High inter‐rater reliability was found for estimated and actual gram amounts of food consumed (r= 0.957, p<0.001). Paired samples t‐test of calories and macronutrients indicated significant differences (p<0.05) for meals eaten when not at school with those provided by the NSLP lunch meal. Overall, the NSLP meals provided fewer calories and macronutrients.ConclusionPhotography can be used to determine the food consumption of the NSLP participants in future studies. Contrary to earlier studies, the NSLP meals provided fewer calories and macronutrients than did the meals eaten when not at school. Future studies using a larger sample size are needed.
Weight Loss with Liquid Formula Diets in Obese Patients with and without Diabetes
Authors: Leslie Wilma , Taylor Roy , Lean Michael 
Published: June 2021

AimTo provide a data synthesis, using systematic methods, of published results comparing weight losses following Very Low Calorie (<800kcal/day VLCD) or Low Energy Liquid‐formula (>800kcal/day LELD) diets in patients with type 2 diabetes and non‐diabetic patients.MethodsMedline (1946‐2013) search to identify published studies of VLDC/LELDs. All studies in T2D patients were included. Limits for studies in non‐diabetic patients<em style=”mso‐bidi‐font‐style: normal;”> were: VLCD/LELD <em style=”mso‐bidi‐font‐style: normal;”>>8weeks duration, sample size >50 and within last 7 years. Mean weekly rate of weight loss during VLCD/LELD were weighted for study number.ResultsWeighted rates of weight loss in the 16 studies included were as follows: 6 in T2DM patients alone, (n=130, mean BMI 33.6‐40kg/m2) ‐1.4kg/week; non‐diabetic obese (n=1031, mean BMI 30‐48 kg/m2) ‐1.3kg/week, and 4 studies that included both patient groups (n=116, mean BMI 36.6‐42.6 kg/m2) ‐1.4kg/week (diabetic) and ‐1.4kg/week (non‐diabetic).ConclusionsThese data show that weight loss with LELD is very similar for obese patients with and without T2DM and support the use of LELD to achieve new weight loss/ maintenance targets of >15‐20% for those with severe and medically complicated obesity.
Influence of High Fat Diet on Macrophage Behavior and Metabolic Processes in Obesity Sensitive and Resistant Mice: Role of Monocyte Chemoattractant Protein 1
Authors: Cranford Taryn , Enos Reilly , Velazquez Kandy , McClellan Jamie , Nagarkatti Mitzi , Murphy E 
Published: June 2021

PURPOSEAdipose tissue macrophages are thought to be a major player in the dysregulation of metabolic and inflammatory processes associated with high fat diet (HFD)‐induced obesity. Monocyte chemoattractant protein 1 (MCP‐1) is a chemokine that recruits macrophages to adipose tissue (AT) and has been implicated in manifestations associated with obesity. The purpose of this study was to examine the role of MCP‐1 in response to HFD feedings. We used both C57Bl/6 (obesity sensitive) and FVB/N (obesity resistant) mice in order to determine obesity versus HFD effects. A HFD that was designed by our laboratory and is similar to a standard American diet was used given its clinical relevance.METHODSC57Bl/6 and FVB/N wild‐type (WT) and MCP1‐/‐ mice at 4 wks of age were fed either a HFD or a low fat diet (LFD) for 16 wks. Body composition was analyzed monthly and HOMA index was determined at mice sacrifice at 20 wks of age. AT mRNA expression of chemokines, macrophage markers and protein expression of inflammatory markers were measured using qPCR and western blot, respectively. Macrophage infiltration into AT were examined using immunohistochemistry and flow cytometry.RESULTSAs expected, HFD induced obesity and increased insulin resistance (IR) comparably in the obesity sensitive C57Bl/6 WT and MCP1‐/‐ mice. Surprisingly, the HFD MCP1‐/‐ in the supposed diet resistant FVB/N strain exhibited increased obesity and IR whereas the HFD WT mice were not different from LFD mice. AT macrophage accumulation increased in HFD WT groups for both strains, regardless of obesity status. Interestingly, the response in HFD MCP1‐/‐ was dependent on the strain.
A randomized control trial of sugar‐sweetened and artificially sweetened beverages and intrahepatic fat in overweight subjects
Authors: Campos V , Despland C , Schneiter Ph , Brandejsky V , Kreis R , Boesch Ch , Tappy L 
Published: June 2021

We conducted a 12‐week trial involving 28 overweight male and female adults (BMI >25) consuming 2 or more servings/day sugar‐sweetened beverages (SSB). Participants were randomized to a 12‐week intervention replacing SSB with artificially sweetened beverages (ASB, 8M, 7F) or to a control arm without intervention (C, 6M, 7F). Intrahepatocellular lipid concentration (IHCL, 1H‐magnetic resonance spectroscopy), visceral adipose tissue volume (VAT, magnetic resonance imaging) and fasting concentrations of selected metabolic parameters were measured before and after intervention.Consumption of SSB was similar in both groups at inclusion. IHCL was reduced to 74 ±10.7% of initial values in the ASB (p<0.01) but did not change in C. ASB however did not significantly change body weight (‐1.4±1.6 vs +0.8±0.6 kg in C), VAT (‐98.4 ±78.4 vs ‐11.8 ±73.4 cc in C) or fasting glucose, insulin, uric acid, ASAT, ALAT, TG, cholesterol or HDL‐cholesterol concentrations.15 participants (8 in the ASB group, 7 in the C group) had hepatic steatosis as defined by IHCL > 60 mmol/L. They had significantly higher body weight, VAT, TG, uric acid, ASAT, ALAT, and lower HDL‐cholesterol than 13 participants without hepatic steatosis. In this subgroup, ASB was associated with significant (p<0.05) decreases in IHCL (‐43.0±12.6%), ALAT (‐19.7±7.8%) and ASAT (‐9.9±4.0 %).These results indicate that, in high SSB consumers 1) body weight and visceral fat volume are major determinants of hepatic steatosis, 2) replacing SSB with ASB reduces intrahepatic fat, most likely due to decreased total energy intake.This work was supported by the Swiss National Science Foundation and the Raymond Berger Foundation.
RGS2 Protein Degradation is Mediated by a Novel Cullin 4B/F‐box 44 E3 Ligase Complex
Authors: Sjogren Benita , Storaska Andrew , Swaney Steven , Neubig Richard 
Published: June 2021

Hypertension and heart failure are major health issues and there is a need to identify more effective treatments. Regulator of G protein Signaling 2 (RGS2) is highly expressed in both heart and vascular smooth muscle and regulates Gαq signaling. RGS2‐/‐ mice are hypertensive, show enhanced responses to vasoconstrictors and lower tolerability to pressure overload. Over‐expression of RGS2 reduces cardiac hypertrophy and we have shown that pharmacologically enhanced RGS2 protein expression has functional effects on G protein signaling. Therefore, we hypothesize that enhancing RGS2 protein expression is a novel route in treating cardiovascular disease.RGS2 is rapidly degraded through the proteasome, but the specific enzymes involved are not known. Thus, the goal of the current study was to identify the molecular machinery responsible for RGS2 protein degradation. A siRNA screen to identify genes that regulate RGS2 protein levels identified components of a putative cullin‐RING E3 ligase (CRL). siRNA knock‐down of F‐box 44 (FBXO44) or cullin 4B (CUL4B), but not CUL4A or CUL1, led to a significant increase in RGS2 protein levels and stability. Overexpression of FBXO44 and CUL4B decreased RGS2 protein levels. FBXO44 associates with both RGS2, CUL4B and the adaptor protein DDB‐1 in cells resulting in a novel E3 ligase protein complex able to degrade RGS2. Studies are now underway to characterize the molecular mechanisms of the FBXO44‐RGS2 interaction.RGS proteins are difficult drug targets due to their mode of action being through protein‐protein interactions. Inhibiting a more druggable protein within the degradation pathway could be a way to increase RGS2 protein levels and function.
Towards the Identification of Small Molecule Inhibitors of the GIV‐Gi Interaction as Potential Anti‐metastatic Drugs
Authors: DiGiacomo Vincent , Nguyen Lien , Blanco‐Canosa Juan , Garcia‐Marcos Mikel 
Published: June 2021

Trimeric G proteins are classically activated by G protein‐coupled receptors (GPCRs). Dysregulation of this mechanism contributes to myriad pathologies from cancer to psychiatric disorders. Indeed, GPCRs are the target for >30% of marketed drugs. G proteins can also be activated by non‐receptor proteins, however, representing a novel signaling mechanism for which no pharmacological modulators exist. Among these newly identified regulators, the GIV protein is a particularly attractive pharmacological target owing to its tight association with cancer metastasis. Moreover, GIV‐Gi coupling underlies the molecular basis of metastasis. The available structural and biochemical information indicates the interface is a tractable target for small molecules. Here, we report the development of assays compatible with high throughput screening (HTS) to monitor the direct interaction between GIV and the Gαi3 G protein. We mapped the minimal Gαi3‐binding sequence in GIV and used it to develop a fluorescence polarization‐based assay that performed robustly in a 384‐well format (Z’ ~0.7). We also developed an orthogonal counterscreen (AlphaScreen) compatible with HTS (384‐well, Z’ ~0.8). A pilot screen of the LOPAC1280 library identified 2 validated hits. Based on literature searches and in silico docking, we provide further evidence that these 2 molecules can directly target the GIV‐Gαi interface. Taken together, these results indicate that the GIV‐Gαi interface is a “druggable” protein‐protein interaction and set the framework for identifying potent anti‐metastatic agents.Support: NIH R01GM112631
Identification of MicroRNAs Involved in the Regulation of Human UGT1A, UGT2B7 and UGT2B15 Gene Expression
Authors: Papageorgiou Ioannis , Court Michael 
Published: June 2021

BackgroundThe human UGT enzymes are responsible for the glucuronidation and elimination of a large number of clinically important drugs. Large interindividual variations of human hepatic drug glucuronidation have been observed in vitro, which cannot be entirely explained by the mechanisms studied so far. MicroRNAs (miRs) are small, non‐coding RNAs that regulate gene expression in a sequence‐specific manner. At present little is known about their role in the regulation of human UGTs. Here we sought to identify miRs that bind to the 3′UTR sequences of UGT1A (identical in all functional UGT1A isoforms), UGT2B7, and UGT2B15, representing the majority of UGTs involved in human drug glucuronidation.MethodsHEK293 cells were co‐transfected with a plasmid carrying the Firefly luciferase gene under the control of the respective UGT 3′UTR (wild‐type allele), a plasmid with Renilla luciferase (normalizer) and a miR. Using this approach we screened a commercial library containing all known human miRs.ResultsThe analysis identified multiple miRs (6 each for UGT1A and 2B7; 16 for 2B15) that down‐regulated reporter activity. Deletion of the predicted miR response element (MRE) abrogated this down‐regulation. Interestingly two miR‐21‐3p MREs in the UGT1A 3′UTR contained two different SNPs (one in each) that did not affect miR binding. Finally UGT1A and 2B15 shared common MREs for miR‐376b‐3p and miR‐103b.ConclusionsWe have identified 28 unique miRs that are likely involved in the regulation of expression of the majority of UGTs responsible for drug glucuronidation in humans. Further experiments will verify the effect of these miRs on UGT gene expression and enzyme activity in human cell lines and primary cultures.
Effect of Pentoxifylline on the hypertension in insulin resistance: Role of low‐grade inflammation
Authors: Azhar Ahmad , El‐Nassossy Hany 
Published: June 2021

ObjectivesThe present study investigated the protective effect of pentoxifylline (PTX) from hypertension in IR.MethodsMetS was induced in rats by high fructose (10% in drinking water) high fat diet for 12 weeks to develop hypertension. PTX was daily administered (30 mg.kg‐1) in the last 4 weeks of study before the blood pressure (BP) parameters were fully investigated in anaesthetized rats via a microtip catheter inserted in the aorta via an opening in the carotid artery. In addition, serum levels of glucose, fructosamine, insulin, tumor necrosis factor alpha (TNFα), adiponectin and lipid profile were determined.ResultsFeeding the rats with high fructose high fat diet led to development of IR as evidenced by the significant hyperinsulinemia and hyperglycemia while PTX administration did not affected the developed IR. IR animals were characterized by significant increase in the mean BP while PTX administration completely prevented the IR associated elevations in mean BP. PTX prevented the elevations in systolic, diastolic and notch BP in IR animals. However, PTX did not affect the elevations in pulse BP seen in IR animals. Neither IR nor PTX affected the heart rate or ejection or non‐ejection durations. In addition, PTX alleviated the low grade inflammation associated with IR as reflected by the significant decrease in serum TNFα and increase in serum adiponectin levels compared with IR animals.ConclusionsPTX directly alleviates hypertension in IR via amelioration of peripheral resistance and the low grade inflammation
Role of Krüppel Like Factor (Klf4) in Pulmonary Fibrosis
Authors: Perkins Tina , Wary Kishore 
Published: June 2021

RationaleThe mature endothelial cells (ECs) encounter with and must respond to several ‘intrinsic’ and ‘extrinsic’ endothelial disrupting factors and inflammatory cytokines. In this regard, KLF4 has been suggested to play an EC protective role; however, the underlying mechanisms are poorly understood.GoalTo address the role of Klf4 in adult lung microvessels by deleting EC‐Klf4 in a timed manner.Methods and ResultsImmunostaining showed significant reduction of KLF4 expression in human emphysema lung tissue samples. Cigarette smoke condensate decreased expression of KLF4 in ECs. Therefore, to address the role EC‐Klf4 in mice, male Klf4fl/+ were crossed with female Klf4fl/+ mice to generate Klf4fl/fl (25%)offspring. Next, Klf4fl/fl mice were crossed with male tg‐Cdh5CreERT2 mice to produce heterozygous Klf4fl::tg‐Cdh5CreERT2 offspring (100%). Thereafter, male Klf4fl::tg‐Cdh5CreERT2 were crossed with female Klf4fl::tg‐Cdh5CreERT2 to produce Klf4fl/fl::tg‐Cdh5CreERT2 siblings (25%). Groups of these mice received tamoxifen (TAM) everyday for 5 days, and on the 10th day (called Klf4ECKO mice) were sacrificed. Accordingly, TAM treatment decreased EC‐Klf4‐protein expression significantly in the Klf4ECKO lug compared to Klf4fl/fl ortg‐Cdh5CreERT2 controls. Klf4ECKO lung tissue showed increased recruitment of neutrophils and inflammatory cells. Trichrome staining revealed increased collagen deposition in the Klf4ECKO versus wild‐type mice. Importantly, the number of lung alveoli decreased (p<0.05 vs control, n=5), hemorrhage in Klf4ECKO mice compared to the control. Accordingly, the number of CD31+ vascular structures decreased in these mice.SummaryKlf4 protect ECs from ‘intrinsic’ and ‘extrinsic’ endothelial disrupting factors, therefore its loss results in pulmonary fibrosis.
ER‐α Receptors Have More Prominent Depressor Role On Vasoconstractile Sensitivity in Treated And Untreated‐Ovariectomized Rat Mesentery Artery
Authors: Akhayeva Tamila , Ozansoy Gulgun 
Published: June 2021

Natural estrogens have cardioprotective effects in premenopausal women. Nuclear estrogen receptors mediate genomic effects of estrogens. G‐protein coupled estrogen receptors are also defined cardiovascular system and mediate rapid non‐genomic effects. The role of estrogen receptor ER‐α and ER‐β on vasculature system is not completely understood. In this study, we investigated the effects of ER‐α and β on α1‐adrenergic receptor mediated vasoconstraction in control (C), 16‐week ovariectomized (O) and 17‐β estradiol (E2) treatment ovariectomized‐rat (OE) mesentery arteries. Phe‐mediated concentration response curves (CRC) were obtained from C, O and OE groups in the presence and absence of non‐selective ER agonist E2 (0.1μM), selective ER‐α agonist PPT (0.1μM) and selective ER‐β agonist DPN (0.1μM). Phe‐mediated CRC shifted to right presence of PPT in C (pD2:6.32 vs pD2: 5.80), O (pD2:6.40 vs pD2: 5.79) and OE (pD2: 6.52 vs pD2:6.08) groups. After E2 incubation, Phe‐mediated CRC only shifted to right in C group (pD2:6.40; pD2: 5.80), but insignificantly shifted to right in O (pD2:6.40 vs pD2:6.13) and OE (6.52 vs. 6.21) groups. On the other hand, DPN incubation did not change Phe‐mediated CRC in all groups.In conclusion, ER‐α receptor mediated responsiveness have more prominent depressor effects on vasocontractile sensitivity in all group.
Identification of a Prostate Cancer Cell Proteinase Activated Receptor/MMP Signaling Axis
Authors: HOLLENBERG Morley , Mihara Koichiro , Liu Yu , Ramachandran Rithwik 
Published: June 2021

HypothesisProteinase Activated Receptors (PARs) are G‐Protein‐Coupled Receptors activated by the proteolytic unmasking of a cryptic tethered ligand. We proposed that prostate cancer cells produce PAR‐regulating enzymes.MethodsWe monitored PC3, DU145 and LNCaP cell supernatant cleavage of PARs in intact cells by fluorescence imaging using N‐ and C‐ terminus dual‐fluorophore‐tagged PARs. Loss of the N‐terminal tag denotes receptor cleavage‐activation. We also used N‐terminal nano‐luciferase‐tagged PARs that, via luciferase release, document PAR‐cleaving proteinase activity in cell‐derived supernatants. MAPKinase and transwell migration assays were used to monitor PAR activation in PC3 cells.ResultsActivation of PARs 1 & 2 triggered MAPKinase signaling and cancer cell migration. Distinct PAR‐cleaving proteinases in conditioned media from all three prostate cancer cell lines were observed with the N‐Luc‐Luciferase assay, with differential inhibition by matrix metalloproteinase inhibitors and soya trypsin inhibitor. Further, when expressed in PC3 cells, the dual‐fluorophore‐tagged PAR1 was seen in a constitutively cleaved‐active state, with an absent N‐terminus.ConclusionOur data show that prostate cancer‐derived cells secrete PAR‐regulating proteinases that, via an autocrine process, can regulate PAR function in the tumour microenvironment. Since PAR activation triggers prostate cell migration and MAPKinase, PAR antagonists and inhibitors of PAR‐activating proteinases represent therapeutic targets for treating prostate cancer.FundingProstate Cancer Canada Movember Discovery grant.