bluuebunny/crossref_metadata_2025_split huggingface dataset:
The activation of calcineurin pathway contributes to the cardiomyocyte hypertrophy induced by thyroid hormone
Authors: Carneiro‐Ramos Marcela Sorelli , Shibata Caroline Emy Rodrigues , Moriscot Anselmo Sigari , Barreto‐Chaves Maria Luiza Morais 
Published: June 2021

ObjectiveT3 exerts strong influence over cardiac trophism and contractility. However, the possible intracellular pathways modulated by T3 are poorly understood. In this manner, several studies have shown that Calcineurin pathway is closely related to hypertrophic process in heart in different experimental models. Thus, the present was undertaken to evaluate Calcineurin pathway contribution to the cardiac hypertrophy triggered by T3 in vivo and in vitro.MethodsHyperthyroidism was induced in rats by i.p. T3 administration (7ug/100g BW) for 12hs, 24hs and 7 days. At the end of treatment, animals were decapitated, hearts collected for gene expression by Real Time PCR or calcineurin protein levels by Western‐Blotting. To evaluate a direct effect of T3, cardiomyocytes primary cultures treated for 24hs with T3 (10nM) or T3 conjugated with a calcineurin inhibitor (Ciclosporin A 500nM) were used. ANF mRNA levels were used as cardiac hypertrophy marker. The results were expressed as mean±SD and P0.05 (n=3).ResultsAs expected, 7d T3 treatment was able to induce significant cardiac mass gain and increase calcineurin levels. Data from in vitro experiments showed that concomitant administration of T3 and Cyclosporin A prevented partially cardiomyocytes hypertrophy when compared to that treated with T3 alone. (1,6 ± 0,21 vs. 2,16 ± 0,35, p<0,05). These results suggest an important role for calcineurin in T3 mediated cardiac hypertrophy.Financial Support: FAPESP.
AT1 receptor knockdown prevents the T3‐induced cardiomyocyte hypertrophy: modulation of the Akt Signaling Pathway
Authors: Diniz Gabriela Placoná , Carneiro‐Ramos Marcela Sorelli , Barreto‐Chaves Maria Luiza Morais 
Published: June 2021

ObjectiveThe Renin Angiotensin System plays an important role in TH‐induced cardiac hypertrophy. We recently demonstrated the in vivo participation of Angiotensin II Type 1 receptor (AT1R) in TH‐induced cardiac hypertrophy. The aim of this study was to analyze the contribution of AT1R to the development of T3‐induced cardiomyocyte hypertrophy, as well as to evaluate the role that AT1R plays in the activation of the Akt/GSK‐3β/mTOR signaling pathway promoted by T3.MethodsCardiomyocytes cultures were prepared from ventricles of neonatal Wistar rats by colagenase/pancreatin method. The cells were transfected with a selective AT1R small interfering RNA (AT1R siRNA). The AT1R knockdown was evaluated by Real Time PCR and Western Blot. Protein synthesis was quantified by tritiated leucine incorporation. The Akt/GSK‐3β/mTOR signaling pathway was analyzed by Western Blot. The results were expressed as mean±SD and P0.05 (n=3).ResultsThe cells transfected with the AT1R siRNA presented reduced mRNA and protein expression of AT1R (P<0.01). AT1R knockdown prevented the T3‐induced cardiomyocyte hypertrophy, as evidenced by the reduction in protein synthesis (P<0.01). In addition, the AT1R siRNA diminished the T3‐induced activation of the Akt/GSK‐3β/mTOR (P<0.01). These data demonstrate for the first time that the AT1R contributes to the trophic effect that T3 has on cardiomyocytes, as well as in the nongenomic T3 effect on the Akt/GSK‐3β/mTOR signaling pathway in cardiomyocytes.Financial Support: FAPESP.
Aging increases resting oxygen consumption in type‐II skeletal muscle
Authors: Villarin Jason J , Conley Kevin E , Kushmerick Martin J , Marcinek David J 
Published: June 2021

Aging affects skeletal muscle metabolic function. Other studies have shown greater generation of reactive oxygen species and the resulting oxidative damage in Type‐II muscle fibers. We tested whether the greater oxidative stress results in a greater effect of age on metabolism in type II muscle. We compared oxidative metabolism in soleus muscle (SOL, primarily type I fibers) and extensor digitorum longus muscle (EDL, primarily type II fibers) of the C57BL/6 mouse in two age groups, 7‐9 months old (adult) and 27‐29 months old (aged). Saponin‐permeabilized muscles were measured with high‐resolution respirometry under ADP‐limiting (state 4) and activated (state 3) conditions. We found no difference in resting oxygen consumption (state 4) between SOL and EDL in adult muscle when normalized to wet weight. Aged muscle showed an elevation in resting oxygen consumption in EDL (62%, p<0.05), but not in soleus. Stimulated state‐3 respiration was higher in SOL, however there was no effect of age. These results suggest there is increased proton leak in the mitochondria during state‐4 metabolism in the predominantly type‐II EDL muscle with aging.Supported by NIH F32‐AG029052, RO1‐AG028455.
Short‐term aerobic exercise training decreases Glucagon‐like Peptide‐1 in obese individuals
Authors: Baynard Tracy , Carhart Robert L , Kanaley Jill A 
Published: June 2021

Glucagon‐like peptide 1 (GLP‐1) is a relatively new incretin hormone secreted by the gut and is associated with appetite regulation and satiety. It is actively under study as a therapy for obesity and diabetes, yet the effects of exercise on this hormone are unknown.ObjectiveTo determine if a short‐term training period would influence GLP‐1 in obese individuals.MethodsSedentary, obese individuals (n=17; 51 yrs; body mass index 33.8 kg/m2) underwent 10 continuous d of exercise training. Training consisted of treadmill walking for 1 h/d at 75% of peak aerobic capacity. GLP‐1, glucose tolerance (2 hr meal test; 2 cans of Ensure(tm)) and aerobic capacity were assessed pre and post‐training.ResultsBody weight did not change with training, but peak aerobic capacity improved (24.1 ± 1.7 vs. 26.0 ± 1.8 mL/kg/min; p<0.05). Fasting insulin concentrations decreased (41.6 ± 5.3 vs. 31.0 ± 2.9 pmol/L; p<0.05), while glucose tolerance was unchanged (glucose AUC). GLP‐1 decreased by 27% following training (3.01 ± 0.32 vs. 2.20 ± 0.22 pmol/L; p<0.05). The change in fasting insulin was correlated to the change in GLP‐1 (r = 0.97; p<0.001).ConclusionsAerobic exercise training appears to elicit early changes in the gut hormone, GLP‐1, in obese persons. This may be an important mechanism by which exercise improves glucose tolerance and insulin sensitivity independent of changes in skeletal muscle. Research support: NIH R21 DK063179 (JAK); Sigma Delta Epsilon Fellowship; American College of Sports Science Mid‐Atlantic Chapter.
Gonadectomy enhances cocaine‐induced locomotor activity in male rats
Authors: Menendez‐Delmestre Raissa , Segarra Annabell 
Published: June 2021

Sex differences in cocaine addiction have been demonstrated in clinical as well as animal studies. However, studies on the role of testosterone in mediating these sex differences are inconclusive. This study investigated the effect of gonadectomy on cocaine‐induced hyperactivity. Adult male rats (300‐320g) were gonadectomized (GDX) or left gonadally intact (INT). Six days later, animals were placed in automated activity cages and their basal locomotor activity (LMA) was recorded. The next day, rats received a saline or cocaine (10, 15 or 30 mg/kg) injection and LMA was recorded.Cocaine increased LMA in both groups. However, this increase was higher in GDX than in INT rats, particularly at doses of 15 and 30 mg/kg. Also, while INT rats show a progressive increase in LMA to the same dose of cocaine (sensitization) at all doses tested, only a dose of 10 mg/kg was effective in inducing behavioral sensitization to cocaine in GDX males. These results show that in the male rat, gonadal hormones participate in mediating neuroadaptations to repeated cocaine administration.Support contributed by: NIH grants: U54NS39405 (SNRP); SO6‐GM08224 (MBRS/SCORE); and R25GM61838 (MBRS/RISE)]
Hepatocyte growth factor (HGF) suppresses hepcidin expression in hepatocytes by interfering with BMP‐mediated hepcidin induction
Authors: Goodnough Julia , Nemeth Elizabeta , Ganz Tomas 
Published: June 2021

Hepcidin, a 25‐amino‐acid peptide synthesized in the liver, is the iron regulatory hormone. Suppression of hepcidin in the setting of anemia appears to depend on a soluble factor expressed in the bone marrow during erythropoiesis. Here we describe that hepatocyte growth factor (HGF) suppresses hepcidin mRNA in a dose‐response fashion, in the concentration range detected in human serum. A similar response is observed in primary cells or cell lines transfected with a hepcidin promoter‐luciferase reporter. HGF is a ligand for the receptor tyrosine kinase (RTK), Met. In HepG2 cells transfected with the hepcidin‐luciferase reporter, the luminescence is specifically suppressed by HGF and not other ligands of RTKs, PDGF and EGF. BMPs 2, 4, 6 and 9 are strong inducers of hepcidin, and the BMP2/4 pathway also mediates hepcidin induction by iron‐transferrin. We report that HGF prevents a BMP2‐mediated increase in hepcidin mRNA and the activity of the luciferase reporter. HGF also prevented the induction by BMP2 of an unrelated BMP‐sensitive luciferase reporter. In primary hepatocytes, the BMP‐sensitive transcript ID1 is also downregulated by HGF. Thus, HGF may be a novel suppressor of hepcidin, modulating hepcidin expression by interfering with BMP signaling. We speculate that HGF could mediate the suppression of hepcidin in chronic liver diseases.This research is supported by National Institutes of Health grant #1F30DK082151.
Interferon‐gamma (IFN γ) induced epithelial barrier dysfunction in T84 human intestinal epithelial cells (IECs) occurs via phosphatidylinositol 3‐kinase (PI3‐K) mediated activation of adenosine monophosphate‐activated protein kinase (AMPK)
Authors: Scharl Michael , Barrett Kim E. , McCole Declan F. 
Published: June 2021

IFNγ plays a pivotal role in the pathophysiology of inflammatory bowel disease (IBD) and some of the effects of IFNγ on intestinal epithelial barrier function are dependent on PI3‐K. Here we investigated the role of the cellular energy sensor, AMPK in the effects of IFNγ on epithelial barrier function.Protein analysis was performed by Western blotting and AMPK knock‐down by siRNA. Transepithelial resistance (TER) was assessed by voltohmeter and epithelial permeability as the flux rate of 10 kDa FITC‐dextran.IFNγ (1000 U/ml) induced activation of AMPK in a manner independent of changes in intracellular ATP (the classic trigger for AMPK activation). Inhibition of PI3‐K with LY294002 (20 µM) blocked IFNγ‐induced AMPK phosphorylation (n=9; p<0.05; 6h), and mitigated the IFNγ‐induced decrease in TER across T84 monolayers (n=9; p<0.05). Knock‐down of AMPK diminished the IFNγ‐induced increase in epithelial permeability (72 h; n=3; p<0.05) and prevented IFNγ‐induced downregulation of the tight junction proteins occludin and ZO‐1 (n=3; p<0.05).We conclude that IFNγ sequentially activates PI3‐K and AMPK to impair tight junction protein expression and thus barrier function in IECs. Our data indicate a novel role for AMPK as a mediator of cytokine signalling, independent of changes in cellular energy status, and thus may have implications for mucosal defense in the setting of inflammation.Supported by the CCFA and NIH.
Age‐dependent regulation of sodium‐potassium adenosinetriphosphatase and sodium‐hydrogen exchanger mRNAs in equine nonglandular mucosa
Authors: Andrews Frank Michael , Peretich Amanda Lynn , Abbott Lora L , Dhar Madhu S 
Published: June 2021

Gastric ulcers, common in young race horses, occur in the nonglandular (NG) region. Histopathology in tissues exposed to hydrochloric acid (HCl) and volatile fatty acids (VFAs) in vitro showed compromised sodium transport, likely regulated by cellular Na‐K‐ATP (NAKA) and/or Na‐H exchanger (NHE) enzymes. We determined relative gene expressions of NAKA and NHE in equine NG mucosa exposed to HCl and VFAs at three different pHs (1.5, 4, or 7.0). Five (≤5 yrs of age) and five (≥12 yrs of age) mixed breed and gender horses were used. NG mucosa near the Margo plicatus was collected and exposed in Ussing chambers to normal Ringer's solution (control), solution with a mixture of VFAs and solution with 40 mM of acetic acid (treatment). NAKA and NHE mRNA expressions were determined using real time PCR.We found increased expression of both target genes after exposure to VFAs, compared to controls. Interestingly, younger horses had significantly (P≤0.05) down‐regulated NAKA, while expression of NHE was increased. On the contrary, both NAKA and NHE were up‐regulated in older horses. Young horses may be more susceptible to gastric ulcers due to an inability to up‐regulate active sodium transport through NG mucosal cells. Novel treatments could be developed to enhance NG cell function and prevent gastric ulcers in horses. Funded by Centers of Excellence in Livestock Disease and Human Health, University of Tennessee.
Efflux of cAMP in the exocrine pancreas: Role of secretin and atrial natriuretic factor (ANF)
Authors: Lange Maria S , Rodriguez Myriam , Ventimiglia Maria , Vatta Marcelo , Davio Carlos , Bianciotti Liliana 
Published: June 2021

We previously reported that ANF dose‐dependently reduces secretin secretory response and intracellular signaling in the exocrine pancreas through NPR‐C receptors coupled to the PLC/PKC pathway. In this study we evaluated if cAMP efflux contributed to ANF response in isolated pancreatic acini. Results showed that ANF favored the efflux of cAMP formed by secretin. The export of cAMP was also observed with secretin alone but not with isoproterenol or amthamine. Evoked‐cAMP efflux was blocked by probenecid, suggesting that multidrug resistant proteins (MRPs) were involved. RT‐PCR analysis revealed that MRP‐4 but not MRP‐5 (transport cyclic nucleotides) was expressed in the pancreas. Time course studies in the presence of phosphodiesterase (PDE) activity showed that tissue cAMP increased peaking at 3 min onset secretin stimulation and declining thereafter. With ANF, secretin‐induced cAMP augmented reaching a peak at 1 min after which it rapidly decreased. Extracellular cAMP accumulation occurred earlier and persisted throughout the experiment whereas with secretin alone it was delayed. Similar findings were observed in studies with PDE inhibition, except that cAMP content was higher and its appearance delayed in tissues and the media. Present findings suggest that cAMP export is likely mediated by MRP‐4 and further it may contribute to ANF‐induced desensitization of secretin response in the exocrine pancreas.(CONICET PIP5929/ UBACYT B079)
Baicalein protects hippocampal neuronal HT22 cells from ER stress‐induced apoptosis by modulating unfolded protein responses‐associated proteins
Authors: Choi Ji Hyun , Kang Insug 
Published: June 2021

The endoplasmic reticulum (ER) stress has been suggested to be involved in various neurodegenerative diseases including Alzheimer's disease and Parkinson's disease. Here we examined the effect of baicaleine (BE), a flavonoid, on ER stress‐induced apoptosis in mouse hippocampal neuronal cell line HT22. BE inhibited apoptosis induced by the ER stress inducers thapsigargin (TG) and brefeldin A (BFA). We showed that BE blocked ER stress‐induced unfolded protein response (UPR), including expression of CHOP and spliced form of XBP‐1 and phosphorylation of eIF2α, JNK, p38MAPK and the cleavage of ATF‐6. Moreover, we showed that BE blocked the accumulation of ROS and maintained mitochondrial membrane potential in ER stress‐induced HT 22 cells. Similar to the effect of BE, N‐acetyl‐cysteine (NAC), an antioxidant inhibited TG‐ or BFA‐induced apoptosis and changes in UPR‐associated proteins. Taken together, these results suggest that BE protects HT22 neuronal cells against ER stress‐induced apoptosis by inhibition of ROS accumulation and regulation of ER stress associated proteins.This work was supported by grants from KOSEF of Korea Government.
Shotgun Proteomics Identifies Protein Biomarkers Specific for Acute Renal Transplant Rejection
Authors: Sigdel Tara , Kaushal Amit , Gritsenko Marina , Norbeck Angela , Xiao Wenzhong , Qian Wei‐Jun , Camp David G. , Sarwal Minnie M 
Published: June 2021

BackgroundProteomics can help in biomarker discovery for different diseases. Since, acute rejection (AR) remains the primary risk factor for the renal transplant outcome; development of non‐invasive biomarkers for AR diagnosis is important.Materials and Method: We analyzed 82 urine samples that included patients with biopsy proven AR and stable grafts (STA), nonspecific proteinuria (NS), and healthy controls (HC). The samples were fractionated and processed by LC‐MS. Protein‐level spectral counts were obtained by which were used to get quantitative information. ELISA assays on UMOD, pigment epithelium‐derived factor (PEDF) and CD44 were performed to verify the trueness of the discovery.ResultsA total of 1446 unique urinary proteins were identified including 22 AR specific proteins a number of which were involved in antigen presentation and complement system pathways. We performed ELISA assay on UMOD, PEDF and CD44 on an independent set of urine samples and demonstrated that the observation from LCMS experiment was true and these proteins can serve as potential biomarkers to discriminate AR urine from STA, NS and HC.ConclusionA number of potentially AR specific biomarkers were identified. The validation of these potential biomarkers will offer a clinically applicable, non‐invasive urinary diagnostic assay for AR.
Development of a novel fluorescent nitric oxide synthase assay
Authors: Stahler Shannon Lynn , Huang Xinyi , Suhadnolik Lorraine , Bray Jennifer , Wilhelm James , Aulabaugh Ann , Harrison Richard 
Published: June 2021

Many therapeutic indications exist for nitric oxide synthase (NOS) making it a target of interest for drug discovery. The NOS family of enzymes, which include inducible NOS, endothelial NOS, and neuronal NOS, regulate functions such as cell communication, immune defense, and vasodilation. NOS enzymes convert L‐arginine and NADPH to L‐citrulline and nitric oxide (NO). NO is the agent responsible for the downstream physiological effects of NOS. In the absence of any downstream targets, NO is rapidly converted to nitrite in in vitro assays.Several NOS assays exist, but lack the sensitivity required for HTS‐compatible formats. This poster describes a new NOS assay using a fluorescent detection system. The detection system utilizes 4,5 Diaminofluorescein (DAF2) in which the probe reacts with nitrite under acidic conditions. Initial concentrations of various assay components were selected based on each component's individual effect on the detection system. The assay was further optimized based on the overall effect of all components on both the enzyme reaction and the detection system. Several known NOS inhibitors were evaluated under the final assay conditions resulting in a validated, robust HTS‐compatible assay.
Does immortalization of primary endothelial cells by overexpression of human telomere‐reverse transcriptase impair typical endothelial characteristics?
Authors: Baumer Yvonne , Schlosshauer Burkhard 
Published: June 2021

To study endothelial remodelling or regeneration in vitro mostly primary endothelial cells (EC) are used. However, primary EC are short living and often high priced. It would be desirable to use immortalized EC that are not transformed by oncogenes and display most EC‐characteristics of naïve EC. Therefore, we have used human telomere‐reverse transcriptase (hTERT)‐transduced primary human umbilical vein EC (HUVEC). Thus, we studied both HUVEC cell types. Comparative analysis included: 1) cell adhesion to 14 different extracellular matrix proteins analysed on a multisubstrate array (MSA), 2) protein‐expression of 9 endothelial and non‐endothelial markers by fluorescence‐microscopy (IF) and western blot‐analysis 3) uptake of Dil‐Ac‐LDL as revealed by IF and fluorescence activated cell sorting (FACS)‐analysis, 4) upregulation of ICAM‐1 after TNFa / interferon y‐treatment, 5) tube formation in hydrogel cultures, and 6) permeability properties. We conclude that immortalization of primary EC by hTERT retains the typical endothelial characteristics. Thus, hTERT‐EC represent a valuable tool for in vitro research.Supported by the German Ministry of Education and Research (BMBF) #01GU0727. Cells kindly provided by Dr. C. Fricke.
Metalloproteinases as key molecules to target invasiveness in solid tumors
Authors: Karamanos Nikos K , Staxtea Xanthi , Gialeli Chrisostomi 
Published: June 2021

Metastasis of cancer cells requires degradation of the extracellular matrix (ECM). Matrix metalloproteinases (MMPs) are the principal ECM‐degrading enzymes. Paracrine interactions between stroma fibroblasts and cancer cells are also involved in the expression and activity of MMPs. The aim of this study was to examine whether gene expression of MMPs is related with the invasive potential of colon cancer. In vitro studies on a panel of cell lines in the presence specific tyrosine kinase inhibitors and monoclonal antibodies were performed.The results showed that the invasive properties of cancer cells are associated with significant changes in gene expression of MT1‐MMP and MMP‐2. The expression of MT1 from HT‐29 colon cancer cells is increased by the conditioned media (CM) obtained by fibroblasts. MT1 contributes to the activation of proMMP‐2. STI571 suppressed the expression of MT1 in HT‐29 colon cancer cells as well as pro‐ and active MMP‐2. Panitumumab that target EGF‐R in colon cancer affects also the expression of MMPs.It is concluded that paracrine interactions is a determining factor for tumor cell behavior and suppression of certain MMPs may be of great value in molecular targeted therapy.
Modulation of PHLPP activity: screening for specific inhibitors
Authors: Sierecki Emma , Newton Alexandra C 
Published: June 2021

PHLPP (PH domain Leucin repeats Protein Phosphatase) has been shown to directly dephosphorylate Akt and PKC, leading to the desactivation of the Akt pathway and degradation of PKC. PHLPP also inhibits the Erk pathway and regulates the levels of tyrosine kinase receptors, in an indirect manner. In order to decipher the role of PHLPP in these different pathways, we need chemical tools to modulate PHLPP activity. However, the phosphatase domain of PHLPP belongs to the PP2C family of phosphatases that are characterized by their requirement of manganese/magnesium for their activity and by their resistance to commonly used phosphatase inhibitors such as okadaic acid or calyculin. To find specific inhibitors and activators, we screened the Diversity Set of the NCI for inhibitors of PHLPP 2. We designed a medium throughput colorimetric assay, using pNPP as a substrate. Compounds were tested at 100 µM and cutoff was set at 30% activity. We then evaluated the specificity of the compounds against the other member of the PHLPP family, PHLPP 1 and against another member of the PP2C family, PP2Cα. We determined the IC50 for the most promising compounds. Molecules with a IC50 of ≈1µM were also tested in cells and their effect on PHLPP was accessed by monitoring the phosphorylation state of Akt.This work was supported by the JDFR 3‐2008‐478 and NIH 067946.