scholarly journals Increased O-GlcNAcylation rapidly decreases GABAAR currents in hippocampus yet depresses neuronal output

2019 ◽  
Author(s):  
Luke T. Stewart ◽  
Kavitha Abiraman ◽  
John C. Chatham ◽  
Lori L. McMahon
Keyword(s):  
Post Translational Modification ◽  
Variable Effect ◽  
Excitatory Transmission ◽  
Hippocampal Ca3 ◽  
Health And Disease ◽  
Acute Increase ◽  
Neuronal Output

AbstractO-GlcNAcylation, a post-translational modification involving O-linkage of β-N-acetylglucosamine to Ser/Thr residues on target proteins, is increasingly recognized as a critical regulator of brain function in health and disease. Enzymes that catalyze O-GlcNAcylation are found at both presynaptic and postsynaptic sites, and O-GlcNAcylated proteins localize to synaptosomes. An acute increase in O-GlcNAcylation induces long-term depression (LTD) of excitatory transmission at hippocampal CA3-CA1 synapses, and depresses hyperexcitable circuits in vitro and in vivo. Yet, no study has investigated how O-GlcNAcylation modulates the efficacy of inhibitory neurotransmission. Here we show an acute increase in O-GlcNAc dampens GABAergic currents onto principal cells in rodent hippocampus likely through a postsynaptic mechanism, and has a variable effect on the excitation/inhibition balance. The overall effect of increased O-GlcNAc is reduced synaptically-driven spike probability via synaptic depression and decreased intrinsic excitability. Our results position O-GlcNAcylation as a novel regulator of the overall excitation/inhibition balance and neuronal output.

scholarly journals SETD3 protein is the actin-specific histidine N-methyltransferase

eLife ◽  
2018 ◽  
Vol 7 ◽  
Author(s):  
Sebastian Kwiatkowski ◽  
Agnieszka K Seliga ◽  
Didier Vertommen ◽  
Marianna Terreri ◽  
Takao Ishikawa ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Substrate Specificity ◽  
Post Translational Modification ◽  
Set Domain ◽  
Health And Disease ◽  
New Research ◽  
Glycolytic Phenotype

Protein histidine methylation is a rare post-translational modification of unknown biochemical importance. In vertebrates, only a few methylhistidine-containing proteins have been reported, including β-actin as an essential example. The evolutionary conserved methylation of β-actin H73 is catalyzed by an as yet unknown histidine N-methyltransferase. We report here that the protein SETD3 is the actin-specific histidine N-methyltransferase. In vitro, recombinant rat and human SETD3 methylated β-actin at H73. Knocking-out SETD3 in both human HAP1 cells and in Drosophila melanogaster resulted in the absence of methylation at β-actin H73 in vivo, whereas β-actin from wildtype cells or flies was > 90% methylated. As a consequence, we show that Setd3-deficient HAP1 cells have less cellular F-actin and an increased glycolytic phenotype. In conclusion, by identifying SETD3 as the actin-specific histidine N-methyltransferase, our work pioneers new research into the possible role of this modification in health and disease and questions the substrate specificity of SET-domain-containing enzymes.

The depressing (negative) effect of oestradiol benzoate on the in vitro secretion of LH and FSH by the pituitary gland of the LRH-pretreated long-term ovariectomized rat changes into the augmentative (positive) effect after discontinuation of the LRH-pretreatment

1985 ◽  
Vol 110 (3) ◽  
pp. 329-337 ◽  
Author(s):  
G. A. Schuiling ◽  
H. Moes ◽  
T. R. Koiter
Keyword(s):  
Depressing Effect ◽  
Ovx Rats ◽  
Gonadotrophin Secretion ◽  
Oestradiol Benzoate ◽  
Negative Effect ◽  
Positive Effect ◽  
Perifusion System

Abstract. The effect of pretreatment in vivo with oestradiol benzoate on in vitro secretion of LH and FSH was studied in long-term ovariectomized (OVX) rats both at the end of a 5-day continuous in vivo pretreatment with LRH and 4-days after cessation of such LRH pretreatment. Rats were on day 0 sc implanted with osmotic minipumps which released LRH at the rate of 250 ng/h. Control rats were implanted with a piece of silicone elastomer with the dimensions of a minipump. On days 2 and 4 the rats were injected with either 3 μg EB or with oil. On day 5 part of the rats were decapitated and the in vitro autonomous (i.e. non-LRH-stimulated) and 'supra-maximally' LRHstimulated release of LH and FSH was studied using a perifusion system. From other rats the minipumps were removed on day 5 and perifusion was performed on day 9. On the 5th day of the in vivo LRH pretreatment the pituitary LH/FSH stores were partially depleted; the pituitaries of the EB-treated rats more so than those of the oil-injected rats. EB alone had no significant effect on the content of the pituitary LH- and FSH stores. On day 9, i.e. 4 days after removal of the minipumps, the pituitary LH and FSH contents had increased in both the oil- and the EB injected rats, but had not yet recovered to control values. In rats not subjected to the 5-days pretreatment with LRH EB had a positive effect on the supra-maximally LRH-stimulated secretion of LH and FSH as well as on the non-stimulated secretion of LH. EB had no effect on the non-stimulated secretion of FSH. After 5 days of in vivo pretreatment with LRH only, the in vitro non-stimulated and supra-maximally LRH-stimulated secretion of both LH and FSH were strongly impaired, the effect correlating well with the LRH-induced depletion of the pituitary LH/FSH stores. In such LRH-pretreated rats EB had on day 5 a negative effect on the (already depressed) LRH-stimulated secretion of LH (not on that of FSH). EB had no effect on the non-stimulated LH/FSH secretion. It could be demonstrated that the negative effect of the combined LRH/EB pretreatment was mainly due to the depressing effect of this treatment on the pituitary LH and FSH stores: the effect of oestradiol on the pituitary LRH-responsiveness (release as related to pituitary gonadotrophin content) remained positive. In LRH-pretreated rats, however, this positive effect of EB was smaller than in rats not pretreated with LRH. Four days after removal of the minipumps there was again a positive effect of EB on the LRH-stimulated secretion of LH and FSH as well as on the non-stimulated secretion of LH. The positive effect of EB on the pituitary LRH-responsiveness was as strong as in rats which had not been exposed to exogenous LRH. The non-stimulated secretion of FSH was again not affected by EB. The results demonstrate that the effect of EB on the oestrogen-sensitive components of gonadotrophin secretion consists of two components: an effect on the pituitary LRH-responsiveness proper, and an effect on the pituitary LH/FSH stores. The magnitude of the effect of EB on the LRH-responsiveness is LRH dependent: it is very weak (almost zero) in LRH-pretreated rats, but strong in rats not exposed to LRH as well as in rats of which the LRH-pretreatment was stopped 4 days previously. Similarly, the effect of EB on the pituitary LH and FSH stores is LRH-dependent: in the absence of LRH, EB has no influence on the contents of these stores, but EB can potentiate the depleting effect of LRH on the LH/FSH-stores. Also this effect disappear after cessation of the LRH-pretreatment.

THE ROLE OF POLYETHYLENIMINE IN ENHANCING PERFORMANCE OF GLUTAMATE BIOSENSORS

2018 ◽  
Vol 8 (3) ◽  
pp. 36-41
Author(s):  
Diep Do Thi Hong ◽  
Duong Le Phuoc ◽  
Hoai Nguyen Thi ◽  
Serra Pier Andrea ◽  
Rocchitta Gaia
Keyword(s):  
First Generation ◽  
Good Reproducibility ◽  
Complex Matrices ◽  
Long Term Stability ◽  
In Vitro Characterization ◽  
Glutamate Oxidase

Background: The first biosensor was constructed more than fifty years ago. It was composed of the biorecognition element and transducer. The first-generation enzyme biosensors play important role in monitoring neurotransmitter and determine small quantities of substances in complex matrices of the samples Glutamate is important biochemicals involved in energetic metabolism and neurotransmission. Therefore, biosensors requires the development a new approach exhibiting high sensibility, good reproducibility and longterm stability. The first-generation enzyme biosensors play important role in monitoring neurotransmitter and determine small quantities of substances in complex matrices of the samples. The aims of this work: To find out which concentration of polyethylenimine (PEI) exhibiting the most high sensibility, good reproducibility and long-term stability. Methods: We designed and developed glutamate biosensor using different concentration of PEI ranging from 0% to 5% at Day 1 and Day 8. Results: After Glutamate biosensors in-vitro characterization, several PEI concentrations, ranging from 0.5% to 1% seem to be the best in terms of VMAX, the KM; while PEI content ranging from 0.5% to 1% resulted stable, PEI 1% displayed an excellent stability. Conclusions: In the result, PEI 1% perfomed high sensibility, good stability and blocking interference. Furthermore, we expect to develop and characterize an implantable biosensor capable of detecting glutamate, glucose in vivo. Key words: Glutamate biosensors, PEi (Polyethylenimine) enhances glutamate oxidase, glutamate oxidase biosensors

Strategies to Protect Hematopoietic Stem Cells from Culture-Induced Stress Conditions

2020 ◽  
Vol 15 ◽  
Author(s):  
Fatima Aerts-Kaya
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Ex Vivo ◽  
Stress Conditions ◽  
Stress Factors ◽  
Hematopoietic Stem ◽  
Induced Stress

: In contrast to their almost unlimited potential for expansion in vivo and despite years of dedicated research and optimization of expansion protocols, the expansion of Hematopoietic Stem Cells (HSCs) in vitro remains remarkably limited. Increased understanding of the mechanisms that are involved in maintenance, expansion and differentiation of HSCs will enable the development of better protocols for expansion of HSCs. This will allow procurement of HSCs with long-term engraftment potential and a better understanding of the effects of the external influences in and on the hematopoietic niche that may affect HSC function. During collection and culture of HSCs, the cells are exposed to suboptimal conditions that may induce different levels of stress and ultimately affect their self-renewal, differentiation and long-term engraftment potential. Some of these stress factors include normoxia, oxidative stress, extra-physiologic oxygen shock/stress (EPHOSS), endoplasmic reticulum (ER) stress, replicative stress, and stress related to DNA damage. Coping with these stress factors may help reduce the negative effects of cell culture on HSC potential, provide a better understanding of the true impact of certain treatments in the absence of confounding stress factors. This may facilitate the development of better ex vivo expansion protocols of HSCs with long-term engraftment potential without induction of stem cell exhaustion by cellular senescence or loss of cell viability. This review summarizes some of available strategies that may be used to protect HSCs from culture-induced stress conditions.

scholarly journals Plant-Derived Extracellular Vesicles: Current Findings,Challenges, and Future Applications

Membranes ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 411
Author(s):  
Nader Kameli ◽  
Anya Dragojlovic-Kerkache ◽  
Paul Savelkoul ◽  
Frank R. Stassen
Keyword(s):  
Human Health ◽  
Extracellular Vesicles ◽  
Preliminary Results ◽  
In Vivo Experiments ◽  
Health And Disease ◽  
Conducting Research ◽  
Protocol Standardization ◽  
Insight Into

In recent years, plant-derived extracellular vesicles (PDEVs) have gained the interest of many experts in fields such as microbiology and immunology, and research in this field has exponentially increased. These nano-sized particles have provided researchers with a number of interesting findings, making their application in human health and disease very promising. Both in vitro and in vivo experiments have shown that PDEVs can exhibit a multitude of effects, suggesting that these vesicles may have many potential future applications, including therapeutics and nano-delivery of compounds. While the preliminary results are promising, there are still some challenges to face, such as a lack of protocol standardization, as well as knowledge gaps that need to be filled. This review aims to discuss various aspects of PDEV knowledge, including their preliminary findings, challenges, and future uses, giving insight into the complexity of conducting research in this field.

scholarly journals An ARF GTPase module promoting invasion and metastasis through regulating phosphoinositide metabolism

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Marisa Nacke ◽  
Emma Sandilands ◽  
Konstantina Nikolatou ◽  
Álvaro Román-Fernández ◽  
Susan Mason ◽  
...  
Keyword(s):  
Metastatic Cancer ◽  
Cellular Responses ◽  
Signalling Pathways ◽  
External Signal ◽  
Phosphoinositide Metabolism ◽  
Invasion And Metastasis ◽  
3 Dimensional

AbstractThe signalling pathways underpinning cell growth and invasion use overlapping components, yet how mutually exclusive cellular responses occur is unclear. Here, we report development of 3-Dimensional culture analyses to separately quantify growth and invasion. We identify that alternate variants of IQSEC1, an ARF GTPase Exchange Factor, act as switches to promote invasion over growth by controlling phosphoinositide metabolism. All IQSEC1 variants activate ARF5- and ARF6-dependent PIP5-kinase to promote PI(3,4,5)P3-AKT signalling and growth. In contrast, select pro-invasive IQSEC1 variants promote PI(3,4,5)P3 production to form invasion-driving protrusions. Inhibition of IQSEC1 attenuates invasion in vitro and metastasis in vivo. Induction of pro-invasive IQSEC1 variants and elevated IQSEC1 expression occurs in a number of tumour types and is associated with higher-grade metastatic cancer, activation of PI(3,4,5)P3 signalling, and predicts long-term poor outcome across multiple cancers. IQSEC1-regulated phosphoinositide metabolism therefore is a switch to induce invasion over growth in response to the same external signal. Targeting IQSEC1 as the central regulator of this switch may represent a therapeutic vulnerability to stop metastasis.

scholarly journals Long-Acting Risperidone Dual Control System: Preparation, Characterization and Evaluation In Vitro and In Vivo

Pharmaceutics ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 1210
Author(s):  
Xieguo Yan ◽  
Shiqiang Wang ◽  
Kaoxiang Sun
Keyword(s):  
Drug Loading ◽  
Entrapment Efficiency ◽  
In Vitro Dissolution ◽  
Dissolution Behavior ◽  
In Vivo Evaluation ◽  
Long Term Treatment ◽  
Long Acting

Schizophrenia, a psychiatric disorder, requires long-term treatment; however, large fluctuations in blood drug concentration increase the risk of adverse reactions. We prepared a long-term risperidone (RIS) implantation system that can stabilize RIS release and established in-vitro and in-vivo evaluation systems. Cumulative release, drug loading, and entrapment efficiency were used as evaluation indicators to evaluate the effects of different pore formers, polymer ratios, porogen concentrations, and oil–water ratios on a RIS implant (RIS-IM). We also built a mathematical model to identify the optimized formulation by stepwise regression. We also assessed the crystalline changes, residual solvents, solubility and stability after sterilization, in-vivo polymer degradation, pharmacokinetics, and tissue inflammation in the case of the optimized formulation. The surface of the optimized RIS microspheres was small and hollow with 134.4 ± 3.5 µm particle size, 1.60 SPAN, 46.7% ± 2.3% implant drug loading, and 93.4% entrapment efficiency. The in-vitro dissolution behavior of RIS-IM had zero-order kinetics and stable blood concentration; no lag time was released for over three months. Furthermore, the RIS-IM was not only non-irritating to tissues but also had good biocompatibility and product stability. Long-acting RIS-IMs with microspheres and film coatings can provide a new avenue for treating schizophrenia.

scholarly journals Expression Profile of New Marker Genes Involved in Differentiation of Canine Adipose-Derived Stem Cells into Osteoblasts

2021 ◽  
Vol 22 (13) ◽  
pp. 6663
Author(s):  
Maurycy Jankowski ◽  
Mariusz Kaczmarek ◽  
Grzegorz Wąsiatycz ◽  
Claudia Dompe ◽  
Paul Mozdziak ◽  
...  
Keyword(s):  
Stem Cells ◽  
In Vitro Culture ◽  
Osteogenic Differentiation ◽  
Molecular Mechanisms ◽  
Marker Genes ◽  
P Value ◽  
Illumina Platform

Next-generation sequencing (RNAseq) analysis of gene expression changes during the long-term in vitro culture and osteogenic differentiation of ASCs remains to be important, as the analysis provides important clues toward employing stem cells as a therapeutic intervention. In this study, the cells were isolated from adipose tissue obtained during routine surgical procedures and subjected to 14-day in vitro culture and differentiation. The mRNA transcript levels were evaluated using the Illumina platform, resulting in the detection of 19,856 gene transcripts. The most differentially expressed genes (fold change >|2|, adjusted p value < 0.05), between day 1, day 14 and differentiated cell cultures were extracted and subjected to bioinformatical analysis based on the R programming language. The results of this study provide molecular insight into the processes that occur during long-term in vitro culture and osteogenic differentiation of ASCs, allowing the re-evaluation of the roles of some genes in MSC progression towards a range of lineages. The results improve the knowledge of the molecular mechanisms associated with long-term in vitro culture and differentiation of ASCs, as well as providing a point of reference for potential in vivo and clinical studies regarding these cells’ application in regenerative medicine.

scholarly journals ADP-ribosylation of integrin α7 modulates the binding of integrin α7β1 to laminin

2004 ◽  
Vol 385 (1) ◽  
pp. 309-317 ◽  
Author(s):  
Zhefeng ZHAO ◽  
Joanna GRUSZCZYNSKA-BIEGALA ◽  
Anna ZOLKIEWSKA
Keyword(s):  
C2c12 Myotubes ◽  
Post Translational Modification ◽  
Integrin Β1 ◽  
Adp Ribosylation ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
C2c12 Skeletal Muscle Cells ◽  
Adp Ribosyltransferase

The extracellular domain of integrin α7 is ADP-ribosylated by an arginine-specific ecto-ADP-ribosyltransferase after adding exogenous NAD+ to intact C2C12 skeletal muscle cells. The effect of ADP-ribosylation on the structure or function of integrin α7β1 has not been explored. In the present study, we show that ADP-ribosylation of integrin α7 takes place exclusively in differentiated myotubes and that this post-translational modification modulates the affinity of α7β1 dimer for its ligand, laminin. ADP-ribosylation in the 37-kDa ‘stalk’ region of α7 that takes place at micromolar NAD+ concentrations increases the binding of the α7β1 dimer to laminin. Increased in vitro binding of integrin α7β1 to laminin after ADP-ribosylation of the 37-kDa fragment of α7 requires the presence of Mn2+ and it is not observed in the presence of Mg2+. In contrast, ADP-ribosylation of the 63-kDa N-terminal region comprising the ligand-binding site of α7 that occurs at approx. 100 μM NAD+ inhibits the binding of integrin α7β1 to laminin. Furthermore, incubation of C2C12 myotubes with NAD+ increases the expression of an epitope on integrin β1 subunit recognized by monoclonal antibody 9EG7. We discuss our results based on the current models of integrin activation. We also hypothesize that ADP-ribosylation may represent a mechanism of regulation of integrin α7β1 function in myofibres in vivo when the continuity of the membrane is compromised and NAD+ is available as a substrate for ecto-ADP-ribosylation.

scholarly journals MODL-16. ABEMACICLIB, A SELECTIVE CDK4/6 INHIBITOR, RESTRICTS GROWTH OF PEDIATRIC GLIAL-LINEAGE TUMORS IN VITRO AND IN VIVO

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii414-iii414
Author(s):  
Muh-Lii Liang ◽  
Tsung-Han Hsieh ◽  
Tai-Tong Wong
Keyword(s):  
Dna Repair ◽  
Therapeutic Strategy ◽  
Target Genes ◽  
Rna Seq ◽  
Downstream Target ◽  
Rb Phosphorylation ◽  
Glial Lineage

Abstract BACKGROUND Glial-lineage tumors constitute a heterogeneous group of neoplasms, comprising gliomas, oligodendrogliomas, and ependymomas, which account for 40%–50% of all pediatric central nervous system tumors. Advances in modern neuro-oncological therapeutics are aimed at improving neoadjuvant chemotherapy and deferring radiotherapy because radiation exposure may cause long-term side effects on the developing brain in young children. Despite aggressive treatment, more than half the high-grade gliomas (pHGGs) and one-third of ependymomas exhibit recurrence within 2 years of initial treatment. METHODS By using integrated bioinformatics and through experimental validation, we found that at least one gene among CCND1, CDK4, and CDK6 was overexpressed in pHGGs and ependymomas. RESULTS The use of abemaciclib, a highly selective CDK4/6 inhibitor, effectively inhibited cell proliferation and reduced the expression of cell cycle–related and DNA repair–related gene expression, which was determined through RNA-seq analysis. The efficiency of abemaciclib was validated in vitro in pHGGs and ependymoma cells and in vivo by using subcutaneously implanted ependymoma cells from patient-derived xenograft (PDX) in mouse models. Abemaciclib demonstrated the suppression of RB phosphorylation, downstream target genes of E2F, G2M checkpoint, and DNA repair, resulting in tumor suppression. CONCLUSION Abemaciclib showed encouraging results in preclinical pediatric glial-lineage tumors models and represented a potential therapeutic strategy for treating challenging tumors in children.

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