TY  - CONF
AU  - Laketa, Danijela
AU  - Josipović, Nataša
AU  - Lavrnja, Irena
AU  - Bjelobaba, Ivana
AU  - Jakovljević, Marija
AU  - Božić, Iva
AU  - Savić, Danijela
AU  - Dacić, Sanja
AU  - Peković, Sanja
AU  - Nedeljković, Nadežda
PY  - 2017
UR  - http://radar.ibiss.bg.ac.rs/handle/123456789/5988
AB  - Introduction. Ecto-5'–nucleotidase (eN) catalyzes terminal step of extracellular ATP hydrolysis, producing anti-inflammatory adenosine. We reported significantly increased eN activity in lumbar spinal cord during experimental autoimmune encephalomyelitis (EAE), together with increased protein expression connected mainly with reactive astrocytes and appearance of new isoform at ~75kDa at the peak of the disease, besides usual ~71kDa isoform. Since eN is glycoprotein with five potential N-glycosylation sites and 
redicted molecular weight of 57-59 kDa, we hypothesized that occurrence of second isoform during EAE is due to changes in glycosylation pattern, possibly affecting kinetic properties of the enzyme. Methods. Lumbar parts of the spinal cords were obtained from Dark Agouti rats at the onset (Eo), peak (Ep) and the end of symptoms (Er) during EAE and from naïve control animals (C). Results. We here report significant changes of kinetic properties regarding AMP-hydrolysis during EAE, with almost 50% increase of maximal velocity at Ep (92.35±1.86nmolPi/min/mg) and Er (90.68±2.17nmolPi/min/mg), compared to C, whilst Km increased double at Ep (0.041±0.003mmol/l). Enzymatic deglycosylation caused triple decrease of Vmax (33.6±1.8nmolPi/mg/min) at Ep, and double decrease of Km (0.022±0.008mmol/l), whilst immunoblot
probed with anti-eN antibody revealed triple protein band at ~60kDa at all investigated time-points. Conclusion. Our results show that changes of kinetic properties during EAE, at least partially, are governed by modification of glycosylation pattern. Also, appearance of new isoform at the peak of EAE is direct consequence of glycosylation changes. In summary, besides gene and protein expression changes of eN, glycosylation might be additional route of inflammation control conducted by astrocytes.
PB  - Belgrade: Serbian Neuroscience Society
C3  - Book of Abstract: 7th Congress of Serbian Neuroscience Society with international participation; 2017 Oct 25-27; Belgrade, Serbia
T1  - Appearance of second ecto-5'-nucleotidase isoform during experimental autoimmune encephalomyelitis is caused by changes in glycosylation pattern
SP  - 70
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_5988
ER  - 

@conference{
author = "Laketa, Danijela and Josipović, Nataša and Lavrnja, Irena and Bjelobaba, Ivana and Jakovljević, Marija and Božić, Iva and Savić, Danijela and Dacić, Sanja and Peković, Sanja and Nedeljković, Nadežda",
year = "2017",
abstract = "Introduction. Ecto-5'–nucleotidase (eN) catalyzes terminal step of extracellular ATP hydrolysis, producing anti-inflammatory adenosine. We reported significantly increased eN activity in lumbar spinal cord during experimental autoimmune encephalomyelitis (EAE), together with increased protein expression connected mainly with reactive astrocytes and appearance of new isoform at ~75kDa at the peak of the disease, besides usual ~71kDa isoform. Since eN is glycoprotein with five potential N-glycosylation sites and 
redicted molecular weight of 57-59 kDa, we hypothesized that occurrence of second isoform during EAE is due to changes in glycosylation pattern, possibly affecting kinetic properties of the enzyme. Methods. Lumbar parts of the spinal cords were obtained from Dark Agouti rats at the onset (Eo), peak (Ep) and the end of symptoms (Er) during EAE and from naïve control animals (C). Results. We here report significant changes of kinetic properties regarding AMP-hydrolysis during EAE, with almost 50% increase of maximal velocity at Ep (92.35±1.86nmolPi/min/mg) and Er (90.68±2.17nmolPi/min/mg), compared to C, whilst Km increased double at Ep (0.041±0.003mmol/l). Enzymatic deglycosylation caused triple decrease of Vmax (33.6±1.8nmolPi/mg/min) at Ep, and double decrease of Km (0.022±0.008mmol/l), whilst immunoblot
probed with anti-eN antibody revealed triple protein band at ~60kDa at all investigated time-points. Conclusion. Our results show that changes of kinetic properties during EAE, at least partially, are governed by modification of glycosylation pattern. Also, appearance of new isoform at the peak of EAE is direct consequence of glycosylation changes. In summary, besides gene and protein expression changes of eN, glycosylation might be additional route of inflammation control conducted by astrocytes.",
publisher = "Belgrade: Serbian Neuroscience Society",
journal = "Book of Abstract: 7th Congress of Serbian Neuroscience Society with international participation; 2017 Oct 25-27; Belgrade, Serbia",
title = "Appearance of second ecto-5'-nucleotidase isoform during experimental autoimmune encephalomyelitis is caused by changes in glycosylation pattern",
pages = "70",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_5988"
}

Laketa, D., Josipović, N., Lavrnja, I., Bjelobaba, I., Jakovljević, M., Božić, I., Savić, D., Dacić, S., Peković, S.,& Nedeljković, N.. (2017). Appearance of second ecto-5'-nucleotidase isoform during experimental autoimmune encephalomyelitis is caused by changes in glycosylation pattern. in Book of Abstract: 7th Congress of Serbian Neuroscience Society with international participation; 2017 Oct 25-27; Belgrade, Serbia
Belgrade: Serbian Neuroscience Society., 70.
https://hdl.handle.net/21.15107/rcub_ibiss_5988

Laketa D, Josipović N, Lavrnja I, Bjelobaba I, Jakovljević M, Božić I, Savić D, Dacić S, Peković S, Nedeljković N. Appearance of second ecto-5'-nucleotidase isoform during experimental autoimmune encephalomyelitis is caused by changes in glycosylation pattern. in Book of Abstract: 7th Congress of Serbian Neuroscience Society with international participation; 2017 Oct 25-27; Belgrade, Serbia. 2017;:70.
https://hdl.handle.net/21.15107/rcub_ibiss_5988 .

Laketa, Danijela, Josipović, Nataša, Lavrnja, Irena, Bjelobaba, Ivana, Jakovljević, Marija, Božić, Iva, Savić, Danijela, Dacić, Sanja, Peković, Sanja, Nedeljković, Nadežda, "Appearance of second ecto-5'-nucleotidase isoform during experimental autoimmune encephalomyelitis is caused by changes in glycosylation pattern" in Book of Abstract: 7th Congress of Serbian Neuroscience Society with international participation; 2017 Oct 25-27; Belgrade, Serbia (2017):70,
https://hdl.handle.net/21.15107/rcub_ibiss_5988 .

TY  - JOUR
AU  - Adžić, Marija
AU  - Stevanović, Ivana
AU  - Josipović, Nataša
AU  - Laketa, Danijela
AU  - Lavrnja, Irena
AU  - Bjelobaba, Ivana
AU  - Božić, Iva
AU  - Jovanović, Marija
AU  - Milošević, Milena
AU  - Nedeljković, Nadežda
PY  - 2017
UR  - http://doi.wiley.com/10.1002/jnr.23950
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/2560
AB  - It is widely accepted that adenosine triphosphate (ATP) acts as a universal danger-associated molecular pattern with several known mechanisms for immune cell activation. In the central nervous system, ATP activates microglia and astrocytes and induces a neuroinflammatory response. The aim of the present study was to describe responses of isolated astrocytes to increasing concentrations of ATP (5 µM to 1 mM), which were intended to mimic graded intensity of the extracellular stimulus. The results show that ATP induces graded activation response of astrocytes in terms of the cell proliferation, stellation, shape remodeling, and underlying actin and GFAP filament rearrangement, although the changes occurred without an apparent increase in GFAP and actin protein expression. On the other hand, ATP in the range of applied concentrations did not evoke IL-1β release from cultured astrocytes, nor did it modify the release from LPS and LPS+IFN-γ–primed astrocytes. ATP did not promote astrocyte migration in the wound-healing assay, nor did it increase production of reactive oxygen and nitrogen species and lipid peroxidation. Instead, ATP strengthened the antioxidative defense of astrocytes by inducing Cu/ZnSOD and MnSOD activities and by increasing their glutathione content. Our current results suggest that although ATP triggers several attributes of activated astrocytic phenotype with a magnitude that increases with the concentration, it is not sufficient to induce full-blown reactive phenotype of astrocytes in vitro. © 2016 Wiley Periodicals, Inc.
T2  - Journal of Neuroscience Research
T1  - Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro
IS  - 4
VL  - 95
DO  - 10.1002/jnr.23950
SP  - 1053
EP  - 1066
ER  - 

@article{
author = "Adžić, Marija and Stevanović, Ivana and Josipović, Nataša and Laketa, Danijela and Lavrnja, Irena and Bjelobaba, Ivana and Božić, Iva and Jovanović, Marija and Milošević, Milena and Nedeljković, Nadežda",
year = "2017",
abstract = "It is widely accepted that adenosine triphosphate (ATP) acts as a universal danger-associated molecular pattern with several known mechanisms for immune cell activation. In the central nervous system, ATP activates microglia and astrocytes and induces a neuroinflammatory response. The aim of the present study was to describe responses of isolated astrocytes to increasing concentrations of ATP (5 µM to 1 mM), which were intended to mimic graded intensity of the extracellular stimulus. The results show that ATP induces graded activation response of astrocytes in terms of the cell proliferation, stellation, shape remodeling, and underlying actin and GFAP filament rearrangement, although the changes occurred without an apparent increase in GFAP and actin protein expression. On the other hand, ATP in the range of applied concentrations did not evoke IL-1β release from cultured astrocytes, nor did it modify the release from LPS and LPS+IFN-γ–primed astrocytes. ATP did not promote astrocyte migration in the wound-healing assay, nor did it increase production of reactive oxygen and nitrogen species and lipid peroxidation. Instead, ATP strengthened the antioxidative defense of astrocytes by inducing Cu/ZnSOD and MnSOD activities and by increasing their glutathione content. Our current results suggest that although ATP triggers several attributes of activated astrocytic phenotype with a magnitude that increases with the concentration, it is not sufficient to induce full-blown reactive phenotype of astrocytes in vitro. © 2016 Wiley Periodicals, Inc.",
journal = "Journal of Neuroscience Research",
title = "Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro",
number = "4",
volume = "95",
doi = "10.1002/jnr.23950",
pages = "1053-1066"
}

Adžić, M., Stevanović, I., Josipović, N., Laketa, D., Lavrnja, I., Bjelobaba, I., Božić, I., Jovanović, M., Milošević, M.,& Nedeljković, N.. (2017). Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro. in Journal of Neuroscience Research, 95(4), 1053-1066.
https://doi.org/10.1002/jnr.23950

Adžić M, Stevanović I, Josipović N, Laketa D, Lavrnja I, Bjelobaba I, Božić I, Jovanović M, Milošević M, Nedeljković N. Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro. in Journal of Neuroscience Research. 2017;95(4):1053-1066.
doi:10.1002/jnr.23950 .

Adžić, Marija, Stevanović, Ivana, Josipović, Nataša, Laketa, Danijela, Lavrnja, Irena, Bjelobaba, Ivana, Božić, Iva, Jovanović, Marija, Milošević, Milena, Nedeljković, Nadežda, "Extracellular ATP induces graded reactive response of astrocytes and strengthens their antioxidative defense in vitro" in Journal of Neuroscience Research, 95, no. 4 (2017):1053-1066,
https://doi.org/10.1002/jnr.23950 . .