29 results on '"Tolić, Anja"'
Search Results
2. Centaurium erythraea methanol extract improves the functionality of diabetic liver and kidney by mitigating hyperglycemia-induced oxidative stress
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Đorđević, Miloš M., Tolić, Anja, Rajić, Jovana, Mihailović, Mirjana, Arambašić Jovanović, Jelena, Uskoković, Aleksandra, Grdović, Nevena, Đorđević, Marija B., Mišić, Danijela, Šiler, Branislav, Vidaković, Melita, and Dinić, Svetlana
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- 2022
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3. TET-mediated DNA hydroxymethylation is negatively influenced by the PARP-dependent PARylation
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Tolić, Anja, Ravichandran, Mirunalini, Rajić, Jovana, Đorđević, Marija, Đorđević, Miloš, Dinić, Svetlana, Grdović, Nevena, Jovanović, Jelena Arambašić, Mihailović, Mirjana, Nestorović, Nataša, Jurkowski, Tomasz P., Uskoković, Aleksandra S., and Vidaković, Melita S.
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- 2022
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4. PARylation, DNA (De)methylation, and Diabetes
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Vidaković, Melita, Tolić, Anja, Grdović, Nevena, Ravichandran, Mirunalini, Jurkowski, Tomasz P., Patel, Vinood B., editor, and Preedy, Victor R., editor
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- 2019
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5. DNA methylation of miR-200 clusters promotes epithelial to mesenchymal transition in human conjunctival epithelial cells
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Rajić, Jovana, Dinić, Svetlana, Uskoković, Aleksandra, Arambašić Jovanović, Jelena, Tolić, Anja, Đorđević, Marija, Đorđević, Miloš, Poznanović, Goran, Mihailović, Mirjana, Inic-Kanada, Aleksandra, Barisani-Asenbauer, Talin, Grdović, Nevena, and Vidaković, Melita
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- 2020
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6. Centaurium erythraea extract reduces redox imbalance and improves insulin expression and secretion in pancreatic β-cells exposed to oxidative and nitrosative stress
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Đorđević Miloš, Grdović Nevena, Mihailović Mirjana, Arambašić-Jovanović Jelena, Uskoković Aleksandra, Rajić Jovana, Đorđević Marija, Tolić Anja, Mišić Danijela, Šiler Branislav, Poznanović Goran, Vidaković Melita, and Dinić Svetlana
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oxidative stress ,nitrosative stress ,β-cells ,centaurium erythraea ,antioxidant ,cytoprotective ,Biology (General) ,QH301-705.5 - Abstract
Oxidative stress is one of the major mechanisms that underlies the damage of pancreatic β-cells and defects in insulin secretion in diabetes. As herbal preparations can alleviate oxidative stress through their redox-active secondary metabolites, in this study we investigated the cytoprotective effects of Centaurium erythraea extract (CEe) against H2O2- and SNP-induced oxidative/nitrosative stress in Rin-5F β-cells. The antioxidant activity of CEe and its effect on cell survival and insulin expression/secretion were evaluated. The CEe increased cell viability and ameliorated the disturbance of redox homeostasis in H2O2- and SNP-treated cells by decreasing DNA damage, lipid peroxidation and protein S-glutathionylation. The CEe restored GSH homeostasis in H2O2-treated β-cells and attenuated the SNP-induced disturbance of the GSH/ GSSG ratio. The H2O2- and SNP-induced disruption of CAT, GPx, GR, MnSOD and CuZnSOD activities was adjusted by the CEe towards control values, as well as mRNA and protein levels of GPx, MnSOD and CAT. The CEe increased insulin expression/secretion particularly in H2O2-treated β-cells, which was in accordance with the more pronounced antioxidant effect of the CEe observed in H2O2-treated β-cells as compared to SNP-treated cells. These findings support the beneficial effect of the CEe in preventing or slowing down β-cell damage and dysfunction caused by oxidative/nitrosative stress during diabetes development. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 173020]
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- 2020
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7. Enrichment of Cxcl12 promoter with TET2: A possible link between promoter demethylation and enhanced gene expression in the absence of PARP-1
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Tolić Anja Z., Rajić Jovana J., Đorđević Marija B., Đorđević Miloš M., Dinić Svetlana S., Grdović Nevena M., Arambašić-Jovanović Jelena D., Mihailović Mirjana V., Poznanović Goran Đ., Jurkowski Tomasz P., Vidaković Melita S., and Uskoković Aleksandra S.
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dna demethylation ,5-aza ,tet2 ,parp-1 ,cxcl12 ,Biology (General) ,QH301-705.5 - Abstract
Previously, we described the link between C-X-C motif chemokine 12 (Cxcl12) gene induction and DNA hypomethylation in the absence of poly(ADP-ribose) polymerase 1 (PARP-1). We have now firmly established that demethylation is the primary cause of gene induction on the basis of Cxcl12 gene upregulation upon treatment with the demethylating agent 5-azacytidine (5-aza). Since the demethylation state of Cxcl12 is favored by PARP-1 absence, we investigated the presence of ten-eleven translocation (TET) proteins on the Cxcl12 promoter in order to corroborate the relationship between the demethylation process and increased gene expression that occurs in the absence of PARP-1. Analysis was performed on the promoter region within CpG islands of Cxcl12 from control mouse embryonic fibroblasts (NIH3T3) and PARP-1 knock-out mouse embryonic fibroblasts (PARP1-/-). The lack of PARP-1 increased the abundance of TET2 on the Cxcl12 promoter, suggesting that TET-mediated demethylation provoked by the absence of PARP-1 could account for the observed increased expression of this chemokine. Deciphering the regulation of DNA (de)methylation factors that control Cxcl12 expression may provide an additional therapeutic approach in pharmacological interventions where gene switching on or off based on targeted stimulation or inhibition is necessary. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 173020]
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- 2019
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8. CXC chemokine ligand 12α-mediated increase in insulin secretion and survival of mouse pancreatic islets in response to oxidative stress through modulation of calcium uptake
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Vidaković Melita, Caballero-Garrido Ernesto, Mihailović Mirjana, Arambašić-Jovanović Jelena, Sinadinović Marija, Rajić Jovana, Uskoković Aleksandra, Dinić Svetlana, Grdović Nevena, Đorđević Miloš, Tolić Anja, and Poznanović Goran
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diabetes ,calcium ,CXC chemokine ligand 12α ,insulin ,pancreatic islet cells ,voltage-gated calcium channels ,Biology (General) ,QH301-705.5 - Abstract
We examined whether CXCL12α improves insulin secretion by influencing the Ca2+ oscillation pattern and Ca2+ influx ([Ca2+]i), thereby enhancing the viability of pancreatic islet cells in oxidative stress. The islets of Langerhans were isolated from male OF1 mice and pretreated with 40 ng/mL of CXCL12α prior to exposure to 7.5 μM hydrogen peroxide, which served to induce oxidative stress. Incubation of islets with CXCL12α induced pancreatic β-cell proliferation and improved the ability of β-cells to withstand oxidative stress. Consecutive treatments of isolated islets with hydrogen peroxide caused a decline in β-cell functioning over time, while the CXCL12α pretreatment of islets exhibited a physiological response to high glucose that was comparable to control islets. The attenuated response of islets to a high D-glucose challenge was observed as a partial to complete abolishment of [Ca2+]i. Treatments with increasing concentrations of CXCL12α decreased the number of Ca2+ oscillations that lasted longer, thus pointing to an overall increase in [Ca2+]i, which was followed by increased insulin secretion. In addition, treatment of islets with CXCL12α enhanced the transcription rate for insulin and the CXCR4 gene, pointing to the importance of CXCL12/CXCR4 signaling in the regulation of Ca2+ intake and insulin secretion in pancreatic islet cells. We propose that a potential treatment with CXCL12α could help to remove preexisting glucotoxicity and associated temporary β-cell stunning that might be present at the time of diabetes diagnosis in vivo. [Project of the Serbian Ministry of Education, Science and Technological Development, Grant no. 173020]
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- 2018
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9. Association of CXCL12 gene promoter methylation with periodontitis in patients with diabetes mellitus type 2
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Grdović, Nevena, Rajić, Jovana, Petrović, Sanja Matić, Dinić, Svetlana, Uskoković, Aleksandra, Mihailović, Mirjana, Jovanović, Jelena Arambašić, Tolić, Anja, Pucar, Ana, Milašin, Jelena, and Vidaković, Melita
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- 2016
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10. Biochemical indicators and biomarkers in chub (Squalius cephalus L.) from the Sava River
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Mihailović, Mirjana, Blagojević, Duško, Ogrinc, Nives, Simonović, Predrag, Simić, Vladica, Vidaković, Melita, Dinić, Svetlana, Uskoković, Aleksandra, Grdović, Nevena, Arambašić-Jovanović, Jelena, Đorđević, Miloš, Tolić, Anja, Kračun-Kolarević, Margareta, Kolarević, Stoimir, Piria, Marina, and Paunović, Momir
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- 2016
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11. EpiCRISPR targeted methylation of Arx gene initiates transient switch of mouse pancreatic alpha to insulin-producing cells
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Đorđević, Marija, primary, Stepper, Peter, additional, Feuerstein-Akgoz, Clarissa, additional, Gerhauser, Clarissa, additional, Paunović, Verica, additional, Tolić, Anja, additional, Rajić, Jovana, additional, Dinić, Svetlana, additional, Uskoković, Aleksandra, additional, Grdović, Nevena, additional, Mihailović, Mirjana, additional, Jurkowska, Renata Z., additional, Jurkowski, Tomasz P., additional, Jovanović, Jelena Arambašić, additional, and Vidaković, Melita, additional
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- 2023
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12. EpiCRISPR targeted methylation of Arx gene initiates transient switch of mouse pancreatic alpha to insulin-producing cells
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Đorđević, Marija, Stepper, Peter, Feuerstein-Akgoz, Clarissa, Gerhauser, Clarissa, Paunović, Verica, Tolić, Anja, Rajić, Jovana, Dinić, Svetlana, Uskoković, Aleksandra, Grdović, Nevena, Mihailović, Mirjana, Jurkowska, Renata Z., Jurkowski, Tomasz P., Jovanović, Jelena Arambašić, and Vidaković, Melita
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Endocrinology, Diabetes and Metabolism - Abstract
IntroductionBeta cell dysfunction by loss of beta cell identity, dedifferentiation, and the presence of polyhormonal cells are main characteristics of diabetes. The straightforward strategy for curing diabetes implies reestablishment of pancreatic beta cell function by beta cell replacement therapy. Aristaless-related homeobox (Arx) gene encodes protein which plays an important role in the development of pancreatic alpha cells and is a main target for changing alpha cell identity.ResultsIn this study we used CRISPR/dCas9-based epigenetic tools for targeted hypermethylation of Arx gene promoter and its subsequent suppression in mouse pancreatic αTC1-6 cell line. Bisulfite sequencing and methylation profiling revealed that the dCas9-Dnmt3a3L-KRAB single chain fusion constructs (EpiCRISPR) was the most efficient. Epigenetic silencing of Arx expression was accompanied by an increase in transcription of the insulin gene (Ins2) mRNA on 5th and 7th post-transfection day, quantified by both RT-qPCR and RNA-seq. Insulin production and secretion was determined by immunocytochemistry and ELISA assay, respectively. Eventually, we were able to induce switch of approximately 1% of transiently transfected cells which were able to produce 35% more insulin than Mock transfected alpha cells.ConclusionIn conclusion, we successfully triggered a direct, transient switch of pancreatic alpha to insulin-producing cells opening a future research on promising therapeutic avenue for diabetes management.
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- 2023
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13. PARylation, DNA (De)methylation, and Diabetes
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Vidaković, Melita, primary, Tolić, Anja, additional, Grdović, Nevena, additional, Ravichandran, Mirunalini, additional, and Jurkowski, Tomasz P., additional
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- 2017
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14. Oxidative stress-mediated beta cell death and dysfunction as a target for diabetes management
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Dinić, Svetlana, primary, Arambašić Jovanović, Jelena, additional, Uskoković, Aleksandra, additional, Mihailović, Mirjana, additional, Grdović, Nevena, additional, Tolić, Anja, additional, Rajić, Jovana, additional, Đorđević, Marija, additional, and Vidaković, Melita, additional
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- 2022
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15. Nucleofection as an Efficient Method for Alpha TC1-6 Cell Line Transfection
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Đorđević, Marija, primary, Paunović, Verica, additional, Jovanović Tucović, Maja, additional, Tolić, Anja, additional, Rajić, Jovana, additional, Dinić, Svetlana, additional, Uskoković, Aleksandra, additional, Grdović, Nevena, additional, Mihailović, Mirjana, additional, Marković, Ivanka, additional, Arambašić Jovanović, Jelena, additional, and Vidaković, Melita, additional
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- 2022
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16. Additional file 1 of TET-mediated DNA hydroxymethylation is negatively influenced by the PARP-dependent PARylation
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Tolić, Anja, Ravichandran, Mirunalini, Rajić, Jovana, Đorđević, Marija, Đorđević, Miloš, Dinić, Svetlana, Grdović, Nevena, Jovanović, Jelena Arambašić, Mihailović, Mirjana, Nestorović, Nataša, Jurkowski, Tomasz P., Uskoković, Aleksandra S., and Vidaković, Melita S.
- Abstract
Additional file 1: Figures S1. In vitro PARylation of TETs. Figure S2. Co-immunoprecipitation of TET1 and PARP-1 from NIH3T3 cell lysates. Figure S3. MTT viability assay. Supplementary Figure S4. Level of 5hmC in control and ascorbic acid treated NIH3T3cells.
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- 2022
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17. Additional file 3 of TET-mediated DNA hydroxymethylation is negatively influenced by the PARP-dependent PARylation
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Tolić, Anja, Ravichandran, Mirunalini, Rajić, Jovana, Đorđević, Marija, Đorđević, Miloš, Dinić, Svetlana, Grdović, Nevena, Jovanović, Jelena Arambašić, Mihailović, Mirjana, Nestorović, Nataša, Jurkowski, Tomasz P., Uskoković, Aleksandra S., and Vidaković, Melita S.
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Additional file 3: Materials and methods: ELISA-based plate assay for DNA hydroxymethylation analysis, determination of 5hmC level by immunocytochemistry and co-immunoprecipitation.
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- 2022
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18. Additional file 2 of TET-mediated DNA hydroxymethylation is negatively influenced by the PARP-dependent PARylation
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Tolić, Anja, Ravichandran, Mirunalini, Rajić, Jovana, Đorđević, Marija, Đorđević, Miloš, Dinić, Svetlana, Grdović, Nevena, Jovanović, Jelena Arambašić, Mihailović, Mirjana, Nestorović, Nataša, Jurkowski, Tomasz P., Uskoković, Aleksandra S., and Vidaković, Melita S.
- Abstract
Additional file 2: Table S1. Statistical significance of 5hmC level differences between groups evaluated by nested ANOVA followed by Tukey post hoc test.
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- 2022
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19. α-Lipoic Acid Increases Collagen Synthesis and Deposition in Nondiabetic and Diabetic Rat Kidneys
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Grdović, Nevena, primary, Rajić, Jovana, additional, Arambašić Jovanović, Jelena, additional, Dinić, Svetlana, additional, Tolić, Anja, additional, Đorđević, Miloš, additional, Đorđević, Marija, additional, Trifunović, Svetlana, additional, Vidaković, Melita, additional, Uskoković, Aleksandra, additional, and Mihailović, Mirjana, additional
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- 2021
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20. Functional analysis of interactions between TET-mediated oxidation 5-methylcytosine and PARP-dependent ADP-ribosylation in the process of DNA demethylation
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Tolić, Anja Z., Uskoković, Aleksandra, Vidaković, Melita, and Savić-Pavićević, Dušanka
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PARylation ,TET2 ,DNA methylation ,Cxcl12 gen ,PAR polymers ,DNA demethylation ,Cxcl12 gene ,PARP-1 ,demetilacija DNK ,PAR polimeri ,metilacija DNK ,PARilacija ,TET1 - Abstract
Važan aspekt u rasvetljavanju procesa demetilacije DNK predstavlja identifikacija faktora koji regulušu aktivnost TET proteina. Cilj ove doktorske disertacije bio je da se ispita funkcionalna veza između TET-posredovane oksidacije 5mC i PARP-zavisne PARilacije u procesu demetilacije DNK, na globalnom i lokalnom nivou. Rezultati ovog istraživanja ukazuju da PARP-1 interaguje sa TET1 i TET2. Pokazano je i da oba proteina podležu in vitro PARilaciji. PARP-1-zavisna PARilacija TET1 proteina negativno utiče na kinetiku aktivnosti TET1 in vitro. Rezultati in cellulo eksperimenata pokazali su da u odsustvu PARP-1 dolazi do porasta ekspresije Tet1 i Tet2, a da prilikom inhibicije PARilacije i u odsustvu PARP-1 dolazi do pada globalnog nivoa metilacije i porasta hidroksimetilacije DNK. Za analizu lokalnih efekata TET-PARP interakcije izabran je gen za hemokin CXCL12 koji učestvuje u brojnim fiziološkim i patološkim procesima pa je ispitivanje regulacije njegove ekspresije važno za buduće terapijske pristupe. Pokazano je da u odsustvu PARP-1 dolazi do povećane ekspresije kao i do pada u metilaciji Cxcl12. Nakon tretmana aktivatorom/inhibitorom TET aktivnosti, u odsustvu PARP-1, ekpresija Cxcl12 je povećana/smanjena. Prisustvo TET1 i TET2 detektovano je na promotoru Cxcl12 uz povećano regrutovanje TET2 u odsustvu PARP-1. U ovoj disertaciji pokazan je inhibitorni uticaj PARP-1 i PARilacije na aktivnost TET enzima u procesu demetilacije DNK na globalnom nivou, a funkcionalnost lokalnih efekata povezanosti TET i PARP enzima u procesu (de)metilacije ustanovljena je na primeru Cxcl12. Demetilacija DNK nalazi se u osnovi brojnih fizioloških i patoloških stanja, zato rasvetljavanje mehanizama regulacije ovog procesa predstavlja važan korak u potencijalnoj primeni stečenih saznanja u medicini. elucidating the process of DNA demethylation. The aim of this doctoral dissertation was to investigate the functional relationship between TET-mediated oxidation of 5mC and PARP-dependent PARylation in the process of DNA demethylation at both global and local levels. This thesis shows that PARP-1 interacts with TET1 and TET2, and that both TET proteins can undergo in vitro PARylation. PARP-1-dependent PARylation of TET1 negatively affected the kinetics of TET1 activity in vitro. The results of in cellulo experiments revealed that the expression of Tet1 and Tet2 increased in the absence of PARP-1, whereas when PARylation was inhibited or in the absence of PARP-1, there was a decrease in the global level of DNA methylation and an increase in DNA hydroxymethylation. The Cxcl12 gene was selected for the analysis of the local effects of TET-PARP interaction since chemokine CXCL12 participates in numerous physiological and pathological processes, and exploring the regulation of expression of this gene is important for potential clinical intervention. Increased expression and decreased methylation of Cxcl12 were observed in the absence of PARP-1. After treatment with an activator or inhibitor of TET activity in the absence of PARP-1, Cxcl12 expression was respectively increased or decreased. The presence of TET1 and TET2 was detected on the Cxcl12 promoter, with enhanced recruitment of TET2 in the absence of PARP-1. This dissertation describes the inhibitory effects of PARP-1 and PARylation on the activity of TET enzymes in the process of DNA demethylation at the global level, and the local effects of TET-PARP interplay on DNA (de)methylation of Cxcl12 gene. DNA demethylation is at the root of numerous physiological and pathological conditions, and elucidating the mechanisms that regulate this process is an important step in the potential application of acquired knowledge in medicine.
- Published
- 2019
21. Funkcionalna analiza interakcija TET-posredovane oksidacije 5-metilcitozina i PARP-zavisne ADP-ribozilacije u procesu demetilacije DNK
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Tolić, Anja, Uskoković, Aleksandra, and Vidaković, Melita
- Subjects
PARylation ,TET2 ,DNA methylation ,Metilacija DNK ,Cxcl12 gen ,PAR polymers ,DNA demethylation ,Cxcl12 gene ,PARP-1 ,PAR polimeri ,PARilacija ,Demetilacija DNK ,TET1 - Abstract
Važan aspekt u rasvetljavanju procesa demetilacije DNK predstavlja identifikacija faktora koji regulušu aktivnost TET proteina. Cilj ove doktorske disertacije bio je da se ispita funkcionalna veza između TET-posredovane oksidacije 5mC i PARP-zavisne PARilacije u procesu demetilacije DNK, na globalnom i lokalnom nivou. Rezultati ovog istraživanja ukazuju da PARP-1 interaguje sa TET1 i TET2. Pokazano je i da oba proteina podležu in vitro PARilaciji. PARP-1-zavisna PARilacija TET1 proteina negativno utiče na kinetiku aktivnosti TET1 in vitro. Rezultati in cellulo eksperimenata pokazali su da u odsustvu PARP-1 dolazi do porasta ekspresije Tet1 i Tet2, a da prilikom inhibicije PARilacije i u odsustvu PARP-1 dolazi do pada globalnog nivoa metilacije i porasta hidroksimetilacije DNK. Za analizu lokalnih efekata TET-PARP interakcije izabran je gen za hemokin CXCL12 koji učestvuje u brojnim fiziološkim i patološkim procesima pa je ispitivanje regulacije njegove ekspresije važno za buduće terapijske pristupe. Pokazano je da u odsustvu PARP-1 dolazi do povećane ekspresije kao i do pada u metilaciji Cxcl12. Nakon tretmana aktivatorom/inhibitorom TET aktivnosti, u odsustvu PARP-1, ekpresija Cxcl12 je povećana/smanjena. Prisustvo TET1 i TET2 detektovano je na promotoru Cxcl12 uz povećano regrutovanje TET2 u odsustvu PARP-1. U ovoj disertaciji pokazan je inhibitorni uticaj PARP-1 i PARilacije na aktivnost TET enzima u procesu demetilacije DNK na globalnom nivou, a funkcionalnost lokalnih efekata povezanosti TET i PARP enzima u procesu (de)metilacije ustanovljena je na primeru Cxcl12. Demetilacija DNK nalazi se u osnovi brojnih fizioloških i patoloških stanja, zato rasvetljavanje mehanizama regulacije ovog procesa predstavlja važan korak u potencijalnoj primeni stečenih saznanja u medicini. Identification of factors that regulate the activity of TET proteins is important for elucidating the process of DNA demethylation. The aim of this doctoral dissertation was to investigate the functional relationship between TET-mediated oxidation of 5mC and PARP-dependent PARylation in the process of DNA demethylation at both global and local levels. This thesis shows that PARP-1 interacts with TET1 and TET2, and that both TET proteins can undergo in vitro PARylation. PARP-1-dependent PARylation of TET1 negatively affected the kinetics of TET1 activity in vitro. The results of in cellulo experiments revealed that the expression of Tet1 and Tet2 increased in the absence of PARP-1, whereas when PARylation was inhibited or in the absence of PARP-1, there was a decrease in the global level of DNA methylation and an increase in DNA hydroxymethylation. The Cxcl12 gene was selected for the analysis of the local effects of TET-PARP interaction since chemokine CXCL12 participates in numerous physiological and pathological processes, and exploring the regulation of expression of this gene is important for potential clinical intervention. Increased expression and decreased methylation of Cxcl12 were observed in the absence of PARP-1. After treatment with an activator or inhibitor of TET activity in the absence of PARP-1, Cxcl12 expression was respectively increased or decreased. The presence of TET1 and TET2 was detected on the Cxcl12 promoter, with enhanced recruitment of TET2 in the absence of PARP-1. This dissertation describes the inhibitory effects of PARP-1 and PARylation on the activity of TET enzymes in the process of DNA demethylation at the global level, and the local effects of TET-PARP interplay on DNA (de)methylation of Cxcl12 gene. DNA demethylation is at the root of numerous physiological and pathological conditions, and elucidating the mechanisms that regulate this process is an important step in the potential application of acquired knowledge in medicine.
- Published
- 2019
22. Centaurium erythraea extract mediates prosurvival pathways and insulin expression in STZ-treated beta-cells
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Đorđević, Miloš, Grdović, Nevena, Mihailović, Mirjana, Arambašić Jovanović, Jelena, Uskoković, Aleksandra, Rajić, Jovana, Sinadinović, Marija, Tolić, Anja, Mišić, Danijela, Šiler, Branislav, Poznanović, Goran, Vidaković, Melita, and Dinić, Svetlana
- Abstract
Diabetes is characterized by hyperglycaemia resulting from a deficiency in insulin secretion and/or action leading to severe diabetic complications. Despite numerous efforts, recovery and maintenance of functional beta-cells is still an unresolved task in diabetes therapy. Considering anti-diabetic properties of medicinal herb Centaurium erythraea Rafn (CE), this study aimed to analyze protective effects of the CE extract on Rin-5F beta-cell line exposed to diabetogenic agent streptozotocin (STZ). Cytoprotective concentration of CE extract (0.25 mg/mL) and IC50 dose of STZ (12mM) were determined using cell viability assay (MTT). The level of insulin mRNA and the concentration of insulin released from beta-cells in a culture medium were analyzed by RT-qPCR and ELISA, respectively. Activity of Akt, ERK and p38 kinases, as well as nuclear levels of islet-enriched Pdx1 and MafA proteins were assessed by Western blot analysis. In comparison to STZ-treated cells, CE extract/STZ co-treatment increased viability of Rin- 5F cells for 12%. STZ-treated beta-cells displayed reduced mRNA level of insulin to 63% and reduced insulin secretion to 76% in comparison to controls, while application of CE extract improved insulin mRNA level to 77% and insulin secretion to 90% of the control level. Improved viability and functionality of beta-cells could be ascribed to a CE extractmediated modulation of the activities of pro-survival Akt, ERK and p38 kinases and Pdx- 1 and MafA factors involved in regulation of beta-cell proliferation and insulin expression/secretion. The results of this study suggest that CE extract promotes proliferative and pro-survival pathways in beta-cells and improves their functional properties.
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- 2019
23. Funkcionalna analiza interakcija TET-posredovane oksidacije 5-metilcitozina i PARP-zavisne ADP-ribozilacije u procesu demetilacije DNK
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Uskoković, Aleksandra, Vidaković, Melita, Savić-Pavićević, Dušanka, Tolić, Anja Z., Uskoković, Aleksandra, Vidaković, Melita, Savić-Pavićević, Dušanka, and Tolić, Anja Z.
- Abstract
Važan aspekt u rasvetljavanju procesa demetilacije DNK predstavlja identifikacija faktora koji regulušu aktivnost TET proteina. Cilj ove doktorske disertacije bio je da se ispita funkcionalna veza između TET-posredovane oksidacije 5mC i PARP-zavisne PARilacije u procesu demetilacije DNK, na globalnom i lokalnom nivou. Rezultati ovog istraživanja ukazuju da PARP-1 interaguje sa TET1 i TET2. Pokazano je i da oba proteina podležu in vitro PARilaciji. PARP-1-zavisna PARilacija TET1 proteina negativno utiče na kinetiku aktivnosti TET1 in vitro. Rezultati in cellulo eksperimenata pokazali su da u odsustvu PARP-1 dolazi do porasta ekspresije Tet1 i Tet2, a da prilikom inhibicije PARilacije i u odsustvu PARP-1 dolazi do pada globalnog nivoa metilacije i porasta hidroksimetilacije DNK. Za analizu lokalnih efekata TET-PARP interakcije izabran je gen za hemokin CXCL12 koji učestvuje u brojnim fiziološkim i patološkim procesima pa je ispitivanje regulacije njegove ekspresije važno za buduće terapijske pristupe. Pokazano je da u odsustvu PARP-1 dolazi do povećane ekspresije kao i do pada u metilaciji Cxcl12. Nakon tretmana aktivatorom/inhibitorom TET aktivnosti, u odsustvu PARP-1, ekpresija Cxcl12 je povećana/smanjena. Prisustvo TET1 i TET2 detektovano je na promotoru Cxcl12 uz povećano regrutovanje TET2 u odsustvu PARP-1. U ovoj disertaciji pokazan je inhibitorni uticaj PARP-1 i PARilacije na aktivnost TET enzima u procesu demetilacije DNK na globalnom nivou, a funkcionalnost lokalnih efekata povezanosti TET i PARP enzima u procesu (de)metilacije ustanovljena je na primeru Cxcl12. Demetilacija DNK nalazi se u osnovi brojnih fizioloških i patoloških stanja, zato rasvetljavanje mehanizama regulacije ovog procesa predstavlja važan korak u potencijalnoj primeni stečenih saznanja u medicini., elucidating the process of DNA demethylation. The aim of this doctoral dissertation was to investigate the functional relationship between TET-mediated oxidation of 5mC and PARP-dependent PARylation in the process of DNA demethylation at both global and local levels. This thesis shows that PARP-1 interacts with TET1 and TET2, and that both TET proteins can undergo in vitro PARylation. PARP-1-dependent PARylation of TET1 negatively affected the kinetics of TET1 activity in vitro. The results of in cellulo experiments revealed that the expression of Tet1 and Tet2 increased in the absence of PARP-1, whereas when PARylation was inhibited or in the absence of PARP-1, there was a decrease in the global level of DNA methylation and an increase in DNA hydroxymethylation. The Cxcl12 gene was selected for the analysis of the local effects of TET-PARP interaction since chemokine CXCL12 participates in numerous physiological and pathological processes, and exploring the regulation of expression of this gene is important for potential clinical intervention. Increased expression and decreased methylation of Cxcl12 were observed in the absence of PARP-1. After treatment with an activator or inhibitor of TET activity in the absence of PARP-1, Cxcl12 expression was respectively increased or decreased. The presence of TET1 and TET2 was detected on the Cxcl12 promoter, with enhanced recruitment of TET2 in the absence of PARP-1. This dissertation describes the inhibitory effects of PARP-1 and PARylation on the activity of TET enzymes in the process of DNA demethylation at the global level, and the local effects of TET-PARP interplay on DNA (de)methylation of Cxcl12 gene. DNA demethylation is at the root of numerous physiological and pathological conditions, and elucidating the mechanisms that regulate this process is an important step in the potential application of acquired knowledge in medicine.
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- 2019
24. Protective effect of Centaurium erythraea methanol extract against oxidative challenge in red blood cells of diabetic rats
- Author
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Đorđević, Miloš, Mihailović, Mirjana, Arambašić Jovanović, Jelena, Grdović, Nevena, Uskoković, Aleksandra, Sinadinović, Marija, Rajić, Jovana, Tolić, Anja, Poznanović, Goran, Vidaković, Melita, and Dinić, Svetlana
- Subjects
Centaurium erythraea ,antioxidant ,diabetes ,oxidative stress ,red blood cells - Abstract
In light of increasing prevalence of diabetes over the past few decades and increasing lifespan of human population, there is a need to better monitor the quality of life of diabetic patients. Epidemiological evidence shows protective effect of regular consumption of diets rich in plant polyphenols against development of diabetes. Centaurium erythraea Rafn (CE) is traditionally used in Serbia for diabetes treatment. Previous phytochemical studies with aerial parts of CE showed it can substantially alleviate oxidative stress, one of the major pathogenic factors that lead to diabetes and its complications. Chronic hyperglycemia, the hallmark of diabetes, leads to increased production of free radicals which affect red blood cells (RBCs) structure and function. Considering RBCs role as oxygen transporters and consequences of impaired oxygen delivery in diabetes, the main goal of this research was to evaluate the protective effect of the methanol extract of aerial parts of CE against oxidative challenge in RBCs of rats with streptozotocin (STZ)-induced diabetes. The CE extract (100 mg/kg) was given daily and orally two weeks before, during diabetes induction (i.p. injection of STZ (40 mg/kg) for five consecutive days), and for four weeks after the STZ injections (animals designated as pre-treated group), or for four weeks after diabetes induction (post-treated group). Daily application of CE extract to STZ-induced diabetic rats improved the redox status of RBCs, observed as reduced lipid peroxidation and alleviated oxidative damage due to improved glutathione system and antioxidant enzyme activity, such as catalase (CAT), superoxide dismutase (SOD) and glutathione reductase (GR). Ameliorated RBCs’ redox status was accompanied with improvement of major biochemical indicators of diabetes. CE extract increased serum insulin level, reduced blood glucose and glycated hemoglobin concentrations and improved lipid profile of diabetic rats. Furthermore, the CE extract reduced non-enzymatic glycation and enzymatic glycosylation and improved parameters which correlate with RBC aggregation and deformability. The protective effect of CE extract was more pronounced in pre-treated diabetic group. According to these results, Centaurium erythraea methanol extract has a great potential for use as dietary supplement in diabetes management. Since plenty of bioactive compounds, including polyphenols were determined in CE extract, identification of active metabolites from CE and their tissue distribution after intake could provide a useful source of potential novel antidiabetic pharmaceutical entities.
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- 2018
25. Centaurium erythraea extract improves survival and functionality of pancreatic beta-cells in diabetes through multiple routes of action
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Đorđević, Miloš, primary, Grdović, Nevena, additional, Mihailović, Mirjana, additional, Arambašić Jovanović, Jelena, additional, Uskoković, Aleksandra, additional, Rajić, Jovana, additional, Sinadinović, Marija, additional, Tolić, Anja, additional, Mišić, Danijela, additional, Šiler, Branislav, additional, Poznanović, Goran, additional, Vidaković, Melita, additional, and Dinić, Svetlana, additional
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- 2019
- Full Text
- View/download PDF
26. Absence of PARP‐1 affects Cxcl12 expression by increasing DNA demethylation
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Tolić, Anja, primary, Grdović, Nevena, additional, Dinić, Svetlana, additional, Rajić, Jovana, additional, Đorđević, Miloš, additional, Sinadinović, Marija, additional, Arambašić Jovanović, Jelena, additional, Mihailović, Mirjana, additional, Poznanović, Goran, additional, Uskoković, Aleksandra, additional, and Vidaković, Melita, additional
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- 2019
- Full Text
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27. Chlamydia trachomatis infection and development of epithelial mesenchymal transition in conjunctiva : possible epigenetic mechanisms unveiled
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Rajić, Jovana, Grdović, Nevena, Inic-Kanada, Aleksandra, Stein, Elisabeth, Schuerer, Nadine, Uskoković, Aleksandra, Ghasemian, Ehsan, Dinić, Svetlana, Mihailović, Mirjana, Arambašić Jovanović, Jelena, Tolić, Anja, Sinadinović, Marija, Đorđević, Miloš, Poznanović, Goran, Barisani-Asenbauer, Talin, Vidaković, Melita, Brajušković, Goran, and Đorđević, Ana
- Abstract
Introduction: Trachoma is the most common cause of infectious blindness worldwide, initiated by repeated infection of the conjunctiva with Chlamydia trachomatis (Ct). The resulting chronic inflammation and formation of fibrotic tissue eventually lead to corneal damage. Based on the facts that epithelial to mesenchymal transition (EMT) plays an important role in the development of fibrosis and that EMT is epigenetically regulated process, the aims of this study were to reveal the capacity of Ct to induce EMT in vitro and to unveil potential underlying epigenetic mechanisms. Methods: Human conjunctival epithelial (HCjE) cells were infected with 107 IFU of Ct for 72 h. EMT-inducing signaling pathways, as well as mRNA and protein expression of EMT markers (E-cadherin, fibronectin and α-SMA) were evaluated by RT-qPCR, Immunoblotting and Immunocytochemistry. DNA methylation patterns of selected regions of E-cadherin, fibronectin and α-SMA genes were examined by Methylation-Specific PCR, High Resolution Melting analysis and Bisulfite Sequencing. Results: Infection with Ct was accompanied with the activation of EMT-inducing signaling pathways, downregulation of epithelial marker E-cadherin and upregulation of mesenchymal markers fibronectin and α-SMA. While DNA methylation status of E-cadherin gene promoter correlated with its expression, methylation status of fibronectin and α-SMA genes couldn’t be related to their expression levels. Conclusion: Ct infection of HCjE cells triggers EMT that goes along with changes in the methylation profile of the E-cadherin promoter. Sequence of events described herein could contribute to scarring process in trachoma and open up possibilities for development of new therapeutic strategies in trachoma treatment. Brajušković G, Đorđević A, editors. CoMBoS. 1st Congress of Molecular Biologists of Serbia; 2017 Sep 20-22; Belgrade, Serbia. Belgrade: University of Belgrade, Faculty of Biology; 2017. p. 70.
- Published
- 2017
28. Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation
- Author
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Sinadinović, Marija, Arambašić Jovanović, Jelena, Tolić, Anja, Đorđević, Miloš, Mihailović, Mirjana, Grdović, Nevena, Uskoković, Aleksandra, Rajić, Jovana, Dinić, Svetlana, Jurkowski, Tomasz P., and Vidaković, Melita
- Abstract
Introduction: Since diabetes is characterized by impaired ability of pancreatic betacells to respond and/or produce insulin, new approaches for renewal and replacement of deficient beta-cells are indispensable. The aim of this study is direct pancreatic alpha- to beta-cells transdifferentiation by using a new synthetic epigenetic tool, Epi-CRISPR system. Using Epi-CRISPR system we aim to introduce targeted DNA methylation and subsequent repression of genes responsible for maintaining alpha-cell identity. Methods: AlphaTC1-6 cells (α-cells) were transiently transfected with dCas9-Dnmt3a- Dnmt3L constructs and one or four different vectors containing guide RNA components for specific targeting the promoter region of aristaless-related homeobox gene (Arx). The success of α-cells transdifferentiation into insulinproducing cells was evaluated by measuring Arx and insulin mRNA level, amount of secreted insulin and by immunostaining of insulin/glucagon in the cells. Results: We observed Arx transcriptional repression in α-cell transfected with Epi- CRISPR construct that targets the Arx gene promoter inducing subsequent methylation. At fifth day post-transfection the expression of Arx was decreased in α- cells followed by consequent increase in insulin (mRNA and protein level). At the same time, the glucagon levels remained unchanged. At twelfth day posttransfection the transfected cells start to lose glucagon while still secreting insulin. Conclusion: This study is near to confirm Epi-CRISPR system functionality and to verify the concept of cell transdifferentiation through silencing of genes responsible for maintaining cell phenotype. The obtained results will be valuable for later Epi- CRISPRs use in mouse in vivo models of diabetes and eventually as a future therapy for diabetes attenuation in humans. Brajušković G, Đorđević A, editors. CoMBoS. 1st Congress of Molecular Biologists of Serbia; 2017 Sep 20-21; Belgrade, Serbia. Belgrade: University of Belgrade, Faculty of Biology; 2017. p. 73.
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- 2017
29. Centaurium erythraea methanol extract protects red blood cells from oxidative damage in streptozotocin-induced diabetic rats
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Đorđević, Miloš, primary, Mihailović, Mirjana, additional, Arambašić Jovanović, Jelena, additional, Grdović, Nevena, additional, Uskoković, Aleksandra, additional, Tolić, Anja, additional, Sinadinović, Marija, additional, Rajić, Jovana, additional, Mišić, Danijela, additional, Šiler, Branislav, additional, Poznanović, Goran, additional, Vidaković, Melita, additional, and Dinić, Svetlana, additional
- Published
- 2017
- Full Text
- View/download PDF
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