TY  - CONF
AU  - Đorđević, Marija
AU  - Arambašić Jovanović, Jelena
AU  - Tolić, Anja
AU  - Đorđević, Miloš
AU  - Mihailović, Mirjana
AU  - Grdović, Nevena
AU  - Uskoković, Aleksandra
AU  - Rajić, Jovana
AU  - Dinić, Svetlana
AU  - Jurkowski, Tomasz P.
AU  - Vidaković, Melita
PY  - 2017
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/2868
UR  - https://radar.ibiss.bg.ac.rs/handle/123456789/2865
AB  - 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.
PB  - Belgrade: University of Belgrade, Faculty of Biology, Belgrade
C3  - 1st Congress of Molecular Biologists of Serbia
T1  - Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation
SP  - 73
EP  - 73
UR  - https://hdl.handle.net/21.15107/rcub_ibiss_2868
ER  - 

@conference{
editor = "Brajušković, Goran, Đorđević, Ana",
author = "Đorđević, Marija and Arambašić Jovanović, Jelena and Tolić, Anja and Đorđević, Miloš and Mihailović, Mirjana and Grdović, Nevena and Uskoković, Aleksandra and Rajić, Jovana and Dinić, Svetlana and Jurkowski, Tomasz P. and Vidaković, Melita",
year = "2017",
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.",
publisher = "Belgrade: University of Belgrade, Faculty of Biology, Belgrade",
journal = "1st Congress of Molecular Biologists of Serbia",
title = "Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation",
pages = "73-73",
url = "https://hdl.handle.net/21.15107/rcub_ibiss_2868"
}

Brajušković, G., Đorđević, A., Đorđević, M., Arambašić Jovanović, J., Tolić, A., Đorđević, M., Mihailović, M., Grdović, N., Uskoković, A., Rajić, J., Dinić, S., Jurkowski, T. P.,& Vidaković, M.. (2017). Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation. in 1st Congress of Molecular Biologists of Serbia
Belgrade: University of Belgrade, Faculty of Biology, Belgrade., 73-73.
https://hdl.handle.net/21.15107/rcub_ibiss_2868

Brajušković G, Đorđević A, Đorđević M, Arambašić Jovanović J, Tolić A, Đorđević M, Mihailović M, Grdović N, Uskoković A, Rajić J, Dinić S, Jurkowski TP, Vidaković M. Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation. in 1st Congress of Molecular Biologists of Serbia. 2017;:73-73.
https://hdl.handle.net/21.15107/rcub_ibiss_2868 .

Brajušković, Goran, Đorđević, Ana, Đorđević, Marija, Arambašić Jovanović, Jelena, Tolić, Anja, Đorđević, Miloš, Mihailović, Mirjana, Grdović, Nevena, Uskoković, Aleksandra, Rajić, Jovana, Dinić, Svetlana, Jurkowski, Tomasz P., Vidaković, Melita, "Transdifferentiation of pancreatic alpha to beta cells using Epi-CRISPR directed DNA methylation" in 1st Congress of Molecular Biologists of Serbia (2017):73-73,
https://hdl.handle.net/21.15107/rcub_ibiss_2868 .