Uloga transkripcijskih čimbenika Gata4/6 u diferencijaciji humanih embrionalnih matičnih stanica in vitro

doctoral thesis

• Title (english): Role of Gata4/6 transcription factors in differentiation of human embryonic stem cells in vitro
• Author: Pavle Josipović
• Mentor: Inga Marijanović (mentor)
• Mentor: Nikica Zaninović (sumentor); MBZ: 140911
• Committee member: Inga Marijanović (član povjerenstva)
• Committee member: Gordana Lacković-Venturin (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2013, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology
• Universal decimal classification (UDC): 576 - Cellular and subcellular biology. Cytology

Abstract

Ljudske embrionalne matične stanice (prema eng. human embryonal stem cells, hESC) mogu poslužiti kao model za istraživanje razvoja primitivnog izvanembrionalnog endoderma. Spontana diferencijacija stanica embrioblasta je posljedica uzgoja hESC u kulturi. GATA transkripcijski čimbenici su od presudnog značaja u razvoju izvanembrionalnog primitivnog endoderma. Istražili smo ekspresiju transkripcijskih čimbenika Gata4 i Gata6, markera primitivnog endoderma Sox17 i gena pluripotentnosti Oct4, Sox2 i Nanog. Kvantitativni PCR je pokazao pojačanu ekspresiju Gata4, Gata6 i Sox17 tijekom spontane diferencijacije u svim istraživanim sustavima. Istražen je utjecaj tri različita diferencijacijska sustava- matrigel, mišji embrionalni fibroblasti i embrioidna tijela, na razvoj primitivnog endoderma i na ekspresiju navedenih biljega. Imunocitokemijskim bojanjem dokazana je prisutnost GATA4 i GATA6 proteina u jezgrama diferenciranih embrionalnih matičnih stanica kao i njihova specifična prostorna raspodjela. U svrhu istraživanja mehanizama rane embrionalne diferencijacije generirane su transgenične linije hESC (VprOrgGata6GFP, VprOrgGata6mCherry, VprOrgCD41aGFP). Uzgojem transgeničnih kultura dokazano je da genetski modificirane hESC mogu poslužiti kao model rane embriogeneze in vitro.

Abstract (english)

Human embryonic stem cells (hESC) can serve as a model for studying the development of primitive extraembryonic endoderm. Spontaneous differentiation of inner cell mass cells is a result of growing hESC in a cell culture. GATA transcription factors are crucial elements in the development of primitive extraembryonal endoderm. In our study we have focused on the expression of transcription factors Gata4 and Gata6, the primitive endoderm marker Sox17, as well as the expression of pluripotency genes Oct4, Sox2 i Nanog. Real time PCR has showed high expression Gata4, Gata6 and Sox17 during the spontaneous differentiation in all differentiating systems. We have explored the impact of three diverse differentiation system - matrigel, murine embryonic fibroblasts and embryoid bodies - on the development of primitive endoderm and expression of above mentioned markers. The presence of GATA4 and GATA6 proteins in the nuclei of differentiated embryonic stem cells and their specific spatial distribution was demonstrated by immunostaining. In order to investigate the mechanisms of early embryonic differentiation we generated hESC transgenic lines (VprOrgGata6GFP, VprOrgGata6mCherry, VprOrgCD41aGFP). Genetically modified hESC can serve as a model of early embryogenesis in vitro.

• Keywords: GATA4; GATA6; Primitivni endoderm; hESC; spontana diferencijacija
• Keywords (english): GATA4; GATA6; Primitive endoderm; hESC; spontaneous differentiation
• Language: croatian
• URN:NBN: urn:nbn:hr:217:719273
• Study programme: Title: Biology; Study programme type: university; Study level: postgraduate; Academic / professional title: doktor/doktorica znanosti, područje prirodnih znanosti, polje biologija (doktor/doktorica znanosti, područje prirodnih znanosti, polje biologija)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Closed access
• Created on: 2017-05-19 08:38:05