Replikacija DNA u sekundarnom limfnom tkivu

master's thesis

• Title (english): DNA replication in secondary lymphoid tissue
• Author: Vanja Tadić
• Mentor: Petra Korać (mentor)
• Committee member: Petra Korać (predsjednik povjerenstva)
• Committee member: Antun Alegro (član povjerenstva)
• Committee member: Damjan Franjević (član povjerenstva)
• Committee member: Mladen Kučinić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2014-09-22, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Germinativni centri (GC) folikula sekundarnog limfnog tkiva iznimno su dinamičan okoliš gdje B‐limfociti prolaze klonalnu ekspanziju, somatsku hipermutaciju, pozitivnu i negativnu selekciju te class‐switch rekombinaciju. To su mjesta gdje oni postaju dugoživuće memorijske B‐stanice i plazma stanice koje luče visokoafinitetna antitijela. Zbog svoje funkcije, germinativni centri imaju visoku proliferacijsku stopu, što čini replikaciju ključnim procesom u tim dijelovima tonzila. Osim germinativnih centara u sekundarnom limfnom tkivu uočljiva je i plaštana zona, u kojoj se nalaze B‐stanice koje ulaze u GC i interfolikularna zona, u kojoj su pretežno T‐limfociti u potrazi za dendritičkim stanicama sa specifičnim antigenom. Cilj je ovog istraživanja analiza količine transkripata gena koji kodiraju za ključne proteine u procesu replikacije u tri funkcionalno različita dijela tonzile. Korišteno je 10 uzoraka tonzila dobivenih tonzilektomijom od djece s čestim upalama. Funkcionalno različiti dijelovi tonzila izdvojeni su laserskom mikrodisekcijom, izolirana je RNA i reverznom transkripcijom pripremljena za analizu transkripata gena DNMT1, PCNA, MCM2, MCM7, CDT1, EZH2, geminina i p300 lančanom reakcijom polimeraze u stvarnom vremenu. Dobiveni rezultati pokazuju da stanice germinativnog centra eksprimiraju značajno veću količinu EZH2 od stanica plaštene zone i značajno veću količinu geminina od stanica interfolikularne zone. Ovakve razlike u ekspresiji EZH2 i geminina upućuju na procese u kojima ti proteini imaju ulogu u tumorskim stanicama porijekla B‐limfocita germinativnog centra i tako čine potencijalne mete daljnjih istraživanja u svrhu razvoja ciljanih terapija.

Abstract (english)

Germinal centers (GC) of secondary lymphoid tissue follicles are extremely dynamic microenvironments, where B‐lymphocytes undergo clonal expansion, somatic hypermutation, positive and negative selection and class‐switch recombination. These are places where B‐lymphocytes grow to become long‐living memory B cells and plasma cells, which produce highly specific antibodies. Because of their function, GC cells have a high proliferating rate, which makes replication a key process in these areas of tonsils. In secondary lymphoid tissue, aside from germinal centers, it is easy to observe mantle zones which contain Blymphocytes entering germinal center and interfollicular zones which contain mostly T‐lymphocytes in search for dendritic cells with specific antigen. Goal of this study was to analyze the transcript levels of genes that code for replication proteins in three functionally different parts of tonsil. Ten tonsils gathered after the tonsillectomy from children with recurrent tonsillitis were used. Functionally different parts of tonsil were collected using laser‐microdissection technology; RNA was isolated and reversely transcribed into cDNA for QPCR analysis of DNMT1, PCNA, MCM2, MCM7, CDT1, EZH2, geminin and p300 transcripts. Results show that GC cells express significantly higher amount of EZH2 compared to mantle zone cells, and significantly higher amount of geminin compared to interfollicular zone cells. These differences in EZH2 and geminin expression point to processes in which these proteins take part in tumor cells originating from GC B‐lymphocytes, that could be used as targets in future therapies.

• Keywords: replikacija; germinativni centar; plaštena zona; interfolikularna zona; EZH2; geminin
• Keywords (english): replication; germinal center; mantle zone; interfollicular zone; EZH2; geminin
• Language: croatian
• URN:NBN: urn:nbn:hr:217:433255
• Study programme: Title: Molecular Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra molekularne biologije (magistar/magistra molekularne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Access restricted to students and staff of home institution
• Created on: 2017-04-26 12:35:18