Title Uloga iglC gena u unutarstaničnom životu Francisella tularensis : doktorski rad
Title (english) The role of iglC gene in intracellular life of Francisella Tularensis
Author Gordana Pavoković
Mentor Marina Šantić (mentor)
Mentor Nije poznat komentor Nije poznat komentor (komentor)
Committee member Tomislav Rukavina (predsjednik povjerenstva)
Committee member Ana Budimir (član povjerenstva)
Committee member Astrid Krmpotić (član povjerenstva) MBZ: 00000000001
Granter University of Rijeka Faculty of Medicine Rijeka
Defense date and country 2012-06-15, Croatia
Scientific / art field, discipline and subdiscipline BIOMEDICINE AND HEALTHCARE Clinical Medical Sciences
Universal decimal classification (UDC ) 616 APPLIED SCIENCES. MEDICINE. TECHNOLOGY Pathology. Clinical medicine
Abstract CILJEVI
Odrediti ulogu iglC gena u unutarstanicnom životu F. tularensis subsp. novicida u humanim
makrofagima i amebama putem kinetike rasta, modulacije apoptoze, biogeneze fagosoma i
ultrastrukturalne organizacije stanica nakon infekcije.
METODE
Stanice A. castellanii i H. vermiformis te humani makrofagi inficirane su s F. tularensis
subsp. novicida i njenom mutantom iglC. U odreenim vremenskim periodima nakon
infekcije stanice su procesuirane za odreivanje ukupnog broja bakterija metodom kinetike
rasta, stupnja zakiseljavanja fagosoma metodom konfokalne mikroskopije dok je
ultrastrukturalana organizacija stanice odreivana elektronskom mikroskopijom.
Otpornost na apoptozu te aktivacija kaspaze-1 i 3 u humanih makrofaga inficiranih s F.
tularensis subsp. novicida ili iglC mutantom odreivana je imunofluorescentim bojenjem
putem konfokalane mikroskopije.
REZULTATI
F. tularensis subsp. novicida pokrece jezgrinu translokaciju p65 podjedinice NF-B u
humanim makrofagima, no translokacija NF-B je defektna kod iglC mutante. Iako se
kaspaza-1 i kaspaza-3 pokrecu tijekom ranih stadija infekcije humanih makrofaga s F.
tularensis subsp. novicida, stanicna smrt je odgoena, što je u korelaciji sa simultanom
aktivacijom NF-B.
F. tularensis subsp. novicida razmnožava se unutar ameba u fagosomu u kojem nije došlo do
acidifikacije dok je iglC mutanta defektna za razmnožavanje u ovim dvjema amebama.
ZAKLJUCCI
Ovo je istraživanje prvi puta pokazalo simultanu modulaciju kaspaze-1, kaspaze-3 i antiapoptoticnog
jezgrinog transkripcijskog cimbenika NF-B i osjetljivu ravnotežu meu njima
kako bi se ocuvala sposobnost za život inficirane stanice.
Francisella može rasti i preživjeti unutar amebe, a iglC je potreban za unutarstanicni rast u A.
castellanii i H. vermiformis. Za razliku od stanica sisavaca gdje se bakterija razmnožava u
citoplazmi u amebama se F. tularensis subsp. novicida razmnožava u fagosomu.
Abstract (english) AIMS
To determine the role of the iglC gene in intracellular life of F. tularensis subsp. novicida
within human macrophages and amoeba through growth kinetics, modulation of apoptosis,
biogenesis of the phagosome and ultrastructural organisation of the cells after the infection.
METHODS
The cells of A. castellanii, H. vermiformis and human macrophages were infected with F.
tularensis subsp. novicida and its mutant iglC. In certain time periods after infection the cells
were processed in order to determine colony forming units by growth kinetics, to examine the
level of phagosome acidification by confocal microscopy and ultrastructural organisation of
the cell was determined by electron microscopy.
VII
Resistance to apoptosis and activation of caspase-1 and caspase-3 in human macrophages
infected with F. tularensis subsp. novicida or iglC mutant was determined with
immunoflourescence staining by confocal microscopy.
RESULTS
F. tularensis triggers nuclear translocation of the p65 subunit of NF-B in hMDMs, but
sustained nuclear translocation of NF-B is defective in the iglC mutant. Although caspase-1
and caspase-3 are triggered within F. tularensis-infected hMDMs during early stages of
infection, cell death is delayed, which is correlated with simultaneous activation of NF-B.
In vitro studies showed intracellular replication of F. tularensis subsp. novicida within A.
castellanii and H. vermiformis, but iglC mutant is defective for survival and replication in
these amoeba.
CONCLUSIONS
This study, for the first time showed simultaneous modulation of caspase-1, caspase-3 and the
anti-apoptosis nuclear transcription factor NF-B and sensitive balance among them in order
to preserve capability for survival of the infected cells.
Francisella is capable to grow and survive in amoeba, and iglC is necessary for intracellular
growth in A. castellanii and H. vermiformis. In contrast to mammalian cells where bacteria
replicate in the cytosol in amoeba F. tularensis subsp. novicida replicates within pahgosomes.
Keywords
Amebe
Apotoza
Francisella tularensis
Humani makrofagi
iglC
Keywords (english)
Amoebae
Apoptosis
Francisella tularensis
Human macrophages
iglC
Language croatian
URN:NBN urn:nbn:hr:188:705189
Study programme Title: Biomedicine Postgraduate (doctoral) study programme Study programme type: university Study level: postgraduate Academic / professional title: doktor/doktorica znanosti, područje biomedicine i zdravstvo (doktor/doktorica znanosti, područje biomedicine i zdravstvo)
Type of resource Text
Extent 125 str; 30 cm
File origin Born digital
Access conditions Closed access
Terms of use
Repository Repository of the University of Rijeka Library
Created on 2017-01-19 19:18:02