Uticaj bioaktivnih molekula laktobacila na procese autofagije i inflamacije u in vitro i in vivo sistemima
Influence of bioactive molecules of lactobacilli on autophagy and inflamation processes in in vitro and in vivo systems
Author
Dinić, Miroslav R.
Mentor
Milenković, Marina
Committee members
Lukić, JovankaĐokić, Jelena
Begović, Jelena

Stepanović-Petrović, Radica

Metadata
Show full item recordAbstract
Izolacija i karakterizacija bakterijskih biomolekula koji interaguju sa receptorima
ćelija domaćina predstavlja kljuĉ za razumevanje mehanizama probiotiĉkog dejstva
laktobacila. Savremena istraţivanja probiotiĉkih bakterija usmerena su ka identifikaciji
biomolekula koji mogu modulisati razliĉite signalne puteve u humanim ćelijama. Svi
molekuli koji su poreklom iz probiotika i odgovorni su za njihov pozitivan efekat na
zdravlje domaćina nazvani su postbiotici. Zbog slabijeg imunostimulišućeg potencijala,
njihova primena predstavlja bezbednu alternativu primeni ţivih bakterija.
Ovo istraţivanje je imalo za cilj da testira mogućnost primene postbiotika u
ublaţavanju simptoma bola i neţeljenih efekata koji nastaju kao posledica primene
analgetika. Taĉnije, definisana su dva specifiĉna cilja istraţivanja: (i) ispitati uticaj
bioaktivnih molekula odabranih sojeva laktobacila na proces autofagije u hepatocitima in
vitro, kao i protektivan efekat ovih molekula kod toksiĉnosti izazvane paraceta...molom i (ii)
ispitati potencijalni imunomodulatorni efekat egzopolisaharida (EPS-CG11) izolovanog iz
soja Lactobacillus paraplantarum BGCG11, u in vivo eksperimentalnim modelima
inflamacije. Eksperimenti u kulturi hepatocita ukljuĉili su praćenje stepena oštećenja
HepG2 ćelija izloţenih toksiĉnoj koncentraciji paracetamola i procesa autofagije, sa ciljem
identifikacije potencijalnog mehanizma delovanja postbiotika. Metodološki, vijabilnost
HepG2 ćelija analizirana je MTT i LDH esejima. Autofagija je praćena Western blot
analizom odreĊivanjem ekspresije p62/SQSTM1 proteina i akumulacijom liposolubilne
forme LC3 proteina. Dodatno, autofagija je analizirana praćenjem ekspresije BECN1, Atg5,
p62/SQSTM1 i PINK1 gena i autofagnog fluksa. Za analizu imunomodulatornog efekta
EPS-CG11 korišćena su dva in vivo modela inflamacije izazvane karageninom: model
inflamatornog bola i model peritonitisa kod pacova Wistar soja. U eksperimentu
inflamatornog bola praćeni su: vremenski tok razvoja hiperalgezije i edema šapica nakon
profilaktiĉke sistemske primene EPS-CG11, ekspresija medijatora inflamacije (IL-1β,
TNF-α, IL-6 i iNOS), infiltracija neutrofila (ekspresija MPO enzima) i aktivacija/infiltracija
monocita (ekspresija CD14 markera)...
Isolation and characterization of bacterial biomolecules involved in the interaction
with the receptors of the host cells represent the key factor for understanding the
mechanisms of probiotic action of lactobacilli. Novel studies regarding probiotic bacteria
have been focused on the identification of biomolecules which can modulate different
signaling pathways in human cells. All molecules that originate from probiotics which are
responsible for its positive effects on the host’s health are called postbiotics. Their
application represents the safe alternative to the use of live bacteria and its
immunostimulating potential.
This research aimed to test the possibility of using postbiotics in alleviation of pain
symptoms and analgesics side effects. More precisely, two main objectives of this research
were: (i) to examine the influence of bioactive molecules of selected strains of lactobacilli
on the autophagy process in the hepatocytes, in vitro, as well as protective effect of these
mo...lecules in paracetamol-induced toxicity and (ii) to examine the potential
immunomodulatory effect of exopolysaccharide (EPS-CG11) isolated from Lactobacillus
paraplantarum BGCG11 strain, in in vivo experimental models of inflammation.
Experiments in the hepatocytes culture included monitoring the degree of damage of
HepG2 cells exposed to the toxic paracetamol concentration and the autophagy process,
with the aim of identification of potential mechanism of postbiotic action.
Methodologically, the cell viability was monitored by MTT and LDH assays. Autophagy
was monitored by Western blot analysis, in order to determine the expression of the
p62/SQSTM1 protein and the accumulation of the liposoluble form of the LC3 protein.
Further, the autophagy was analyzed by monitoring the expression of BECN1, Atg5,
p62/SQSTM1 and PINK1 genes and the autophagy flux. For the analysis of the
immunomodulatory effect of EPS-CG11, two in vivo models of carrageenan-induced
inflammation were used: an inflammatory pain model and a peritonitis model in the Wistar
rats...
Faculty:
Универзитет у Београду, Фармацеутски факултетDate:
11-06-2018Projects:
- Molecular designing of nanoparticles with controlled morphological and physicochemical characteristics and functional materials based on them (RS-45004)
- Genes and molecular mechanisms promoting probiotic activity of lactic acid bacteria from Western Balkan (RS-173019)
- Examination of mechanisms of action, toxicity and interactions of adjuvant analgesics (RS-175045)