Utjecaj upale na adheziju probiotskih i patogenih bakterija na epitelne stanice kolona

master's thesis

• Title (english): The impact of inflammation on adhesion of probiotic and pathogenic bacteria to colon epithelial cells
• Author: Martina Mustač
• Mentor: Ana Savić Mlakar (mentor)
• Mentor: Silvija Černi (sumentor)
• Committee member: Silvija Černi (predsjednik povjerenstva)
• Committee member: Renata Šoštarić (član povjerenstva)
• Committee member: Jasna Lajtner (član povjerenstva)
• Committee member: Petra Peharec Štefanić (član povjerenstva)
• Granter: University of Zagreb; Faculty of Science; (Department of Biology); Zagreb
• Defense date and country: 2021-02-22, Croatia
• Scientific / art field, discipline and subdiscipline: NATURAL SCIENCES; Biology

Abstract

Narušavanjem crijevne mikrobiote smanjenjem brojnosti bakterija vrsta Lactobacillus sp. i Bifidobacterium sp., istodobnom kolonizacijom patogenim sojevima bakterije Escherichia coli te prekomjernim upalnim procesom dolazi do narušavanja integriteta crijevne barijere debelog crijeva. U ovakvim uvjetima smanjuje se ekspresija proteina zonula occludens (ZO-1), jednog od ključnih proteina interstaničnih čvrstih spojeva (proteinski spojni kompleksi koji reguliraju selektivno kretanje otopljenih tvari kroz epitel), što rezultira poremećajem integriteta crijevne barijere, a povezuje se s nastankom upalnih bolesti crijeva. Posljedično dolazi i do povećanja metabolizma sulfida što dodatno pridonosi pogoršanju imunopatološkog procesa. Cilj ovog istraţivanja bio je ispitati utjecaj probiotske bakterije (Lactobacillus reuteri) na modulaciju strukture epitelne barijere i kolonizacije enteropatogenog soja bakterije Escherichia coli na upalno kondicioniranu staničnu liniju kolorektalnog adenokarcinoma, Caco-2, koja je uzgajana u hipoksičnim uvjetima te podvrgnuta tretmanu medijem s natrijevim sulfidom i proupalnim citokinima (IFN-γ, TNF-α i IL-1β). Fluorescentno obiljeţena probiotska bakterija (L. reuteri) i enteropatogena bakterija E. coli 3014 nasaĎene su na stanice Caco-2 uz različito vrijeme inkubacije. Potom su kulturama dodana fluorescentno obiljeţena monoklonska antitijela za protein ZO-1. Analiza ekspresije proteina ZO-1 ukazala je na trend povećanja u pristunosti natrijeva sulfida, trend smanjenja u prisutnosti enteropatogene bakterije E. coli te trend povećanja u prisutnosti bakterije L. reuteri. Rezultati sugeriraju da enteropatogena bakterija E. coli 3014 negativno utječe na integritet epitelne barijere dok L. reuteri, kao probiotska bakterija, utječe na rekonstituiranje upalom narušene strukture epitela.

Abstract (english)

Disruption of the intestinal microbiota by reducing the number of bacterial species of Lactobacillus sp. and Bifidobacterium sp., simultaneous colonization by pathogenic strains of Escherichia coli and excessive inflammatory process leads to disruption of the integrity of the intestinal barrier of the colon. Under such conditions, the expression of the protein zonula occludens (ZO-1), one of the key proteins of intercellular solid compounds (protein binding complexes that regulate the selective movement of solutes through the epithelium), decreases, resulting in a disruption of the integrity of the intestinal barrier, and is associated with the development of inflammatory bowel disease. Consequently, there is an increase in sulfide metabolism, which further contributes to the deterioration of the immunopathological process. The aim of this study was to examine the influence of probiotic bacteria (Lactobacillus reuteri) on the modulation of the epithelial barrier structure and colonization of the enteropathogenic strain of Escherichia coli on the inflammatory-conditioned colorectal adenocarcinoma cell line, Caco-2, which was grown under hypoxic conditions and treated with sodium sulfide medium and proinflammatory cytokines (IFN-γ, TNF-α and IL-1β). Fluorescently labeled probiotic bacteria (L. reuteri) and enteropathogenic bacteria E. coli 3014 were implanted into Caco-2 cells at different incubation times. Fluorescently labeled monoclonal antibodies for the ZO-1 protein were then added to the cultures. Analysis of ZO-1 protein expression indicated an increasing trend in the presence of sodium sulfide, a decreasing trend in the presence of the enteropathogenic bacterium E. coli, and an increasing trend in the presence of L. reuteri. The results suggest that the enteropathogenic bacterium E. coli 3014 negatively affects the integrity of the epithelial barrier while L. reuteri, as a probiotic bacterium, affects the reconstitution of the inflammatory-damaged epithelial structure.

• Keywords: Caco-2; Lactobacillus reuteri; Escherichia coli; ZO-1; proupalni citokini; natrijev sulfid
• Keywords (english): Caco-2; Lactobacillus reuteri; Escherichia coli; ZO-1; pro-inflammatory cytokines; sodium sulfide
• Language: croatian
• URN:NBN: urn:nbn:hr:217:568928
• Study programme: Title: Experimental Biology; Study programme type: university; Study level: graduate; Academic / professional title: magistar/magistra eksperimentalne biologije (magistar/magistra eksperimentalne biologije)
• Type of resource: Text
• File origin: Born digital
• Access conditions: Open access
• Created on: 2021-02-23 13:09:21