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Gut microbiome and its changes related to therapy of chronic diseases
Hurych, Jakub ; Cinek, Ondřej (advisor) ; Kolář, Milan (referee) ; Falt, Přemysl (referee)
This work examines the response of the gut microbiome to therapeutic interventions in three longitudinal studies of chronic gastrointestinal diseases: Crohn's disease, celiac autoimmunity and irritable bowel syndrome. Multiple methods of stool microbiome analysis (especially massively parallel 16S rDNA or 18S rDNA sequencing and metagenomic sequencing) followed by bioinformatic and statistical analysis were used. In Crohn's disease, we detected a previously undescribed secondary nature of changes in the gut bacteriome after anti-TNF treatment. In celiac disease autoimmunity, where previous works described an effect of probiotic intervention on serological markers of the disease, the gut bacteriome and metabolome, we described the absence of significant changes in beneficial gut protozoa. In irritable bowel syndrome, we observed a significant response of the bacteriome after administering four doses of mixed microbiota transplantation but no response in the reduction of clinical symptoms. The results of these studies could contribute to a better understanding of the gut microbiome's role in the pathogenesis of these serious diseases. Keywords: microbiome, Crohn's disease, celiac disease, irritable bowel syndrome
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Genetic aspects of Crohn's disease
Hradský, Ondřej ; Cinek, Ondřej (advisor) ; Petřek, Martin (referee) ; Hubáček, Jaroslav (referee)
JednosmmJw.ý SOWlrn vanglitrlné Crohn'. _ oe (COl h.. been ohown ..._Ied YIiIh Ihe variant. o, NOD2, ATGI51.! ond IL23R gene. , o, Ihe IB051<>1:...... _ .. o, oIher gene> 0,_ in the irnm..... respoooe_ T"" ~cies <>f Ihe von..l> pro!oundIy <line< _ popuIoIiooo """ 10 doeo the .osoe..ted IN_ Moreo_, II!e """ <>f the IB05 Iocu. _ ClU_ gene .. de",Iopm",,' ol Crohn'. <lise.... h...... .,.,.., c_We e. omined Ihe ~. ol ,onanl> .. the N002, ATGI51.! , 1L2JR, TIIFA _ PTPl/22 gene roI V3t\aI1" in IBOS _ CTLH d"romooomol fegions wiIh ped;atric-on...' and odul-onoet co .. ..., C,ecl> """"",boo. The ge_, phenoly;>o, _ . _ Irequencies '01<= compe<ed between .59 unrelo1ed P'_ wiIh CO 000 HO on<eI3led _ y cootrols_ The strongest oooo<..tion wiIh CD >on found .. NOD2 gene (1fIret> V_I, loIowed by "'" v_nto o, 1B05 1<>1:.... (IGR206Jb_ ' , ,.65960151, _.~'" asoociotion YIiIh vorian1 .. 1L23R ond ATGlfiLl _ , _ "" ~ .osoc:i3tion,.,.. _for the p.R62OW vofi.ant .. the PTPN22 ge"" or forthe 9_2306G>A variant .. the TIIFA gene. We hove rep<lfled . ~h ~t"j ol the minor'" ol trie NODll007fs poIymorpIlism o, "e Czecl> popuIrioo _ . otrong e!leC1 ol """ alele on " e _ .1 diagnoo.ia ond .,.. Iofm ol <Iise,...,_ o..,. OILl<Iy confi'm. the mpoflonce ol IB05 in det""" .....g CO . UK<'ptibMy, ond _"'Ol,," lhal IWO indepe_'"...
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Genetics MODY diabetes
Dušátková, Petra ; Cinek, Ondřej (advisor) ; Haluzík, Martin (referee) ; Gašperíková, Daniela (referee)
The most common form of monogenic diabetes is MODY (Maturity-Onset Diabetes of the Young). It ranks among genetic defects of the β cell. It is clinically heterogenous group of disorders characterised with non insulin-dependent diabetes mellitus with autosomal dominant inheritance and age at diagnosis up to 40 years. We specified the diagnosis of MODY in more than 240 Czech families using molecular-genetic approach. The most common subtype of MODY is GCK-MODY which was proved in 376 subjects from 175 families. The risk of macrovascular complications in patiens with GCK-MODY was not detected. Additionally, we tested the ancestral origin of 4 recurrent mutations in the GCK gene causing GCK-MODY using genetic and statistical methods. We showed that three mutations (p.Glu40Lys, p.Leu315His, p.Gly318Arg) spread approximately 82-110 generations ago due to a founder effect. We also dealed with the impact of the rs560887 polymorphism on the phenotype of the patients with GCK-MODY. The genotype GG was associated with higher level of glycated haemoglobin. Analysis of patients suspect for HNF1A- or HNF4A-MODY revealed the probably first evidence of patient with HNF1A-MODY present with macrosomia and recurring ketotic hypoglycaemias in the childhood. The rare MODY genes were investigated in patients fulfilling the...
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The stool bacteriome during therapy for paediatric non-infectious diseases
Vodolánová, Lucie ; Cinek, Ondřej (advisor) ; Hrabák, Jaroslav (referee)
The intestinal microbiota is composed of up to 100 trillion microorganisms of which bacteria are overwhelming majority. The microbiota affects the development of the immune system, defence against pathogens, host nutrition, vitamin synthesis or fat storage and its composition is changing throughout life. Some studies point to an association between microbiota composition and the development of inflammatory bowel disease. One of the treatment options is anti-TNFα antibodies therapy, which uptake or antagonize the TNFα cytokine that otherwise mediates inflammation in the intestinal mucosa. The aim of the thesis was to examine how this treatment affects the composition of the intestinal bacteriome in paediatric patients with Crohn's disease, and to find specific bacterial taxa, whose abundance changes during the treatment. By inclusion of patients with juvenile idiopathic arthritis, also treated with anti-TNFα, the study aims to discern specific effects of therapeutically induced intestinal restitution (observable in patients with Crohn's disease) from general effects of anti-TNFα therapy. Stool samples from healthy children were used to determine "healthy" bacteriome. The composition of the bacteriome was studied by profiling the variable region of the V4 gene of 16S rDNA from patients stool samples...
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Role of enterovirus and adenovirus infection in the pathogenesis of celiac disease
Chudá, Kateřina ; Cinek, Ondřej (advisor) ; Kverka, Miloslav (referee)
Celiac disease, a chronic immune-mediated disorder of the small intestine, manifests in a fraction of individuals with genetic predispositions consuming gluten. Environmental factors play an essential role in its triggering. The environmental stimuli may include dietary factors, infections etc. Identification of specific triggers could help in celiac disease prevention. Our research project focused on common intestinal infections in infancy. We investigated adenoviruses and enteroviruses in stool specimens of children carrying a high-risk HLA genotype for celiac disease. We aimed to determine whether these infections are associated with early markers of celiac autoimmunity, and to identify virus genotypes. To distinguish multiple infections, massive parallel amplicon sequencing was utilized. During 2001-2007, nearly 50.000 Norwegian newborns were screened within the MIDIA study for the presence of the HLA DR3-DQ2/DR4-DQ8 genotype, which is known to significantly increase the risk of celiac disease. The risk genotype was identified in 912 babies. Up to three years of children's age, monthly stool specimens were collected and archived. Blood sampling was done every three months up to the age of a year, and then annually. Periodical questionnaires on children's thrive were collected. During 2014-2016,...
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Highly multiplexed virus detection in research of multifactorial diseases
Kunteová, Kateřina ; Cinek, Ondřej (advisor) ; Saláková, Martina (referee)
Next generation sequencing, which allows concurrent parallel sequencing of many samples and makes it possible to distinguish the infection from multiple viral types in the sample, is well suited as a detection format for such assays described below. The aim of the thesis was to develop a method that could detect all known types of human adenoviruses, human enteroviruses, and bacteriophages selected for their presence in the intestine. Using the next- generation sequencing. The first step was to design primers capable of detecting all known types of viruses, covering the area that is capable of distinguishing these viruses. This method was tested on a set of 47 human adenovirus samples and 30 human enterovirus samples of known serotype. Samples with two serotypes in different proportions were also created. After amplification of the target genome, the samples were purified and sequenced on MiSeq, Illumina. The method was further used in the typing of adenoviruses, enteroviruses and bacteriophages in pre-diabetic cohorts of DIPP, MIDIA, and a cohort of diabetics from African and Asian countries. The tested sample was RNA / DNA isolated from the stool specimen. We have demonstrated that the method is capable to detect all tested virus types, including infections with two different types, even if the...
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Viruses in the pathogenesis of coeliac disease
Chudá, Kateřina ; Cinek, Ondřej (advisor) ; Šroller, Vojtěch (referee)
Celiac disease is a chronic inflammatory disorder affecting the small bowel. It develops in genetically susceptible individuals upon yet unknown environmental stimuli. Environmental triggers such as infections, dietary change or other "hits" are clearly required for disease development, as only a tiny fraction of genetically susceptible subjects develops celiac disease upon gluten exposure. This thesis aims to summarize the current evidence on viruses in the pathogenesis of celiac disease regarding their relevance in population or their involvement in immune processes leading to celiac disease. Rotavirus, orthoreovirus, adenovirus, astrovirus, respiratory syncytial virus, hepatitis viruses and herpesviruses are discussed. In addition, prospective cohort studies are presented that investigate environmental triggers of type 1 diabetes and celiac disease, two diseases sharing genetic predispositions. Keywords: celiac disease, orthoreovirus, rotavirus, adenovirus, astrovirus, respiratory syncytial virus, hepatitis C virus, hepatitis B virus, prospective cohort study
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