Seznam témat doktorských projektů, nabízených školiteli pro akademický rok 2024/2025

OR Imunologie 

Vypsané doktorské práce pro akademický rok 2024/2025 

The function of a new interacting partner of ORMDL3, LAIR-1, in mast cell physiology

RNDr. Petr Dráber, DrSc. 

ID 277388 

The human and mouse ORMDL3 genes encode proteins comprised of 153 amino acids. Genetic studies have linked ORMDL3 with asthma and inflammatory diseases [1]. Asthma and associated allergies stem from the coordinated release of cytokines, chemokines, lipid mediators, proteolytic enzymes, and vasoactive mediators originating from activated tissue mast cells during degranulation  [2] . Our recent studies found that various antigen-mediated signaling pathways are affected by the  activity of the serine-palmitoyl transferase (SPT), the activity of which is inhibited by interaction with  ORMDL3 [3- 6] . Our unpublished immunoprecipitation and mass spectrometry data showed that  ORMDL3 binds several proteins, including LAIR-1 (CD305), and that this binding affects the inhibitory  activity of ORMDL3. In this project, we will investigate the effect of ORMDL3 interacting partner LAIR 1 in mast cell signaling and asthma-related inflammatory diseases. We will cross ORMDL3 knockout  (KO) mice, generated in our laboratory and maintained on the C57BL/6 genetic background, with  LAIR-1 KO mice to generate ORMDL3/LAIR-1 double KO mice and determine airway  hyperresponsiveness to ovalbumin – induced inflammation and antigen-induced allergies in vivo.  Based on previous findings that collagen binds to LAIR-1 [7], we will prepare new bispecific tools  formed by TNP-modified collagen. We propose that the TNP-modified collagen will bind to both LAIR 1 (via collagen) and TNP-specific-IgE-receptor complexes to inhibit the FcεRI signaling. We aim to  prepare constructs that will inhibit FcεRI signaling by crosslinkíng LAIR 1 with FcεRI complexes under  in vivo and in vitro conditions. The project will use various methods, including high-resolution  microscopy, lentiviral transduction, mass spectrometry, immunochemistry, and transcriptomics. At  least three publications in high-impact journals will be published based on the data from this project. 

References 

1. Paulenda, T.; Draber, P. The role of ORMDL proteins, guardians of cellular sphingolipids, in asthma. Allergy 2016, 71, 918-930. 
2. James, B.; Milstien, S.; Spiegel, S. ORMDL3 and allergic asthma: From physiology to pathology. J. Allergy Clin. Immunol. 2019, 144, 634-640. 
3. Bugajev, V.; Halova, I.; Draberova, L.; Bambouskova, M.; Potuckova, L.; Draberova, H.; Paulenda, T.; Junyent, S.; Draber, P. Negative regulatory roles of ORMDL3 in the FcεRI-triggered expression of proinflammatory mediators and chemotactic response in murine mast cells. Cell Mol. Life. Sci. 2016, 73, 1265-1285. 
4. Bugajev, V.; Halova, I.; Demkova, L.; Cernohouzova, S.; Vavrova, P.; Mrkacek, M.; Utekal, P.; Draberova, L.; Kuchar, L.; Schuster, B. et al. ORMDL2 deficiency potentiates the ORMDL3- dependent changes in mast cell signaling. Front Immunol. 2020, 11, 591975. 
5. Bugajev, V.; Paulenda, T.; Utekal, P.; Mrkacek, M.; Halova, I.; Kuchar, L.; Kuda, O.; Vavrova, P.; Schuster, B.; Fuentes-Liso, S. et al. Crosstalk between ORMDL3, serine palmitoyltransferase, and 5-lipoxygenase in the sphingolipid and eicosanoid metabolic pathways. J. Lipid. Res. 2021, 62,
6. Demkova, L.; Bugajev, V.; Utekal, P.; Kuchar, L.; Schuster, B.; Draber, P.; Halova, I. Simultaneous reduction of all ORMDL proteins decreases the threshold of mast cell activation. Sci. Rep. 2023, 13, 9615. 
7. Borst, R.; Meyaard, L.; Pascoal Ramos, M.I. Understanding the matrix: collagen modifications in tumors and their implications for immunotherapy. J. Transl. Med. 2024, 22, 382.

Effect of gluten-free diet (GFD) on immune parameters, microbiome and metabolome in  inflammatory bowel disease and primary sclerosing cholangitis 

MUDr. David Funda, Ph.D. 

ID 246642 

Idiopathic inflammatory bowel diseases (IBD) are chronic, relapsing, immune-mediated inflammatory  disorders of the gastrointestinal tract. Primary sclerosing cholangitis (PSC) is a progressive disease of  the bile ducts, which in advanced stages of the disease represents one of the most common  indications for orthotopic liver transplantation in developed countries. While genetic factors  contribute to the development of these diseases, environmental factors (such as diets) and  subsequent changes in the microbiome likely have a significant impact on the etiopathogenesis of  these diseases. PSC is associated with IBD in approximately 70 % of patients. Recently, independent  evidence has emerged regarding the role of chronic intestinal inflammation in a wide range of  immune-mediated diseases. Gluten and its components have a pro-inflammatory effect similar to  lipopolysaccharide. Beneficial effects of a gluten-free diet (GFD) have already been demonstrated in  several autoimmune diseases (type 1 diabetes, certain neurological disorders). 

The aim of this project is to elucidate the mechanisms of interaction between a gluten-free diet, the  microbiome, and the metabolome, and to study the impact of GFD on mucosal and innate immunity  in IBD and PSC. The project is based on both human randomized clinical trials (the effect of a gluten free diet on clinical symptoms of PSC and IBD; ClinicalTrials.gov Identifier: NCT06026449) and on  mouse-induced models of IBD (DSS-induced colitis model in BALB/c mice) and PSC (DSS-induced PSC  model in Mdr2-/- KO mice). 

Mechanistic studies include the analysis of immune parameters, changes in metabolomes, and  changes in bacteriomes in relation to a gluten-free diet. Methods used in this project include  isolation of peripheral blood leukocytes from patients and cells from lymphoid tissues in mice, flow  cytometry, histology, immunohistochemistry, tissue culture, ELISA, metabolome characterization (in  collaboration with the Laboratory for Molecular Structure Characterization), sample preparation for  bacteriome sequencing, data analysis, and eventually also single cell transcriptomics. The project  involves international collaborations.

Intestinal barrier disruption as a consequence of modulation of Th17 pathway in obese psoriatic  patients: mechanisms of action and microbiota involvement 

RNDr. Zuzana Jirásková Zákostelská, Ph.D. 

ID 275513 

prezenční 

Goals: Our aim is to determine whether the specific composition of the intestinal microbiota of  psoriasis patients can influence the intestinal mucosal immune system and the Th17 immune  response in the pathogenesis of psoriasis. 

Intensive efforts are being made to determine what triggers intestinal barrier dysfunction in patients  with psoriasis and whether the composition of the gut microbiota plays a role in this process.  Another unknown is how the gut-skin axis influences the pathogenesis of the disease. In addition, we  do not know why some patients with psoriasis do not respond appropriately to therapy or in whom  the adverse effects are seen. It could be connected to the role of IL-17 in maintenance of intestinal  barrier integrity on one hand and its role in intestinal inflammation during pathogenesis of psoriasis  on the other. For all these reasons, we would like to try to achieve our proposed goals using unique  gnotobiological techniques, genetically modified mice, high-throughput sequencing and  transcriptomics. 

1) To elucidate the impact of patients’ microbiota in mouse models of psoriasis and by colonization  of mice with samples collected from patients with or without an attenuated Th17 immune response  (using gnotobiotic models). 

2) To determine the consequences of Th17 pathway attenuation on immune system regulation and  intestinal barrier integrity in experimental mouse models of psoriasis using knockout mice or IL-17  inhibitors. 

3) To characterize cellular pathways triggered by specific bacterial consortia isolated from the  patients with psoriasis prior and during anti Th17 treatment by in vitro experiments in human  primary cells and cell lines 

To evaluate the role of obesity in disease pathogenesis using spatial transcriptomics and unique  gnotobiotic techniques.

Primary microbiota and immunocompromised host 

doc. Ing. Bc. Igor Šplíchal, CSc. 

ID 246727 

Multicellular hosts are settled with simple, often unicellular, organisms called microorganisms, and  their pool is called microbiota. The microbiota is accumulated mainly in the lower gastrointestinal  tract and “communicates” with hosts on the epithelial cell interface and through its metabolites. The  primary bacterial colonization of the newborns and the establishment of a balanced microbiota are  essential for the next life of the host. Immunocompromised hosts, e.g., preterm infants and elders,  are sensitive to the disbalance of microbiota (dysbiosis) that facilitates infection with pathogens and  overgrowth of pathobionts. 

During the doctoral study, the student will work with gnotobiotic piglets, which will be associated  with a defined microbiota. Signs of enterocolitis (fever, anorexia, diarrhea), intestinal barrier (mucin,  claudin-1, occludin, villin) and histology of the intestine, pattern recognition receptors (TLR and NLR)  signaling, and inflammatory/sepsis markers (interleukins 1, 6, 8, 10, 12, 18, TNF-alpha, IFN-gamma,  and HMGB1) will be evaluated to monitor the host response to bacterial colonization. 

The doctoral thesis will be prepared at the Institute of Microbiology of the Czech Acad Sci in Nový  Hrádek, and the doctoral student will collaborate on a grant from the MEYS. 

The student will use Real-Time PCR (RNA isolation, cDNA synthesis, qPCR), Western blot, histology,  immunohistochemistry, hemocytometry, and xMAP technology (Luminex) in his/her work.

Cancer Immunotherapy Based on Drug Delivery Systems for IDO Inhibitors in Combination with  Chemotherapy 

Mgr. David Větvička, Ph.D. 

ID 277385 

The aim of this dissertation is to explore the potential and benefits of using targeted drug delivery  systems for the administration of IDO inhibitors in combination with chemotherapy, with a focus on  enhancing the efficacy of immunotherapy in the treatment of cancer. The work will focus on the  development and optimization of IDO inhibitor delivery using modern technologies, such as  nanoparticles and other carriers, and will assess their effectiveness in combination with conventional  anticancer therapies, particularly chemotherapy and immune checkpoint blockade.