Book of Abstracts - CarDia National Institute Annual Meeting

The project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. edited by Dobromila Kozlerová ISBN: 978-80-908332-9-6 Published by: Institut klinické a experimentální medicíny, Vídeňská 1958/9, 140 21 Praha 4 2 Dear colleagues, it is my great pleasure to welcome you to the CarDia National Institute Annual Meeting. Over the past years, the collaboration of the five participating institutions has created a strong foundation for excellent research, innovation, and mutual learning. Although this conference marks the conclusion of the implementation period, it does not mark the end of our cooperation. In the upcoming three years (2026–2028), the sustainability phase will ensure that the established partnership continues to thrive and further develop its scientific and practical impact. As this is our concluding meeting, all Work Packages (WP1–WP7) will present a comprehensive summary of their activities and achievements. In addition, each WP will also provide a perspective on the sustainability phase, outlining the main research themes to be further developed and the forms of collaboration planned within the consortium. We hope that this meeting will allow us not only to reflect on what has been achieved so far, but also to strengthen our commitment to continuing our joint research and fostering innovations that will benefit science, healthcare and society in the long term in the field of metabolic and cardiovascular diseases. I wish you inspiring discussions and a successful meeting. Martin Haluzík, Principal Investigator of the CarDia National Institute project 3 CONTENT WP1 Prevention of obesity, diabetes and cardiovascular complications ............................................. 4 WP2 New treatment options for diabetes and obesity and their complications ................................. 9 WP3 Complex analytical methods and bioinformatics ....................................................................... 15 WP4 Heart rhythm disorders and their relationship to metabolic diseases ...................................... 18 WP5 Heart failure due to obesity and diabetes .................................................................................. 21 WP 6 Early atherosclerosis and atherothrombosis as its acute reversal ............................................ 25 WP 7 Influencing acute and chronic complications of diabetes by organ and tissue transplantation ..................................................................................................................................... 29 Posters section ...................................................................................................................................... 34 4 WP1 Prevention of obesity, diabetes and cardiovascular complications Work Package 1 Name of the lecture: Adiposity, Age, or Acute Exercise: What does affect the FAHFA Levels in Healthy Women? Name of the group: Laboratory of Physiology and Pathophysiology of Adipose Tissue, Department of Pathophysiology, Third Faculty of Medicine, Charles University, Prague Principal Investigator: Lenka Rossmeislová, Ph.D. Author(s): Lenka Rossmeislová, Viktor Šebo, Jan Gojda, Michal Koc, Marek Wilhelm, Martin Riečan, Tomáš Čajka, Ezequiel Harnichar, Jana Neubert, Michaela Šiklová, Martin Rossmeisl Abstract: Adipose tissue (AT) is a source of insulin-sensitizing lipokines FAHFAs. While exercise training is known to increase FAHFA levels in elderly women, it is unclear how a single bout of exercise affects FAHFA levels in women of different ages and adiposity, and what physiological pathways are involved. We addressed these questions by combining a clinical trial with in vitro experiments on human adipocytes. Sixty healthy women were divided into four groups: young lean (BMI 21,5; %FM 28, n=20) vs young obese (BMI 34,1; %FM 45,2, n=20), and young (age 29,7; %FM 37.7) vs. older (age 72,2; %FM 37.3, n=20). All participants underwent clinical assessments, including body composition analysis, an acute bout of exercise (50-60% VO2max), and a non-exercise control intervention. Insulin sensitivity and circulating FAHFA levels were measured throughout the study, while AT FAHFA was measured at baseline. We found that both lower adiposity and higher age were associated with higher total AT-FAHFA levels (lean: 113.0 ±74.2 vs obese: 69.0 ± 22.1 pmol/g, young: 87.0 ±49.0 vs. older 122.9 ± 65.5 pmol/g). Thus, cardiovascular fitness level was positively correlated with IS indices and total AT-FAHFA only when older women were excluded from the calculation. In plasma, total FAHFA levels were similar across all groups, with only a small fraction of regioisomers affected by adiposity and none by age. Acute exercise did not increase circulating FAHFA levels and, in fact, led to a decrease in some regioisomers, an effect that was also seen in the control group. In human adipocytes, FAHFA levels were modulated by IBMX, but not dexamethasone, although both treatments increased expression of PRDX6 and ATGL, i.e genes implicated in FAHFA synthesis. IBMX treatment selectively upregulated several β oxidation genes and FAHFA content strongly correlated with PGC1 expression. Summary: Both, the adiposity and age appear to be important regulators of FAHFA content in AT but not in circulation, which questions whether subcutaneous AT is the primary source of circulating FAHFA in women. An acute bout of exercise was not sufficient to induce any rise of FAHFA in circulation above the level observed in non-exercising women, suggesting that chronic training, rather than single sessions, is required for this effect. Stimulation of FAHFA levels in adipocytes in vitro by IBMX suggest that FAHFA synthesis is dependent on lipolysis combined with increased β oxidation. What drives higher AT FAHFA levels in older women remains to be investigated. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------------------- 5 Work Package 1 Name of the lecture: Integrative metabolomic and genetic mapping for identification of genes regulating liver metabolism and blood pressure in the HXB/BXH rat panel Name of the group: Laboratory Genetics of Model Diseases, Institute of Physiology, CAS, Praha Principal Investigator: Ing. Michal Pravenec, DrSc. Author(s): Ing. Michal Pravenec, DrSc. Abstract: The spontaneously hypertensive rat (SHR) exhibits several features of the metabolic syndrome. To facilitate the genetic analysis of complex traits in the SHR, we developed a panel of recombinant inbred (RI) strains derived from SHR and Brown Norway progenitors, the HXB/BXH RI strains (N=30). In this study, we performed LC-MS-based metabolomic profiling of liver samples, detecting a total of 761 metabolites. Using GeneNetwork, we identified over 50 significant metabolomic quantitative trait loci (mQTL) (LOD score ˃4). Several illustrative examples highlight the utility of this approach: (1) A significant mQTL on chromosome 11 (LOD score 7.5) associated with hepatic proline concentration correlates with hepatic expression of Prodh1 (r=-0.82, p=3.55e-09). A Prodh1 cis-eQTL overlaps this mQTL, supporting its role in proline catabolism. (2) Hepatic levels of N-alpha-acetylarginine are regulated by an mQTL on chromosome 4 (LOD score 8.5) that colocalizes with a delta L-NAME blood pressure QTL and a cis-eQTL for Nat8f1, a candidate gene involved in arginine acetylation. This highlights a potential link between N-alpha-acetylarginine levels and blood pressure regulation. (3) Hepatic concentrations of several lipid species are linked to an mQTL near the Cd36 gene on chromosome 4. The Cd36 expression is cis-regulated, and the mQTL overlaps with lipolysis QTL, suggesting a mechanistic role for Cd36 in hepatic lipid metabolism. (4) Hepatic mQTLs on chromosome 3, associated with phenylacetylglutamine, phenaceturic acid, and phenylglucuronic acid, microbial-derived metabolites, colocalize with a QTL for epididymal fat weight, implicating the gut-liver axis in adiposity regulation. Summary: Integrative genetic and metabolomic analysis in HXB/BXH RI strains identified key genes and pathways linking liver metabolism, gut microbial metabolites, and cardiometabolic traits. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------------------- Work Package 1 Name of the lecture: Pancreatic β-Cell Redox Signaling: A Double-Edged Sword Name of the group: Laboratory of Pancreatic Islet Research, IPHYS, CAS, Prague Principal Investigator: RNDr. Lydie Plecitá-Hlavatá, Ph.D. Author(s): Blanka Holendová, Štěpánka Benáková, Monika Křivonosková, Soňa Štemberková-Hubáčková, Linda Stokičová, Lydie Plecitá-Hlavatá Abstract: Redox regulation is a central determinant of pancreatic β-cell physiology, linking nutrient metabolism to insulin secretion and systemic glucose control. β-cells are highly sensitive to metabolic 6 fluctuations, with reactive oxygen species (ROS) functioning both as signaling mediators and potential stressors. We show that glucose stimulation induces hydrogen peroxide production via NADPH oxidase 4 (NOX4), which modulates key metabolic proteins and serves as an essential coupling signal for insulin release. However, persistent NOX4 activity under nutrient excess shifts redox balance toward prooxidative signaling, promoting local and systemic inflammation characteristic of type 2 diabetes. Paradoxically, longterm β-cell–specific NOX4 inactivation accelerates the presence of senescent cells in islets of mice, indicating a complex role for redox signaling in disease progression. These findings underscore the dual nature of ROS as both regulators and disruptors of β-cell homeostasis, and highlight redox pathways as potential but challenging targets for therapeutic intervention in diabetes. These should be time-limited and selectively targeted. Summary: NOX4-derived ROS are essential for insulin secretion but, when persistent, drive oxidative stress and inflammation in diabetes. Long-term NOX4 inactivation also promotes β-cell senescence, highlighting the double-edged role of redox signaling in β-cell life. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ------------------------- Work Package 1 Name of the lecture: The associations of maternal liver and lipid biomarkers in pregnancy with gestational diabetes and adverse pregnancy outcomes Name of the group: Dept. of Pathophysiology, FoM Masaryk University, Brno Principal Investigator: prof. MUDr. Kateřina Kaňková, Ph.D. Author(s): Vendula Bartáková, Lukáš Pácal, Eliška Boženková, Petr Žák, Veronika Ťápalová, Petr Janků, Kateřina Kaňková Abstract: Some studies (incl. our pilot results) suggest that women with GDM have a higher prevalence of metabolic syndrome compared to healthy pregnant women. The liver – namely metabolic dysfunction-associated liver disease (MAFLD) - plays a central role in orchestrating metabolic changes associated with metabolic syndrome. The aim of this study was (i) to investigate differences in lipids and liver enzyme levels (LEL) between pregnant women with and without GDM, (ii) to examine the correlation of lipids and LEL during pregnancy with the risk of conversion of GDM to persistent pre-/diabetes after delivery and (iii) to find a possible correlation with adverse peripartal events. The study included 109 healthy pregnant women and 163 GDM patients. In 77% birth data were available. Repeated oGTT after delivery underwent 42% (n = 68) GDM patients and in 13% (n = 9) the conversion to PGI was confirmed. Parameters: Age at the time of dg. GDM, pre-gestational BMI, lipid profile at dg. GDM (2nd trim.) and before delivery, liver enzymes at the time of dg. GDM, glycemia during oGTT test, delivery data. Patients with GDM had significantly lower values of total cholesterol (TC), LDL and HDL before birth compared to controls, yet, the parameters did not differ in the 2nd trimester (at the time of GDM dg.). Lipid values in the GDM group of PGI patients did not differ from the rest of the GDM patients. Pregestational BMI negatively correlated with HDL in all three samples, the fetal weight positively correlated with LDL and TC in the 2nd trimester. Patients with induced labour had significantly higher TAG and lower HDL values before birth, the HDL was lower in the 2nd trimester. With the exception of ALP (higher in GDM group, P=0.01, Man Whitney), no other differences between groups or correlations were found in LEL. 7 Summary: Lipid spectrum values in patients with and without GDM did not differ significantly during the pregnancy. Patients with GDM had - hypothetically due to the effective dietary education and subsequent lifestyle changes - lower LDL and TC compared to healthy women before delivery. LEL were also similar in both groups, only ALP in GDM patient was higher during pregnancy, but it has no influence on peripartal outcomes or PGI conversion. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------- Work Package 1 Name of the lecture: Omics signatures of plant-based diets and their potential to predict remission of diabetes Name of the group: CRU, Department of Internal Medicine, Third Faculty of Medicine, Charles University, Prague Principal Investigator: Jan Gojda, doc., MUDr., Ph.D. Author(s): Jan Gojda, Anna Ouřadová, Eliška Selinger, Marina Heniková, Filip Tichánek, Petra Polakovičová, Pavel Dlouhý, Dana Hrnčířová, Marek Kuzma, Martin Světnička, Eva El-Lababidi, Monika Cahová Abstract: Dietary choices influence both human health and the sustainability of planetary systems. Global food production is a major driver of greenhouse gas emissions, land degradation, and biodiversity loss, and international policy agendas emphasize transitions toward more plant-based diets to mitigate these impacts. Yet such transitions raise questions about nutrient adequacy, inter-individual variability in metabolic responses, and the need for evidence-based frameworks to guide public health and policy. At the same time, the burden of the metabolic disease is on an unprecedented rise globally. First, in a cross-sectional study of 95 Czech families (47 vegan, 23 vegetarian, 25 omnivore; 187 adults and 142 children), we compared growth, cardiovascular health, bone turnover, iodine, and micronutrient status. Vegans exhibited the most favorable cardiometabolic profiles (lower LDL and total cholesterol), with no significant differences in children’s growth across dietary groups. Bone turnover indices were comparable, vitamin D was highest and urinary phosphate lowest among vegans, and lower urinary iodine in vegan children was not associated with altered thyroid function. Familial clustering of nutritional biomarkers highlighted shared dietary and environmental influences. These findings suggest that plant-based diets can support healthy growth when critical nutrients (iodine, vitamin B12) are monitored. Second, in a case-control study of 147 participants—49 with pre/diabetes (DM), 66 overweight/obese (OB), and 32 lean healthy (LH)—we integrated dietary intake, gut microbiota, and fecal/serum metabolome (MIME) profiles. MIME composition differed significantly across metabolic spectra: DM and LH represented opposite extremes, with OB closer to DM. In a subsequent 3-month intervention in 27 DM patients receiving dietary inulin (10 g/day), glycemic indices improved overall, but responses were highly variable. Favorable outcomes were associated with baseline glycemic status, gut microbial taxa (e.g., Blautia, Eubacterium halii group, Lachnoclostridium, Ruminiclostridium, Dialister, Phascolarctobacterium), serum branched-chain amino acid derivatives, asparagine, and fecal indole and other volatile compounds. These results indicate that obesity is a stronger determinant of MIME variation than glycemic status, and that baseline MIME signatures can predict responsiveness to dietary interventions. 8 Together, these studies illustrate the dual imperative of planetary health: population-level dietary shifts toward plant-based eating patterns to reduce environmental burden, and precision-nutrition strategies that leverage microbiome and metabolome profiling to tailor interventions for individuals at risk of metabolic disease. Integrating both approaches into policy and practice—through school catering reform, targeted fortification, and microbiome-guided dietary personalization—offers a pathway to healthier populations and a more sustainable food system. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. 9 WP2 New treatment options for diabetes and obesity and their complications Work Package 2 Name of the lecture: Bone marrow adipose tissue: a hidden player in health and disease Name of the group: Molecular Physiology of Bone, Institute of Physiology, CAS, Prague Principal Investigator: Mgr. Michaela Tencerova, Ph.D. Author(s): Mgr. Michaela Tencerova, Ph.D. Abstract: Obesity, type 2 diabetes, and osteoporosis are metabolic diseases often accompanied by increased bone marrow adipose tissue (BMAT), which is closely linked to complications in both fat and bone metabolism. While obesity management strategies—particularly anti-diabetic agents and dietary interventions— generally improve systemic metabolic parameters, they can also exert adverse effects on bone health. Our research has identified unique characteristics of BMAT under obesogenic conditions, which appear to be differentially regulated across mouse strains with varying susceptibility to obesity, as well as in transgenic models with targeted inactivation of metabolic enzymes relevant to bone and fat biology. In this presentation, I will highlight our findings on BMAT phenotypic alterations in diverse mouse models, the influence of sex dimorphism on bone and metabolic phenotypes in transgenic models, and the resulting impact on the bone marrow microenvironment and stem cell function in metabolic disease. I will also present the development of novel models of bone impairment within the framework of the Cardia Project. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------------------- Work Package 2 Name of the lecture: COX6B1 secures a redox-sensitive step in early cytochrome c oxidase assembly Kristýna Čunátová1,2,3, Marek Vrbacký1, Michal Knězů1, Alena Pecinová1, Lukáš Alán1,4, Josef Houštěk1, Erika Fernández-Vizarra2,3,5, Petr Pecina1, Tomáš Mráček1 1 Laboratory of Bioenergetics, Institute of Physiology, Czech Academy of Sciences, Prague, 14200, Czech Republic 2 Department of Biomedical Sciences, University of Padova, Padova, 35131, Italy 3 Veneto Institute of Molecular Medicine, Padova, 35129, Italy 4 Department of Biology, University of Padova, Padova, 35131, Italy 5 Department of Biochemistry and Molecular and Cellular Biology, University of Zaragoza, Huesca, 22002, Spain COX6B1 is a nuclear-encoded subunit of the human mitochondrial cytochrome c oxidase (cIV) located in its intermembrane space-facing region. The relevance of COX6B1 in mitochondrial 16 identification and amyloid protein typing in patient tissue samples. While progressively introduced into practice in the last years, specific therapies are available only for a fraction of amyloidoses, leaving a number of patients (predominantly those with rare or ultra-rare types) only with symptomatic treatment options. We have recently documented clinical, laboratory and tissue pathologies in a four-generation family affected by hereditary Amyloid A (AA) amyloidosis lacking any underlying inflammatory condition. By combined use of genomic, proteomic and functional studies, we managed to link this phenotype to a specific SAA1 gene promotor variant that triggers increased circulating levels of the amyloidogenic SAA1 protein. Important, anti-interleukin 6 receptor treatment by monoclonal antibody (tocilizumab) proved beneficial in the affected family members. In a second family with primary hereditary AA amyloidosis a pathogenic structural variant triggering SAA1 aggregation was detected. N-of-1 clinical trial using anti-sense oligo therapy has been recently conducted by our collaborator at Mayo Clinic (Rochester, MN, USA). Last, we identified APOA4 pathogenic variants as the cause of autosomal dominant tubulointerstitial kidney disease with renal medullary amyloidosis in several families. Only sporadic (non-hereditary) occurrence of ApoA4 amyloid has been known prior our report. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. -------------------------------------- Work Package 3 Name of the lecture: α-Galactosidase A Misprocessing-Induced Unfolded Protein Response: Lysosomal Storage-Independent Mechanism of Fabry Disease Pathogenesis Name of the group: Research Unit for Rare Diseases, First Faculty of Medicine, Charles University in Prague Principal Investigator: prof. Ing. Stanislav Kmoch, CSc. Author(s): Ing. Klára Svojšová, Mgr. Martina Živná, Ph.D., prof. MUDr. Aleš Linhart, Dr.Sc., MUDr. Jakub Sikora, Ph.D., prof. Ing. Stanislav Kmoch, CSc. Abstract: Classic Fabry disease (FD) is caused by GLA mutations that result in enzymatic deficiency of α-galactosidase A, lysosomal storage of globotriaosylceramide (lyso-Gb3Cer), and a resulting multisystemic disease. In nonclassic FD, patients have some preserved α-galactosidase A activity and a milder disease course, though heterozygous female carriers may also be affected. While FD pathogenesis has been mostly attributed to catalytic deficiency of mutated α-galactosidase A, lysosomal storage, and impairment of lysosomal functions, other pathogenic factors may contribute, especially in non-classic FD. We characterized the clinical, biochemical, genetic, molecular, cellular, and organ pathology correlates of the p.L394P α-galactosidase A variant that was identified initially in six individuals with kidney failure by the Czech national screening program for FD and by further screening in an additional 24 family members. Clinical findings in affected male patients revealed a milder clinical course, with approximately 15% residual α-galactosidase A activity with normal plasma lyso-Gb3Cer levels and abnormally low ratio of these values. Kidney biopsies did not show lysosomal storage. Laboratory investigations documented intracellular retention of mutated α-galactosidase A with resulting endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), which were alleviated with BRD4780, a small molecule clearing misfolded proteins from the early secretory compartment. We observed similar findings of ER stress and UPR with several other classic and non-classic FD missense GLA variants. Summary: 17 We identified defective proteostasis of mutated α-galactosidase A resulting in chronic ER stress and UPR of α-galactosidase A expressing cells as a contributor to FD disease pathogenesis. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU; MULTIOMICS_CZ (ID Project CZ.02.01.01/00/23_020/0008540) funded by the European Union - Next Generation EU from the Ministry of Education, Youth and Sports of the Czech Republic; grant from the Ministry of Education, Youth and Sports of the Czech Republic - LUAUS24087; Institutional programmes: LM2023067 and UNCE/24/MED/022 and SVV 260631 ------------------- Work Package 3 Name of the lecture: Rapid integrative diagnostics in cardiogenetics Name of the group: Research Group of Inherited Disorders, KPDPM, 1. LF UK Principal Investigator: Prof. Ing. Stanislav Kmoch, CSc. Author: Ing. Lenka Piherová, Ph.D. Abstract: Inherited cardiovascular diseases are often difficult to recognize, and delayed diagnosis may have severe consequences. The use of modern scientific methods enables rapid integration of clinical evaluation with molecular-genetic and protein-based analyses, significantly accelerating the diagnostic process. By combining clinical assessment, advanced imaging, endomyocardial biopsy, and state-of-the-art genetic technologies such as WES and RNASeq, an accurate diagnosis can be established within a short timeframe. Parallel analysis at both protein and DNA levels further supports causal variant identification and helps to differentiate from clinically similar conditions. Summary: This approach represents a robust and integrated diagnostic algorithm that translates scientific knowledge into clinical practice within days. It allows for the rapid identification of rare diseases and timely implementation of therapeutic measures, thereby minimizing the risk of sudden cardiac death and improving patient outcomes. This concept will be illustrated by two cases: rapid diagnosis of Danon disease and reclassification of a variant of uncertain significance (VUS) in MYBPC3 to a pathogenic variant. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. 18 WP4 Heart rhythm disorders and their relationship to metabolic diseases Work Package 4 Name of the lecture: Individualization of CRT treatment in heart failure patients Name of the group: Arrhytmology Department, 3rd Medical Faculty, Charles University in Prague Principal Investigator: doc. MUDr. Ing. Karol Čurila, PhD. Author(s): Karol Curila Introduction: The standard treatment for patients with heart failure and indications for CRT is biventricular pacing (BVP). Thirty to forty patients do not respond adequately to this treatment, and therefore new ways of optimally selecting patients for CRT and CRT treatment methods that would yield better results are being sought. Objective: The aim of our research was to determine whether non-invasive assessment of ventricular dyssynchrony using UHF-ECG can provide a better understanding of the differences in ventricular synchrony between BVP and new CRT methods such as left bundle branch pacing (LBBP) and left ventricular septal pacing (LVSP). Another objective was to determine whether UHF-ECG could be used to better identify the site of conduction block in the left ventricle, thereby identifying patients for whom LBBP may be the first-line treatment and who would benefit most from CRT. Methodology: We compared the effects of different CRT methods on ventricular synchrony and LV efficiency using UHFECG and a special hemodynamic protocol in 35 patients with heart failure. We investigated whether UHFECG is better than standard ECG at detecting the site of left ventricular conduction disturbance in 80 patients with an indication for CRT. Results: We demonstrated that all three methods of resynchronization therapy (BVP, LVSP, and LBBP) lead to excellent resynchronization of the left ventricle and increase its efficiency. We also found that BVP and LVSP are completely comparable, while LBBP leads to both better ventricular synchrony and better LVEF. We also found that UHF-ECG is much more accurate in detecting left ventricular conduction block than standard RKG (97% vs. 76%). Summary: The results of our research have significantly contributed to our understanding of how to individualize treatment for patients with heart failure and indications for resynchronization therapy. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------- 19 Work Package 4 Name of the lecture: Arrhythmogenic substrate in rat models of heart failure Name of the group: Developmental Cardiology, IPHYS; Anatomy, LF1, CUNI; Cardiology, IKEM Principal Investigator: Prof. MUDr. Frantisek Kolar, Ph.D. Author(s): Prof. MUDr. David Sedmera, DSc. Abstract: Heart failure is associated with an increased risk of arrhythmic sudden death, especially if associated with other co-morbidities such as diabetes or hypertension. Our goal is to explore various rat models of heart failure, optically map their conduction properties and correlate those with morphological substrates of arrhythmias, such as fibrosis or gap junctional remodeling. During the project period, several different models were analyzed in detail. Some data are complete in form of manuscripts under review (SUGEN-hypoxia model of pulmonary hypertension with right ventricular failure), some are near being accepted for publication (neonatal model of left ventricular pressure overload), while others are still being analyzed and interpreted (e.g., double-hit model of high-fat diet and L-NAME administration). Summary: No rat model fully recapitulates all the aspects of particular human conditions, such as HFpEF; this is likely due to physiological differences between rodents and primates. However, carefully selected models are able to reproduce particular human symptoms, and are useful for testing potential therapeutic interventions. Support of the CarDia project enabled development of various experimental models that will be useful in the upcoming years to study combinations of risk factors leading to, or worsening, the heart failure in humans. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. -------------------- Work Package 4 Name of the lecture: Diabetic Cardiomyopathy as a Stem and Progenitor Cell Disorder Name of the group: - Stem Cell Disease Modeling (SCDM) group, FNUSA ICRC and MUNI, Brno, Principal Investigator: Mgr. Vladimír Rotrekl, Ph.D. Author(s): MUDr. Martin Pešl, Ph.D., Mgr. Deborah Beckerová, Mgr. Šimon Vrana Klimovič Ph.D., Mgr. Jan Přibyl, Ph. D. and Mgr. Vladimír Rotrekl, Ph.D. Abstract: Hyperglycemia increased ROS production in stem cells, activated PI3K/AKT signaling, induced DNA damage, and drove premature differentiation. This led to fibrotic remodeling and loss of progenitor populations, explaining mechanisms of diabetic cardiomyopathy. A biobank of patient-derived iPSC and fibroblast lines was established, including mutations in SCN5A, MYH6, MYH7, MYBPC3, PKP2, AKAP9, TTN, RBM20, and others. Multimodal phenotyping combining microelectrode arrays, atomic force microscopy, and AFM–Raman spectroscopy was standardized. Gene 20 expression, Troponin T release, and collagen quantification provided complementary readouts of differentiation and turnover. A 30-day hyperglycemic culture model in 2D and 3D embryoid body systems demonstrated altered cardiomyocyte development, increased fibrosis, and reduced stability. Tunable hyaluronic acid–based hydrogels improved contractility and maturation. A novel base excision repair inhibitor modulated DNA damage responses under oxidative stress. Cardiac organoids enabled arrhythmogenicity testing. Salbutamol attenuated aminophyllineinduced arrhythmias, validating the system for pharmacological screening. A method to discriminate conductive vs focal arrhythmias was developed using AFM lateral force measurements. Summary: The project created experimental resources for mechanistic studies of metabolism and electrophysiology, platforms for drug testing, and translational tools for precision medicine in inherited cardiomyopathy and diabetes-related heart disease. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU and by the European Union’s Horizon European Innovation Council Transition Open No. 101214165 (CResPulse) ------------------------ 21 WP5 Heart failure due to obesity and diabetes Work Package 5 Name of the lecture: Comparison of left ventricular function recovery in diabetic and non-diabetic patients with RODCM Name of the group: Principal Investigator: MUDr. Karolina Motilova Author(s): MUDr. Karolina Motilova Abstract: The study evaluated the effect of type 2 diabetes mellitus on left ventricular (LV) reverse remodeling in patients with newly diagnosed dilated cardiomyopathy (DCM). It is known that glucose metabolism disorders (diabetes or prediabetes) can negatively affect LV function, though this is more often associated with heart failure with preserved ejection fraction (HFpEF). In this analysis, only patients with newly developed systolic heart failure due to non-ischemic DCM were included. A total of 49 patients were analyzed (13 with diabetes/prediabetes, 36 without). LV ejection fraction (EF) was assessed echocardiographically at baseline, 3 months, and 6 months. Baseline EF was comparable between groups (25.5 ± 8.4 % vs 27.5 ± 9.4 %), as was the improvement after 6 months (40.6 ± 12.7 % vs 40.4 ± 10.5 %, all p = n.s.). End-diastolic LV volume decreased similarly in both groups, and diastolic function also improved comparably. Summary: In conclusion, the presence of diabetes or prediabetes was not associated with a worse recovery of LV systolic function in patients with recent-onset non-ischemic DCM. The study is limited by small sample size and relatively low prevalence of metabolic disorders, reflecting the younger population referred for advanced heart failure therapy evaluation. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------------------- Work Package 5 Name of the lecture: A New Dual-Hit Rat Model of Type 2 Pulmonary Hypertension in Ischemic Heart Failure Author(s): Matúš Miklovič (1), Matej Molnár (1), Petr Kala (1, 2), Kateřina Kroupová (1), Zdenka Vaňourková (1), Dominik Havlíček (1), Daniel Jirák (1), Matúš Sýkora (3), Barbara Szeiffová Bačová (3), Laura Silva Sousa (4), Attila Kiss (4), Vojtěch Melenovský (1, 5) Authors institution: 1. Center for Clinical and Experimental Medicine – IKEM, Prague, Czech Republic 2. University Hospital Motol and 2nd Faculty of Medicine, Charles University, Prague, Czech Republic 22 3. Centre of Experimental Medicine, Institute for Heart Research, Slovak Academy of Sciences, Dúbravská Cesta 9, 841 04, Bratislava, Slovakia 4. Ludwig Boltzmann Institute for Cardiovascular Research at the Center for Biomedical Research and Translational Surgery, Medical University of Vienna, Austria 5. Department of Cardiology, Institute for Clinical and Experimental Medicine – IKEM, Prague, Czech Republic Introduction Heart failure is frequently associated with right ventricular (RV) dysfunction, primarily due to the development of pulmonary hypertension. RV performance often determines the overall prognosis in heart failure, yet the RV remains underappreciated compared to the left ventricle (LV) in both research and clinical settings. Moreover, there is a lack of robust experimental models that reliably replicate biventricular heart failure. Hypothesis We hypothesized that combining ligation of the left anterior descending (LAD) coronary artery with administration of a vascular endothelial growth factor receptor (VEGFr) inhibitor (sugen, semaxinib) would produce a model of LV dysfunction accompanied by precapillary pulmonary hypertension. Goals The goal of this study was to establish a novel model of heart failure characterized by LV dysfunction induced through high-throughput myocardial infarction (IM) in combination with precapillary pulmonary arterial hypertension. Methods During the first week, LV dysfunction was induced in normotensive HanSD rats via LAD ligation. One week later, pulmonary hypertension was triggered by intraperitoneal injection of sugen (100 mg/kg). In the seventh week, cardiac function was assessed using echocardiography, magnetic resonance imaging (MRI), and pressure–volume (PV) analysis. Hearts and other organs were then harvested for weighing, histological examination, and molecular analyses. Results Two weeks post-infarction, ejection fraction dropped significantly from 84 ± 5% to 36 ± 15% (p < 0.001). LV anterior wall thickness decreased, while LV internal diameters at end-systole and end-diastole increased. Heart weight was markedly elevated in both IM/placebo and IM/sugen groups compared with sham controls, largely due to RV hypertrophy (+70 ± 27% in IM/sugen vs. sham/placebo, p < 0.001). The IM/sugen group also showed pulmonary congestion, reflected by increased lung weight (+37 ± 26%, p < 0.01 vs. sham/placebo). Cardiac output was reduced in the LV of IM/placebo rats and in both ventricles of IM/sugen animals, accompanied by elevated LV end-diastolic volume. RV systolic pressure rose substantially in IM/sugen rats (78 ± 10 mmHg, p < 0.001) compared to sham/placebo (32 ± 9 mmHg), but remained lower in IM/placebo (40 ± 3 mmHg). LV contractility parameters (end-systolic elastance, preload recruitable stroke work) were 23 decreased in both infarcted groups. Interestingly, RV contractility (dP/dt max) increased in IM/sugen rats (2149 ± 400 vs. 1271 ± 348, p = 0.002), suggesting a compensatory mechanism in response to elevated RV load, while remaining unchanged in IM/placebo animals. Gene expression analysis demonstrated upregulation of Nppa, Adrb1, Thbs4, Bmpr2, Fgf23, Gja, Nox4, and Pln in both ventricles of IM/placebo rats compared to sham controls. Serum ACE2 levels were significantly higher in IM/placebo animals, indicating activation of the renin–angiotensin system. Conversely, IM/sugen rats exhibited markedly reduced ACE2 concentrations, consistent with sugen-induced pulmonary endothelial damage via VEGFr blockade. Conclusion These findings confirm the successful establishment of a reproducible rat model of myocardial infarction– induced LV failure accompanied by pulmonary hypertension, suitable for studying biventricular heart failure. Supported by MH CZ - DRO („Institute for Clinical and Experimental Medicine – IKEM, IN 00023001“) and Ministry of Health of the Czech Republic, grant no. NU22-02-00161, NV19-02-00130 and NU21-02-00402. All rights reserved. Project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, Project no. LX22NPO5104) - Funded by the European Union - Next Generation EU. ----------------------------- Work Package 5 Name of the lecture: HIF1α and mitochondria in cardioprotection induced by chronic hypoxia Name of the group: Laboratory of Developmental Cardiology IPHYS, Prague Principal Investigator: RNDr. Petra Alanova, Ph.D. Authors: Alanova P, Alan L, Opletalova B, Bohuslavova R, Abaffy P, Matejkova K, Holzerova K, Benak D, Kaludercic N, Menabo R, Di Lisa F, Ostadal B, Hlavackova M, Kolar F, Pavlinkova G Abstract: Transcriptional factor HIF-1α is known to contribute to cardioprotection against ischemia/reperfusion (I/R) injury. Adaptation to chronic hypoxia (CH) is a cardioprotective phenomenon associated with HIF-1α stabilization. Nevertheless, its precise role in protective changes induced by CH remains incompletely understood. This study aimed to elucidate whether partial Hif1a deficiency would nullify the cardioprotective benefits of CH, while also investigating its impact on mitochondrial function and dynamics. Male wild-type (WT) mice and mice with partial Hif1a deficiency (Hif1a+/−) were exposed to CH for 4 weeks, while their respective controls were kept under normoxic conditions. Subsequently, their isolated perfused hearts were subjected to I/R to determine infarct size, while RNA-sequencing of isolated cardiomyocytes was performed. Mitochondrial respiration was measured to evaluate mitochondrial function, and western blots were performed to assess mitophagy. We demonstrated enhanced ischemic tolerance in WT mice induced by adaptation to CH compared with their normoxic controls and chronically hypoxic Hif1a+/− mice. Through cardiomyocyte bulk mRNA-sequencing analysis, we unveiled significant reprogramming of cardiomyocytes induced by CH emphasizing mitochondrial processes. CH reduced mitochondrial content and respiration and altered mitochondrial ultrastructure. Notably, the reduced mitochondrial content correlated with enhanced autophagosome formation exclusively in chronically hypoxic WT mice, supported by an increase in the LC3-II/LC3-I ratio, expression of PINK1, and degradation of SQSTM1/p62. Furthermore, 24 pretreatment with the mitochondrial division inhibitor (mdivi-1) abolished the infarct size-limiting effect of CH in WT mice, highlighting the key role of mitophagy in CH-induced cardioprotection. Summary: These findings provide new insights into the contribution of HIF-1α to cardiomyocyte survival during acute I/R injury by activating the selective autophagy pathway. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------------- 25 WP 6 Early atherosclerosis and atherothrombosis as its acute reversal Work Package 6 Name of the lecture: The role of sex, reproductive age, and (pre)diabetes in the development and progression of cardiovascular diseases (IKEM) Name of the group: Vascular changes and reproductive status – experimental and human perspectives Principal Investigator: prof. MUDr. Jan Piťha, CSc. Author(s): prof. MUDr. Jan Piťha, CSc. Abstract: Vascular diseases are the main cause of mortality and morbidity not only in men, but also in women. Despite that risk factors leading to vascular impairment are identical in women and men, in women several specific features could modify the effect of these factors. Among the most frequent modifiers could be those associated with menopause. In this respect, transition to menopause and time short after menopause seem to be periods with increased sensitivity to smoking, hypertension, dyslipidemia and diabetes and, thus, the most sensitive period for intervention of these risk factors. We tested hypothesis on experimental models of menopause/ovariectomy, but also in pilot human study if early intervention of dyslipidemia and other risk factors could lead to better improvement of cardiovascular health. This was tested on hypertriglyceridemic (HHTg) female rats after ovariectomy. In addition, we analyzed data in subgroup of women from population sample from study of pre-, peri and postmenopausal females from Prague (3PMFs). In experimental model only hormonal substitution with estradiol substantially improved cardiovascular profile, but no substantial effect was detected for early treatment by statins, similarly inconsistent results were obtained for metformin. In pilot human study, physical activity led to better improvement in vascular health in women earlier after menopause (6 years at maximum). Recently are tested spontaneously hypertensive (SHR) female rats after ovariectomy for early treatment of hypertension. Summary: Data obtained in this study did not show substantial effect of early intervention by statins and metformin after menopause in experimental model of prediabetes. More optimistic results were obtained in women with improved physical activity earlier after menopause. Studies in hypertension in experimental model are under way. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ------------------------------------- Work Package 6 Name of the lecture: Overview of Mechanism of Microbiome Action in the Development of Inflammation and Atherosclerosis Name of the group: Group of Mechanism of Microbiome Action in the Development of Inflammation and Atherosclerosis, Sci MU, Brno Principal Investigator: prof. Mgr. Lukáš Kubala, Ph.D. Authors: Turková Kristýna, Šindelář Martin, Berka Vratislav, Romana Šínová, Šimek Matěj, Kubala Lukáš 32 possible long innovative development path for liraglutide-conjugated magnetic contrast agents. The specific accumulation of liraglutide-conjugated nanoparticles in pancreas was demonstrated after intramuscular application of nanoparticles. Moreover, for research purposes, nanoparticles were made so that they emitted also visible light after irradiation by 800 or 980 nm (so-called light-upconversion nanoparticles), which enables their optical tracking (Shapoval et al., ACS Appl Mater Interfaces. 2025; 17(30):42863-42876). Thus, liraglutide (LGL)-conjugated poly(methyl vinyl ether-alt-maleic acid) (PMVEMA)-coated core−shell NaYF4:Yb,Er,Fe@NaYF4:Nd upconversion nanoparticles (CS-UCNPs) have been developed, thoroughly physicochemically characterized, and evaluated in vivo. Novel codoping of Fe2+, Yb3+, and Er3+ ions in the host NaYF4 induced upconversion emission in the red region at both 980 and 808 nm excitation, making the particles suitable for deep-tissue imaging. These nanoparticles enabled targeted binding to GLP-1 receptors on pancreatic β-cells, increasing glucose-stimulated insulin secretion from isolated Langerhans islets. Next steps will involve liragluitideplus polyfluorine-polymer-conjugated nanoparticles for 19F-NMR. Summary: Contrast agents based on nanoparticle conjugates of liraglutide are to be developed for magnetic resonance imaging (MRI) and possible semi-quantification of pancreatic beta cell mass. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------------------- Work Package 7 Name of the lecture: IsletNet: web service for automatic image analysis with highly accurate model for islet volume estimation before transplantation. Author: David Habart Effective transplantation of isolated pancreatic islets required sufficient islet dose quantified in terms for the number and volume per weight of the recipient. At present, the-state-of-the-art quantification method is fully manual, making it subjective, and based on islet size using arbitrary spherical model which makes it inaccurate. To facilitate the comparison of transplantation programs worldwide, we created a dedicated web service for AI-based image analysis and presented it in 2017 at IPITA meeting in Oxford. Over the time many centres across Europe and USA have joint the project by participating the their images for IsletNet training. Cardia supported networking with collaborating centres at IPITA and EPITA conferences and by means of visit of a student in one European centre, helping them establish IsletNet into routine use for Clinical Islet transplantation. To improve accuracy of the model, we collected thousand of rat and human isolated islets, studied their real 3D geometries and measured volumes. We hypothesised that microscopic contour in additional to the size can be harnessed for alternative model development. Cardia funding facilitated the finalising the development of the model Sphiracle, including its validation on independent dataset. The model improved accuracy by reducing the volume overestimation form ~1.45-fold to ~1.05fold. The model is implemented in publicly available professional image analysis tools. Summary: 33 IsletNet is was proposed at IPITA 2025 meeting by the Oxford Centre to serve the the new standard for islet graft assessment in Europe. Sphiracle model with improved accuracy is available for professional SW Fiji and in IsletNet (so far the private IKEM version). ------------------------------- Work Package 7 Name of the lecture: Nanofibers modified with platelet lysate for chronic wound healing. Name of the group: Group of Laboratory of Biomaterials and Tissue Engineering of the Institute of Physiology, CAS, Prague Principal Investigator: doc. MUDr. Lucie Bacakova, Ph.D. Author(s): Mgr. Elena Filova, Ph.D., Andreu Blanquer, Ph.D., Adam Eckhardt, Ph.D., Jana Musilkova, Ph.D., Johanka Taborska , Ph.D. Eduard Brynda, Ph.D., Tomas Riedel, Ph.D., Ing. Andrea Sidova, Ph.D., doc. Ing. Vera Jencova, Ph.D., Jana Mullerova , MUDr. Renata Prochazkova, Ph.D., Jana Sabova, MSc., RNDr. Zuzana Demcisakova, Ph.D., Ing. Zuzana Tirpakova, Bc., Prof. MVDr. Eva Petrovova, Ph.D. doc. MUDr. Lucie Bacakova, Ph.D. Abstract: Chronic diabetic ulcers are present in 15% of the diabetic population. Generally, 1-2% of population will suffer from a chronic wound during their lifetime. Chronic wounds contain decreased levels of growth factors, increased ROS production, degradation of ECM, delayed re-epithelialization, etc. Platelet lysate (PL) contains about 200 bioactive chemical compounds, e.g. growth factors that could stimulate wound healing. Therefore, various nanofibers based on poly (vinyl alcohol) (PVA) or copolymer poly(L-lactide-co-Ɛ-caprolactone) (PLCL), with incorporated or attached PL and/or VEGF/FGF2 growth factors were prepared. The materials were evaluated with various skin cells, e.g. keratinocytes, endothelial cells, and fibroblasts in vitro in monocultures and co-cultures, using a 3D collagen wound model in vitro, and finally were evaluated with the chorioallantoic membrane of chick embryo ex ovo. All PL-based materials proved the ability to stimulate skin cell proliferation, differentiation or migration, the effects were improved, when the materials contained both VEGF/FGF2 and PL. PVA_PL materials with higher concentration of PL stimulated wound defect healing in the 3D collagen wound model and the most stimulated angiogenesis in mesodermal layer of chorioallantoic membrane in ex ovo experiment. High hemocompatibility of the materials was proved as well. The advantage of these materials was sustained release of many bioactive compounds in relatively low concentrations within days without any adverse effects observed. Summary: PVA and PLCL nanofibers modified with PL or with both PL and VEGF/FGF2 stimulated skin cell proliferation, differentiation, and migration in vitro, healing the wound in a 3D collagen wound model, and the angiogenic response of the chorioallantoic membrane of the chick embryo. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. 34 Posters section Work Package 2 Name of the poster: Towards development of IR-B specific insulins Name of the group: Chemistry and Biology of Insulin and Insulin-like growth factors, IOCB, Prague Principal Investigator: Jiří Jiráček, Ph.D. Author(s): Maxmilián Hejl Abstract: Modern insulin therapies aim to improve the quality of life for patients. Among these, one of the least invasive options involves administering insulin subcutaneously. However, this method leads to uneven insulin distribution in the body, as the hormone is first absorbed by peripheral tissues and its liver action is delayed. This contrasts with natural insulin production, where the liver is the primary target. In the liver, insulin not only regulates glucose uptake but also promotes glycogen synthesis and suppresses gluconeogenesis. Our long-term goal is to develop an insulin analogue that bypasses peripheral tissues and acts predominantly in the liver. By directly targeting gluconeogenesis, this analogue could mitigate the common side effects of current therapies, such as hypoglycemia, weight gain, and cardiovascular risks. One approach to creating a hepatoselective insulin analogue involves leveraging the distinct distribution of insulin receptor isoforms (IR-A and IR-B) across different organs. Notably, the liver predominantly expresses the IR-B isoform while peripheral tissues express different proportions of both isoforms. This unique distribution provides a crucial clue for our research. The challenge lies in understanding the subtle differences between these two receptor isoforms and their interactions with hormones — an area where, as the saying goes, "The devil, as always, is in the detail." Despite their similarities as 320 kDa proteins, IRB differs from IR-A by a mere 12 amino acids at its C-terminus. While structural studies have been conducted, no one has yet resolved the structure within these crucial 12 amino acids. Building on the extensive expertise of Dr. Jiráček’s laboratory, we hypothesize that the C-terminus of the insulin B-chain is crucial for modulating its ability to bind selectively to IR-B. By combining this knowledge with synthetic methods, we aim to modify insulin precisely in this region. This work systematically explores modifications in the final eight positions of the insulin B-chain C-terminus to investigate its role in IR-B binding specificity and its potential contribution to hepatoselectivity. Summary: Modern insulin therapies rely on subcutaneous delivery, which delays liver action and causes side effects such as hypoglycemia and weight gain. Our goal is to develop hepatoselective insulin analogs that act mainly in the liver by targeting the IR-B isoform, the predominant receptor there. Although IR-A and IR-B differ by only 12 amino acids, these differences may determine tissue selectivity. Building on structural and synthetic expertise, we systematically modify the C-terminus of the insulin B-chain to enhance IR-B binding and achieve liver-targeted insulin action. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ------------------------ 35 Work Package 1 Name of the poster: The Cav1.2 paradox: severe epilepsy without arrhythmia in a child with CACNA1C mutation Name of the group: Pathophysiology of Ion Channels, Department of Pathophysiology, Third faculty of Medicine, Charles University, Prague Principal Investigator: Norbert Weiss, Ph.D. Author(s): Robin N Stringer, Xuechen Tang, Bohumila Jurkovicova-Tarabova, Mary Murphy, Klaus R Liedl, Norbert Weiss Abstract: Mutations in CACNA1C, encoding the Cav1.2 voltage-gated calcium channel, underlie a wide phenotypic spectrum ranging from Timothy syndrome to isolated cardiac arrhythmias and neurodevelopmental disorders. This pleiotropy highlights the central role of Cav1.2 in both cardiac and neuronal excitability. We describe a child with a de novo heterozygous missense variant (c.1973T > C; L658P) in CACNA1C, presenting with refractory epilepsy, global developmental delay, hypotonia, and systemic abnormalities, but notably without overt cardiac dysfunction. Electrophysiological studies of Cav1.2 L658P revealed profound gating alterations, including a hyperpolarizing shift in voltage dependence of activation and inactivation. Structural modeling suggested that the L658P substitution disrupts interactions within the IIS5 transmembrane segment, lowering the energetic threshold for channel opening at negative potentials. Summary: These results identify L658P as a pathogenic CACNA1C variant driving severe neurological disease in the absence of arrhythmia or cardiomyopathy. Importantly, this case refines the emerging genotype-phenotype correlations in CACNA1C-related disorders and underscores the need for cardiovascular surveillance even when initial presentation appears neurologically restricted. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ------------------------- Work Package 2 Name of the poster: The impact of muscle non-shivering thermogenesis on energy expenditure during physical activity Name of the group: Institute of Physiology of the Czech Academy of Sciences, Videnska 1083, 142 20 Prague, Czech Republic Principal Investigator: MUDr. Jan Kopecky, CSc. Author(s): Anna Vavrova, Kristina Bardova, Sara Stanic, Petr Zouhar, Jan Kopecky Abstract: Acute cold exposure induces heat production, mainly through muscle shivering. During prolonged cold acclimation, acute mechanisms of heat production are replaced in most mammalian species by other 36 mechanisms, such as non-shivering (NST) brown adipose tissue (BAT) thermogenesis. However, mice lacking UCP1 (UCP1-KO), the crucial protein for BAT NST, are cold-resistant when gradually cold-adapted, suggesting that other heat-generating mechanisms are involved. These mechanisms could include prolonged muscle shivering period, but also muscle NST and mechanisms involving futile cycling in BAT, muscle or other tissues. Muscle NST might be based based on uncoupling of endoplasmatic SERCA Ca+II pumping activity by sarcolipin. The fact that sarcolipin affects calcium pumping efficiency could have an effect on energy expenditure during muscle contraction. This needs to be better characterized. The objective of this project was to quantify energy expenditure during forced physical activity using gradually cold-adapted UCP1-KO and control mice. A unique combination of treadmills, indirect calorimetry and a climate chamber was employed in the study. The experimental configuration facilitated the determination of the cost of physical activity under thermoneutral conditions in mice that had been pre-acclimatised to different temperatures. The energy expenditure of the control mice remained unaltered by cold acclimation; however, a gradual increase was observed in cold-acclimated UCP1-KO mice. Furthermore, UCP1-KO mice acclimated to cold exhibited reduced endurance, potentially attributable to overheating, a hypothesis that will be tested in the subsequent experiment. Summary: The results of this study provide further insight into the impact of muscle NST on the efficiency of muscle contraction. These data obtained with CarDia support demonstrated the interaction between muscle contractility and heat production. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------------------------- Work Package 1 Laboratory of Pancreatic Islets Research, IPHYS, Prague Principal Investigator: RNDr. Lydie Plecitá, Ph.D. Author: Mgr. Blanka Holendová, Ph.D. Abstract: Pancreatic β-cells are central to blood glucose control, secreting insulin in response to glucose metabolism. This process naturally generates reactive oxygen species (ROS), which play a crucial, yet incompletely understood, role in maintaining healthy β-cell function. However, in conditions of chronic high glucose and nutrient excess, often seen in the progression to Type 2 Diabetes, this ROS production can become excessive. This leads to oxidative stress, a key factor in β-cell damage and dysfunction, ultimately contributing to the impaired insulin secretion characteristic of the disease. Our research reveals a specific mechanism by which glucose influences β-cell function: it promotes ROS production, which then selectively and reversibly modifies cysteine residues within key β-cell proteins. We have identified numerous such "redox-sensitive" proteins involved in fundamental metabolic processes, including glucose breakdown (glycolysis), energy production (TCA cycle, oxidative phosphorylation), and proper protein handling (ER protein processing). Dysfunction in these pathways is directly linked to impaired insulin secretion. Importantly, we show that this reversible cysteine modification acts in concert with other vital protein regulations (like acetylation and phosphorylation), providing a dynamic system that allows β-cells to adapt, or maladapt, to metabolic changes. Understanding this mechanism offers new insights into how glucose overload damages β-cells and could pave the way for novel therapeutic strategies targeting β-cell health in diabetes. 37 Summary: This research, supported by the CarDia project, identifies a specific mechanism where glucose promotes ROS production that reversibly modifies key proteins, offering new insights into regulation of βcell metabolism. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------- Work Package 5 Name of the poster: Water-Soluble Star Polymers as Safe and Effective 19F MRI Contrast Agents Name of the group: ZRIR IKEM, Prague Principal Investigator: prof. Ing. Daniel Jirák, PhD. Author(s): Mgr. Michal Franc, Ing. Ladislav Androvič, Ing. Richard Laga, PhD., prof. Ing. Daniel Jirák, PhD. Abstract: Magnetic resonance imaging (MRI) is widely regarded as a cornerstone of clinical and experimental diagnostics, with 1H MRI providing high spatial resolution and excellent soft tissue contrast. Nevertheless, the reliance on endogenous hydrogen nuclei gives rise to a high background signal and limited specificity, particularly in the context of targeted cancer imaging. Conventional gadolinium-based contrast agents enhance sensitivity but raise safety concerns due to tissue retention and lack of molecular specificity. As a promising alternative, ¹⁹F MRI enables "hotspot imaging" without biological background interference, while its favourable magnetic properties allow straightforward integration into existing MRI systems. However, current 19F tracers, such as perfluorocarbons, are hydrophobic and require encapsulation, which complicates formulation and limits translational potential. In order to overcome these limitations, a new class of water-soluble, biocompatible star-shaped polymers was designed for use in 19F MRI and potential theranostic applications. In this study, RAFT polymerisation was utilised to synthesise of a porphyrin-based star polymers. The polymers comprised a non-fluorinated reference polymer (porf-MPC-TTC) and a fluorinated analogue (porf-19FMCP-TTC) with trifluoromethylmodified side chains, with the objective of generating a strong 19F signal. The molecular weights of the polymers were found to be in the range of 60–65 kDa, as determined by SEC and DLS, which also confirmed narrow dispersity (Đ < 1.2) and hydrodynamic diameters of approximately 20 nm. The characterisation of the polymers was conducted using NMR spectroscopy (1H, 19F, 31P). The relaxation properties were assessed on a 1.5 T relaxometer, and high-field studies were performed on a 7 T Bruker BioSpec using custom-built coils. Complementary fluorescence imaging was achieved via Cy7-labeling, and the in vitro evaluation of the anticancer properties of the samples was conducted using AlamarBlue assays on 4T1 breast carcinoma and HEK cell lines. The biodistribution of the compound in vivo was investigated in 4T1 tumour-bearing mice using a combination of MRI and fluorescence imaging techniques. The fluorinated polymers demonstrated exceptional solubility, colloidal stability, and a robust 19F signal even at concentrations as low as 1.5 mg/mL. The nanoscale size of the particles favours passive tumour targeting, with both MRI and fluorescence imaging confirming efficient tumour accumulation and signal persistence for up to nine days post-injection in all tested animals. Assays for determining cellular toxicity indicated no significant toxicity across the range of concentrations that were examined. These findings underscore the considerable potential of porphyrin-based star polymers as dual-functional MRI probes, integrating diagnostic imaging with therapeutic versatility. 38 Summary: In summary, the results demonstrate that fluorinated star polymers represent a highly promising platform for safe and effective 19F MRI cellular contrast agents, combining high sensitivity, long circulation time, and intrinsic multifunctionality. The objective of the present research is to optimise polymer dimensions for clinical translation, extend the platform to additional nuclei (11B and 31P), and incorporate pH-responsive drug release mechanisms. This strategy establishes the basis for the development of modular theranostic nanomaterials that have the potential to integrate advanced imaging and targeted therapy. The authors acknowledge the financial support from the the Ministry of Health of the Czech Republic (MH CZ-DRO, Institute for Clinical and Experimental Medicine IKEM, IN 00023001) and the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, Project no. LX22NPO5104) Funded by the European Union Next Generation EU. -------------------------------- Work Package 2 Name of the poster: Proliferative and signaling properties of IGF and insulin analogs with promiscuous binding to both insulin and insulin-like growth factor 1 receptors Name of the group: Chemistry and Biology of Insulin and Insulin-like growth factors, IOCB, Prague Principal Investigator: Jiří Jiráček, Ph.D. Author(s): Irena Selicharová, Ph.D. Abstract: Insulin and insulin-like growth factor 1 (IGF-1) are closely related hormones involved in the regulation of metabolism and growth. They elicit their functions through activation of tyrosine kinase-type receptors: insulin receptors (IR-A and IR-B) and IGF-1 receptor (IGF-1R). Despite similarity in primary and threedimensional structures, insulin and IGF-1 bind the noncognate receptor with substantially reduced affinity. In this study we focused on closer characterization of our previously prepared analogs with promiscuous binding to both the IR and IGR-1R and their potential impact on cells of neurological origin. We employed our analogs [His49]-IGF-1 and [His48]-IGF-2 with increased binding to IR-A compared to the wild-type hormones. Of special relevance, we prepared [GluB10, D-HisB24, GlyB31, TyrB32]-insulin, which binds all three receptors with unusually high affinity: 209 or 950 % binding affinity to IR-A, respectively to IR-B, relative to insulin and, remarkably, 79 % binding affinity to IGF-1R relative to IGF-1. We show the increased proliferative activity of the promiscuous analogs together with potentiation of the insulin/IGF-1 signaling pathway in the cells of neuronal origin. We suggest that the new super-potent insulin analog might be an ideal candidate for use where the growth potential of both insulin and IGF-1 is needed, which may be, for example, media for stimulating cell growth and differentiation, and where a single derivative could replace the action of two native hormones with greater efficiency and at lower cost. Summary: We developed and studied new insulin/IGF-1 dual agonist that binds insulin and IGF-1 receptors with very high affinity and stimulates neuronal cell growth more effectively than native hormones. Such molecules may replace both hormones in cell culture, offering greater efficiency and lower cost. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ---------------- 39 Work Package 2 Name of the poster: Central regulation of body temperature and energy expenditure by FGF21 Name of the group: 1 Laboratory of adipose tissue biology, Institute of physiology of the Czech academy of sciences, Prague, Czech Republic, 142 00, 2 Department of physiology, Faculty of science, Charles university, Prague, Czech Republic, 142 00 Principal Investigator: MUDr. Jan Kopecky, CSc. Author(s): Sara Stanic,1,2 Kristina Bardova,1 Petra Janovska,1 Jan Kopecky,1 Petr Zouhar*,1 Abstract: Fibroblast growth factor 21 (FGF21) is a promising metabolic regulator with well-documented effects in rodents, yet several of its benefits, most notably weight loss, fail to fully translate to humans. To better understand this discrepancy, we investigated the role of FGF21 in thermoregulation and its impact on energy expenditure (EE) and body weight. Using C57BL/6J mice and mice deficient in Uncoupling protein 1 (UCP1), an essential mediator of classical non-shivering thermogenesis, we explored whether increased body temperature precedes or follows elevated EE, and the relevance of UCP1 in this process. Our findings indicate that FGF21 acts primarily through the central nervous system to shift the body's temperature setpoint upward, with EE increasing secondarily to reach this new setpoint.1 The elevated body temperature can be reached rapidly by increased heat conservation and activation of UCP1. Prolonged FGF21 administration may also induce some alternative mechanisms of non-shivering thermogenesis, namely futile cycling of triacylglycerols and fatty acids in brown adipose tissue.2 In contrast to UCP1 activity, induction of UCP1-independent mechanisms does not seem to be mediated by a sympathetic nervous system. Although sympathetically independent EE has limited thermogenic capacity, it contributes to weight loss when coupled with reduced food intake. Summary: These results highlight a central role for FGF21 as a metabolic manager that directs the brain to adjust body temperature and body weight using available physiological mechanisms. These data obtained with CarDia support may help explain the species-specific outcomes of FGF21 treatment and guide more effective therapeutic strategies in humans. 1 Zouhar, Petr, et al. "A pyrexic effect of FGF21 independent of energy expenditure and UCP1." Molecular Metabolism 53 (2021): 101324. 2 Stanic, Sara, et al. "Prolonged FGF21 treatment increases energy expenditure and induces weight loss in obese mice independently of UCP1 and adrenergic signaling." Biochemical pharmacology 221 (2024): 116042. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------------- Work Package 2 Name of the poster: Temperature Imprinting in Skeletal Muscle of Mice Differing in Their Propensity to Obesity Name of the group: 40 1 Laboratory of Adipose Tissue Biology, Institute of Physiology of the Czech Academy of Sciences, Videnska 1083, 142 00 Prague, Czech Republic 2 Metabolomics Service Laboratory, Institute of Physiology of the Czech Academy of Sciences, Videnska 1083, 142 00 Prague, Czech Republic 3 Proteomics service laboratory, Institute of Physiology of the Czech Academy of Sciences, Videnska 1083, 142 00 Prague, Czech Republic 4 Research Unit for Rare Diseases, Department of Pediatrics and Inherited Metabolic Disorders, 1st Faculty of Medicine Charles University, Ke Karlovu 2, 128 00 Praha 2, Czech RepublicPrincipal Investigator: MUDr. Jan Kopecky, CSc. Author(s): Eliska Haasova1, Tatyana Kobets2, Marek Vrbacky3, Lenka Steiner Mrazova4, Viktor Stranecky4, Jan Kopecky1, Petra Janovska1 Abstract: Mammals maintain body temperature via shivering and non-shivering thermogenesis (NST). While NST is primarily dependent on UCP1 in brown adipose tissue (BAT), the UCP1-independent mechanisms remain unclear. In cold, obesity-prone C57BL/6J (B6) mice primarily activate NST in BAT, while obesity-resistant A/J mice activate skeletal muscle (SM) NST via sarcolipin and SERCA uncoupling, which likely helps A/J mice tolerate cold and resist obesity. This study was focused on effect of ambient temperature on muscle NST and metabolism in mice with different susceptibility to obesity. A/J and B6 mice were maintained at 30°C or 20°C from birth to 6 weeks of age and then at 30°C for up to 3 months. Proteomic, lipidomic and transcriptomic analyses were performed in muscle gastrocnemius. Early life ambient temperature permanently affected SM lipid metabolism in A/J but not in B6. A/J maintained at 30°C upregulated enzymes for both lipid synthesis and lipolysis, suggesting a futile lipid cycle, whereas A/J maintained at 20°C had higher levels of TAG in the SM. DEXA revealed temperature-related changes in A/J body composition. These findings suggest that perinatal ambient temperature may imprint SM metabolism and affect thermogenesis, body composition, and obesity risk. Summary: Further research should investigate the molecular basis of this imprinting and the pathways responsible for A/J resistance to obesity, with potential applications for obesity prevention. These data obtained with CarDIA support enable us to define the fundamental importance of early postnatal development for later susceptibility to obesity. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. ----------------- Work Package 7 Name of the poster: Cooperation with the Institute of Physiology of the Czech Academy of Sciences Radiation-Modulated Decellularized Pericardium Vascular Patches with Wharton's Jelly Stem Cells Name of the group: Group of Experimental Vascular Surgery and Tissue Engineering, IKEM, Prague Principal Investigator: MUDr. Jaroslav Chlupáč, Ph.D. Author(s): MUDr. Jan Frank, MUDr. Jaroslav Chlupáč, Ph.D., Zuzana Šimůnková, Ing. Iveta Mrázová, MUC. Tomáš Novák, doc. MUDr. Ondřej Fabián, Ph.D., RNDr. Martin Květoň, Ing. Zdeňka Vaňourková, Ph.D., prof. MUDr. Jiří Froněk, Ph.D. 41 Abstract: The development of novel vascular substitutes remains a key challenge in vascular surgery and tissue engineering. Recent studies have demonstrated the potential of Wharton's Jelly-derived mesenchymal stem cells (WJSCs) for tissue colonization. In collaboration with the Institute of Physiology of the Czech Academy of Sciences, we investigated the feasibility of colonizing decellularized porcine pericardia with WJSCs and their use as vascular patches in a porcine carotid artery model. In the first phase, carried out in vitro, decellularized porcine pericardia were modified using various surface treatments, including radiation, vascular growth factors, heparin, and fibrin. Four types of patches were prepared: two seeded with WJSCs and two unseeded controls. In the second phase, conducted at IKEM, the prepared patches were implanted into porcine carotid arteries. We evaluated four types of vascular patches, each tested in triplicate. Intraoperative measurements of arterial blood flow were performed before and after implantation to assess patency. One-month post-implantation, angiography was used to confirm graft patency, followed by explantation of the grafts and adjacent artery. Macroscopic analysis was performed on cross-sections of the explanted samples. These were subsequently fixed in formalin and submitted for histological examination. During microscopy examination of preimplant histology samples, we find surface layer of human WJSC. As our metric we chose thickness of intimal hyperplasia (NIH) which is known as one of the negative factors in vascular remodeling and in most cases reason for arterial stenosis. We also measured thickness of arterial wall. As expected, all patches remained patent, likely due to their positioning, which covered only half of the arterial lumen. Two clinically silent aneurysms were observed, each in a different group (Non-irradiated + WJC and Irradiated + WJC). Neointimal hyperplasia in the Non-irradiated + WJC group measured 862 ± 77.21 μm²/μm, which was significantly greater than in the Non-irradiated group (637 ± 32.69 μm²/μm, p < 0.05). Although this increase was statistically significant, it did not translate into any apparent clinical consequences. In the Irradiated + WJC and Irradiated groups, NIH measured 786 ± 97.82 μm²/μm and 613 ± 118 μm²/μm, respectively. No other significant differences were observed among the groups. Summary: Only limited differences were observed in vivo among decellularized pericardial vascular patches prepared using irradiation, vascular growth factors, heparin, fibrin, or cell seeding. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU. --------------- Work Package 1 Name of the poster: Effects of acute exercise on FAHFA metabolism in mice Name of the group: Laboratory of Adipose Tissue Biology FGÚ, Prague Principal Investigator: MUDr. Martin Rossmeisl, Ph.D. Author(s): Marko Mitrovic, Olga Horakova, Martin Riecan, Veronika Kleinova, Tomas Cajka, Lenka Rossmeislova, Ondrej Kuda, Martin Rossmeisl Abstract: Improving the metabolic functions of adipose tissue (AT) may contribute to the beneficial health effects of exercise. AT is a source of various fatty acid esters of hydroxy fatty acids (FAHFA), which may have potent anti-inflammatory and insulin-sensitizing properties. We have previously shown that exercise training improves insulin sensitivity and raises levels of PAHSA lipokines, representing one of the FAHFA 48 compared to other countries (MEST-C score in patients with baseline proteinuria › 0,5 g/d: Czech Republic - C1 70 %, C2 6,3 %; Argentina - C 1 35.5 %; Romania - C1 27.9 %,C2 12.6 %; India C1 24.7%, C2 4.1 %). Summary: In our study we analyzed clinical and histological data of a huge number of patients with IgAN from three continents with different renal prognosis which might be affected by various indication for renal biopsy. Renal prognosis of patients in different countries will be assessed by means of propensity-score analysis. Fig 1 Renal survival of patients with IgA nephropathy from Argentina, India, Romania and Czech Republic (follow up of five years) 0 – patients with preserved renal function; 1 – patients with >50 % decline of eGFR during FU; 2 – patients with ESKD during FU; 3 – patients with death during FU Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU, MH CZ DRO VFN 64165, and the research project SVV 260374. ------------- Name of the poster: Volume reduction rate in human and rat pancreatic islets in culture after isolation Name of the group: Laboratory for the Islets of Langerhans (LIL) IKEM, Prague Principal Investigator: doc. MUDr, David Habart, Ph.D. Author(s): Sarah Suergiu, MSc Abstract: 89,0 64,9 65,2 43,5 2,0 5,0 3,9 7,7 7,0 30,2 27,8 41,7 2,0 0,0 3,0 7,1 0 10 20 30 40 50 60 70 80 90 100 Argentina India Romania Czech R Relative frequency Relative frequency (Primary outcome) 0 1 2 3 49 Accurate pancreatic islet volume estimation is essential for graft dose calculation and in vitro studies. Isolated islets undergo changes in morphology, which can impact data normalization in metabolic or secretory assays. Failure to account for early post-isolation shrinkage may lead to misinterpretation of results. However, individual islets volume dynamics remain uncharacterized. Our aim is to quantify the volumetric changes of individual rat and human pancreatic islets during the first hours in culture. Human and rat islets were isolated and seeded in a μ-Slide with supplemented medium, within an hour after the end of isolation. Time-lapse imaging was performed using a Leica DMi8 microscope (37°C, 5% CO2), acquiring images every 20 minutes for the first 3 hours, then hourly up to 17 hours. Images were segmented using the GraphCut plug-in in Fiji, and volumes were estimated from islet sizes and projection shapes using our newly developed volume model. A total of 65 Wistar rat islets from four donors exhibited an average volume reduction of 12% (CV 0.07), while 49 human islets from a single donor showed an 18% reduction (CV 0.06). In both cases, 50% of the total volume loss occurred within the first 3 hours. Summary: Freshly isolated pancreatic islets exhibit a rapid volume reduction, reaching half of that change within the first 3 hours. Great variability was observed among the individual islets, irrespective of the species or size. Supported by the project National Institute for Research of Metabolic and Cardiovascular Diseases (Programme EXCELES, ID Project No. LX22NPO5104) - Funded by the European Union – Next Generation EU.