Ústav biomedicínského inženýrství

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Pigment Formation by Monascus pilosus DBM 4361 in Submerged Liquid Culture
(2025-10-08) Husakova, Marketa; Bezdicek, Matej; Branska, Barbora; Sedlář, Karel; Patakova, Petra
Monascus pilosus is usually cultivated on rice because of monacolin K. We focused on pigment production in submerged liquid culture (SLC) where M. pilosus produced different pigments compared to M. purpureus and M. ruber. From the group of classic Monascus pigments, there were formed mostly compounds with a five-carbon side chain, and the dominant pigment was monascuspiloin, a yellow pigment structurally similar to monascin. In SLC, previously undescribed patterns affecting pigment formation were observed, such as the Crabtree effect, carbon catabolite repression of pigments caused by glucose and other mono-/disaccharides, as well as nitrogen regulation, particularly repression of pigment formation by ammonium sulfate. The highest pigment concentration in the extract was obtained using an organic nitrogen source, specifically 340 mg/L for yellow pigments utilizing a combination of sucrose and tryptone, 346 mg/L for orange pigments using starch and tryptone, and 75 mg/L for red pigments using lactose and tryptone.
Fully automated Bayesian analysis for quantifying the extent and distribution of pulmonary perfusion changes on CT pulmonary angiography in CTEPH
(2025-05-28) Suchánek, Vojtěch; Jakubíček, Roman; Hrdlička, Jan; Novák, Matěj; Miksová, Lucie; Jansa, Pavel; Burgetová, Andrea; Lambert, Lukáš
Objectives This work aimed to develop an automated method for quantifying the distribution and severity of perfusion changes on CT pulmonary angiography (CTPA) in patients with chronic thromboembolic pulmonary hypertension (CTEPH) and to assess their associations with clinical parameters and expert annotations. Materials and methods Following automated segmentation of the chest, a machine-learning model assuming three distributions of attenuation in the pulmonary parenchyma (hyperemic, normal, and oligemic) was fitted to the attenuation histogram of CTPA images using Bayesian analysis. The proportion of each component, its spatial heterogeneity (entropy), and center-to-periphery distribution of the attenuation were calculated and correlated with the findings on CTPA semi-quantitatively evaluated by radiologists and with clinical function tests. Results CTPA scans from 52 patients (mean age, 65.2 +/- 13.0 years; 27 men) diagnosed with CTEPH were analyzed. An inverse correlation was observed between the proportion of normal parenchyma and brain natriuretic propeptide (proBNP, rho = -0.485, p = 0.001), mean pulmonary arterial pressure (rho = -0.417, p = 0.002) and pulmonary vascular resistance (rho = -0.556, p < 0.0001), mosaic attenuation (rho = -0.527, p < 0.0001), perfusion centralization (rho = -0.489, p = < 0.0001), and right ventricular diameter (rho = -0.451, p = 0.001). The entropy of hyperemic parenchyma showed a positive correlation with the pulmonary wedge pressure (rho = 0.402, p = 0.003). The slope of center-to-periphery attenuation distribution correlated with centralization (rho = -0.477, p < 0.0001), and with proBNP (rho = -0.463, p = 0.002). Conclusion This study validates an automated system that leverages Bayesian analysis to quantify the severity and distribution of perfusion changes in CTPA. The results show the potential of this method to support clinical evaluations of CTEPH by providing reproducible and objective measures.
Acidogenesis, solventogenesis, metabolic stress response and life cycle changes in Clostridium beijerinckii NRRL B-598 at the transcriptomic level
(Springer Nature, 2019-02-04) Patáková, Petra; Branská, Barbora; Sedlář, Karel; Vasylkivska, Maryna; Šabatová, Kateřina; Kolek, Jan; Cicková, Pavlína; Provazník, Valentýna
Clostridium beijerinckii NRRL B-598 is a sporulating, butanol and hydrogen producing strain that utilizes carbohydrates by the acetone-butanol-ethanol (ABE) fermentative pathway. The pathway consists of two metabolic phases, acidogenesis and solventogenesis, from which the latter one can be coupled with sporulation. Thorough transcriptomic profiling during a complete life cycle and both metabolic phases completed with flow cytometry, microscopy and a metabolites analysis helped to find out key genes involved in particular cellular events. The description of genes/operons that are closely involved in metabolism or the cell cycle is a necessary condition for metabolic engineering of the strain and will be valuable for all C. beijerinckii strains and other Clostridial species. The study focused on glucose transport and catabolism, hydrogen formation, metabolic stress response, binary fission, motility/chemotaxis and sporulation, which resulted in the composition of the unique image reflecting clostridial population changes. Surprisingly, the main change in expression of individual genes was coupled with the sporulation start and not with the transition from acidogenic to solventogenic metabolism. As expected, solvents formation started at pH decrease and the accumulation of butyric and acetic acids in the cultivation medium.
A transcriptional response of Clostridium beijerinckii NRRL B-598 to a butanol shock
(BioMed Central, 2019-10-13) Sedlář, Karel; Kolek, Jan; Gruber, Markus; Šabatová, Kateřina; Branská, Barbora; Csaba, Gergely; Vasylkivska, Maryna; Zimmer, Ralf; Patáková, Petra; Provazník, Valentýna
Background One of the main obstacles preventing solventogenic clostridia from achieving higher yields in biofuel production is the toxicity of produced solvents. Unfortunately, regulatory mechanisms responsible for the shock response are poorly described on the transcriptomic level. Although the strain Clostridium beijerinckii NRRL B-598, a promising butanol producer, has been studied under different conditions in the past, its transcriptional response to a shock caused by butanol in the cultivation medium remains unknown. Results In this paper, we present a transcriptional response of the strain during a butanol challenge, caused by the addition of butanol to the cultivation medium at the very end of the acidogenic phase, using RNA-Seq. We resequenced and reassembled the genome sequence of the strain and prepared novel genome and gene ontology annotation to provide the most accurate results. When compared to samples under standard cultivation conditions, samples gathered during butanol shock represented a well-distinguished group. Using reference samples gathered directly before the addition of butanol, we identified genes that were differentially expressed in butanol challenge samples. We determined clusters of 293 down-regulated and 301 up-regulated genes whose expression was affected by the cultivation conditions. Enriched term “RNA binding” among down-regulated genes corresponded to the downturn of translation and the cluster contained a group of small acid-soluble spore proteins. This explained phenotype of the culture that had not sporulated. On the other hand, up-regulated genes were characterized by the term “protein binding” which corresponded to activation of heat-shock proteins that were identified within this cluster. Conclusions We provided an overall transcriptional response of the strain C. beijerinckii NRRL B-598 to butanol shock, supplemented by auxiliary technologies, including high-pressure liquid chromatography and flow cytometry, to capture the corresponding phenotypic response. We identified genes whose regulation was affected by the addition of butanol to the cultivation medium and inferred related molecular functions that were significantly influenced. Additionally, using high-quality genome assembly and custom-made gene ontology annotation, we demonstrated that this settled terminology, widely used for the analysis of model organisms, could also be applied to non-model organisms and for research in the field of biofuels.
The Effect of Rhodamine-Derived Superparamagnetic Maghemite Nanoparticles on the Motility of Human Mesenchymal Stem Cells and Mouse Embryonic Fibroblast Cells
(MDPI, 2019-03-27) Chmelíková, Larisa; Skopalík, Josef; Chmelík, Jiří; Zumberg, Inna; Čmiel, Vratislav; Poláková, Kateřina; Provazník, Valentýna
Nanoparticles have become popular in life sciences in the last few years. They have been produced in many variants and have recently been used in both biological experiments and in clinical applications. Due to concerns over nanomaterial risks, there has been a dramatic increase in investigations focused on safety research. The aim of this paper is to present the advanced testing of rhodamine-derived superparamagnetic maghemite nanoparticles (SAMN-R), which are used for their nontoxicity, biocompatibility, biodegradability, and magnetic properties. Recent results were expanded upon from the basic cytotoxic tests to evaluate cell proliferation and migration potential. Two cell types were used for the cell proliferation and tracking study: mouse embryonic fibroblast cells (3T3) and human mesenchymal stem cells (hMSCs). Advanced microscopic methods allowed for the precise quantification of the function of both cell types. This study has demonstrated that a dose of nanoparticles lower than 20 microg-cm -2 per area of the dish does not negatively affect the cells’ morphology, migration, cytoskeletal function, proliferation, potential for wound healing, and single-cell migration in comparison to standard CellTracker Green CMFDA (5-chloromethylfluorescein diacetate). A higher dose of nanoparticles could be a potential risk for cytoskeletal folding and detachment of the cells from the solid extracellular matrix.

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