Transient receptor potential (TRPs) channels are crucial downstream targets of calcium signalling cascades. They can be modulated either by calcium itself and/or by calcium-binding proteins (CBPs). Intracellular messengers usually interact with binding domains present at the most variable TRP regions-N- and C-cytoplasmic termini. Calmodulin (CaM) is a calcium-dependent cytosolic protein serving as a modulator of most transmembrane receptors. Although CaM-binding domains are widespread within intracellular parts of TRPs, no such binding domain has been characterised at the TRP melastatin member-the transient receptor potential melastatin 6 (TRPM6) channel. Another CBP, the S100 calcium-binding protein A1 (S100A1), is also known for its modulatory activities towards receptors. S100A1 commonly shares a CaM-binding domain. Here, we present the first identified CaM and S100A1 binding sites at the N-terminal of TRPM6. We have confirmed the L520-R535 N-terminal TRPM6 domain as a shared binding site for CaM and S100A1 using biophysical and molecular modelling methods. A specific domain of basic amino acid residues (R526/R531/K532/R535) present at this TRPM6 domain has been identified as crucial to maintain non-covalent interactions with the ligands. Our data unambiguously confirm that CaM and S100A1 share the same binding domain at the TRPM6 N-terminus although the ligand-binding mechanism is different.

• Klíčová slova
• CaM and S100A1, TRPM6, binding domain, calmodulin binding motif, fluorescence anisotropy, molecular modelling,
• MeSH
• kalmodulin chemie   MeSH
• kationtové kanály TRPM chemie   MeSH
• lidé MeSH
• molekulární modely * MeSH
• proteinové domény MeSH
• proteiny S100 chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• kalmodulin MeSH
• kationtové kanály TRPM MeSH
• proteiny S100 MeSH
• S100A1 protein MeSH Prohlížeč
• TRPM6 protein, human MeSH Prohlížeč

The spring-loaded camming device (SLCD), also known as "friend", is a simple mechanism used to ensure the safety of the climber through fall prevention. SLCD consists of two pairs of opposing cams rotating separately, with one (single-axle SLCD) or two (dual-axle SLCD) pins connecting the opposing cams, a stem, connected to the pins, providing the attachment of the climbing rope, springs, which simultaneously push cams to a fully expanded position, and an operating element controlling the cam position. The expansion of cams is thus adaptable to allow insertion or removal of the device into/from a rock crack. While the pins, stem, operating element, and springs can be considered optimal, the (especially internal) shape of the cam allows space for improvement, especially where the weight is concerned. This paper focuses on optimizing the internal shape of the dual-axle SLCD cam from the perspective of the weight/stiffness trade-off. For this purpose, two computational models are designed and multi-step topology optimization (TOP) are performed. From the computational models' point of view, SLCD is considered symmetric and only one cam is optimized and smoothened using parametric curves. Finally, the load-bearing capacity of the new cam design is analyzed. This work is based on practical industry requirements, and the obtained results will be reflected in a new commercial design of SLCD.

• Klíčová slova
• ANSYS, FEM, MATLAB, NURBS, SIMP, SLCD, cam, friend, topology optimization,
• Publikační typ
• časopisecké články MeSH

OBJECTIVES: During the physiological ageing process atrophy of the alveolar bone appears in vertical direction. This bone resorption causes pushing the limits of the maxillary sinus at the expense of a degraded bone. The sinus volume increases due to the facial development in children and adolescents or during the ageing process due to the loss of teeth and bone mass. The main aim of this study is to determine the sinus shape and sinus floor morphology related to age. MATERIALS AND METHODS: Human adult male and female cadaveric heads (aged 37 to 83 years) with different dental status were used. The three-dimensional CAD/CAM software was used to scan the solid impressions of the maxillary sinus to visualize the real sinus shape and sinus floor. Subsequently, other findings are shown in tables and evaluated graphically. RESULTS: The maxillary sinus morphology, its relationship to the nasal cavity, the sub sinus alveolar bone height, displacement of the lowest and highest points of sinus, and the sinus relationship to the roots of the upper teeth were studied and evaluated. Some septa, crests, and the prominent infraorbital canal were also found in the area of the sinus floor. CONCLUSIONS: This paper provides a unique view on the maxillary sinus and its changes during the ageing process with preserved topographical relations in a representative sample of the Slovak population. The visualization of the maxillary sinus anatomy is necessary in the diagnosis and treatment plans for dental implants and during current surgical procedures.

• Klíčová slova
• 3D imaging, 3D scanning, Ageing process, Alveolar recess, Maxillary sinus, Morphology,
• MeSH
• anatomické modely MeSH
• design s pomocí počítače * MeSH
• dospělí MeSH
• kosti a kostní tkáň anatomie a histologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mrtvola MeSH
• nosní dutina anatomie a histologie   růst a vývoj   MeSH
• počítačové zpracování obrazu MeSH
• processus alveolaris anatomie a histologie   MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• sinus maxillaris anatomie a histologie   růst a vývoj   MeSH
• stárnutí fyziologie   MeSH
• zobrazování trojrozměrné metody   MeSH
• zuby anatomie a histologie   růst a vývoj   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Slovenská republika MeSH

Cancer cells facilitate tumor growth by creating favorable tumor micro-environments (TME), altering homeostasis and immune response in the extracellular matrix (ECM) of surrounding tissue. A potential factor that contributes to TME generation and ECM remodeling is the cytoskeleton-associated human death-associated protein kinase 1 (DAPK1). Increased tumor cell motility and de-adhesion (thus, promoting metastasis), as well as upregulated plasminogen-signaling, are shown when functionally analyzing the DAPK1 ko-related proteome. However, the systematic investigation of how tumor cells actively modulate the ECM at the tissue level is experimentally challenging since animal models do not allow direct experimental access while artificial in vitro scaffolds cannot simulate the entire complexity of tissue systems. Here, we used the chorioallantoic membrane (CAM) assay as a natural, collagen-rich tissue model in combination with all-optical experimental access by multiphoton microscopy (MPM) to study the ECM remodeling potential of colorectal tumor cells with and without DAPK1 in situ and even in vivo. This approach demonstrates the suitability of the CAM assay in combination with multiphoton microscopy for studying collagen remodeling during tumor growth. Our results indicate the high ECM remodeling potential of DAPK1 ko tumor cells at the tissue level and support our findings from proteomics.

• Klíčová slova
• ECM remodeling, MPM, collagen, colon cancer, uPAR,
• Publikační typ
• časopisecké články MeSH

OBJECTIVE: To describe the epidemiology and time spectrum of delirium using Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition criteria and to validate a tool for delirium assessment in patients in the acute poststroke period. DESIGN: A prospective observational cohort study. SETTING: The stroke unit of a university hospital. PATIENTS: A consecutive series of 129 patients with stroke (with infarction or intracerebral hemorrhage, 57 women and 72 men; mean age, 72.5 yrs; age range, 35-93 yrs) admitted to the stroke unit of a university hospital were evaluated for delirium incidence. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Criterion validity and overall accuracy of the Czech version of the Confusion Assessment Method for the Intensive Care Unit (CAM-ICU) were determined using serial daily delirium assessments with CAM-ICU by a junior physician compared with delirium diagnosis by delirium experts using the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition criteria that began the first day after stroke onset and continued for at least 7 days. Cox regression models using time-dependent covariate analysis adjusting for age, gender, prestroke dementia, National Institutes of Stroke Health Care at admission, first-day Sequential Organ Failure Assessment, and asphasia were used to understand the relationships between delirium and clinical outcomes. An episode of delirium based on reference Diagnostic and Statistical Manual assessment was detected in 55 patients with stroke (42.6%). In 37 of these (67.3%), delirium began within the first day and in all of them within 5 days of stroke onset. A total of 1003 paired CAM-ICU/Diagnostic and Statistical Manual of Mental Disorders daily assessments were completed. Compared with the reference standard for diagnosing delirium, the CAM-ICU demonstrated a sensitivity of 76% (95% confidence interval [CI] 55% to 91%), a specificity of 98% (95% CI 93% to 100%), an overall accuracy of 94% (95% CI 88% to 97%), and high interrater reliability (κ = 0.94; 95% CI 0.83-1.0). The likelihood ratio of the CAM-ICU in the diagnosis of delirium was 47 (95% CI 27-83). Delirium was an independent predictor of increased length of hospital stay (hazard ratio 1.63; 95% CI 1.11-2.38; p = .013). CONCLUSIONS: Poststroke delirium may frequently be detected provided that the testing algorithm is appropriate to the time profile of poststroke delirium. Early (first day after stroke onset) and serial screening for delirium is recommended. CAM-ICU is a valid instrument for the diagnosis of delirium and should be considered an aid in delirium screening and assessment in future epidemiologic and interventional studies in patients with stroke.

• MeSH
• cévní mozková příhoda diagnóza   mortalita   terapie   MeSH
• delirium diagnóza   epidemiologie   terapie   MeSH
• dospělí MeSH
• hodnocení rizik MeSH
• jednotky intenzivní péče * MeSH
• kohortové studie MeSH
• lidé středního věku MeSH
• lidé MeSH
• míra přežití MeSH
• následné studie MeSH
• nemocnice univerzitní MeSH
• neparametrická statistika MeSH
• neuropsychologické testy * MeSH
• péče o pacienty v kritickém stavu metody   MeSH
• progrese nemoci MeSH
• proporcionální rizikové modely MeSH
• prospektivní studie MeSH
• reprodukovatelnost výsledků MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• stupeň závažnosti nemoci MeSH
• zmatenost klasifikace   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Molecular determinants of the binding of various endogenous modulators to transient receptor potential (TRP) channels are crucial for the understanding of necessary cellular pathways, as well as new paths for rational drug designs. The aim of this study was to characterise interactions between the TRP cation channel subfamily melastatin member 4 (TRPM4) and endogenous intracellular modulators-calcium-binding proteins (calmodulin (CaM) and S100A1) and phosphatidylinositol 4, 5-bisphosphate (PIP2). We have found binding epitopes at the N- and C-termini of TRPM4 shared by CaM, S100A1 and PIP2. The binding affinities of short peptides representing the binding epitopes of N- and C-termini were measured by means of fluorescence anisotropy (FA). The importance of representative basic amino acids and their combinations from both peptides for the binding of endogenous TRPM4 modulators was proved using point alanine-scanning mutagenesis. In silico protein-protein docking of both peptides to CaM and S100A1 and extensive molecular dynamics (MD) simulations enabled the description of key stabilising interactions at the atomic level. Recently solved cryo-Electron Microscopy (EM) structures made it possible to put our findings into the context of the entire TRPM4 channel and to deduce how the binding of these endogenous modulators could allosterically affect the gating of TRPM4. Moreover, both identified binding epitopes seem to be ideally positioned to mediate the involvement of TRPM4 in higher-order hetero-multimeric complexes with important physiological functions.

• Klíčová slova
• CaM, PIP2, S100A1, TRPM4 channel, binding epitope, docking, fluorescence anisotropy, molecular dynamics simulations,
• MeSH
• akvaporiny chemie   metabolismus   MeSH
• interakční proteinové domény a motivy * MeSH
• kalmodulin chemie   metabolismus   MeSH
• kationtové kanály TRPM chemie   metabolismus   MeSH
• kinetika MeSH
• konformace proteinů MeSH
• lidé MeSH
• molekulární modely MeSH
• multiproteinové komplexy chemie   metabolismus   MeSH
• peptidové fragmenty MeSH
• proteiny S100 chemie   metabolismus   MeSH
• sekvence aminokyselin MeSH
• vazba proteinů MeSH
• vazebná místa * MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• akvaporiny MeSH
• kalmodulin MeSH
• kationtové kanály TRPM MeSH
• multiproteinové komplexy MeSH
• peptidové fragmenty MeSH
• proteiny S100 MeSH
• S100A1 protein MeSH Prohlížeč
• TRPM4 protein, human MeSH Prohlížeč

BACKGROUND: The aim of this paper was to design a finite element model for a hinged PROSPON oncological knee endoprosthesis and to verify the model by comparison with ankle flexion angle using knee-bending experimental data obtained previously. METHOD: Visible Human Project CT scans were used to create a general lower extremity bones model and to compose a 3D CAD knee joint model to which muscles and ligaments were added. Into the assembly the designed finite element PROSPON prosthesis model was integrated and an analysis focused on the PEEK-OPTIMA hinge pin bushing stress state was carried out. To confirm the stress state analysis results, contact pressure was investigated. The analysis was performed in the knee-bending position within 15.4-69.4° hip joint flexion range. RESULTS: The results showed that the maximum stress achieved during the analysis (46.6 MPa) did not exceed the yield strength of the material (90 MPa); the condition of plastic stability was therefore met. The stress state analysis results were confirmed by the distribution of contact pressure during knee-bending. CONCLUSION: The applicability of our designed finite element model for the real implant behaviour prediction was proven on the basis of good correlation of the analytical and experimental ankle flexion angle data.

• Klíčová slova
• Endoprosthesis, Finite element method, Finite element model, Knee joint, Knee-bending, Oncological implant,
• MeSH
• algoritmy MeSH
• analýza metodou konečných prvků MeSH
• analýza selhání vybavení MeSH
• biologické modely * MeSH
• design s pomocí počítače MeSH
• kolenní kloub patofyziologie   MeSH
• kosterní svaly patofyziologie   MeSH
• lidé MeSH
• mechanický stres MeSH
• modul pružnosti MeSH
• nádory kostí patofyziologie   chirurgie   MeSH
• pevnost v tahu MeSH
• pevnost v tlaku MeSH
• počítačová simulace MeSH
• protetické vybavení metody   MeSH
• protézy - design MeSH
• protézy kolene * MeSH
• šlachy patofyziologie   MeSH
• software MeSH
• svalová kontrakce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• validační studie MeSH

The combination of in ovo and ex ovo chorioallantoic membrane (CAM) assay provides an excellent platform which extends its relevance in studying carcinogenesis to the field of screening of anticancer activity of platinum nanoparticles (PtNPs) and further study of the amino acids' fluctuations in liver and brain. PtNPs are promising candidates for replacing cisplatin (CDDP); however, insufficient data of their antitumor efficiency and activity on the cancer-related amino acid metabolism are available, and the assessment of the in vivo performance has barely scratched the surface. Herein, we used CAM assay as in vivo model for screening of novel therapeutic modalities, and we conducted a comparative study of the effects of CDDP and polyvinylpyrrolidone coated PtNPs on MDA-MB-231 breast cancer xenograft. PtNPs showed a higher efficiency to inhibit the tumor growth and metastasis compared to CDDP. The amino acids profiling in the MDA-MB-231 ​cells revealed that the PtNPs had an overall depleting effect on the amino acids content. Noteworthy, more side effects to amino acid metabolism were deduced from the depletion of the amino acids in tumor, brain, and liver upon CDDP treatment. Different sets of enzymes of the tricarboxylic acid (TCA) cycle were targeted by PtNPs and CDDP, and while mRNA encoding multiple enzymes was downregulated by PtNPs, the treatment with CDDP affected only two TCA enzymes, indicating a different mechanism of action. Taken together, CAM assay represents and invaluable model, demonstrating the PtNPs capability of repressing angiogenesis, decrease amino acid contents and disrupt the TCA cycle.

• Klíčová slova
• Amino acids metabolism, Breast cancer, CAM assay, Cisplatin, Platinum nanoparticles, TCA cycle,
• Publikační typ
• časopisecké články MeSH

In recent years, the chorioallantoic membrane (CAM) has emerged as a crucial component of biocompatibility testing for biomaterials designed for regenerative strategies and tissue engineering applications. This study explores angiogenic potential of an innovative acellular and porous biopolymer scaffold, based on polyhydroxybutyrate and chitosan (PHB/CHIT), using the ex ovo quail CAM assay as an alternative to the conventional chick CAM test. On embryonic day 6 (ED6), we placed the tested biomaterials on the CAM alone or soaked them with various substances, including vascular endothelial growth factor (VEGF-A), saline, or the endogenous angiogenesis inhibitor Angiostatin. After 72 h (ED9), we analyzed blood vessels formation, a sign of ongoing angiogenesis, in the vicinity of the scaffold and within its pores. We employed marker for cell proliferation (PHH3), embryonic endothelium (WGA, SNA), myofibroblasts (α-SMA), and endothelial cells (QH1) for morphological and histochemical analysis. Our findings demonstrated the robust angiogenic potential of the untreated scaffold without additional influence from the angiogenic factor VEGF-A. Furthermore, gene expression analysis revealed an upregulation of pro-angiogenic growth factors, including VEGF-A, ANG-2, and VE-Cadherin after 5 days of implantation, indicative of a pro-angiogenic microenvironment. These results underscore the inherent angiogenic potential of the PHB/CHIT composite. Additionally, monitoring of CAM microvilli growing to the scaffold provides a methodology for investigating the biocompatibility of materials using the ex ovo quail CAM assay as a suitable alternative model compared to the chicken CAM platform. This approach offers a rapid screening method for biomaterials in the field of tissue repair/regeneration and engineering.

• Klíčová slova
• Angiogenesis, Avian animal model, Bone regeneration, Chitosan, Polyhydroxybutyrate,
• MeSH
• biokompatibilní materiály * farmakologie   MeSH
• chitosan * farmakologie   MeSH
• chorioalantoická membrána * účinky léků   MeSH
• fyziologická neovaskularizace účinky léků   MeSH
• křepelky a křepelovití embryologie   MeSH
• testování materiálů MeSH
• tkáňové podpůrné struktury chemie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• biokompatibilní materiály * MeSH
• chitosan * MeSH

TRPM3 has been reported to play an important role in Ca(2+) homeostasis, but its gating mechanisms and regulation via Ca(2+) are unknown. Ca(2+) binding proteins such as calmodulin (CaM) could be probable modulators of this ion channel. We have shown that this protein binds to two independent domains, A35-K124 and H291-G382 on the TRPM3 N-terminus, which contain conserved hydrophobic as well as positively charged residues in specific positions, and that these residues have a crucial impact on its binding. We also showed that the other Ca(2+) binding protein, S100A1, is able to bind to these regions and that CaM and S100A1 compete for these binding sites on the TRPM3 N-terminus. Moreover, our results suggest that another very important TRP channel activity modulator, PtdIns(4,5)P(2), interacts with the CaM/S100A1 binding sites on the TRPM3 N-terminus with high affinity.

• MeSH
• fluorescenční polarizace MeSH
• fosfatidylinositol-4,5-difosfát metabolismus   MeSH
• kalmodulin metabolismus   MeSH
• kationtové kanály TRPM chemie   metabolismus   MeSH
• liposomy metabolismus   MeSH
• molekulární modely MeSH
• povrchová plasmonová rezonance MeSH
• proteiny S100 metabolismus   MeSH
• terciární struktura proteinů MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fosfatidylinositol-4,5-difosfát MeSH
• kalmodulin MeSH
• kationtové kanály TRPM MeSH
• liposomy MeSH
• proteiny S100 MeSH
• S100A1 protein MeSH Prohlížeč