Collagen I-based foams were modified with calcined or noncalcined hydroxyapatite or calcium phosphates with various particle sizes and pores to monitor their effect on cell interactions. The resulting scaffolds thus differed in grain size, changing from nanoscale to microscopic, and possessed diverse morphological characteristics and resorbability. The materials' biological action was shown on human bone marrow MSCs. Scaffold morphology was identified by SEM. Using viability test, qPCR, and immunohistochemical staining, we evaluated the biological activity of all of the materials. This study revealed that the most suitable scaffold composition for osteogenesis induction is collagen I foam with calcined hydroxyapatite with a pore size of 360 ± 130 µm and mean particle size of 0.130 µm. The expression of osteogenic markers RunX2 and ColI mRNA was promoted, and a strong synthesis of extracellular protein osteocalcin was observed. ColI/calcined HAP scaffold showed significant osteogenic potential, and can be easily manipulated and tailored to the defect size, which gives it great potential for bone tissue engineering applications.

• Klíčová slova
• bioceramics, collagen, osteogenesis,
• MeSH
• buněčná diferenciace MeSH
• hydroxyapatit * chemie   farmakologie   MeSH
• kolagen typu I genetika   MeSH
• kultivované buňky MeSH
• lidé MeSH
• osteogeneze * MeSH
• tkáňové inženýrství metody   MeSH
• tkáňové podpůrné struktury chemie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• hydroxyapatit * MeSH
• kolagen typu I MeSH

Wound healing is a process regulated by a complex interaction of multiple growth factors including fibroblast growth factor 2 (FGF2). Although FGF2 appears in several tissue engineered studies, its applications are limited due to its low stability both in vitro and in vivo. Here, this shortcoming is overcome by a unique nine-point mutant of the low molecular weight isoform FGF2 retaining full biological activity even after twenty days at 37 °C. Crosslinked freeze-dried 3D porous collagen/chitosan scaffolds enriched with this hyper stable recombinant human protein named FGF2-STAB® were tested for in vitro biocompatibility and cytotoxicity using murine 3T3-A31 fibroblasts, for angiogenic potential using an ex ovo chick chorioallantoic membrane assay and for wound healing in vivo with 3-month old white New Zealand rabbits. Metabolic activity assays indicated the positive effect of FGF2-STAB® already at very low concentrations (0.01 µg/mL). The angiogenic properties examined ex ovo showed enhanced vascularization of the tested scaffolds. Histological evaluation and gene expression analysis by RT-qPCR proved newly formed granulation tissue at the place of a previous skin defect without significant inflammation infiltration in vivo. This work highlights the safety and biocompatibility of newly developed crosslinked collagen/chitosan scaffolds involving FGF2-STAB® protein. Moreover, these sponges could be used as scaffolds for growing cells for dermis replacement, where neovascularization is a crucial parameter for successful skin regeneration.

• Klíčová slova
• FGF2, chitosan, collagen, scaffold, skin regeneration, tissue engineering,
• Publikační typ
• časopisecké články MeSH

Vitiligo is the most common depigmentation disorder of the skin. Currently, its therapy focuses on the halting of the immune response and stimulation of the regenerative processes, leading to the restoration of normal melanocyte function. Platelet-rich plasma (PRP) represents a safe and cheap regenerative therapy option, as it delivers a wide spectrum of native growth factors, cytokines and other bioactive molecules. The aim of this study was to develop a simple delivery system to prolong the effects of the bioactive molecules released from platelets. The surface of electrospun and centrifugally spun poly-ε-caprolactone (PCL) fibrous scaffolds was functionalized with various concentrations of platelets; the influence of the morphology of the scaffolds and the concentration of the released platelet-derived bioactive molecules on melanocytes, was then assessed. An almost two-fold increase in the amount of the released bioactive molecules was detected on the centrifugally spun vs. electrospun scaffolds, and a sustained 14-day release of the bioactive molecules was demonstrated. A strong concentration-dependent response of melanocyte to the bioactive molecules was observed; higher concentrations of bioactive molecules resulted in improved metabolic activity and proliferation of melanocytes. This simple system improves melanocyte viability, offers on-site preparation and is suitable for prolonged topical PRP administration.

• Klíčová slova
• centrifugal spinning, electrospinning, melanocyte, platelets, vitiligo,
• Publikační typ
• časopisecké články MeSH

This study presents a toxicological evaluation of two types of carbon dots (CD), similar in size (<10 nm) but differing in surface charge. Whole-genome mRNA and miRNA expression (RNAseq), as well as gene-specific DNA methylation changes, were analyzed in human embryonic lung fibroblasts (HEL 12469) after 4 h and 24 h exposure to concentrations of 10 and 50 µg/mL (for positive charged CD; pCD) or 10 and 100 µg/mL (for negative charged CD, nCD). The results showed a distinct response for the tested nanomaterials (NMs). The exposure to pCD induced the expression of a substantially lower number of mRNAs than those to nCD, with few commonly differentially expressed genes between the two CDs. For both CDs, the number of deregulated mRNAs increased with the dose and exposure time. The pathway analysis revealed a deregulation of processes associated with immune response, tumorigenesis and cell cycle regulation, after exposure to pCD. For nCD treatment, pathways relating to cell proliferation, apoptosis, oxidative stress, gene expression, and cycle regulation were detected. The expression of miRNAs followed a similar pattern: more pronounced changes after nCD exposure and few commonly differentially expressed miRNAs between the two CDs. For both CDs the pathway analysis based on miRNA-mRNA interactions, showed a deregulation of cancer-related pathways, immune processes and processes involved in extracellular matrix interactions. DNA methylation was not affected by exposure to any of the two CDs. In summary, although the tested CDs induced distinct responses on the level of mRNA and miRNA expression, pathway analyses revealed a potential common biological impact of both NMs independent of their surface charge.

• Klíčová slova
• DNA methylation, carbon dots, gene expression, human lung fibroblasts, surface charge,
• MeSH
• apoptóza účinky léků   genetika   MeSH
• exprese genu účinky léků   genetika   MeSH
• extracelulární matrix genetika   MeSH
• fibroblasty účinky léků   MeSH
• kultivované buňky MeSH
• lidé MeSH
• messenger RNA genetika   MeSH
• metylace DNA účinky léků   genetika   MeSH
• mikro RNA genetika   MeSH
• nádory genetika   MeSH
• oxidační stres účinky léků   genetika   MeSH
• plíce účinky léků   MeSH
• proliferace buněk účinky léků   genetika   MeSH
• signální transdukce účinky léků   genetika   MeSH
• stanovení celkové genové exprese metody   MeSH
• uhlík farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• messenger RNA MeSH
• mikro RNA MeSH
• uhlík MeSH

We investigated the transcriptomic response and epigenetic changes in the lungs of mice exposed to inhalation of copper(II) oxide nanoparticles (CuO NPs) (8 × 105 NPs/m3) for periods of 3 days, 2 weeks, 6 weeks, and 3 months. A whole genome transcriptome and miRNA analysis was performed using next generation sequencing. Global DNA methylation was assessed by ELISA. The inhalation resulted in the deregulation of mRNA transcripts: we detected 170, 590, 534, and 1551 differentially expressed transcripts after 3 days, 2 weeks, 6 weeks, and 3 months of inhalation, respectively. Biological processes and pathways affected by inhalation, differed between 3 days exposure (collagen formation) and longer treatments (immune response). Periods of two weeks exposure further induced apoptotic processes, 6 weeks of inhalation affected the cell cycle, and 3 months of treatment impacted the processes related to cell adhesion. The expression of miRNA was not affected by 3 days of inhalation. Prolonged exposure periods modified miRNA levels, although the numbers were relatively low (17, 18, and 38 miRNAs, for periods of 2 weeks, 6 weeks, and 3 months, respectively). Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways analysis based on miRNA-mRNA interactions, revealed the deregulation of processes implicated in the immune response and carcinogenesis. Global DNA methylation was not significantly affected in any of the exposure periods. In summary, the inhalation of CuO NPs impacted on both mRNA and miRNA expression. A significant transcriptomic response was already observed after 3 days of exposure. The affected biological processes and pathways indicated the negative impacts on the immune system and potential role in carcinogenesis.

• Klíčová slova
• DNA methylation, copper(II) oxide nanoparticles, gene expression, inhalation, mouse,
• Publikační typ
• časopisecké články MeSH

Regulatory T cells (Tregs) play a critical role in the maintenance of a pregnancy. While the kinetics of the number of peripheral blood Tregs has been satisfactorily described in mouse models, analysis of these cell populations in human pregnancy is complicated by high variability in the quantity of Tregs and inconsistencies in the markers used for detecting different types of Treg. In the light of this, we set out to investigate the kinetics of various types of Treg, including CD45RA, GARP and PD-1(+) Tregs, in the peripheral blood of pregnant women in the first, second and third trimester, and at the time of delivery. Tregs, defined as a CD4(+)CD25(++)CD127(dim)Foxp3(+) population of leucocytes, were detected using flow cytometry. Natural thymus-derived Tregs and induced Tregs in the peripheral blood were distinguished by the expression or absence of a Helios marker, respectively. Our results showed that during normal pregnancy the sizes of various Treg subpopulations varied across women and also in an individual woman did not remain constant but varied significantly, most notable being the decrease observed at the time of delivery. Helios(-) cells were significantly less frequent in the peripheral blood of healthy pregnant women than Helios(+) cells, and the majority of Tregs were Helios(+)PD-1(+) Tregs.

• Klíčová slova
• CD45RA, GARP, Helios, PD-1, Treg, normal pregnancy,
• MeSH
• buněčná diferenciace MeSH
• forkhead transkripční faktory metabolismus   MeSH
• imunofenotypizace MeSH
• kultivované buňky MeSH
• lidé MeSH
• počet lymfocytů MeSH
• průtoková cytometrie MeSH
• regulační T-lymfocyty imunologie   MeSH
• T-lymfocyty - podskupiny imunologie   MeSH
• těhotenství imunologie   MeSH
• thymus imunologie   MeSH
• transformující růstový faktor beta krev   MeSH
• transkripční faktor Ikaros metabolismus   MeSH
• zdraví dobrovolníci pro lékařské studie MeSH
• Check Tag
• lidé MeSH
• těhotenství imunologie   MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• forkhead transkripční faktory MeSH
• FOXP3 protein, human MeSH Prohlížeč
• IKZF2 protein, human MeSH Prohlížeč
• transformující růstový faktor beta MeSH
• transkripční faktor Ikaros MeSH