By using indirect immunofluorescence we demonstrated the localisation of extracellular matrix (ECM) proteins (laminin--LAM, collagen IV--COL IV, fibronectin--FN) and the basic fibroblast growth factor (bFGF) in rabbit and mouse primary embryonic fibroblasts (PEF). Proliferating mitotically arrested PEF (by mitomycin C) were compared in both species. The stability of protein expression was ascertained during the first five successive passages. In addition, STO cells (i.e. permanent line of irradiated mouse fibroblasts) were similarly analysed. Rabbit PEF showed very high extracellular staining for FN and a negligible cytoplasmic positivity for LAM and COL IV. A totally reversed staining pattern for ECM proteins was found in mouse PEF. A dense cytoplasmic granulation (concentrated around the nucleus) was revealed for LAM and COL IV and almost no reaction for FN. The staining patterns were very stable at the culture conditions we applied. They were maintained during the first five successive passages in proliferating as well as non-proliferating mouse and rabbit PEF and were independent of cell concentration (individually dispersed cells versus cells in a confluent layer). STO cells showed the same staining for ECM proteins as the mouse PEF, thus confirming their origin from the same animal species. Light granular staining for bFGF was found in the cytoplasm of proliferating and mitotically arrested rabbit and mouse PEF and STO cells. The differences in expression of ECM proteins between the rabbit and mouse PEF, as well as the synthesis of bFGF, should be taken into consideration when these cells are used in vitro as a feeder layer for various cells (e.g. embryonic stem cells).

• MeSH
• buněčné dělení MeSH
• embryo savčí * MeSH
• fibroblastový růstový faktor 2 analýza   MeSH
• fibroblasty chemie   MeSH
• fibronektiny analýza   MeSH
• fluorescenční protilátková technika nepřímá MeSH
• imunohistochemie * MeSH
• inhibitory syntézy nukleových kyselin farmakologie   MeSH
• keratiny analýza   MeSH
• kolagen analýza   MeSH
• králíci MeSH
• laminin analýza   MeSH
• mitomycin farmakologie   MeSH
• mitóza účinky léků   MeSH
• myši inbrední ICR MeSH
• myši MeSH
• vimentin analýza   MeSH
• zvířata MeSH
• Check Tag
• králíci MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fibroblastový růstový faktor 2 MeSH
• fibronektiny MeSH
• inhibitory syntézy nukleových kyselin MeSH
• keratiny MeSH
• kolagen MeSH
• laminin MeSH
• mitomycin MeSH
• vimentin MeSH

Suspension culture of mouse fibroblast cell line L-A 115 was used to test beryllium toxicity in the presence of magnesium ions. Beryllium added to the MEM cultivation medium was bound in a complex with sulphosalicylic acid BeSSA complex, because the use of beryllium chloride turned out to yield ineffective beryllium phosphate that formed macroscopically detectable insoluble opacities. The BeSSA complex was used in the concentration range: 10(-3)--10(-9)M, magnesium was used in 3 concentrations: 10(-1)M, 5 x 10(-2)M and 10(-2)M. Growth curve analysis revealed pronounced beryllium toxicity at the concentration of 10(-3)M, magnesium-produced toxic changes were observed only at the concentration of 10(-1)M. No competition between the beryllium and magnesium ions was recorded. It is assumed that the possible beryllium-magnesium competition was significantly modified by the use of BeSSA complex-bound beryllium.

• MeSH
• beryllium farmakologie   toxicita   MeSH
• buněčné dělení účinky léků   MeSH
• buněčné linie MeSH
• fibroblasty MeSH
• myši MeSH
• síran hořečnatý farmakologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• beryllium MeSH
• síran hořečnatý MeSH

BACKGROUND INFORMATION: Multipotent mesenchymal stem cells can participate in the formation of a microenvironment stimulating the aggressive behaviour of cancer cells. Moreover, cells exhibiting pluripotent ESC (embryonic stem cell) markers (Nanog and Oct4) have been observed in many tumours. Here, we investigate the role of cancer-associated fibroblasts in the formation of stem cell supporting properties of tumour stroma. We test the influence of fibroblasts isolated from basal cell carcinoma on mouse 3T3 fibroblasts, focusing on the expression of stem cell markers and plasticity in vitro by means of microarrays, qRT-PCR (quantitative real-time PCR) and immunohistochemistry. RESULTS: We demonstrate the biological activity of the cancer stromal fibroblasts by influencing the 3T3 fibroblasts to express markers such as Oct4, Nanog and Sox2 and to show differentiation potential similar to mesenchymal stem cells. The role of growth factors such as IGF2 (insulin-like growth factor 2), FGF7 (fibroblast growth factor 7), LEP (leptin), NGF (nerve growth factor) and TGFβ (transforming growth factor β), produced by the stromal fibroblasts, is established to participate in their bioactivity. Uninduced 3T3 do not express the stem cell markers and show minimal differentiation potential. CONCLUSIONS: Our observations indicate the pro-stem cell activity of cancer-associated fibroblasts and underline the role of epithelial-mesenchymal interaction in tumour biology.

• MeSH
• bazocelulární karcinom patologie   MeSH
• buněčná diferenciace MeSH
• buňky 3T3 MeSH
• buňky stromatu metabolismus   patologie   MeSH
• epitelo-mezenchymální tranzice MeSH
• fenotyp MeSH
• fibroblasty metabolismus   patologie   MeSH
• imunohistochemie MeSH
• keratinocyty cytologie   MeSH
• kokultivační techniky MeSH
• kultivované buňky MeSH
• lidé MeSH
• mezenchymální kmenové buňky metabolismus   patologie   MeSH
• multipotentní kmenové buňky metabolismus   patologie   MeSH
• myši MeSH
• nádorové biomarkery metabolismus   MeSH
• polymerázová řetězová reakce s reverzní transkripcí MeSH
• separace buněk MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• nádorové biomarkery MeSH

Fibroblast growth factor (FGF) signaling is crucial for mammary gland development. Although multiple roles for FGF signaling in the epithelium have been described, the function of FGF signaling in mammary stroma has not been elucidated. In this study, we investigated FGF signaling in mammary fibroblasts. We found that murine mammary fibroblasts express FGF receptors FGFR1 and FGFR2 and respond to FGF ligands. In particular, FGF2 and FGF9 induce sustained ERK1/2 signaling and promote fibroblast proliferation and migration in 2D cultures. Intriguingly, only FGF2 induces fibroblast migration in 3D extracellular matrix (ECM) through regulation of actomyosin cytoskeleton and promotes force-mediated collagen remodeling by mammary fibroblasts. Moreover, FGF2 regulates production of ECM proteins by mammary fibroblasts, including collagens, fibronectin, osteopontin and matrix metalloproteinases. Finally, using organotypic 3D co-cultures we show that FGF2 and FGF9 signaling in mammary fibroblasts enhances fibroblast-induced branching of mammary epithelium by modulating paracrine signaling, and that knockdown of Fgfr1 and Fgfr2 in mammary fibroblasts reduces branching of mammary epithelium. Our results demonstrate a pleiotropic role for FGF signaling in mammary fibroblasts, with implications for regulation of mammary stromal functions and epithelial branching morphogenesis.

• Klíčová slova
• Branching morphogenesis, Collagen, Extracellular matrix, Fibroblast, Fibroblast growth factor, Mammary gland, Mouse, Stroma,
• MeSH
• fibroblastový růstový faktor 2 metabolismus   MeSH
• fibroblastový růstový faktor 9 metabolismus   MeSH
• fibroblasty cytologie   metabolismus   MeSH
• MAP kinasový signální systém * MeSH
• mléčné žlázy zvířat cytologie   embryologie   MeSH
• myši inbrední ICR MeSH
• myši MeSH
• parakrinní signalizace * MeSH
• receptor fibroblastových růstových faktorů, typ 1 metabolismus   MeSH
• receptor fibroblastových růstových faktorů, typ 2 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• Fgf9 protein, mouse MeSH Prohlížeč
• Fgfr1 protein, mouse MeSH Prohlížeč
• Fgfr2 protein, mouse MeSH Prohlížeč
• fibroblastový růstový faktor 2 MeSH
• fibroblastový růstový faktor 9 MeSH
• receptor fibroblastových růstových faktorů, typ 1 MeSH
• receptor fibroblastových růstových faktorů, typ 2 MeSH

• MeSH
• chromozomy MeSH
• dimethylsulfoxid MeSH
• fibroblasty * MeSH
• glycerol MeSH
• karyotypizace * MeSH
• lyofilizace MeSH
• myši MeSH
• uchovávání tkání MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• dimethylsulfoxid MeSH
• glycerol MeSH

Patch clamp recordings carried out in the inside-out configuration revealed activity of three kinds of channels: nonselective cation channels, small-conductance K(+) channels, and large-conductance anion channels. The nonselective cation channels did not distinguish between Na(+) and K(+). The unitary conductance of these channels reached 28 pS in a symmetrical concentration of 200 mM NaCl. A lower value of this parameter was recorded for the small-conductance K(+) channels and in a 50-fold gradient of K(+) (200 mM/4 mM) it reached 8 pS. The high selectivity of these channels to potassium was confirmed by the reversal potential (-97 mV), whose value was close to the equilibrium potential for potassium (-100 mV). One of the features of the largeconductance anion channels was high conductance amounting to 493 pS in a symmetrical concentration of 200 mM NaCl. The channels exhibited three subconductance levels. Moreover, an increase in the open probability of the channels at voltages close to zero was observed. The anion selectivity of the channels was low, because the channels were permeable to both Cl(-) and gluconate - a large anion. Research on the calcium dependence revealed that internal calcium activates nonselective cation channels and small-conductance K(+) channels, but not largeconductance anion channels.

• MeSH
• buněčná membrána fyziologie   MeSH
• buněčné linie MeSH
• fibroblasty fyziologie   MeSH
• iontové kanály fyziologie   MeSH
• myši MeSH
• napětím ovládané aniontové kanály fyziologie   MeSH
• nízkovodivostní draslíkové kanály aktivované vápníkem fyziologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• iontové kanály MeSH
• napětím ovládané aniontové kanály MeSH
• nízkovodivostní draslíkové kanály aktivované vápníkem MeSH
• nonselective cation channel protein, mouse MeSH Prohlížeč

There are several reports in the literature focusing on regulation of major histocompatibility complex (MHC) class I genes by transcription factors of the jun family. The methods employed in these reports differed in various respects, and their results are inconsistent. In mouse Lewis lung carcinoma, B16-melanoma and F9-teratocarcinoma cell lines, c-jun was characterized as a transcriptional activator of the murine MHC class I H2-Kb gene, while c-jun was identified as a direct transcriptional repressor of the swine class I PD1 gene, and c-jun stably transfected clones of mouse L-fibroblasts markedly reduced their H-2 class I gene expression. In this study, we attempted to reproduce this last effect by means of transient transfection coupled to Northern hybridization, upon transfecting L-fibroblasts with expression vectors for all jun family members as well as with an array of c-jun-derived dominant negative mutants. No change in H-2 class I expression could be identified. Next, we derived two additional fibroblastic cell lines from the fibrosarcoma of the H2-Kk/v-jun transgenic mouse and transfected them with the two most potent c-jun dominant negative mutants, again without eliciting any change in H-2 class I mRNA level. We conclude that the negative regulation of H-2 class I genes by c-jun in cells of the fibroblastic lineage is not a primary effect.

• MeSH
• fibroblasty metabolismus   MeSH
• H-2 antigeny biosyntéza   genetika   MeSH
• myši MeSH
• onkogenní protein p65(gag-jun) genetika   metabolismus   MeSH
• transfekce MeSH
• transkripční faktory metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• H-2 antigeny MeSH
• onkogenní protein p65(gag-jun) MeSH
• transkripční faktory MeSH

Telomere repeats are added onto chromosome ends by telomerase, consisting of two main core components: a catalytic protein subunit (telomerase reverse trancriptase, TERT), and an RNA subunit (telomerase RNA, TR). Here, we report for the first time evidence that HMGB1 (a chromatin-associated protein in mammals, acting as a DNA chaperone in transcription, replication, recombination, and repair) can modulate cellular activity of mammalian telomerase. Knockout of the HMGB1 gene (HMGB1 KO) in mouse embryonic fibroblasts (MEFs) results in chromosomal abnormalities, enhanced colocalization of γ-H2AX foci at telomeres, and a moderate shortening of telomere lengths. HMGB1 KO MEFs also exhibit significantly (>5-fold) lower telomerase activity than the wild-type MEFs. Correspondingly, enhanced telomerase activity is observed upon overexpression of HMGB1 in MEFs. HMGB1 physically interacts with both TERT and TR, as well as with active telomerase complex in vitro. However, direct interaction of HMGB1 with telomerase is most likely not accountable for the observed higher telomerase activity in HMGB1-containing cells, as revealed from the inability of purified HMGB1 protein to stimulate telomerase activity in vitro. While no transcriptional silencing of TERT is observed in HMGB1 KO MEFs, levels of TR are diminished (~3-fold), providing possible explanation for the observed lower telomerase activity in HMGB1 KO cells. Interestingly, knockout of the HMGB2 gene elevates telomerase activity (~3-fold) in MEFs, suggesting that the two closely related proteins of the HMGB family, HMGB1 and HMGB2, have opposite effects on telomerase activity in the cell. The ability of HMGB1 to modulate cellular activity of telomerase and to maintain telomere integrity can help to understand some aspects of the protein involvement in chromosome stability and cancer.

• MeSH
• buněčné linie MeSH
• chromozomální aberace MeSH
• down regulace MeSH
• fibroblasty cytologie   metabolismus   MeSH
• fluorescenční mikroskopie MeSH
• fragmentace DNA MeSH
• genový knockout * MeSH
• histony genetika   metabolismus   MeSH
• hybridizace in situ fluorescenční MeSH
• myši MeSH
• poškození DNA MeSH
• protein HMGB1 genetika   metabolismus   MeSH
• protein HMGB2 genetika   metabolismus   MeSH
• replikace DNA MeSH
• RNA genetika   metabolismus   MeSH
• telomerasa genetika   metabolismus   MeSH
• telomery metabolismus   patologie   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• gamma-H2AX protein, mouse MeSH Prohlížeč
• histony MeSH
• protein HMGB1 MeSH
• protein HMGB2 MeSH
• RNA MeSH
• telomerasa MeSH
• telomerase RNA MeSH Prohlížeč
• Tert protein, mouse MeSH Prohlížeč

The administration of antioxidants has been shown to enhance repair and healing processes in cutaneous tissue. Silymarin, an extract from Silybum marianum has been reported to be beneficial in the treatment of chemically-induced oxidative stress in mouse. In this study, we investigated the protective effects of silymarin, its flavonolignans silybin and dehydrosilybin and flavonoids quercetin and taxifolin against hydrogen peroxide-induced damage to human keratinocytes and mouse fibroblasts. The results showed that the cytotoxicity of hydrogen peroxide was dose-dependent in both cell lines. Pre-treatment with test compounds decreased oxidative injury. Dehydrosilybin and quercetin were the most powerful protectants. Silymarin was comparable to silybinin, its main component. This correlates with the antioxidant potential of the compounds. Our findings suggest that silymarin, flavonolignans and flavonoids may be useful as agents for improving skin tissue regeneration.

• MeSH
• fibroblasty metabolismus   MeSH
• flavonolignany chemie   farmakologie   MeSH
• flavonoly chemie   farmakologie   MeSH
• fytoterapie MeSH
• keratinocyty metabolismus   MeSH
• kultivované buňky MeSH
• lidé MeSH
• myši inbrední BALB C MeSH
• myši MeSH
• ochranné látky farmakologie   MeSH
• oxidační stres účinky léků   MeSH
• oxidancia škodlivé účinky   MeSH
• peroxid vodíku škodlivé účinky   MeSH
• quercetin analogy a deriváty   chemie   farmakologie   MeSH
• rostlinné extrakty chemie   farmakologie   MeSH
• silibinin MeSH
• silymarin chemie   farmakologie   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• dehydrosilybin MeSH Prohlížeč
• flavonolignany MeSH
• flavonoly MeSH
• ochranné látky MeSH
• oxidancia MeSH
• peroxid vodíku MeSH
• quercetin MeSH
• rostlinné extrakty MeSH
• silibinin MeSH
• silymarin MeSH
• taxifolin MeSH Prohlížeč

The tumorigenic potential of mouse polyomavirus (MPyV) has been studied for decades in cell culture models and has been mainly attributed to nonstructural middle T antigen (MT), which acts as a scaffold signal adaptor, activates Src tyrosine kinases, and possesses transforming ability. We hypothesized that MPyV could also transform mouse cells independent of MT via a Toll-like receptor 4 (TLR4)-mediated inflammatory mechanism. To this end, we investigated the interaction of MPyV with TLR4 in mouse embryonic fibroblasts (MEFs) and 3T6 cells, resulting in secretion of interleukin 6 (IL-6), independent of active viral replication. TLR4 colocalized with MPyV capsid protein VP1 in MEFs. Neither TLR4 activation nor recombinant IL-6 inhibited MPyV replication in MEFs and 3T6 cells. MPyV induced STAT3 phosphorylation through both direct and MT-dependent and indirect and TLR4/IL-6-dependent mechanisms. We demonstrate that uninfected mouse fibroblasts exposed to the cytokine environment from MPyV-infected fibroblasts upregulated the expressions of MCP-1, CCL-5, and α-SMA. Moreover, the cytokine microenvironment increased the invasiveness of MEFs and CT26 carcinoma cells. Collectively, TLR4 recognition of MPyV induces a cytokine environment that promotes the cancer-associated fibroblast (CAF)-like phenotype in noninfected fibroblasts and increases cell invasiveness.

• Klíčová slova
• CAF, IL-6, MPyV, TLR4, mouse fibroblasts, mouse polyomavirus, spheroid invasiveness,
• Publikační typ
• časopisecké články MeSH