Glioblastomas are aggressive brain tumors for which effective therapy is still lacking, resulting in dismal survival rates. These tumors display significant phenotypic plasticity, harboring diverse cell populations ranging from tumor core cells to dispersed, highly invasive cells. Neuron navigator 3 (NAV3), a microtubule-associated protein affecting microtubule growth and dynamics, is downregulated in various cancers, including glioblastoma, and has thus been considered a tumor suppressor. In this study, we challenge this designation and unveil distinct expression patterns of NAV3 across different invasion phenotypes. Using glioblastoma cell lines and patient-derived glioma stem-like cell cultures, we disclose an upregulation of NAV3 in invading glioblastoma cells, contrasting with its lower expression in cells residing in tumor spheroid cores. Furthermore, we establish an association between low and high NAV3 expression and the amoeboid and mesenchymal invasive phenotype, respectively, and demonstrate that overexpression of NAV3 directly stimulates glioblastoma invasive behavior in both 2D and 3D environments. Consistently, we observed increased NAV3 expression in cells migrating along blood vessels in mouse xenografts. Overall, our results shed light on the role of NAV3 in glioblastoma invasion, providing insights into this lethal aspect of glioblastoma behavior.

• MeSH
• fenotyp * MeSH
• glioblastom * patologie   genetika   metabolismus   MeSH
• invazivní růst nádoru * genetika   MeSH
• lidé MeSH
• membránové proteiny MeSH
• mikrotubuly metabolismus   MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• nádory mozku * patologie   genetika   metabolismus   MeSH
• pohyb buněk genetika   fyziologie   MeSH
• proteiny nervové tkáně metabolismus   genetika   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

The endothelial-mesenchymal transition (EndMT) of endothelial progenitor cells (EPCs) plays a notable role in pathological vascular remodeling. Emerging evidence indicated that long non-coding RNA-regulator of reprogramming (linc-ROR) can promote epithelial-mesenchymal transition (EMT) in a variety of cancer cells. Nevertheless, the function of linc-ROR in EPC EndMT has not been well elucidated. The present study investigated the effect and possible mechanisms of function of linc-ROR on the EndMT of EPCs. A linc-ROR overexpression lentiviral vector (LV linc-ROR) or a linc-ROR short hairpin RNA lentiviral vector (LV-shlinc-ROR) was used to up or downregulate linc-ROR expression in EPCs isolated from human umbilical cord blood. Functional experiments demonstrated that LV-linc-ROR promoted the proliferation and migration of EPCs, but inhibited EPC angiogenesis in vitro. In the meantime, reverse transcription-quantitative PCR and western blotting results showed that the expression of the endothelial cell markers vascular endothelial-cadherin and CD31 was decreased, while the expression of the mesenchymal cell markers ?-smooth muscle actin and SM22? was increased at both mRNA and protein levels in LV-linc-ROR-treated EPCs, indicating that linc-ROR induced EPC EndMT. Mechanistically, the dual-luciferase reporter assay demonstrated that microRNA (miR/miRNA)-145 was a direct target of linc-ROR, and miR-145 binds to the 3'-untranslated region of Smad3. Moreover, LV-shlinc-ROR increased the expression of miR-145, but decreased the expression of Smad3. In conclusion, linc-ROR promotes EPC EndMT, which may be associated with the miR-145/Smad3 signaling pathway. Keywords: Endothelial progenitor cells, Endothelial to mesenchymal transition, Linc-ROR, MiR-145, Atherosclerosis.

• MeSH
• endotel-mezenchymální transformace MeSH
• endoteliální progenitorové buňky * metabolismus   MeSH
• epitelo-mezenchymální tranzice * MeSH
• kultivované buňky MeSH
• lidé MeSH
• mikro RNA * metabolismus   genetika   MeSH
• pohyb buněk fyziologie   MeSH
• proliferace buněk MeSH
• protein Smad3 * metabolismus   genetika   MeSH
• RNA dlouhá nekódující * genetika   metabolismus   MeSH
• signální transdukce * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Gliomagenesis induces profound changes in the composition of the extracellular matrix (ECM) of the brain. In this study, we identified a cellular population responsible for the increased deposition of collagen I and fibronectin in glioblastoma. Elevated levels of the fibrillar proteins collagen I and fibronectin were associated with the expression of fibroblast activation protein (FAP), which is predominantly found in pericyte-like cells in glioblastoma. FAP+ pericyte-like cells were present in regions rich in collagen I and fibronectin in biopsy material and produced substantially more collagen I and fibronectin in vitro compared to other cell types found in the GBM microenvironment. Using mass spectrometry, we demonstrated that 3D matrices produced by FAP+ pericyte-like cells are rich in collagen I and fibronectin and contain several basement membrane proteins. This expression pattern differed markedly from glioma cells. Finally, we have shown that ECM produced by FAP+ pericyte-like cells enhances the migration of glioma cells including glioma stem-like cells, promotes their adhesion, and activates focal adhesion kinase (FAK) signaling. Taken together, our findings establish FAP+ pericyte-like cells as crucial producers of a complex ECM rich in collagen I and fibronectin, facilitating the dissemination of glioma cells through FAK activation.

• MeSH
• endopeptidasy MeSH
• extracelulární matrix * metabolismus   patologie   MeSH
• fibronektiny * metabolismus   MeSH
• glioblastom patologie   metabolismus   MeSH
• gliom * patologie   metabolismus   MeSH
• kolagen typu I metabolismus   MeSH
• lidé MeSH
• membránové proteiny metabolismus   MeSH
• nádorové buněčné linie MeSH
• nádorové mikroprostředí fyziologie   MeSH
• nádory mozku * patologie   metabolismus   MeSH
• pericyty * metabolismus   patologie   MeSH
• pohyb buněk fyziologie   MeSH
• serinové endopeptidasy metabolismus   MeSH
• želatinasy metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

ETHNOPHARMACOLOGICAL RELEVANCE: The prevalence of different types of chronic wounds, due to the ageing population and increase incidence of diseases, is becoming a worldwide problem. Various medicinal plants used in folk medicine have demonstrated wound healing and antimicrobial properties, and some of these species are currently used in commercial preparations. Despite the well-documented and rich tradition of the use of local herbs for the treatment of skin injuries in Samoan folk medicine, their wound healing potential has not yet been systematically studied. AIM OF THE STUDY: Investigation into the in vitro antibacterial activity of ethanol extracts from 14 medicinal plants used in Samoan traditional medicine for the healing of wounds, burns and sores, and their effects on the proliferation and migration of human fibroblasts. MATERIALS AND METHODS: The antibacterial activity of these extracts was tested against pathogens associated with infected skin injuries, using the broth microdilution method. The effect on migration, proliferation and viability of human dermal fibroblasts was evaluated using wound healing scratch assay, cell proliferation assay, and thiazolyl blue tetrazolium bromide cytotoxicity test. RESULTS: The extracts from Cerbera manghas, Commelina diffusa, Kleinhovia hospita, Mikania micrantha, Omalanthus nutans, Peperomia pellucida, Phymatosorus scolopendria, Piper graeffei, Psychotria insularum, and Schizostachyum glaucifolium inhibited the growth of Staphylococcus aureus at the minimum inhibitory concentration (MIC) of ≥4 μg/mL, whereas C. manghas and P. pellucida produced the same MIC against both Escherichia coli and Pseudomonas aeruginosa. Among the antibacterially active species, C. diffusa, K. hospita, P. scolopendria, P. insularum, and S. glaucifolium did not produce toxicity towards the standard line of normal adult human dermal fibroblasts (IC80 > 128 μg/mL). In addition, extracts from Barringtonia asiatica, C. manghas, M. micrantha, O. nutans, P. insularum, and Piper graeffei stimulated significant migration of dermal fibroblasts, while M. micrantha, O. nutans, and P. insularum did not affect cell proliferation at a concentration of 32 μg/mL. CONCLUSIONS: The results suggest that the above-mentioned species of Samoan medicinal plants can be used for the development of new wound healing agents. However, further phytochemical and pharmacological research is needed regarding the isolation and identification of their active constituents.

• MeSH
• antibakteriální látky izolace a purifikace   farmakologie   MeSH
• fibroblasty účinky léků   fyziologie   MeSH
• léčivé rostliny * MeSH
• lidé MeSH
• mikrobiální testy citlivosti metody   MeSH
• nadzemní části rostlin MeSH
• pohyb buněk účinky léků   fyziologie   MeSH
• proliferace buněk účinky léků   fyziologie   MeSH
• rostlinné extrakty izolace a purifikace   farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Geografické názvy
• Samoa MeSH

Cells attaching to the extracellular matrix spontaneously acquire front-rear polarity. This self-organization process comprises spatial activation of polarity signaling networks and the establishment of a protruding cell front and a non-protruding cell rear. Cell polarization also involves the reorganization of cell mass, notably the nucleus that is positioned at the cell rear. It remains unclear, however, how these processes are regulated. Here, using coherence-controlled holographic microscopy (CCHM) for non-invasive live-cell quantitative phase imaging (QPI), we examined the role of the focal adhesion kinase (FAK) and its interacting partner Rack1 in dry mass distribution in spreading Rat2 fibroblasts. We found that FAK-depleted cells adopt an elongated, bipolar phenotype with a high central body mass that gradually decreases toward the ends of the elongated processes. Further characterization of spreading cells showed that FAK-depleted cells are incapable of forming a stable rear; rather, they form two distally positioned protruding regions. Continuous protrusions at opposite sides results in an elongated cell shape. In contrast, Rack1-depleted cells are round and large with the cell mass sharply dropping from the nuclear area towards the basal side. We propose that FAK and Rack1 act differently yet coordinately to establish front-rear polarity in spreading cells.

• MeSH
• buněčná adheze genetika   fyziologie   MeSH
• buněčné linie MeSH
• fibroblasty cytologie   metabolismus   MeSH
• fokální adhezní tyrosinkinasy genetika   metabolismus   MeSH
• krysa rodu rattus MeSH
• mikroskopie fázově kontrastní MeSH
• pohyb buněk genetika   fyziologie   MeSH
• polarita buněk genetika   fyziologie   MeSH
• receptory pro aktivovanou kinasu C genetika   metabolismus   MeSH
• RNA interference MeSH
• tvar buňky genetika   fyziologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

A microfluidic device made of polydimethylsiloxane was developed for continuous evaluation of natural migration mobility of many eukaryotic cells in relaxed and deformed state. The device was fabricated by standard photolithography and soft lithography techniques using the SU-8 3010 negative photoresist on a glass wafer as the master mold. The simple flow-free device exploits the chemotactic movement of cells through a set of mechanical barriers in the direction of concentration gradients of attractants. The barriers are formed by arrays of circular cross-section pillars with decreasing spacing 7, 5, and 3 μm. To pass through the obstacles, the cells are deformed and change their cytoskeletal architecture. The instantaneous migration velocities of cells are monitored in a time-lapse setup of the scanning confocal microscope. Thus, the cellular deformability and migratory activity can easily be evaluated. The functionality of the device was tested with model HeLa cells stably transfected with fluorescent Premo FUCCI Cell Cycle Sensor. The designed device has the potential to be implemented for testing the tendency of patients' tumors to metastasis.

• MeSH
• buněčné kultury přístrojové vybavení   MeSH
• design vybavení MeSH
• dimethylpolysiloxany chemie   MeSH
• HeLa buňky MeSH
• konfokální mikroskopie MeSH
• lidé MeSH
• mikrofluidní analytické techniky přístrojové vybavení   MeSH
• pohyb buněk fyziologie   MeSH
• tvar buňky fyziologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

A better understanding of the molecular mechanisms leading to mast cell migration and chemotaxis is the long-term goal in mast cell research and is essential for comprehension of mast cell function in health and disease. Various techniques have been developed in recent decades for in vitro and in vivo assessment of mast cell motility and chemotaxis. In this chapter, three microscopy assays facilitating real-time quantification of mast cell chemotaxis and migration are described, focusing on individual cell tracking and data analysis.

• MeSH
• analýza buněčné migrace metody   MeSH
• biotest metody   MeSH
• buněčný tracking metody   MeSH
• chemotaxe fyziologie   MeSH
• fibronektiny metabolismus   MeSH
• lidé MeSH
• mastocyty cytologie   fyziologie   MeSH
• mikroskopie metody   MeSH
• myši MeSH
• počítačové systémy MeSH
• pohyb buněk fyziologie   MeSH
• prostředí kontrolované MeSH
• sefarosa MeSH
• software MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Surface protein CD20 serves as the critical target of immunotherapy in various B-cell malignancies for decades, however its biological function and regulation remain largely elusive. Better understanding of CD20 function may help to design improved rational therapies to prevent development of resistance. Using CRISPR/Cas9 technique, we have abrogated CD20 expression in five different malignant B-cell lines. We show that CD20 deletion has no effect upon B-cell receptor signaling or calcium flux. Also B-cell survival and proliferation is unaffected in the absence of CD20. On the contrary, we found a strong defect in actin cytoskeleton polymerization and, consequently, defective cell adhesion and migration in response to homeostatic chemokines SDF1α, CCL19 and CCL21. Mechanistically, we could identify a reduction in chemokine-triggered PYK2 activation, a calcium-activated signaling protein involved in activation of MAP kinases and cytoskeleton regulation. These cellular defects in consequence result in a severely disturbed homing of B cells in vivo.

• MeSH
• aktiny metabolismus   MeSH
• antigeny CD20 genetika   metabolismus   fyziologie   MeSH
• B-buněčný lymfom metabolismus   patologie   MeSH
• B-lymfocyty patologie   fyziologie   MeSH
• buněčná adheze fyziologie   MeSH
• genový knockdown MeSH
• leukemie B-buněčná metabolismus   patologie   MeSH
• lidé MeSH
• multimerizace proteinu fyziologie   MeSH
• myši inbrední NOD MeSH
• myši SCID MeSH
• myši transgenní MeSH
• myši MeSH
• nádorové buněčné linie MeSH
• pohyb buněk fyziologie   MeSH
• polymerizace MeSH
• receptory antigenů B-buněk metabolismus   MeSH
• signální transdukce imunologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Focal adhesion kinase (FAK) is essential for vascular endothelial growth factor-A (VEGFA)/VEGF receptor-2 (VEGFR2)-stimulated angiogenesis and vascular permeability. We have previously noted that presence of the Src homology-2 domain adapter protein B (SHB) is of relevance for VEGFA-stimulated angiogenesis in a FAK-dependent manner. The current study was conducted in order address the temporal dynamics of co-localization between these components in HEK293 and primary lung endothelial cells (EC) by total internal reflection fluorescence microscopy (TIRF). An early (<2.5 min) VEGFA-induced increase in VEGFR2 co-localization with SHB was dependent on tyrosine 1175 in VEGFR2. VEGFA also enhanced SHB co-localization with FAK. FAK co-localization with VEGFR2 was dependent on SHB since it was significantly lower in SHB deficient EC after VEGFA addition. Absence of SHB also resulted in a gradual decline of VEGFR2 co-localization with FAK under basal (prior to VEGFA addition) conditions. A similar basal response was observed with expression of the Y1175F-VEGFR2 mutant in wild type EC. The distribution of focal adhesions in SHB-deficient EC was altered with a primarily perinuclear location. These live cell data implicate SHB as a key component regulating FAK activity in response to VEGFA/VEGFR2.

• MeSH
• adaptorové proteiny signální transdukční metabolismus   MeSH
• endoteliální buňky metabolismus   MeSH
• fokální adhezní tyrosinkinasy genetika   metabolismus   MeSH
• fyziologická neovaskularizace fyziologie   MeSH
• HEK293 buňky MeSH
• lidé MeSH
• myši inbrední BALB C MeSH
• myši knockoutované MeSH
• myši MeSH
• pohyb buněk fyziologie   MeSH
• protoonkogenní proteiny metabolismus   MeSH
• receptor 2 pro vaskulární endoteliální růstový faktor metabolismus   MeSH
• vaskulární endoteliální růstový faktor A genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• mužské pohlaví MeSH
• myši MeSH
• ženské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Increased levels of the chemokine CCL2 in cancer patients are associated with poor prognosis. Experimental evidence suggests that CCL2 correlates with inflammatory monocyte recruitment and induction of vascular activation, but the functionality remains open. Here, we show that endothelial Ccr2 facilitates pulmonary metastasis using an endothelial-specific Ccr2-deficient mouse model (Ccr2ecKO). Similar levels of circulating monocytes and equal leukocyte recruitment to metastatic lesions of Ccr2ecKO and Ccr2fl/fl littermates were observed. The absence of endothelial Ccr2 strongly reduced pulmonary metastasis, while the primary tumor growth was unaffected. Despite a comparable cytokine milieu in Ccr2ecKO and Ccr2fl/fl littermates the absence of vascular permeability induction was observed only in Ccr2ecKO mice. CCL2 stimulation of pulmonary endothelial cells resulted in increased phosphorylation of MLC2, endothelial cell retraction, and vascular leakiness that was blocked by an addition of a CCR2 inhibitor. These data demonstrate that endothelial CCR2 expression is required for tumor cell extravasation and pulmonary metastasis. IMPLICATIONS: The findings provide mechanistic insight into how CCL2-CCR2 signaling in endothelial cells promotes their activation through myosin light chain phosphorylation, resulting in endothelial retraction and enhanced tumor cell migration and metastasis.

• MeSH
• chemokin CCL2 metabolismus   MeSH
• endoteliální buňky metabolismus   patologie   MeSH
• kapilární permeabilita MeSH
• karcinom plic Lewisové krevní zásobení   metabolismus   patologie   sekundární   MeSH
• lehké řetězce myosinu metabolismus   MeSH
• metastázy nádorů MeSH
• myši inbrední C57BL MeSH
• myši knockoutované MeSH
• myši MeSH
• nádory plic krevní zásobení   metabolismus   patologie   sekundární   MeSH
• pohyb buněk fyziologie   MeSH
• receptory CCR2 metabolismus   MeSH
• zvířata MeSH
• Check Tag
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH