Apico-basal polarity is typical of cells present in differentiated epithelium while front-rear polarity develops in motile cells. In cancer development, the transition from epithelial to migratory polarity may be seen as the hallmark of cancer progression to an invasive and metastatic disease. Despite the morphological and functional dissimilarity, both epithelial and migratory polarity are controlled by a common set of polarity complexes Par, Scribble and Crumbs, phosphoinositides, and small Rho GTPases Rac, Rho and Cdc42. In epithelial tissues, their mutual interplay ensures apico-basal and planar cell polarity. Accordingly, altered functions of these polarity determinants lead to disrupted cell-cell adhesions, cytoskeleton rearrangements and overall loss of epithelial homeostasis. Polarity proteins are further engaged in diverse interactions that promote the establishment of front-rear polarity, and they help cancer cells to adopt different invasion modes. Invading cancer cells can employ either the collective, mesenchymal or amoeboid invasion modes or actively switch between them and gain intermediate phenotypes. Elucidation of the role of polarity proteins during these invasion modes and the associated transitions is a necessary step towards understanding the complex problem of metastasis. In this review we summarize the current knowledge of the role of cell polarity signaling in the plasticity of cancer cell invasiveness.

• Klíčová slova
• AMT, EMT, invasion, plasticity, polarity,
• MeSH
• invazivní růst nádoru patologie   MeSH
• lidé MeSH
• nádory patologie   MeSH
• polarita buněk fyziologie   MeSH
• signální transdukce fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• přehledy MeSH

Melanoma represents a malignant disease with steadily increasing incidence. UV-irradiation is a recognized key factor in melanoma initiation. Therefore, the efficient prevention of UV tissue damage bears a critical potential for melanoma prevention. In this study, we tested the effect of UV irradiation of normal keratinocytes and their consequent interaction with normal and cancer-associated fibroblasts isolated from melanoma, respectively. Using this model of UV influenced microenvironment, we measured melanoma cell migration in 3-D collagen gels. These interactions were studied using DNA microarray technology, immunofluorescence staining, single cell electrophoresis assay, viability (dead/life) cell detection methods, and migration analysis. We observed that three 10 mJ/cm2 fractions at equal intervals over 72 h applied on keratinocytes lead to a 50% increase (p < 0.05) in in vitro invasion of melanoma cells. The introduction cancer-associated fibroblasts to such model further significantly stimulated melanoma cells in vitro invasiveness to a higher extent than normal fibroblasts. A panel of candidate gene products responsible for facilitation of melanoma cells invasion was defined with emphasis on IL-6, IL-8, and CXCL-1. In conclusion, this study demonstrates a synergistic effect between cancer microenvironment and UV irradiation in melanoma invasiveness under in vitro condition.

• Klíčová slova
• Cancer microenvironment, Cancer-associated fibroblasts, Chemokine, Cytokine, Keratinocytes, Melanoma,
• MeSH
• fibroblasty cytologie   patologie   MeSH
• imunohistochemie MeSH
• invazivní růst nádoru * MeSH
• keratinocyty patologie   účinky záření   MeSH
• kokultivační techniky MeSH
• kultivované buňky MeSH
• lidé MeSH
• melanom patologie   MeSH
• ultrafialové záření * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

Tumor cells exhibit at least two distinct modes of migration when invading the 3D environment. A single tumor cell's invasive strategy follows either mesenchymal or amoeboid patterns. Certain cell types can use both modes of invasiveness and undergo transitions between them. This work outlines the signaling pathways involved in mesenchymal and amoeboid types of tumor cell motility and summarizes the molecular mechanisms that are involved in transitions between them. The focus is on the signaling of the Rho family of small GTPases that regulate the cytoskeleton-dependent processes taking place during the cell migration. The multiple interactions among the Rho family of proteins, their regulators and effectors are thought to be the key determinants of the particular type of invasiveness. Mesenchymal and amoeboid invasive strategies display different adhesive and proteolytical interactions with the surrounding matrix and the alterations influencing these interactions can also lead to the transitions.

• MeSH
• cytoskelet MeSH
• invazivní růst nádoru * MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• pohyb buněk * MeSH
• rac proteiny vázající GTP metabolismus   MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• rac proteiny vázající GTP MeSH

Sarcomas are a heterogeneous group of mesenchymal tumours, with a great variability in their clinical behaviour. While our knowledge of sarcoma initiation has advanced rapidly in recent years, relatively little is known about mechanisms of sarcoma progression. JUN-murine fibrosarcoma progression series consists of four sarcoma cell lines, JUN-1, JUN-2, JUN-2fos-3, and JUN-3. JUN-1 and -2 were established from a single tumour initiated in a H2K/v-jun transgenic mouse, JUN-3 originates from a different tumour in the same animal, and JUN-2fos-3 results from a targeted in vitro transformation of the JUN-2 cell line. The JUN-1, -2, and -3 cell lines represent a linear progression from the least transformed JUN-2 to the most transformed JUN-3, with regard to all the transformation characteristics studied, while the JUN-2fos-3 cell line exhibits a unique transformation mode, with little deregulation of cell growth and proliferation, but pronounced motility and invasiveness. The invasive sarcoma sublines JUN-2fos-3 and JUN-3 show complex metabolic profiles, with activation of both mitochondrial oxidative phosphorylation and glycolysis and a significant increase in spared respiratory capacity. The specific transcriptomic profile of invasive sublines features very complex biological relationships across the identified genes and proteins, with accentuated autocrine control of motility and angiogenesis. Pharmacologic inhibition of one of the autocrine motility factors identified, Ccl8, significantly diminished both motility and invasiveness of the highly transformed fibrosarcoma cell. This progression series could be greatly valuable for deciphering crucial aspects of sarcoma progression and defining new prognostic markers and potential therapeutic targets.

• Klíčová slova
• Ccl8, fibrosarcoma, invasiveness, progression series, transcriptome,
• Publikační typ
• časopisecké články MeSH

BACKGROUND: The local invasion of tumor cells into the surrounding tissue is the first and most critical step of the metastatic cascade. Cells can invade either collectively, or individually. Individual cancer cell invasion can occur in the mesenchymal or amoeboid mode, which are mutually interchangeable. This plasticity of individual cancer cell invasiveness may represent an escape mechanism for invading cancer cells from anti-metastatic treatment. METHODS: To identify new signaling proteins involved in the plasticity of cancer cell invasiveness, we performed proteomic analysis of the amoeboid to mesenchymal transition with A375m2 melanoma cells in a 3D Matrigel matrix. RESULTS: In this screen we identified PKCα as an important protein for the maintenance of amoeboid morphology. We found that the activation of PKCα resulted in the mesenchymal-amoeboid transition of mesenchymal K2 and MDA-MB-231 cell lines. Consistently, PKCα inhibition led to the amoeboid-mesenchymal transition of amoeboid A375m2 cells. Next, we showed that PKCα inhibition resulted in a considerable decrease in the invading abilities of all analyzed cancer cell lines. CONCLUSIONS: Our results suggest that PKCα is an important protein for maintenance of the amoeboid morphology of cancer cells, and that downregulation of PKCα results in the amoeboid to mesenchymal transition. Our data also suggest that PKCα is important for both mesenchymal and amoeboid invasiveness, making it an attractive target for anti-metastatic therapies.

• MeSH
• invazivní růst nádoru genetika   patologie   MeSH
• lidé MeSH
• melanom genetika   patologie   MeSH
• mezoderm metabolismus   patologie   MeSH
• nádorové buněčné linie MeSH
• pohyb buněk genetika   MeSH
• proteinkinasa C-alfa biosyntéza   genetika   MeSH
• proteomika MeSH
• regulace genové exprese u nádorů MeSH
• signální transdukce MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• PRKCA protein, human MeSH Prohlížeč
• proteinkinasa C-alfa MeSH

Tumour microenvironment plays a critical role in cell invasion and metastasis. To investigate the role of cancer-associated fibroblasts (CAFs) in melanoma cell invasiveness, we used 3D spheroid invasion assay. The effect of conditioned media from normal fibroblasts and CAFs cultivated alone or co-cultivated with melanoma cells on BLM or A2058 melanoma spheroid invasion was analysed. We found that conditioned media from CAFs and CAFs co-cultured with melanoma cells, especially, promote invasion and migration, without significant effect on melanoma cell proliferation. We further analysed the expression of pro-invasive cytokines IL-8 and IL-6 in media and found that melanoma cells are dominant producers of IL-8 and fibroblasts are dominant producers of IL-6 in 2D monocultures, while co-cultivation of CAFs with melanoma cells induces production/secretion of IL-6 and IL-8 into the media. The analyses of IL-6 levels in 3D cultures and human melanoma samples, however, revealed that at least in some cases IL-6 is also produced directly by melanoma cells. Analysis of the role of IL-6 and IL-8 in CAF-induced melanoma invasion, using neutralising antibodies, revealed that simultaneous blocking of IL-6 and IL-8 is sufficient to fully inhibit CAF-induced human melanoma cell invasiveness. In summary, these experiments indicate the important role of CAFs and IL-8 and IL-6 cytokines in melanoma cell invasiveness.

• Klíčová slova
• 3D culture of melanoma cells, Cancer microenvironment, Cancer-associated fibroblasts, Chemokine, Cytokine, Invasiveness,
• MeSH
• ELISA MeSH
• fibroblasty asociované s nádorem účinky léků   patologie   MeSH
• imunohistochemie MeSH
• interleukin-6 analýza   antagonisté a inhibitory   metabolismus   MeSH
• interleukin-8 analýza   antagonisté a inhibitory   metabolismus   MeSH
• invazivní růst nádoru prevence a kontrola   MeSH
• kokultivační techniky MeSH
• kultivační média speciální farmakologie   MeSH
• kultivované buňky MeSH
• lidé MeSH
• melanom farmakoterapie   metabolismus   patologie   MeSH
• pohyb buněk účinky léků   MeSH
• proliferace buněk účinky léků   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• interleukin-6 MeSH
• interleukin-8 MeSH
• kultivační média speciální MeSH

The aim of this study was to analyze the potential role of NG2 chondroitin sulfate proteoglycan in amoeboid morphology and invasiveness of cancer cells. In the highly metastatic amoeboid cell lines A3 and A375M2, siRNA-mediated down-regulation of NG2 induced an amoeboid-mesenchymal transition associated with decreased invasiveness in 3D collagen and inactivation of the GTPase Rho. Conversely, the expression of NG2 in mesenchymal sarcoma K2 cells as well as in A375M2 cells resulted in an enhanced amoeboid phenotype associated with increased invasiveness and elevated Rho-GTP levels. Remarkably, the amoeboid-mesenchymal transition in A375M2 cells triggered by NG2 down-regulation was associated with increased extracellular matrix-degrading ability, although this was not sufficient to compensate for the decreased invasive capability caused by down-regulated Rho/ROCK signaling. Conversely, in K2 cells with overexpression of NG2, the ability to degrade the extracellular matrix was greatly reduced. Taken together, we suggest that NG2-mediated activation of Rho leading to effective amoeboid invasiveness is a possible mechanism through which NG2 could contribute to tumor cell invasion and metastasis.

• MeSH
• chondroitinsulfát proteoglykany genetika   metabolismus   MeSH
• down regulace MeSH
• extracelulární matrix metabolismus   MeSH
• invazivní růst nádoru MeSH
• kinázy asociované s Rho metabolismus   MeSH
• kolagen chemie   MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• malá interferující RNA MeSH
• membránové proteiny genetika   metabolismus   MeSH
• molekulární konformace MeSH
• nádorové buněčné linie MeSH
• nádory metabolismus   patologie   ultrastruktura   MeSH
• pohyb buněk * MeSH
• Rho proteiny vázající GTP metabolismus   MeSH
• upregulace MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• chondroitinsulfát proteoglykany MeSH
• CSPG4 protein, human MeSH Prohlížeč
• kinázy asociované s Rho MeSH
• kolagen MeSH
• malá interferující RNA MeSH
• membránové proteiny MeSH
• Rho proteiny vázající GTP MeSH

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

Crk-associated substrate (CAS) is a major tyrosine-phosphorylated protein in cells transformed by v-crk and v-src oncogenes and plays an important role in invasiveness of Src-transformed cells. A novel phosphorylation site on CAS, Tyr-12 (Y12) within the ligand-binding hydrophobic pocket of the CAS SH3 domain, was identified and found to be enriched in Src-transformed cells and invasive human carcinoma cells. To study the biological significance of CAS Y12 phosphorylation, phosphomimicking Y12E and nonphosphorylatable Y12F mutants of CAS were studied. The phosphomimicking mutation decreased interaction of the CAS SH3 domain with focal adhesion kinase (FAK) and PTP-PEST and reduced tyrosine phosphorylation of FAK. Live-cell imaging showed that green fluorescent protein-tagged CAS Y12E mutant is, in contrast to wild-type or Y12F CAS, excluded from focal adhesions but retains its localization to podosome-type adhesions. Expression of CAS-Y12F in cas-/- mouse embryonic fibroblasts resulted in hyperphosphorylation of the CAS substrate domain, and this was associated with slower turnover of focal adhesions and decreased cell migration. Moreover, expression of CAS Y12F in Src-transformed cells greatly decreased invasiveness when compared to wild-type CAS expression. These findings reveal an important role of CAS Y12 phosphorylation in the regulation of focal adhesion assembly, cell migration, and invasiveness of Src-transformed cells.

• MeSH
• fokální adheze metabolismus   MeSH
• fokální adhezní tyrosinkinasy metabolismus   MeSH
• fosforylace MeSH
• invazivní růst nádoru MeSH
• lidé MeSH
• molekuly buněčné adheze metabolismus   MeSH
• mutace MeSH
• myši MeSH
• nádorová transformace buněk MeSH
• nádorové buněčné linie MeSH
• pohyb buněk MeSH
• signální transdukce MeSH
• src homologní domény MeSH
• substrátový protein asociovaný s Crk chemie   genetika   metabolismus   MeSH
• transformované buněčné linie MeSH
• tyrosin metabolismus   MeSH
• tyrosinfosfatasa nereceptorového typu 12 metabolismus   MeSH
• zelené fluorescenční proteiny metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• myši MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• fokální adhezní tyrosinkinasy MeSH
• molekuly buněčné adheze MeSH
• PTPN12 protein, human MeSH Prohlížeč
• substrátový protein asociovaný s Crk MeSH
• tyrosin MeSH
• tyrosinfosfatasa nereceptorového typu 12 MeSH
• zelené fluorescenční proteiny MeSH

Migration and invasiveness are phenotypic characteristics of cells that contribute to physiological processes, such as wound healing or embryogenesis and they are involved in serious pathological processes, namely in tumor cell metastasis. Availability of methods for studying migration and invasiveness of the cells is important for understanding molecular basis of these processes. In the case of cancer, migration, invasiveness and metastatic potential of tumor cells are key factors that determine clinical prognosis of the patients. This communication provides an overview of in vitro and in vivo methods which are used to study cell migration, invasion and metastasis. In vitro meth-ods for studying cell migration include simple two dimensional assays (scratch - wound assay and the assay based on the effect of hepatocyte growth factor) and methods based on chemotaxis (Dunns chamber, videomicroscopy of cells, the use of carriers with chemoattractants). Methods for studying both cell migration and invasiveness in vitro include more complex systems based on the principle of the Boyden chamber (transwell migration/ invasive test, analysis of cell migration and invasion in xCELLigence system, confocal microscopy based approaches) as well as analysis of cell migration in microchannels. Our overview of in vivo methods provides an introduction into model organisms and methods used in this field, with an emphasis on the study of cancer metastasis in mouse models. The methods described in this review are mainly involved in larger research projects aiming at developing new diagnostic and therapeutic approaches in oncology.

• MeSH
• analýza buněčné migrace MeSH
• invazivní růst nádoru patofyziologie   MeSH
• lidé MeSH
• metastázy nádorů patofyziologie   MeSH
• pohyb buněk fyziologie   MeSH
• techniky in vitro metody   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• anglický abstrakt MeSH
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH