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Inhibition of Rho-associated kinases disturbs the collective cell migration of stratified TE-10 cells.

Mikami T, Yoshida K, Sawada H, Esaki M, Yasumura K, Ono M - Biol. Res. (2015)

Bottom Line: Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent.Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet.In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion.

View Article: PubMed Central - PubMed

Affiliation: Department of Histology and Cell Biology, Yokohama City University School of Medicine, Yokohama, Kanagawa-ken, Japan. zeong3@mac.com.

ABSTRACT

Background: The collective cell migration of stratified epithelial cells is considered to be an important phenomenon in wound healing, development, and cancer invasion; however, little is known about the mechanisms involved. Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent. Here, we report the development of a novel modified scratch assay that was used to observe the collective cell migration of stratified TE-10 cells derived from a human esophageal cancer specimen.

Results: Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet. The leading edge of cells in the cell sheet formed a simple layer and moved forward regularly; these rows were followed by the stratified epithelium. ROCK inhibitors and ROCK small interfering RNAs (siRNAs) disturbed not only the collective migration of the leading edge of this cell sheet, but also the stratified layer in the rear. In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion.

Conclusions: The data presented in this study suggest that ROCKs play an important role in mediating the collective migration of TE-10 cell sheets. In addition, differences between the effects of siRNAs targeting either RhoA or ROCKs suggested that distinct mechanisms regulate the collective cell migration in the simple epithelium of the wound edge versus the stratified layer of the epithelium.

No MeSH data available.


Related in: MedlinePlus

Images of stratified TE-10 cells in the wound edge and cross sections. a A randomly selected area of the wound edge was observed 24 h after scrape-induced wounding. Cells in the third row and deeper were multilayered, as indicated by the presence of overlapping nuclei. The top panel shows a phase-contrast image, while a nuclear-stained fluorescent image is presented in the middle panel. A merged image is shown in the lower panel. Each inset is the magnified image of the portion indicated in the merged image. The scale bar indicates 100 µm. b Transmission electron micrographs of the cross sections of stratified TE-10 cells plated in a hole of a silicone stencil at high density for 24 h. The cells were stratified into 5–7 layers from the apical to the basal side (the upper region of the image), which was attached to the glass cover slip. Few morphological differences were observed between the apical and basal sides, with the exception of microvillus formation (white arrows) at the surface of most apical cells. The scale bar indicates 2 µm. c Desmosomes were found between the cells (arrowheads). Arrows show cytokeratin bundles. The scale bar indicates 200 nm
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Fig1: Images of stratified TE-10 cells in the wound edge and cross sections. a A randomly selected area of the wound edge was observed 24 h after scrape-induced wounding. Cells in the third row and deeper were multilayered, as indicated by the presence of overlapping nuclei. The top panel shows a phase-contrast image, while a nuclear-stained fluorescent image is presented in the middle panel. A merged image is shown in the lower panel. Each inset is the magnified image of the portion indicated in the merged image. The scale bar indicates 100 µm. b Transmission electron micrographs of the cross sections of stratified TE-10 cells plated in a hole of a silicone stencil at high density for 24 h. The cells were stratified into 5–7 layers from the apical to the basal side (the upper region of the image), which was attached to the glass cover slip. Few morphological differences were observed between the apical and basal sides, with the exception of microvillus formation (white arrows) at the surface of most apical cells. The scale bar indicates 2 µm. c Desmosomes were found between the cells (arrowheads). Arrows show cytokeratin bundles. The scale bar indicates 200 nm

Mentions: TE-10 cells had accumulated between 3 and 5 layers by 24 h after plating in the holes of silicone blocks, as described in the Materials and Methods section. Desmosomes were observed between cells by transmission electron microscopy (Fig. 1b, c; Additional file 1). Although obvious basal laminae could not be observed and apical-basal polarity was morphologically vague between the cells of the upper and lower layers, we observed desmosomes, cytokeratin bundles, and microvilli in the uppermost layer of cells as signs of its differentiation into a stratified squamous epithelium.Fig. 1


Inhibition of Rho-associated kinases disturbs the collective cell migration of stratified TE-10 cells.

Mikami T, Yoshida K, Sawada H, Esaki M, Yasumura K, Ono M - Biol. Res. (2015)

Images of stratified TE-10 cells in the wound edge and cross sections. a A randomly selected area of the wound edge was observed 24 h after scrape-induced wounding. Cells in the third row and deeper were multilayered, as indicated by the presence of overlapping nuclei. The top panel shows a phase-contrast image, while a nuclear-stained fluorescent image is presented in the middle panel. A merged image is shown in the lower panel. Each inset is the magnified image of the portion indicated in the merged image. The scale bar indicates 100 µm. b Transmission electron micrographs of the cross sections of stratified TE-10 cells plated in a hole of a silicone stencil at high density for 24 h. The cells were stratified into 5–7 layers from the apical to the basal side (the upper region of the image), which was attached to the glass cover slip. Few morphological differences were observed between the apical and basal sides, with the exception of microvillus formation (white arrows) at the surface of most apical cells. The scale bar indicates 2 µm. c Desmosomes were found between the cells (arrowheads). Arrows show cytokeratin bundles. The scale bar indicates 200 nm
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4556056&req=5

Fig1: Images of stratified TE-10 cells in the wound edge and cross sections. a A randomly selected area of the wound edge was observed 24 h after scrape-induced wounding. Cells in the third row and deeper were multilayered, as indicated by the presence of overlapping nuclei. The top panel shows a phase-contrast image, while a nuclear-stained fluorescent image is presented in the middle panel. A merged image is shown in the lower panel. Each inset is the magnified image of the portion indicated in the merged image. The scale bar indicates 100 µm. b Transmission electron micrographs of the cross sections of stratified TE-10 cells plated in a hole of a silicone stencil at high density for 24 h. The cells were stratified into 5–7 layers from the apical to the basal side (the upper region of the image), which was attached to the glass cover slip. Few morphological differences were observed between the apical and basal sides, with the exception of microvillus formation (white arrows) at the surface of most apical cells. The scale bar indicates 2 µm. c Desmosomes were found between the cells (arrowheads). Arrows show cytokeratin bundles. The scale bar indicates 200 nm
Mentions: TE-10 cells had accumulated between 3 and 5 layers by 24 h after plating in the holes of silicone blocks, as described in the Materials and Methods section. Desmosomes were observed between cells by transmission electron microscopy (Fig. 1b, c; Additional file 1). Although obvious basal laminae could not be observed and apical-basal polarity was morphologically vague between the cells of the upper and lower layers, we observed desmosomes, cytokeratin bundles, and microvilli in the uppermost layer of cells as signs of its differentiation into a stratified squamous epithelium.Fig. 1

Bottom Line: Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent.Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet.In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion.

View Article: PubMed Central - PubMed

Affiliation: Department of Histology and Cell Biology, Yokohama City University School of Medicine, Yokohama, Kanagawa-ken, Japan. zeong3@mac.com.

ABSTRACT

Background: The collective cell migration of stratified epithelial cells is considered to be an important phenomenon in wound healing, development, and cancer invasion; however, little is known about the mechanisms involved. Furthermore, whereas Rho family proteins, including RhoA, play important roles in cell migration, the exact role of Rho-associated coiled coil-containing protein kinases (ROCKs) in cell migration is controversial and might be cell-type dependent. Here, we report the development of a novel modified scratch assay that was used to observe the collective cell migration of stratified TE-10 cells derived from a human esophageal cancer specimen.

Results: Desmosomes were found between the TE-10 cells and microvilli of the surface of the cell sheet. The leading edge of cells in the cell sheet formed a simple layer and moved forward regularly; these rows were followed by the stratified epithelium. ROCK inhibitors and ROCK small interfering RNAs (siRNAs) disturbed not only the collective migration of the leading edge of this cell sheet, but also the stratified layer in the rear. In contrast, RhoA siRNA treatment resulted in more rapid migration of the leading rows and disturbed movement of the stratified portion.

Conclusions: The data presented in this study suggest that ROCKs play an important role in mediating the collective migration of TE-10 cell sheets. In addition, differences between the effects of siRNAs targeting either RhoA or ROCKs suggested that distinct mechanisms regulate the collective cell migration in the simple epithelium of the wound edge versus the stratified layer of the epithelium.

No MeSH data available.


Related in: MedlinePlus