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Roofed grooves: rapid layer engineering of perfusion channels in collagen tissue models.

Tan NS, Alekseeva T, Brown RA - J Biomater Appl (2014)

Bottom Line: In the second part, this was used for effective fabrication of multi-layered plastically compressed collagen constructs with internal channels by roofing the grooves with a second layer.Resulting µ-channels retained their dimensions and were stable over time in culture with fibroblasts and could be cell seeded with a lining layer by simple transfer of epithelial cells.The results of this study provide a valuable platform for rapid fabrication of complex collagen-based tissues in particular for provision of perfusing microchannels through the bulk material for improved core nutrient supply.

View Article: PubMed Central - PubMed

Affiliation: Tissue Repair & Engineering Centre, Institute of Orthopaedics, University College London, United Kingdom.

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SEM images of the fibroblast-populated microchannelled PC constructs immediately after assembly (a) and after 14 days in culture (b). Insets show the appearance of the grooves at the respective culture times (0 and 14 day) with the roofing layer peeled away. Fibroblasts within the collagen were not normally visible by SEM, but the technique does clearly show that the channel structure was effectively unaltered by the matrix remodelling of cells cultured over this period. (c) Histological section (H&E staining) showing internal matrix structure of constructs after 14 days remodelling by human dermal fibroblasts (contrast with the unremodelled appearance immediately after fabrication (Figure 5). (d) Histological (H&E) cross-section of a roofed channel seeded with HaCat epithelial cells and cultured for 7 days to expand into a cell-lining layer. The roof layer is above the dotted line and the grooved layer (1) is below.
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fig7-0885328214538865: SEM images of the fibroblast-populated microchannelled PC constructs immediately after assembly (a) and after 14 days in culture (b). Insets show the appearance of the grooves at the respective culture times (0 and 14 day) with the roofing layer peeled away. Fibroblasts within the collagen were not normally visible by SEM, but the technique does clearly show that the channel structure was effectively unaltered by the matrix remodelling of cells cultured over this period. (c) Histological section (H&E staining) showing internal matrix structure of constructs after 14 days remodelling by human dermal fibroblasts (contrast with the unremodelled appearance immediately after fabrication (Figure 5). (d) Histological (H&E) cross-section of a roofed channel seeded with HaCat epithelial cells and cultured for 7 days to expand into a cell-lining layer. The roof layer is above the dotted line and the grooved layer (1) is below.

Mentions: Whilst studies so far have established the stability and predictability of roofed channels it is also important to know the effect of cell-matrix remodelling. The final stage of this analysis tested the stability of micro-moulded channels using cell-seeded collagen in culture. Figure 7 shows the structure of the channels, and in particular the surrounding matrix, after 0 and 14 days in culture with human dermal fibroblasts. Channels clearly remained present and patent after culture and retained their initial shape. This was despite clear histological evidence for substantial cell remodelling of the bulk collagen construct matrix over that time period (Figure 7(c)), as reported previously.4 The ability to seed cells onto the channel lumen and expand them to cover the internal surface, as a channel lining, was tested, using epithelial cells. In this case, the epithelial cells (HaCats) were first seeded onto the surface of the micro-moulding template. When this cell-carrying template was used to form roofed channels in a collagen construct, cells successfully transferred from the template to the groove/channel surface. These were subsequently expanded over the following 7 days in culture to produce an epithelial lining layer as shown in Figure 7(d).Figure 7.


Roofed grooves: rapid layer engineering of perfusion channels in collagen tissue models.

Tan NS, Alekseeva T, Brown RA - J Biomater Appl (2014)

SEM images of the fibroblast-populated microchannelled PC constructs immediately after assembly (a) and after 14 days in culture (b). Insets show the appearance of the grooves at the respective culture times (0 and 14 day) with the roofing layer peeled away. Fibroblasts within the collagen were not normally visible by SEM, but the technique does clearly show that the channel structure was effectively unaltered by the matrix remodelling of cells cultured over this period. (c) Histological section (H&E staining) showing internal matrix structure of constructs after 14 days remodelling by human dermal fibroblasts (contrast with the unremodelled appearance immediately after fabrication (Figure 5). (d) Histological (H&E) cross-section of a roofed channel seeded with HaCat epithelial cells and cultured for 7 days to expand into a cell-lining layer. The roof layer is above the dotted line and the grooved layer (1) is below.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2 - License 3
Show All Figures
getmorefigures.php?uid=PMC4230962&req=5

fig7-0885328214538865: SEM images of the fibroblast-populated microchannelled PC constructs immediately after assembly (a) and after 14 days in culture (b). Insets show the appearance of the grooves at the respective culture times (0 and 14 day) with the roofing layer peeled away. Fibroblasts within the collagen were not normally visible by SEM, but the technique does clearly show that the channel structure was effectively unaltered by the matrix remodelling of cells cultured over this period. (c) Histological section (H&E staining) showing internal matrix structure of constructs after 14 days remodelling by human dermal fibroblasts (contrast with the unremodelled appearance immediately after fabrication (Figure 5). (d) Histological (H&E) cross-section of a roofed channel seeded with HaCat epithelial cells and cultured for 7 days to expand into a cell-lining layer. The roof layer is above the dotted line and the grooved layer (1) is below.
Mentions: Whilst studies so far have established the stability and predictability of roofed channels it is also important to know the effect of cell-matrix remodelling. The final stage of this analysis tested the stability of micro-moulded channels using cell-seeded collagen in culture. Figure 7 shows the structure of the channels, and in particular the surrounding matrix, after 0 and 14 days in culture with human dermal fibroblasts. Channels clearly remained present and patent after culture and retained their initial shape. This was despite clear histological evidence for substantial cell remodelling of the bulk collagen construct matrix over that time period (Figure 7(c)), as reported previously.4 The ability to seed cells onto the channel lumen and expand them to cover the internal surface, as a channel lining, was tested, using epithelial cells. In this case, the epithelial cells (HaCats) were first seeded onto the surface of the micro-moulding template. When this cell-carrying template was used to form roofed channels in a collagen construct, cells successfully transferred from the template to the groove/channel surface. These were subsequently expanded over the following 7 days in culture to produce an epithelial lining layer as shown in Figure 7(d).Figure 7.

Bottom Line: In the second part, this was used for effective fabrication of multi-layered plastically compressed collagen constructs with internal channels by roofing the grooves with a second layer.Resulting µ-channels retained their dimensions and were stable over time in culture with fibroblasts and could be cell seeded with a lining layer by simple transfer of epithelial cells.The results of this study provide a valuable platform for rapid fabrication of complex collagen-based tissues in particular for provision of perfusing microchannels through the bulk material for improved core nutrient supply.

View Article: PubMed Central - PubMed

Affiliation: Tissue Repair & Engineering Centre, Institute of Orthopaedics, University College London, United Kingdom.

Show MeSH