Complete Granular-like keratinocytes are rare event in <i>in vitro</i> epidermal cell culture.

<p>Granular-like keratinocytes emerged on the top of the hNEK culture and displayed expression of Involucrin and a polymerized keratin 10 network when cultured at 31°C. A. Image acquired with a LSM780 confocal microscope (parameters 2) shows detection of strong levels of K10 (green) and INV (red) in two cells emerging from a confluent culture of hNEK (top left panel). Bright field image was merged to demonstrate the confluence of the hNEK culture. Orthogonal [X-Z] and [Y-Z] views (top right panels) were performed as indicated by the red cross lines on the main picture and show that the K10 ⁺ /INV⁺ cell (cell 1) is set above a K10^-/INV^- cell (cell 2) detected with its Dapi-stained nucleus. Gallery of Z-sectioning images from the bottom of the dish to the top of cell culture also shows the position of the K10 ⁺ /INV⁺ cell on the top of the cell culture. B. Confocal slide showing a polymerized network of Keratin 10 in a single keratinocyte cultured at 31°C for 5 days (left panel). 3D deconvolution (center image of middle panel) after Z-sectioning (right panel) of the same field shows the concomitant detection of K10 (green) and INV (red). Rotation of the 3D image by 90° from they axis to the Z axis shows the keratinocyte from below (bottom image of middle panel) or from above (top image of middle panel) and demonstrates a shell-like architecture of the polymerized network of K10 and INV.