Page 53 - Read Online

Figure 5: Hair follicle stem cells morphological observation and protein expression. Cells have round or oval nuclei and a high nucleus and cytoplasm ratio.
Immunofluorescence staining of pan-cytokeratin (A), K19 (B), and β1-integrin (C). Magnification, ×200

Figure 6: Effect of different LiCl concentrations on hair follicle stem cells proliferation. Cells were assessed 3 days after treatment of LiCl at different
concentrations. (A) after treatment with 0 mmol/L LiCl, cells have grown well and display a cobblestone-like appearance; (B) after treatment with 0.5 mmol/
L LiCl, cells have become rounded, or occasionally an elongated spindle shape, and show a tendency to aggregate; (C) after treatment with 1.5 mmol/L LiCl,
in general, cells have become larger. Occasionally, cells have developed protuberances, which may be related to cell migration; (D) after treatment with
10 mmol/L LiCl, cell morphology has changed. Cells have become highly aggregated and cell density is reduced; (E) after treatment with 25 mmol/L LiCl,
cell density has significantly decreased. Both the cytoplasm and nuclei show various morphological changes. The proliferation rate has also decreased.
Magnification, ×100

was set at P < 0.05; P < 0.01 indicates that the difference to culture vessel, although most remained in suspension
between experimental groups was significant; P > 0.05 [Figure 1A and B]. After 4-6 days in culture, a few single cells
indicates no difference between experimental groups. or cell clusters had attached to the culture dish; these had
a cobblestone-like appearance, an orderly arrangement, and
RESULTS were highly refractive. Cell numbers increased with increasing
culture time [Figure 1C and D]. After 2 weeks in culture, a
HFSC growth characteristics small number of aging cells were observed.
Primary cells derived from the hair follicle bulge were
spheroid, small, and evenly sized. Live cells were optically Dynamics of cell growth
transparent with a clear boundary and clusters of Growth curves [Figures 1 and 2] show that the cell growth
undissociated cells could be seen [Figure 1]. Trypan blue rate was slow on days 1-3. On days 4-6, cells grew faster and
staining showed that 95% of cells were alive. At the initial stage were in the logarithmic phase. After this point, cell growth
of culture, cell adhesion and activity were weak. After culture became slower and aging cells appeared.
in serum-free K-SFM containing epidermal growth factor (EGF)
for 1-3 days, a few human HFSCs were observed to adhere Cell proliferation varied according to seeding density [Figures
3 and 4]. At seeding densities of 1 × 10 and 1 × 10 , a few
4
3
cells showed clonal growth but cells proliferated slowly and
most died after a short time in culture. When the seeding
5
density was increased to 1 × 10 cells/mL, cells proliferated
well and showed adherent growth at early time points.
6
Cells seeded at a density of 1 × 10 cells/mL rapidly formed
clusters and underwent contact inhibition following rapid
nutrient depletion. These cells showed signs of aging.

Identification of HFSCs and characteristics
of HFSC protein expression
Pan-cytokeratin expression: adherent pan-cytokeratin-
Figure 7: Effect of LiCl on hair follicle stem cells (HFSCs) proliferation labeled HFSCs showed that the cytoplasm of these cells
42 Plast Aesthet Res || Vol 3 || Issue 2 || Feb 29, 2016