Bulge Epithelial Stem Cells

Bulge epithelial stem cells (eHFSCs) are harvested from autologous occipital scalp punches (2 mm) under local anesthesia. After enzymatic dissociation (collagenase IV, 2 mg/mL, 90 min), single-cell suspensions are FACS-sorted on a CD200⁺ / CD34⁻ / integrin α6ʰⁱ / K15⁺ profile. Yield is 1.2–2.0 × 10⁴ eHFSCs per follicular unit.

Cells are expanded on lethally-irradiated 3T3-J2 feeder layers in defined KSFM (keratinocyte serum-free medium) supplemented with EGF (5 ng/mL), bFGF (10 ng/mL), and Rho-kinase inhibitor Y-27632 (10 µM) for the first 48 h to suppress apoptotic signaling at low density. We hold passages ≤ P4 to limit clonogenic potential drift; population doublings tracked in real time.