Abstract:

BACKGROUND: Hair follicle stem cells (HFSCs) based on in vivo role of the bladder and surrounding tissue environment are likely to differentiate into urothelial cells and smooth muscle cells, which become a new stem cell research.
OBJECTIVE: To investigate an effective and simple method of isolation, culture and identification of rat HFSCs.
METHODS: After sterling, the skin of the rat’s beard was cut off, the vibrissa follicles were dissected under a stereocroscope. The dermal sheath was moved by incubated in Dispase Ⅱ firstly, second step with a mixture of trypsin and EDTA. The cells suspension was cultured in 10% fetal bovine serum keratinocyte free serum medium and selected by rapid adhering on collagen Ⅳ. The cultured cells were passaged, when cells were confluent with 70%-80%. Selected cell microstruture was observed under optics microscope and electron microscopy respectively. Cells were characterized by flow cytomitry with antibody against CD34 and β1-integrin.
RESULTS AND CONCLUSION: The selected HFSCs were uniform in shape, more refraction, cobblestone-morph by optics microscope. The form was primitive, small ratio of nucleus and cytoplasm, obvious nucleolus, organelle of developmental immaturity by transmission electron microscope. In adherent cells (experiment groups), CD34 and β1-integrin expression rate was (39.52±19.57)% and (93.46±4.73)%, respectively. In no-adherent cells (control groups) was corresponding (19.20±11.53)% and (63.57±14.42)% respectively. There was a significant difference between two groups (P < 0.05). Highly purified HFSCs can be obtained by dissection under a stereocroscope and two-step enzyme digestion combining with rapid adhering on collagen Ⅳ. CD34 combined with β1-integrin are an ideal marker to identify HFSCs.