The Study Of Cultivation And Identification On Hair Follicle Stem Cells Of The Rat

Objective:It was well known that the stem cells was definited with prematuration and the patent of refreshment itself and differentiation to embryo and adult stem cells, other tissues and organs'cells. The periodicity growth of hair follicle revealed there are a group of cells that were named after hair follicle stem cells(HFSCs)with proliferating potentiality in it. The source of hair follicle stem cells is widespread, moreover, it is easy to draw the materials from the animal's body, these stem cells displayed powerful potential of growth and colonies'formation out of the animal's body, a single cell can fission 1.7X1038, not to involve in the question of the ethics and morality, these stem cells can be differentiated to epiderm, sebaceae, hair follicle, neuron, adipose cell and osteoblast, and so forth, so HFSCs became a ideal seed cells. In presently, there are two main methods on HFSCs'selection and purification, by magnetic activated cell sorting(MACS)and rapid adhering on collagenⅣ. The approach is not an ideal choice because it suffers from expensive, operation complicated, the stem cells could be injuried in the process, only be done in better condition laboratory. In contrast, highly purified HFSCs can be obtained by dissection under a stereocroscope and two-step enzyme digestion combining with rapid adhering on collagenⅣ. Methods:The offsprings of cleaning SD rats was selected in the experiment, After steriling, the skin of the rat's beard was cut off, the vibrissa follicles were dissected under a stereocroscope. the Hair follicle outer sheath was moved by incubated in DispaseⅡfirstly, second step with a mixture of trypsin and EDTA. The cells suspension was cultured in 10% fetal bovine serum(FBS)Keratinocyte free serum medium(K-FSM)and selected by rapid adhering on collagenⅣThe cultured cells were passaged, when cells were confluence with 70-80%. Selected cells microstruture was observed under optics microscope and electron microscopy respectively. The growth curve was drew, to stain of Giemsa and immunofluorescence. Cells were characterized by flow cytomitry with antibody against CD34 andβ1-integrin. Results:The selected HFSCs was 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. Conclusion, Highly purified and better activity HFSCs can be obtained by dissection under a stereocroscope and two-step enzyme digestion combining with rapid adhering on collagenⅣ. CD34 combined withβ1-integrin were more ideal marker to identify hair follic stem cells. To do the groundwork for the next step work about differentiation.