Methods for culturing hair follicle epithelial matrix cells

Abstract

The subject invention relates to methods of isolating, culturing and propagating epithelial matrix cells by removing/isolating the bulb region of a hair follicle directly above the dermal papilla, incubating the bulb in a pre-coated culture dish containing Chang medium supplemented with 10% fetal bovine serum, replacing the Chang medium with keratinocyte growth medium after 3-4 days, propagating the cells to the next passage after adding the keratinocyte growth medium and passing the pure epithelial matrix cells again to obtain a third passage of cells. The epithelial matrix cells may be passed for up to six passages, and are useful for screening potential hair growth actives.

What is claimed is: 
     
       1. A method of isolating and culturing epithelial matrix cells, comprising the steps of: a) micro-dissecting intact hair follicles, wherein the hair follicles comprise a bulb region, the bulb region comprising dermal papilla cells and epithelial matrix cells, wherein the epithelial matrix cells exist above the dermal papilla cells and the dermal papilla cells exist proximate to the base of the bulb region;   b) cutting the bulb region of each hair follicle to remove the bulb region from the follicle, the bulb region being cut within about 50 microns or less above the dermal papilla cells to form a cut bulb region comprising epithelial matrix cells and the dermal papilla cells;   c) placing the cut bulb region in a culture plate containing Chang medium;   d) allowing the cut bulb region to form new epithelial matrix cells for about 4 to 6 days;   e) replacing the Chang medium with a medium suitable for keratinocyte growth which allows the proliferation of new epithelial matrix cells beyond the period of step (d);   f) passing the new epithelial matrix cells 4-6 days following addition of the medium suitable for keratinocyte growth by i) treating the new epithelial matrix cells with trypsin and EDTA to detach the new epithelial matrix cells from the bottom of the culture plate;   ii) neutralizing the trypsin; and   iii) dispensing the new epithelial matrix cells to a new plate to obtain a second passage of pure epithelial matrix cells; and     g) passing the pure epithelial matrix cells obtained from step (f) to a third passage.   
     
     
       2. A method of screening a potential hair growth compound for activity comprising measuring change in proliferation of epithelial matrix cells after addition of a hair growth compound to the epithelial matrix cells, wherein the epithelial matrix cells are obtained according to the method described in claim 1. 
     
     
       3. A method of screening a potential hair growth compound for activity comprising measuring change in level of hair specific keratin ribonucleic acid of epithelial matrix cells after addition of the hair growth compound to the epithelial matrix cells, wherein the epithelial matrix cells are obtained according to the method described in claim 1.