Tonofilament differentiation in human epidermis, isolation and polypeptide chain composition of keratinocyte...

Epidermal tonofilaments are intermediate filaments with an unusually complex polypeptide chain composition which undergoes profound changes during differentiation. Assessment of the possible functional significance of these changes requires their correlation with defined stages of keratinocyte differentiation. Methods have, therefore, been developed for the isolation of the four keratinocyte subpopulations from human epidermis in high yield and purity without detectable damage to tonofilament chains. Trypsinization at 4 degrees C yielded sheets of granular and horny layers and a suspension of basal and spinous cells. Granular cells were removed from the horny layer sheets by mechanical action and basal cells separated from spinous cells by selective attachment to collagen. The living cell fractions were representative, by morphological and ultrastructural criteria, of their counterparts in situ. Analysis of the keratinocyte subpopulations by SDS-PAGE revealed directly their tonofilament chain composition. Basal cells contain two quantitatively major tonofilament chains (Mr 58000 and 50000). A third major chain (Mr 70000) is only synthesised suprabasally when the cells lose their ability to divide. The Mr 50000 chain is removed before the granular cell stage and several minor chains are present in the spinous and granular cells. Two chains appearing for the first time in horny cells probably result from processing during the terminal stages of differentiation. It is suggested that synthesis of the Mr 70000 chain in suprabasal keratinocytes is normally linked with their loss of mitotic ability. This may be significant in the skin disease psoriasis in which synthesis of the Mr 70000 chain is defective and the epidermis hyperproliferates.