NONINTRUSIVE TESTING OF SKIN 19 Table I Non-Intrusive Assessments of Epidermal Proliferative Activity • Transit Time Number of Turnover Rate Site (days) Cell Layers (h/layer) Volar Forearm 35 19.8 + 1.39 17.0 + 0.83 28.3 + 1.2 60 28.1 _+ 2.66 16.8 + 0.66 $0.8 + 3.8 tipper Inner Arm 35 17.7 + 2.02 14.3 + 0.61 30.0 + 2.6 60 25.5 + 2.63 13.9 -+ 0.81 46.6 + 6.5 •Results are means _+ S.E. bound, they move in unison through the stratum corneum. Thus, in this special case, transit time is equivalent to turnover time, i.e., the time required for a compartment to renew or replace itself. This value can be measured nonintrusively by determining with the aid of a Wood's lamp, the time required for a fluorescent marker (Dansyl Chloride) to disappear from fully stained horny layer (10). This dye binds avidly to the horny layer only and its rate of disappearance is not influenced by washing or protecting the surface. Our preliminary results with Dansyl Chloride Disappearing Method (Table I) indicate that for both the volar forearm and upper inner arm, stratum corneum transit times of young adult subjects are approximately 18 to 20 days. In older subjects, this was lengthened about eight to nine days at both sites indicating that epidermal proliferation decreases with age. It should be emphasized that transit time values can be misleading unless they are related to the number of cell layers (11). If cell proliferation is the same, the time for a cell to reach the skin surface will be longer if there are more cell layers. Thus, transit times should be corrected for differences in number of cell layers and converted to turnover rates. Estimating cell layers entails raising up small blisters with ammonium hydroxide, cryostat sectioning the blister roof and treating it with alkali (12). The horny cells swell up and the number of cell layers can easily be counted. Our preliminary results (Table I) reveal a site difference but no age differences with regard to the number of cell layers in the stratum corneum. Thus, the increased transit time values observed in older subjects is truly a reflection of diminished proliferative activity. In fact, the calculated turnover values indicate that on the average it takes about 30 hours to replace each horny cell layer in younger subjects and about 45 hours for older subjects. In addition to providing a stratum corneum sample for cell layer counts, the blister roofs can be subjected to additional types of analyses, such as moisture avidity or physiochemical properties (13). It is apparent that much will be learned about the structural and functional properties of the stratum corneum from such a comprehen- sive examination of blister roof samples. WOUND HEALING The unroofed blister site represents a reasonable standard superficial wound at which the dermatoglyphics have been completely obliterated. Thus, by observing the restoration of the original markings we can objectively evaluate the rate of wound

20 JOURNAL OF THE SOCIETY OF COSMETIC CHEMISTS Week I. Week 2 Week 5 . ß .. ...... :'"- • [:: :' t __ E-]No Dermatoglyphics •Minor Lines Filling In :::•Mojor Lines Returning •Complete Restoration Figure 4. Wound healing following unroofing of NH4OH blisters on volar forearms of human subjects. healing. Figure 4 shows some preliminary results with regard to the volar forearm site. Note that by the end of the first week, all young subjects had begun to reestablish major lines, while half of the older ones still show a complete absence of lines. By the end of the second week, most of the young and many of the old had reestablished all major lines and were beginning to fill in the minor ones. The differences in rate of wound healing become even more pronounced in subsequent weeks. By the fourth week, all young subjects had completely restored their dermatoglyphic pattern. In contrast, only 25% of the older group had achieved this stage, and in fact, one individual had not even completed reestablishment of the major lines. By the end of the sixth week, all but one of the older subjects had completely restored their dermatoglyphics. A similar progression of events was observed in the upper inner arm site. BLISTER FORMATION The ability to raise up blisters has prov&d useful in a variety of ways in this investigation. Frosch and Kligman (14) originally described the induction of blisters by ammonium hydroxide (NH4OH). In this procedure, a 1:1 aqueous solution of NH4OH is placed in a 14-mm well drilled in an acrylic plastic block which is held snugly to the skin surface. During the exposure, the site is examined under good lighting at 30-s intervals. The time required for the appearance of tiny vesicles around the follicles is termed minimal blistering time (MBT). The exposure is continued and the additional time required for a fully tense blister to form, blister filling time (BFT), is also measured. Table II compares blister formation of the two age groups on the volar forearm and upper inner arm. At both sites, the young achieved an MBT response after 13 to 14 min of exposure, while in striking contrast, the older subjects as a group took only half as long. At the present time, it is not clear what accounts for these differences.