Yoshiyuki Kohno

KINETIC STUDY OF QUENCHING REACTION OF SINGLET OXYGEN AND SCAVENGING REACTION OF FREE RADICAL BY SQUALENE IN N-BUTANOL

The rate constant of quenching of singlet oxygen (kQ) by squalene (SQ) is found to be much larger than those of the lipids in human skin surface. SQ is the first target lipid in human skin surface by oxidative stresses such as sun light exposure. kQ of SQ is similar to that of 3,5-di-t-butyl-4-hydroxytoluene (BHT). The large kQ of SQ is due to the small ionization potential. SQ consists of six 2-methyl-2-pentene units and kQ of SQ is about 6-times as large as that of 2-methyl-2-pentene. The electron donating property of methyl groups bonded to quaternary carbons of SQ is essential to the large kQ. SQ is not very susceptible to peroxidation and is stable for attacks by peroxide radicals. The chain reaction of lipid peroxidation is unlikely to be propagated with SQ in human skin surface. It is concluded that SQ functions as an efficient quencher of singlet oxygen and prevents the corresponding part of lipid peroxidation in human skin surface.

Urokinase-type plasminogen activator is activated in stratum corneum after barrier disruption

The plasminogen/plasmin system in epidermis is thought to be the major protease involved in the delay of barrier recovery. However, little is known about the mechanism through which this system is activated. In order to clarify this mechanism, we first determined the distribution of proteolytic activity by using in situ zymography. As a result, plasminogen-activator activity was found to be present in the stratum corneum (SC) after barrier disruption. Next, SC subjected to repeated barrier disruption was collected to identify the protease. The protease was identified as urokinase-type plasminogen activator, because flybrinolytic activity of the collected SC was abolished by addition of anti-urokinase antibody. Urokinase activation in SC was confirmed by means of an in vitro assay, in which the precursor of urokinase (pro-uPA) became active after incubation with the insoluble component of SC homogenate. These findings indicated that urokinase-type plasminogen activator is activated in SC after barrier disruption and this activation might trigger the plasminogen/plasmin system in the epidermis.

Non-invasive study of gelatinases in sun-exposed and unexposed healthy human skin based on measurements in stratum corneum

Gelatinases, which belong to the family of matrix metalloproteinases, degrade various components of skin, and may be involved in photoaging, since they are upregulated by low-dose UV exposure to the skin. However, their behavior in healthy human skin is still unclear. In the present study, gelatinases was specifically detected in stratum corneum (SC) of skin from sun-exposed sites, including the face, in healthy humans, but not in SC of skin from unexposed sites. Following experimental UVB irradiation of the abdomen in volunteers, gelatinases were detected in tape-stripped SC from the site for several weeks, and subsequently disappeared. The appearance of gelatinase in SC after a lag time consistent with SC turnover is considered to reflect upregulation of gelatinase expression in keratinocytes in response to UVB-exposure of the skin. A survey of gelatinases in facial SC samples collected by tape-stripping from the cheeks of 100 healthy women revealed that the enzyme was present in 90% of subjects. These results, taken together, suggest that gelatinase is constantly upregulated by sunlight in the facial epidermis of most women during their daily lives, and may be an etiological factor in photoaging, e.g., by promoting wrinkle formation.