Polarization-resolved second-harmonic-generation imaging of photoaged dermal collagen fiber

ABSTRACT Polarization-resolved second-harmonic-generation (SHG) microscopy is useful for assessment of collagen fiber orientation in tissues. In this paper, we investigated the relation between wrinkle direction and collagen orientation in ultraviolet-B-exposed (UVB-exposed) skin using polarization-resolved SHG microscopy. A polarization anisotropic image of the SHG light indicated that wrinkle direction in UVB-exposed skin is predominantly parallel to the orientation of dermal collagen fibers whereas no-UVB-exposed skin was dominated by collagen orientation parallel to the meridian line of body. The method proposed has the potential to become a powerful non-invasive tool for assessment of cutaneous photoaging. Keywords: Second-harmonic-generation, microscopy, polarization, collagen, orientation, photoaging, wrinkle 1. INTRODUCTION Aging is an irreversible, physiological phenomenon that cannot be avoided. In skin, mechanical properties such as tension and elasticity decline gradually with age, resulting in the appearance of wrinkles and sagging. Furthermore, repeated exposure of ultraviolet B (UVB) rays in sunlight to the skin often accel erates skin aging and causes changes in the skin, such as mottled pigmentation, leathery texture, laxity, sallowness, and deep wrinkle. This is called cutaneous photoaging [1]. Therefore, the need exists for an in vivo assessment technique to estimate the degree of cutaneous photoaging for studies in fields such as skin cosmetics and anti-aging dermatology. Deep wrinkle characteristic of UVB-exposed skin is closely re lated to the quantity and structure of collagen fibers in the superficial dermis because dermal collagen fibers contribute to the morphology and mechan ical properties of the skin. For example, the biochemical, histochemical, and immunohistochemical experiments indicated that the dermal collagen fibers in the photoaged neck skin remarkably decrease in comparison with the intrinsically aged gluteal skin [2]. Therefore, in vivo monitoring of dermal collagen fibers is a key step in assessment of cutaneous photoaging. Optical probe methods are attractive for such assessments because they are simple, rapid, and non-invasive. Furthermore, optical probes can be applied directly because skin is a superficial tissue. Among the optical probe methods available for skin studies [3-5], second-harmonic-generation (SHG) light is attractive for in vivo monitoring of dermal collagen fiber because SHG light is generated specifically by collagen fibers in the dermis [6, 7]. This SHG light provides unique imaging characteristics: high contrast, high spatial resolution, optical three-dimensional (3D) sectioning, non-invasiveness, and deep penetration. By using the naturally endogenous SHG process as a contrast mechanism, the structures of collagen fiber in the tissues can be clearly visualized without additional staining, and so are free from photodamage, phototoxicity, or photobleaching. For the evaluation of cutaneous photoaging, the combination of SHG imaging with two-photon autofluorescence imaging has been applied effectively as an indicator of changes in dermal collagen and elastin content because degr adation of collagen and excessive depos ition of abnormal elastin are observed in photoaged skin [8, 9]. In contrast, combining SHG imag ing with polarization methods can provide additional insights regarding the direction of collagen fibers, i.e., collagen orientation. The efficiency of SHG light is sensitive to collagen orientation when the incident light is linearly polarized, and hence polarization measurements of the SHG light are effective for probing collagen orientation in the dermis [10, 11] and other tissues [11, 12]. However, few reports have examined changes in collagen orientation caused by cutaneous photoaging.