In vivo harmonic generation microscopy for monitoring the height of basal keratinocytes in solar lentigines after laser depigmentation treatment

Abstract

The development of solar lentigines (SLs) is related to chronic ultraviolet exposure-induced cell senescence. We have previously demonstrated that basal keratinocyte enlargement is a morphological hallmark of skin senescence correlated to the process of skin aging, while clinical studies on the long-term monitoring of the cellular morphological changes in SLs after laser treatment are lacking. In this study, we have developed the harmonic generation microscopy (HGM) for in vivo monitoring the height of basal keratinocytes (HBK) and had administered Q-switched ruby laser or picosecond 532-nm Nd:YAG laser treatment on each side of the face of 25 Asian patients with facial SLs, respectively. In vivo HGM imaging was conducted to longitudinally analyze HBK and the horizontal cell size (HCS). Before treatment, the HBK was significantly higher in the SLs lesional area than that in the adjacent normal region, whereas there was no significant difference in the HCS. After treatment, the lesional HBK remained significantly higher than normal skin regardless of the laser treatment used. Our study indicates that the basal keratinocytes remain abnormal after laser treatment and demonstrates the capability of in vivo HGM for longitudinal, quantitative monitoring of cell senescence and therapeutic effect in SLs.

Characterization of picosecond laser‐induced optical breakdown using harmonic generation microscopy

“…A Cr:forsterite-based HGM system [23][24][25][26][27]32 with a central wavelength of ~1260 nm was used for skin imaging. A 1.15 numerical aperture objective (×40; working distance 250 μm; UApo N 340; Olympus) was used to focus the laser beam on the skin to capture high-resolution images.…”

“…Previous studies have shown that combining a ~1260 nm harmonic generation microscopy (HGM) system with second harmonic generation (SHG) and third harmonic generation (THG) modalities can provide a label-free, noninvasive optical virtual skin biopsy with a resolution of 0.4 mm and a penetration depth of 270 mm to observe the epidermis and papillary dermis. [23][24][25][26][27] The image of HGM system is dichromatic and consists of two signals, THG and SHG, respectively. THG signal mainly comes from the components and structures of epidermis, such as melanin pigment, multilayer structures, organelles, and lipids, while SHG mainly comes from the contrast of type I collagen fibers.…”

“…[28][29][30][31] The vivid contrast at the dermal-epidermal junction and cellular resolution of HGM has been shown to be useful in the assessment of intrinsic skin aging, solar lentigines after laser treatment and diseases such as actinic keratosis and basal cell carcinoma. 24,25,27,29 This pilot study aimed to understand the response of human skin to nonablative fractional 1064-nm picosecond laser treatment using an in vivo HGM imaging system, particularly emphasizing dermal changes after laser treatment.…”

A Noninvasive Skin Biopsy of Free Nerve Endings via Realtime Third-Harmonic Microscopy

“…The imaging plane was moved to different depths by tuning the 3D stage along the optical axis. Following a similar pulse width measurement and dispersion control [69], the pulse width after the objective was estimated to be 28 fs. The epi-THG transverse and axial resolutions were approximately 0.4 and 1.1 μm, respectively [70].…”

In vivo microscopy as an adjunctive tool to guide detection, diagnosis, and treatment

“…Furthermore, longitudinal IVM may be used to monitor response to various therapies without biopsy, such as endoscopic mucosal or radiofrequency (RF) ablation, photodynamic therapy, skin pigmentation disorder treatments, and others. 34 – 37 …”