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Background and Objectives: Considering the pulse widths of picosecond and nanosecond lasers used in cutaneous laser surgery differ by approximately one order of magnitude, can nanosecond lasers produce the optical effect in human skin similar to laser-induced optical breakdown (LIOB) caused by picosecond lasers?, Methods: Cutaneous changes induced by a focused fractional nanosecond 1064-nm Nd:YAG laser were evaluated by VISIA-CR imaging, histological examination, and harmonic generation microscopy (HGM)., Results: A focused fractional nanosecond 1064-nm Nd:YAG laser can generate epidermal vacuoles or dermal cavities similar to the phenomenon of LIOB produced by picosecond lasers. The location and extent of photodisruption can be controlled by the laser fluence and focus depth. Moreover, laser-induced shock wave propagation and thermal degeneration of papillary collagen can be observed by HGM imaging., Conclusion: Focused fractional nanosecond lasers can produce an optical effect on human skin similar to LIOB caused by picosecond lasers. With techniques of application, the treatment can induce epidermal and dermal repair mechanisms in a tunable fashion to improve skin texture, wrinkles, scars, and dyspigmentation, without disrupting the epidermal surface., (© 2024 Wiley Periodicals LLC.)

Imaging depth and quality of optical microscopy can be enhanced by optical clearing. Here we investigate the optical clearing of the ex vivo human skin by 50% glycerol topical application, which is allowed for cosmetic usage. Harmonic generation microscopy, by combining second and third harmonic generation (THG) modalities, was utilized to examine the clearing effect. The THG image intensity is sensitive to the improved optical homogeneity after optical clearing, and the second harmonic generation (SHG) image intensity in the dermis could serve as a beacon to confirm the reduction of the scattering in the epidermis layer. As a result, our study supports the OC effect through 50% glycerol topical application. Our study further indicates the critical role of stratum corneum shrinkage for the observed SHG and THG signal recovery.

Third-harmonic generation (THG) microscopy has been reported to provide intrinsic contrast in elastic fibers, cytoplasmic membrane, nucleus, actin filaments, lipid bodies, hemoglobin, and melanin in human skin. For advanced molecular imaging, exogenous contrast agents are developed for a higher structural or molecular specificity. We demonstrate the potential of the commonly adopted tattoo dye as a THG contrast agent for in vivo optical biopsy of human skin. Spectroscopy and microscopy experiments were performed on cultured cells with tattoo dyes, in tattooed mouse skin, and in tattooed human skin to demonstrate the THG enhancement effect. Compared with other absorbing dyes or nanoparticles used as exogenous THG contrast agents, tattoo dyes are widely adopted in human skin so that future clinical biocompatibility evaluation is relatively achievable. Combined with the demonstrated THG enhancement effect, tattoo dyes show their promise for future clinical imaging applications.

Surgical borders of extramammary Paget's disease (EMPD) are difficult to be identified via its clinical appearance. In this study, we propose a new diagnostic technique which combines nonlinear harmonic generation microscopy (HGM) with the deep learning method to instantaneously determine whether the imaged 3D stack is malignant EMPD or surrounding normal skin digitally. To demonstrate our proposal, in this study different locations of fresh EMPD surgical samples were 3D imaged starting from the surface up to a depth of 180 μm using stain-free HGM. With the followed histopathological examination of the same sample, we mapped the gold-standard results to 3D HGM image stacks with labels for the training of the deep learning model. With only 2095 3D image stacks as training and validation data, the results of EMPD and normal skin tissue classification achieve 98.06% sensitivity, 93.18% specificity and 95.81% accuracy. This study supports our proposed 3D convolutional-neural-network-based technique with a high potential to assist physicians to quickly map the EMPD margins by providing noninvasive instant information regarding the imaged sub-millimeter site as malignant or surrounding normal with a high accuracy. [ABSTRACT FROM AUTHOR]

Melanin is known to provide strong third-harmonic generation (THG) contrast in human skin. With a high concentration in basal cell cytoplasm, THG contrast provided by melanin overshadows other THG sources in human skin studies. For better understanding of the THG signals in keratinocytes without the influence of melanin, an in vivo THG microscopy (THGM) study was first conducted on vitiliginous skin. As a result, the THG-brightness ratio between the melanin-lacking cytoplasm of basal cells and collagen fibers is about 1.106 at the dermal–epidermal junctions of vitiliginous skin, indicating high sensitivity of THGM for the presence of melanin. We further applied the in vivo THGM to assist evaluating the therapeutic outcome from the histopathological point of view for those showed no improvement under narrowband ultraviolet B therapy based on the seven-point Physician Global Assessment score. Our clinical study indicates the high potential of THGM to assist the histopathological assessment of the therapeutic efficacy of vitiligo treatments. [ABSTRACT FROM AUTHOR]