Skin dark spot mapping and evaluation of brightening product efficacy using Line-field Confocal Optical Coherence Tomography (LC-OCT).

BACKGROUND: Facial dark spots remain a significant challenge for the cosmetic industry, in terms of providing effective treatment. Using Line-field Confocal Optical Coherence Tomography (LC-OCT), we investigated the internal structural features of photo-aging spot areas and evaluated the efficacy of a skin-brightening cosmetic product. MATERIALS AND METHODS: Twenty-six Asian female volunteers, aged between 29 and 65 years, applied a cosmetic product on their entire face twice a day for 2 months. LC-OCT was used to evaluate the dermal-epidermal junction (DEJ) undulation and the volume density of melanin in the epidermis at D0 and D56. Skin brightening and redness were also assessed by photography (SkinCam). RESULTS: Using LC-OCT technology, various microscopic dark spot morphologies, spanning from minimally deformed DEJ to complex DEJ patterns, were identified. Dark spots characterized by slight deformities in the DEJ were predominantly observed in the youngest age group, while older volunteers displayed a wavier pattern. Furthermore, a total of 44 spots were monitored to evaluate the brightening product efficacy. A statistically significant reduction in melanin volumetric density of 7.3% in the spots and 12.3% in their surrounding area was observed after 56 days of product application. In line with these results, an analysis of color parameters using SkinCam reveals a significant increase in brightening and decrease in redness in both pigmented spots and the surrounding skin following application. CONCLUSIONS: LC-OCT proves to be a valuable tool for in-depth dark spots characterization and assessment of skin brightening products, enabling various applications in the field of dermatological sciences.

Line-field confocal optical coherence tomography coupled with artificial intelligence algorithms to identify quantitative biomarkers of facial skin ageing.

Quantitative biomarkers of facial skin ageing were studied from one hundred healthy Caucasian female volunteers, aged 20-70 years, using in vivo 3D Line-field Confocal Optical Coherence Tomography (LC-OCT) imaging coupled with Artificial Intelligence (AI)-based quantification algorithms. Layer metrics, i.e. stratum corneum thickness (SC), viable epidermal thickness and Dermal-Epidermal Junction (DEJ) undulation, as well as cellular metrics were measured for the temple, cheekbone and mandible. For all three investigated facial areas, minimal age-related variations were observed in the thickness of the SC and viable epidermis layers. A flatter and more homogeneous epidermis (decrease in the standard deviation of the number of layers means), a less dense cellular network with fewer cells per layer (decrease in cell surface density), and larger and more heterogeneous nuclei within each layer (increase in nuclei volume and their standard deviation) were found with significant variations with age. The higher atypia scores further reflected the heterogeneity of nuclei throughout the viable epidermis. The 3D visualisation of fine structures in the skin at the micrometric resolution and the 1200 µm × 500 µm field of view achieved with LC-OCT imaging enabled to compute relevant quantitative biomarkers for a better understanding of skin biology and the ageing process in vivo.

Line-field confocal optical coherence tomography as a tool for three-dimensional in vivo quantification of healthy epidermis: A pilot study.

Epidermal three-dimensional (3D) topography/quantification has not been completely characterized yet. The recently developed line-field confocal optical coherence tomography (LC-OCT) provides real-time, high-resolution, in-vivo 3D imaging of the skin. This pilot study aimed at quantifying epidermal metrics (epidermal thicknesses, dermal-epidermal junction [DEJ] undulation and keratinocyte number/shape/size) using 3D LC-OCT. For each study participant (8 female, skin-type-II, younger/older volunteers), seven body sites were imaged with LC-OCT. Epidermal metrics were calculated by segmentations and measurements assisted by artificial intelligence (AI) when appropriate. Thicknesses of epidermis/SC, DEJ undulation and keratinocyte nuclei volume varied across body sites. Evidence of keratinocyte maturation was observed in vivo: keratinocyte nuclei being small/spherical near the DEJ and flatter/elliptical near the skin surface. Skin microanatomy can be quantified by combining LC-OCT and AI. This technology could be highly relevant to understand aging processes and conditions linked to epidermal disorders. Future clinical/research applications are to be expected in this scenario.

In vivo large-scale analysis of Drosophila neuronal calcium traces by automated tracking of single somata.

How does the concerted activity of neuronal populations shape behavior? Impediments to address this question are primarily due to critical experimental barriers. An integrated perspective on large scale neural information processing requires an in vivo approach that can combine the advantages of exhaustively observing all neurons dedicated to a given type of stimulus, and simultaneously achieve a resolution that is precise enough to capture individual neuron activity. Current experimental data from in vivo observations are either restricted to a small fraction of the total number of neurons, or are based on larger brain volumes but at a low spatial and temporal resolution. Consequently, fundamental questions as to how sensory information is represented on a population scale remain unanswered. In Drosophila melanogaster, the mushroom body (MB) represents an excellent model to analyze sensory coding and memory plasticity. In this work, we present an experimental setup coupled with a dedicated computational method that provides in vivo measurements of the activity of hundreds of densely packed somata uniformly spread in the MB. We exploit spinning-disk confocal 3D imaging over time of the whole MB cell body layer in vivo while it is exposed to olfactory stimulation. Importantly, to derive individual signal from densely packed somata, we have developed a fully automated image analysis procedure that takes advantage of the specificities of our data. After anisotropy correction, our approach operates a dedicated spot detection and registration over the entire time sequence to transform trajectories to identifiable clusters. This enabled us to discard spurious detections and reconstruct missing ones in a robust way. We demonstrate that this approach outperformed existing methods in this specific context and made possible high-throughput analysis of approximately 500 single somata uniformly spread over the MB in various conditions. Applying this approach, we find that learned experiences change the population code of odor representations in the MB. After long-term memory (LTM) formation, we quantified an increase in responsive somata count and a stable single neuron signal. We predict that this method, which should further enable studying the population pattern of neuronal activity, has the potential to uncover fine details of sensory processing and memory plasticity.

Comparison of line-field confocal optical coherence tomography images with histological sections: Validation of a new method for in vivo and non-invasive quantification of superficial dermis thickness.

BACKGROUND: Line-field confocal optical coherence tomography (LC-OCT) is an imaging technique providing "optical biopsies" of the skin in real time and non-invasively. At a center optical wavelength of 1.3 µm, this innovative technology can be applied to dermo-cosmetic product development due to both high image resolution (~2 µm) and sufficient penetration (~0.5 mm). Nevertheless, the precise dermal area analyzed with LC-OCT has never been identified. In this study, the objective was to compare LC-OCT images with histological sections of the same area, in order to validate a new method for in vivo and non-invasive quantification of superficial dermis thickness. Once validated, this standardized and quantitative method was used to assess age-related changes of the superficial dermis. MATERIALS AND METHODS: Ex vivo LC-OCT acquisitions and hematoxylin-eosin-safran staining were performed on a panel of four healthy Caucasian female volunteers. In vivo LC-OCT study of skin aging was performed on a panel of 37 healthy Caucasian female divided into five different age-groups. RESULTS: Comparison with histological sections revealed that LC-OCT images allow the visualization and the quantification of the superficial portion of papillary dermis. Applied to different age-group of volunteers, LC-OCT images show a constant decrease in this superficial dermis thickness with age. CONCLUSIONS: In conclusion, we have introduced LC-OCT as a novel technique for in vivo and non-invasive evaluation of superficial dermis thickness. This approach could be used in the future to demonstrate visually and quantitatively the capacity of a dermo-cosmetic active ingredient to renormalize the structural properties of the dermis.

Sensorless adaptive optics implementation in widefield optical sectioning microscopy inside in vivo Drosophila brain.

We present an implementation of a sensorless adaptive optics loop in a widefield fluorescence microscope. This setup is designed to compensate for aberrations induced by the sample on both excitation and emission pathways. It allows fast optical sectioning inside a living Drosophila brain. We present a detailed characterization of the system performances. We prove that the gain brought to optical sectioning by realizing structured illumination microscopy with adaptive optics down to 50 µm deep inside living Drosophila brain.

UV Photofragmentation Dynamics of Protonated Cystine: Disulfide Bond Rupture.

Disulfide bonds (S-S) play a central role in stabilizing the native structure of proteins against denaturation. Experimentally, identification of these linkages in peptide and protein structure characterization remains challenging. UV photodissociation (UVPD) can be a valuable tool in identifying disulfide linkages. Here, the S-S bond acts as a UV chromophore and absorption of one UV photon corresponds to a σ-σ* transition. We have investigated the photodissociation dynamics of protonated cystine, which is a dimer of two cysteines linked by a disulfide bridge, at 263 nm (4.7 eV) using a multicoincidence technique in which fragments coming from the same fragmentation event are detected. Two types of bond cleavages are observed corresponding to the disulfide (S-S) and adjacent C-S bond ruptures. We show that the S-S cleavage leads to three different fragment ions via three different fragmentation mechanisms. The UVPD results are compared to collision-induced dissociation (CID) and electron-induced dissociation (EID) studies.

Photofragmentation at 263 nm of small peptides containing tyrosine: the role of the charge transfer on CO.

The photofragmentation pathways at 263 nm of several small peptides containing tyrosine as the UV chromophore have been characterized using a multi-coincidence technique. A detailed study of the fragmentation dynamics of protonated Glycine-Tyrosine (GYH(+)), Tyrosine-Glycine (YGH(+)), Glycine-Tyrosine-Glycine (GYGH(+)), Alanine-Tyrosine (AYH(+)) and Tyrosine-Alanine (YAH(+)) is presented in this paper. Fragmentations occurring or initiated in an excited state are distinguished from those occurring after internal conversion to the ground electronic state by their rapid fragmentation times and binary nature. For the studied systems, it is shown that fragmentations occurring after internal conversion to the ground state are the dominant processes compared to fragmentations occurring in the excited state. The low abundances associated with the observed UV photospecific channels, i.e. Cα-Cß bond breakage in YGH(+) and YAH(+) and direct z-type bond breakage in GYGH(+), respectively, can be rationalized upon consideration of charge transfer states accessible after absorption of one UV photon. Indeed, excited state calculations performed at the RI-CC2 level of theory indicate that charge transfer on the active CO group is a prerequisite for photospecific bond ruptures. The fragmentation mechanisms and the localization of the charge on the side chain after fragmentation are discussed in terms of ionization energies of the fragments.