This conference presentation was prepared for SPIE Photonics West BiOS 2024.

Optical coherence tomography angiography (OCTA) has growing application for microvascular assessment in dermatology. Various instruments, imaging protocols, processing methods, and metrics have been used to describe the microvasculature, so comparing different study outcomes is currently not generally feasible. We present a user-friendly, open-source toolbox, OCTAVA, to help remedy this shortcoming. We present three use cases for assessing software accuracy and repeatability, and we investigate how OCTAVA can support wider adoption of OCTA as a clinical tool. Wide adoption of this software will help drive the development of reliable microvascular biomarkers for early detection and treatment guidance of diseases.

Optical coherence tomography (OCT) and speckle imaging are two coherence-based imaging modalities with burgeoning applications in rapid non-invasive measurement of the skin. Both modalities are sensitive to tissue morphology and have polarization-sensitive augmentations. This study compares Polarization Sensitive OCT (PS-OCT) and Polarization Speckle measurements to better understand the relationship between polarization and coherence properties of skin. Volunteers of Fitzpatrick skin type I through VI were recruited and healthy skin was measured at four body sites (palm, inner forearm, forehead, and eye corner). Preliminary results indicate a strong similarity between the skin surface roughness measurements of PS-OCT and polarization speckle. In regards to tissue depolarization, PS-OCT measurements appear minimally affected by skin color, whereas polarization speckle was strongly affected due to differing measurement wavelengths. Among body sites, the palm and face were found to be generally smoother than the forearm; however the epidermis layer of the palm demonstrated notably greater polarization scrambling.

Sex-related skin differences are known to be expressed such as differences in sebum secretion, yet the relationship to skin’s internal structures is still unclear. Recently, a histopathological study revealed differences in micro-vessel density between females and males. Past studies have been limited to histological vascular assessments, but this study aimed to characterize dermal blood vessels in females and males using optical coherence tomography angiography. We found sex-related differences in vessel volume density through inflammatory-related factors. Also, a male group with higher vascular density showed higher trans-epidermal water loss, implying a relationship between dermal vasculature and skin physiological conditions in males.

The key objective of this study is to test the ability of a newly developed laser therapy technique – multiphoton photothermolysis – to generate precise tissue alteration on in vivo human skin, and to demonstrate the efficiency of a prototype multimodal microscopy to monitor skin response in real-time after the laser treatment. Cellular activities such as proliferation, inflammation, exfoliation of injured tissue can be captured over time. This study highlights the potential use of multiphoton photothermolysis for treatment of conditions that preventing collateral damage is critical. Also, multimodal microscopy offers a non-invasive method for monitoring tissue physiology.

Introducing PRESS (Pressure Enhanced Sensing Surgery) – a groundbreaking method to image hypoxia in skin lesions. Chaotic, weak blood vessels in tumors cause microregional chronic and cycling hypoxia, and low-pressure blood flow. PRESS takes advantage of the FDA-approved drug aminolaevulinic acid, to imply the synthesis of Protoporphyrin IX (PpIX) in tissue. PpIX's delayed fluorescence (DF) serves as a unique indicator of local oxygen levels, particularly in mitochondria, making it ideal for tissue oxygen measurement. Enhanced signal resolution is achieved through palpation, capturing both chronic and pressure-induced transient hypoxia. In dermatology, PRESS shows great promise for assessing tissue types and malignancy margins, especially with the widespread use of ALA for skin lesion treatments. Preliminary results from the first clinical trial highlight its potential in dermatological applications.

Tissue expansion is a common technique for acquiring enough tissue to reconstruct large defects; however, the problem of increased risk of flap necrosis in expanded flaps has not yet been adequately addressed. Indocyanine green angiography (ICGA) is a near-infrared laser activated, non-radioactive, real-time fluorescent imaging system that has allowed for the quantitative evaluation of flap perfusion. Nevertheless, a universally accepted threshold value for evaluating the viability of expanded flaps has not yet been established. Since our institution routinely treats patients with massive defects to be reconstructed with an expanded flap, we determined the optimal fluorescent cutoff value for intraoperative decision making to prevent postoperative necrosis.

We propose a smartphone-enabled multispectral microscope system for biopsy imaging and color-enhancement of stained slides. Experimental results on eight cancer-disease slides demonstrate the feasibility of enhancing color contrast in different tissue types. Leveraging smartphone accessibility, our approach promises low-cost biopsy diagnosis and improved medical services in resource-limited settings, potentially revolutionizing healthcare with efficient and affordable biopsy analysis using widely available smartphones.

Non-melanoma skin cancers (NMSC) pose challenges with current clinical approaches. In this context, Multispectral Optoacoustic Tomography (MSOT) offers a promising non-invasive imaging solution. With high isotropic resolution and contrast-enhanced capabilities, MSOT provides a 3D tumor map by resolving melanin and haemoglobin signals. Our feature extraction and automated level-set image segmentation algorithm enables tumor profiling and precise boundary delineation of width, depth, and volume. Validated against histology, these metrics aid preoperative tumor mapping and surgical planning as it fosters a comprehensive understanding of tumor morphology and metabolic activities. Thus, clinicians can optimize NMSC management, revolutionizing diagnostics and treatment for improved patient outcomes.

Natural Moisturizing Factor (NMF) development in newborns; identification of FLG loss-of-function mutation carriers by Raman spectroscopy In vivo Raman spectroscopy was used to identify newborns with a loss-of-function filaggrin gene mutation, who are at a much increased risk of developing atopic dermatitis. This enables targeted preventative treatment, which may significantly reduce this risk.

Scleroderma, or systemic sclerosis (SSc) is a chronic, autoimmune disorder that manifests in the fibrosis of skin and internal organs. The clinical gold standard used to track SSc disease progression is the modified Rodnan skin score (mRSS), which is based on clinical palpation. Spatial Frequency Domain Imaging (SFDI), a widefield, non-contact diffuse optical imaging technology may provide an alternate quantitative and objective method to track progression of SSc by measuring tissue reflectance and optical properties. To investigate this, SFDI measurements were conducted on 10 SSc patients and 8 healthy controls, with 6 patients and 2 controls also being measured longitudinally. We found that there were proportional changes in SFDI metrics (μs' at 851 nm and Rd at 851 nm and 0.2 mm-1) corresponding to SSc progression (measured by mRSS scores and with histopathological metrics), suggesting SFDI could provide an improved method to track SSc progression.

We have recently developed a novel, cavitation-based, highly selective anti-vascular technique, termed photo-mediated ultrasound therapy (PUT). In this study, the effectiveness and safety of PUT on cutaneous vascular malformation was examined through in vivo experiments in a clinically relevant chicken wattle model. The typical results showed perfusion stop of microvessel on OCT angiography and fade of the wattle color on skin imager after treatment. The safety is checked by H&E histology and immunohistochemistry evaluations include: CD31, Caspase-3, and Masson’s Trichrome (MTC) stains. The findings demonstrate that PUT can efficiently and safely remove hypervascular dermal capillaries by using laser fluence at a level which is orders of magnitude smaller than that used in conventional laser treatment of vascular lesions, thus offering a safer alternative technique for clinical management of cutaneous vascular malformations.

We introduce a “mini-phototest” concept, which reduces the skin area exposed to UV light by 20-folds for skin MED (minimal erythema dose) determination. This significantly minimizes cosmetic effects compared to conventional phototest. We validated this concept by Monte-Carlo simulation, and also studied cellular response induced by mini-size UV exposure using in vivo multimodality microscopy. This novel phototest method may enable site specific light dosimetry, facilitating personalized precision phototherapy.

Dermatological lasers are broadly classified as ablative or non-ablative, with tissue absorbance being a key consideration. The primary target in the Infrared (IR) spectrum is water, owing to its high absorption and its high concentration in tissues. Ablative lasers at 10 and 3μm, CO₂ and Erbium:YAG lasers, respectively, leverage this characteristic effectively. Conversely, non-ablative, lasers at 1.5-2μm primarily coagulate tissues without achieving the ablation threshold. Thulium lasers are positioned around a local peak in water absorption at 1.94μm, and exhibits approximately six times lower absorption than CO₂ (10.6μm) and significantly higher absorption than the 1.5μm wavelength. So far they were employed as more superficial non ablative lasers but did not achieve the ablation threshold. This study presents an innovative Tm:YAP laser at 1.94μm as an ablative laser. Employing high-energy, passively Q-switched pulses, at the nanosecond regime enables to reach the ablation threshold. The relatively high absorption characteristics at 1.94μm ensure the laser surpasses the lowered ablation threshold successfully. Experimental demonstrations on porcine skin using a fractional method showcased the creation of clean ablation micro columns. Micro columns with a remarkable thinness of up to 60μm was demonstrated. Deep column of up to 1.9mm was also observed. The 1.94μm Tm:YAP laser, as an innovative addition to the arsenal of ablative lasers, has the potential to revolutionize dermatological practices, providing a safe and reliable solution for skin treatments. Further refinement and development could open new avenues for enhancing patient care in dermatology.

Raman spectroscopy has been evaluated for skin cancer detection. Data augmentation has been used for image processing by deep neural networks. In this study, we proposed and evaluated different data augmentation strategies for spectral augmentation, including added random noise, spectral shift, spectral combination and artificially synthesized spectra using one-dimensional generative adversarial networks (1D-GAN). The stratified samples (n=731) were divided randomly into training (70%), validation (10%) and test dataset (20%), and were repeated 56 times in parallel computing. It was found that data augmentation is not only applicable to deep neural networks, but also applicable to conventional machine learning techniques. When all the strategies were combined to augment the training dataset, the performance of the test dataset could be improved by 2-71%.

In clinical dermatology, understanding collagen remodeling and reorganization are most important. Collagen provides the support matrix underpinning healthy skin and is key to preserving skin firmness and elasticity. Any issue with collagen deposition and deviation would affect the appearance in skin. In this regard, an ability to monitor the morphological changes in collagen organizations along with vascular morphology may improve our understanding in its pathophysiology and, ultimately, prevent these abnormal conditions. In this effort, we developed an advanced single input in house-built polarization-sensitive optical coherence tomography (PSOCT) device to directly visualize collagen organization within skin in vivo in various dermatological conditions like lipoatrophy, Necrobiosis lipoidica diabeticorum, and surgical scars to show the clinical utility of the system.

Accurate and repeatable epidermal thickness (ET) measurement is essential for assessing and quantifying cutaneous characteristics in dermatological disease, skin injury, and cosmetic research. Here, we investigated the integration of pressure sensors with optical coherence tomography (OCT) for improved epidermal thickness (ET) measurements. Traditional contact imaging techniques could induce variations in measurement due to inconsistent pressure. By using pressure sensors, real-time monitoring of applied pressure during OCT imaging provides stable and consistent ET mapping, improving repeatability. The result further demonstrates that increased compression forces can lead to significant deformation of the epidermis. This study demonstrates a pressure sensor integration to allow more accurate ET measurements and offers improved imaging quality, with potential applications in dermatology and cosmetics, underscoring the need for controlled pressure in contact imaging.

Melanoma is responsible for around 10,000 deaths annually in the US. While simple excision can cure most melanomas, some progress into metastatic cancer, necessitating advanced treatment options. However, the current clinical diagnosis methods are insufficient in accurately identifying metastasis. There is a need for better biomarkers for patients at risk of developing metastases to enable timely intervention and appropriate treatment. We present a biomarker based on femtosecond pump-probe microscopy and supervised learning techniques to diagnose metastatic melanoma. Pump-probe microscopy images of primary melanomas reveal the chemical and physical structure of melanin, a naturally occurring pigment in most melanoma tumors. Leveraging supervised learning models, we classify melanin features and utilize them to guide the diagnosis of metastatic disease. Our proposed biomarker is compatible with the current clinical protocol, as it only requires a slice of the primary tumor, which is routinely excised following clinical guidelines. In our preliminary dataset of approximately 50 patients, the biomarker demonstrates encouraging sensitivity and specificity, exceeding 80%.

Fast-speed volumetric imaging methods are desired for both multiphoton and confocal microscopy. A tilted-plane volumetric imaging and reconstruction method was developed to achieve fast volumetric imaging of freshly biopsied/excised human skin tissue based on only xy laser scanning. A volume that covers a surface area of 10 mm × 8 mm and a thickness of 80 um can be acquired in 9 minutes. The large surface area was achieved by stitching multiple volume strips and each volume strip was reconstructed from a sequence of frames acquired along the tissue surface at a frame rate of 198 frames per second and each frame was captured in an imaging plane that is tilted relative to the tissue surface. The imaging method was implemented on a multimodality imaging system to generate confocal reflectance, second harmonic, and two-photon fluorescence volumes simultaneously and give complementary information of the excised tissue. Application in skin cancer surgery guidance will be pursued in the future.

Nonmelanoma skin cancer is the most common malignancy in the US, and while Mohs microsurgery is curative in the majority of cases, inaccurate presurgical visual estimation of tumor margins leads to the need for more than one stage in 30% of cases. Dual wavelength optical polarization imaging (OPI) provides accurate presurgical delineation of tumor margins, but existing OPI devices are not easily translated to the dermatology clinical setting. We found that the clinically ubiquitous dermatascope can be repurposed as a handheld OPI system that has functionality on par with existing OPI devices at a fraction of the cost.

New advanced imaging technologies can improve the accuracy of skin cancer diagnosis, reducing the risk of misdiagnosis and unnecessary excision. We explore reflectance confocal microscopy (RCM) and photoacoustic imaging (PAI) for fast, precise skin cancer diagnosis at the bedside. 75 patients with suspicious pigmented skin lesions were scanned with RCM and PAI prior to excision. Dermoscopy images were obtained prior to scanning. Morphological features and blood vessel characteristics were analysed and diagnostic accuracy assessed. Tumour thickness, blood flow, collagen, lipid and melanin content were measured by PAI. New diagnostic hypotheseses, combining PAI, RCM and dermoscopy, will be presented. The combination of PAI and RCM offers high-resolution and deep penetration visualization of skin structures, facilitating accurate diagnosis and assessment of suspicious skin tumors at first patient visit. This approach can potentially reduce invasive procedures and the risk of misdiagnosis.

In this study, we developed an improved piston-based specimen holder to provide even pressure distribution across an irregular tissue surface. A series of support fixtures are also developed to facilitate the pressure distribution from the piston to image specimens with small contact area relative to thickness such as bisected shave skin biopsies. Using this capability, we demonstrate imaging of tall and narrow biopsy specimens with precise coregistration to conventional histology as well as rapid imaging of Mohs margins during surgery.

We present the first laser speckle imaging device for skin cancer detection that can image and distinguish shallow and deep vessels from speckle frames acquired with a single exposure time. To do this, we have developed a new signal processing technique based on the simultaneous evaluation of two metrics: one based on contrast, and one based on a normalized version of the second order autocorrelation function, to reveal deep and shallow vessels, respectively. We will present data from the pilot studies we are conducting on healthy volunteers and skin cancer patients at MGH skin cancer clinic, in which we will compare the microvasculature images obtained with our device from different skin lesions.

Both new and established technologies are explored in the pursuit of finding an efficient approach for diagnosing malignant melanoma skin cancer. The non-invasive sub-surface imaging technique of optical coherence tomography (OCT) is frequently applied in the ophthalmological clinic, but is still emerging within the field of dermatology. We present our work in testing two supercontinuum OCT systems for characterizing tape strips applied to skin cancer patients. Tape strips were collected in a clinical trial of 75 skin cancer patients at Bispebjerg Hospital, Denmark. After low-temperature storage, samples were scanned by a near-infrared OCT system and a mid-infrared OCT system of wavelengths 1.3 µm and 4 µm. We report on the scanning protocol and how the wavelength dependent OCT scans can be interpreted in order to target malignant melanoma characteristics.

Cutaneous neurofibromas (cNF) appear commonly in neurofibromatosis type I (NF1) patients. cNFs usually appear in the skin at puberty and proliferate with increasing numbers and sizes throughout life. If cNFs can be detected and treated in their nascent stage, patient’s quality of life can be improved. To detect early-stage cNF, we have employed spatial frequency domain imaging (SFDI) and optical coherence tomography (OCT). SFDI has been applied to screen large skin areas on eleven cNF subjects. Suspect lesions invisible to the unaided eye but detected using SFDI were imaged with OCT to observe lesion microstructure. Three lesions were biopsied to compare with SFDI and OCT images. Suspect nascent cNFs which are invisible to the unaided eye were detected as low optical scattering regions in all patients. Large area screening using SFDI confirmed scattering contrast between the suspect nascent cNF and the surrounding uninvolved skin. Abnormal disc-shaped structures with reduced scattering regions detected by SFDI were also observed in OCT cross-sections.