Confocal Laser Endomicroscopy in Resection of Sinonasal Malignant Melanoma-Preliminary Report on Real-Time Margin Assessment and Support in Surgical Decision-Making.

Background/Objectives: Building upon the rising value of Confocal Laser Endomicroscopy (CLE) in squamous cell carcinoma of the head and neck, we present the first application of CLE during the resection of sinonasal malignant melanomas. This study aims to evaluate the potential of CLE to assist surgeons in intraoperative decision-making, with a particular focus on resection margin assessment within the constrained nasal cavity. Methods: Two cases of sinonasal malignant melanoma were included in this study. CLE was employed to examine visible tumors and their margins, both pre- and post-endoscopic resection. The findings were compared to histopathological results as well as data on squamous cell carcinoma, for which malignancy criteria had already been established in prior projects. Results: CLE provided the real-time visualization of sinonasal malignant melanomas and their margins, successfully differentiating between healthy and neoplastic tissue compared to histopathological findings. Conclusion: CLE offers the potential for real-time assessment, aiding surgeons in more precise tumor resection and potentially improving patient outcomes. This study demonstrates the feasibility of using CLE in the resection of sinonasal malignant melanoma, highlighting its ability to differentiate between healthy and neoplastic tissue intraoperatively.

Imaging Biomarkers of Oral Dysplasia and Carcinoma Measured with In Vivo Endoscopic Optical Coherence Tomography.

Optical coherence tomography is a noninvasive imaging technique that provides three-dimensional visualization of subsurface tissue structures. OCT has been proposed and explored in the literature as a tool to assess oral cancer status, select biopsy sites, or identify surgical margins. Our endoscopic OCT device can generate widefield (centimeters long) imaging of lesions at any location in the oral cavity-but it is challenging for raters to quantitatively assess and score large volumes of data. Leveraging a previously developed epithelial segmentation network, this work develops quantifiable biomarkers that provide direct measurements of tissue properties in three dimensions. We hypothesize that features related to morphology, tissue attenuation, and contrast between tissue layers will be able to provide a quantitative assessment of disease status (dysplasia through carcinoma). This work retrospectively assesses seven biomarkers on a lesion-contralateral matched OCT dataset of the lateral and ventral tongue (40 patients, 70 sites). Epithelial depth and loss of epithelial-stromal boundary visualization provide the strongest discrimination between disease states. The stroma optical attenuation coefficient provides a distinction between benign lesions from dysplasia and carcinoma. The stratification biomarkers visualize subsurface changes, which provides potential for future utility in biopsy site selection or treatment margin delineation.

Raman spectroscopy in lung cancer diagnostics: Can an in vivo setup compete with ex vivo applications?

Lung carcinoma remains the leading cause of cancer death worldwide. The tactic to change this unfortunate rate may be a timely and rapid diagnostic, which may in many cases improve patient prognosis. In our study, we focus on the comparison of two novel methods of rapid lung carcinoma diagnostics, label-free in vivo and ex vivo Raman spectroscopy of the epithelial tissue, and assess their feasibility in clinical practice. As these techniques are sensitive not only to the basic molecular composition of the analyzed sample but also to the secondary structure of large biomolecules, such as tissue proteins, they represent suitable candidate methods for epithelial cancer diagnostics. During routine bronchoscopy, we collected 78 in vivo Raman spectra of normal and cancerous lung tissue and 37 samples of endobronchial pathologies, which were subsequently analyzed ex vivo. Using machine learning techniques, namely principal component analysis (PCA) and support vector machines (SVM), we were able to reach 87.2% (95% CI, 79.8-94.6%) and 100.0% (95% CI, 92.1-100.0%) of diagnostic accuracy for in vivo and ex vivo setup, respectively. Although the ex vivo approach provided superior results, the rapidity of in vivo Raman spectroscopy might become unmatchable in the acceleration of the diagnostic process.

Hyperspectral dark-field microscopy of human breast lumpectomy samples for tumor margin detection in breast-conserving surgery.

Significance: Hyperspectral dark-field microscopy (HSDFM) and data cube analysis algorithms demonstrate successful detection and classification of various tissue types, including carcinoma regions in human post-lumpectomy breast tissues excised during breast-conserving surgeries. Aim: We expand the application of HSDFM to the classification of tissue types and tumor subtypes in pre-histopathology human breast lumpectomy samples. Approach: Breast tissues excised during breast-conserving surgeries were imaged by the HSDFM and analyzed. The performance of the HSDFM is evaluated by comparing the backscattering intensity spectra of polystyrene microbead solutions with the Monte Carlo simulation of the experimental data. For classification algorithms, two analysis approaches, a supervised technique based on the spectral angle mapper (SAM) algorithm and an unsupervised technique based on the K-means algorithm are applied to classify various tissue types including carcinoma subtypes. In the supervised technique, the SAM algorithm with manually extracted endmembers guided by H&E annotations is used as reference spectra, allowing for segmentation maps with classified tissue types including carcinoma subtypes. Results: The manually extracted endmembers of known tissue types and their corresponding threshold spectral correlation angles for classification make a good reference library that validates endmembers computed by the unsupervised K-means algorithm. The unsupervised K-means algorithm, with no a priori information, produces abundance maps with dominant endmembers of various tissue types, including carcinoma subtypes of invasive ductal carcinoma and invasive mucinous carcinoma. The two carcinomas' unique endmembers produced by the two methods agree with each other within <2% residual error margin. Conclusions: Our report demonstrates a robust procedure for the validation of an unsupervised algorithm with the essential set of parameters based on the ground truth, histopathological information. We have demonstrated that a trained library of the histopathology-guided endmembers and associated threshold spectral correlation angles computed against well-defined reference data cubes serve such parameters. Two classification algorithms, supervised and unsupervised algorithms, are employed to identify regions with carcinoma subtypes of invasive ductal carcinoma and invasive mucinous carcinoma present in the tissues. The two carcinomas' unique endmembers used by the two methods agree to <2% residual error margin. This library of high quality and collected under an environment with no ambient background may be instrumental to develop or validate more advanced unsupervised data cube analysis algorithms, such as effective neural networks for efficient subtype classification.

Intrathoracic non-tuberculous mycobacteriosis with endobronchial lesion in a child aged 11 with HIV infection diagnosed by bronchoscopic biopsy, EBUS-TBNA and confocal laser endomicroscopy.

The diagnosis of intrathoracic non-tuberculous mycobacteriosis (NTM) is challenging. We report a case of a pediatric pulmonary NTM with endobronchial lesion and lymphadenitis in a child with HIV infection diagnosed by bronchoscopic biopsy, EBUS-TBNA and probe-based confocal laser endomicroscopy (pCLE). The pCLE showed a large number of highly fluorescent cells and zones of density and disorganized elastin fibers at alveolar areas. A combination of diagnostic endoscopic procedures is required to establish the diagnosis of NTM.

Discontinuity third harmonic generation microscopy for label-free imaging and quantification of intraepidermal nerve fibers.

Label-free imaging methodologies for nerve fibers rely on spatial signal continuity to identify fibers and fail to image free intraepidermal nerve endings (FINEs). Here, we present an imaging methodology-called discontinuity third harmonic generation (THG) microscopy (dTHGM)-that detects three-dimensional discontinuities in THG signals as the contrast. We describe the mechanism and design of dTHGM and apply it to reveal the bead-string characteristics of unmyelinated FINEs. We confirmed the label-free capability of dTHGM through a comparison study with the PGP9.5 immunohistochemical staining slides and a longitudinal spared nerve injury study. An intraepidermal nerve fiber (IENF) index based on a discontinuous-dot-connecting algorithm was developed to facilitate clinical applications of dTHGM. A preliminary clinical study confirmed that the IENF index was highly correlated with skin-biopsy-based IENF density (Pearson's correlation coefficient R = 0.98) and could achieve differential identification of small-fiber neuropathy (p = 0.0102) in patients with diabetic peripheral neuropathy.

Detection of NADH and NADPH levels in vivo identifies shift of glucose metabolism in cancer to energy production.

Profound changes in the metabolism of cancer cells have been known for almost 100 years, and many aspects of these changes have continued to be actively studied and discussed. Differences in the results of various studies can be explained by the diversity of tumours, which have differing processes of energy metabolism, and by limitations in the methods used. Here, using fluorescence lifetime needle optical biopsy in a hepatocellular carcinoma (HCC) mouse model and patients with HCC, we measured reduced nicotinamide adenine dinucleotide (NADH) and reduced nicotinamide adenine dinucleotide phosphate (NADPH) in control liver, and in HCC tumours and their adjacent regions. We found that NADH level (mostly responsible for energy metabolism) is increased in tumours but also in adjacent regions of the same liver. NADPH level is significantly decreased in the tumours of patients but increased in the HCC mouse model. However, in the ex vivo tumour slices of mouse HCC, reactive oxygen species production and glutathione level (both dependent on NADPH) were significantly suppressed. Thus, glucose-dependent NADH and NADPH production in tumours changed but with a more pronounced shift to energy production (NADH), rather than NADPH synthesis for redox balance.

Optical Biopsies Using Probe-Based Confocal Laser Endomicroscopy for Autoimmune Pulmonary Alveolar Proteinosis.

INTRODUCTION: Increasing numbers of cases of mild asymptomatic pulmonary alveolar proteinosis (PAP) are being reported with the recent increase in chest computed tomography (CT). Bronchoscopic diagnosis of mild PAP is challenging because of the patchy distribution of lesions, which makes it difficult to obtain sufficient biopsy samples. Additionally, the pathological findings of mild PAP, particularly those that differ from severe PAP, have not been fully elucidated. This study aimed to clarify the pathological findings of mild PAP and the usefulness of optical biopsy using probe-based confocal laser endomicroscopy (pCLE). METHODS: We performed bronchoscopic optical biopsy using pCLE and tissue biopsy in 5 consecutive patients with PAP (three with mild PAP and two with severe PAP). We compared the pCLE images of mild PAP with those of severe PAP by integrating clinical findings, tissue pathology, and chest CT images. RESULTS: pCLE images of PAP showed giant cells with strong fluorescence, amorphous substances, and thin alveolar walls. Images of affected lesions in mild PAP were equivalent to those obtained in arbitrary lung lesions in severe cases. All 3 patients with mild PAP spontaneously improved or remained stable after ≥3 years of follow-up. Serum autoantibodies to granulocyte-macrophage colony-stimulating factor were detected in all 5 cases. CONCLUSION: Optical biopsy using pCLE can yield specific diagnostic findings, even in patients with mild PAP. pCLE images of affected areas in mild and severe PAP showed similar findings, indicating that the dysfunction level of pathogenic alveolar macrophages in affected areas is similar between both disease intensities.

Femtosecond optical Kerr effect in normal and grades of cancerous breast tissues as a new optical biopsy method.

This study reports on the first use of the optical Kerr effect (OKE) in breast cancer tissue. This proposed optical biopsy method utilizes a Femtosecond Optical Kerr Gate to detect changes in dielectric relaxation and conductivity created by a cancerous infection. Here, the temporal behavior of the OKE is tracked in normal and cancerous samples of human and mouse breast. These tissues display a double peaked temporal structure and its decay rate changes depending on the tissue's infection status. The decay of the secondary peak, attributed to ultrafast plasma response, indicates that the tissue's conductivity has doubled once infected. A slower molecular contribution to the Kerr effect can also be observed in healthy tissues. These findings suggest two possible biomarkers for the use of OKE in optical biopsy. Both markers arise from alterations in the infected tissue's cellular structure, which changes the rate at which electronic and molecular processes occur.

Multispectral Raman Differentiation of Malignant Skin Neoplasms In Vitro: Search for Specific Biomarkers and Optimal Wavelengths.

Confocal scanning Raman and photoluminescence (PL) microspectroscopy is a structure-sensitive optical method that allows the non-invasive analysis of biomarkers in the skin tissue. We used it to perform in vitro diagnostics of different malignant skin neoplasms at several excitation wavelengths (532, 785 and 1064 nm). Distinct spectral differences were noticed in the Raman spectra of basal cell carcinoma (BCC) and squamous cell carcinoma (SCC), compared with healthy skin. Our analysis of Raman/PL spectra at the different excitation wavelengths enabled us to propose two novel wavelength-independent spectral criteria (intensity ratios for 1302 cm-1 and 1445 cm-1 bands, 1745 cm-1 and 1445 cm-1 bands), related to the different vibrational "fingerprints" of cell membrane lipids as biomarkers, which was confirmed by the multivariate curve resolution (MCR) technique. These criteria allowed us to differentiate healthy skin from BCC and SCC with sensitivity and specificity higher than 95%, demonstrating high clinical importance in the differential diagnostics of skin tumors.

Articular Cartilage-From Basic Science Structural Imaging to Non-Invasive Clinical Quantitative Molecular Functional Information for AI Classification and Prediction.

This review presents the changes that the imaging of articular cartilage has undergone throughout the last decades. It highlights that the expectation is no longer to image the structure and associated functions of articular cartilage but, instead, to devise methods for generating non-invasive, function-depicting images with quantitative information that is useful for detecting the early, pre-clinical stage of diseases such as primary or post-traumatic osteoarthritis (OA/PTOA). In this context, this review summarizes (a) the structure and function of articular cartilage as a molecular imaging target, (b) quantitative MRI for non-invasive assessment of articular cartilage composition, microstructure, and function with the current state of medical diagnostic imaging, (c), non-destructive imaging methods, (c) non-destructive quantitative articular cartilage live-imaging methods, (d) artificial intelligence (AI) classification of degeneration and prediction of OA progression, and (e) our contribution to this field, which is an AI-supported, non-destructive quantitative optical biopsy for early disease detection that operates on a digital tissue architectural fingerprint. Collectively, this review shows that articular cartilage imaging has undergone profound changes in the purpose and expectations for which cartilage imaging is used; the image is becoming an AI-usable biomarker with non-invasive quantitative functional information. This may aid in the development of translational diagnostic applications and preventive or early therapeutic interventions that are yet beyond our reach.

Femtosecond Optical Kerr Gate in tissues.

The Optical Kerr Effect is investigated for the first time in biological tissues. This nonlinear effect was explored in both human brain and avian breast tissues using a time-resolved femtosecond pump-probe Optical Kerr Gate that looks for phase changes that arise in the probe from the pump induced Kerr refractive index change. The tissue samples produced a unique ultrafast (700-800 fs) doubled peaked temporal signal, which is indicative of interplay between the different ultrafast mechanisms (electronic plasma and molecular) that make up the Kerr index. The unique profile was replicated in theoretical simulations. The properties of the temporal profile varied between samples suggesting that it could be used as a new diagnostic. Understanding this behavior can help improve the scientific understanding of nonlinear spectral diagnostic techniques and potentially create a new Kerr-based optical biopsy method.

Deconvolution of the fluorescence spectra measured through a needle probe to assess the functional state of the liver.

OBJECTIVES: Currently, one of the most pressing issues for surgeons in the treatment of obstructive jaundice is the ability to assess the functional state of the liver and to detect and determine the degree of liver failure in a timely manner with simple and objective techniques. In this regard, the use of fluorescence spectroscopy method can be considered as one of the ways to improve the informativity of existing diagnostic algorithms in clinical practice and to introduce new diagnostic tools. Thus, the aim of the work was to study in vivo the functional state of liver parenchyma by the method of fluorescence spectroscopy implemented through a needle probe and assess the contribution of the main tissue fluorophores to reveal new diagnostic criteria. MATERIALS AND METHODS: We compared data from 20 patients diagnosed with obstructive jaundice and 11 patients without this syndrome. Measurements were performed using a fluorescence spectroscopy method at excitation wavelengths of 365 and 450 nm. Data were collected using a 1 mm fiber optic needle probe. The analysis was based on the comparison of the results of deconvolution with the combinations of Gaussian curves reflecting the contribution of the pure fluorophores in the liver tissues. RESULTS: The results showed a statistically significant increase in the contribution of curves reflecting NAD(P)H fluorescence, bilirubin, and flavins in the group of patients with obstructive jaundice. This and the calculated redox ratio values indicated that the energy metabolism of the hepatocytes may have shifted to glycolysis due to hypoxia. An increase in vitamin A fluorescence was also observed. It may also serve as a marker of liver damage, indicating impaired vitamin A mobilization from the liver due to cholestasis. CONCLUSIONS: The results obtained reflect changes associated with shifts in the content of the main fluorophores characterizing hepatocyte dysfunction caused by accumulation of bilirubin and bile acids and after disturbance of oxygen utilization. The contributions of NAD(P)H, flavins, and bilirubin as well as vitamin A can be used for further studies as promising diagnostic and prognostic markers for the course of liver failure. Further work will include collecting fluorescence spectroscopy data in patients with different clinical effects of obstructive jaundice on postoperative clinical outcome after biliary decompression.

Design and test of a rigid endomicroscopic system for multimodal imaging and femtosecond laser ablation.

Significance: Conventional diagnosis of laryngeal cancer is normally made by a combination of endoscopic examination, a subsequent biopsy, and histopathology, but this requires several days and unnecessary biopsies can increase pathologist workload. Nonlinear imaging implemented through endoscopy can shorten this diagnosis time, and localize the margin of the cancerous area with high resolution. Aim: Develop a rigid endomicroscope for the head and neck region, aiming for in-vivo multimodal imaging with a large field of view (FOV) and tissue ablation. Approach: Three nonlinear imaging modalities, which are coherent anti-Stokes Raman scattering, two-photon excitation fluorescence, and second harmonic generation, as well as the single photon fluorescence of indocyanine green, are applied for multimodal endomicroscopic imaging. High-energy femtosecond laser pulses are transmitted for tissue ablation. Results: This endomicroscopic system consists of two major parts, one is the rigid endomicroscopic tube 250 mm in length and 6 mm in diameter, and the other is the scan-head (10×12×6 cm3 in size) for quasi-static scanning imaging. The final multimodal image accomplishes a maximum FOV up to 650 µm, and a resolution of 1 µm is achieved over 560 µm FOV. The optics can easily guide sub-picosecond pulses for ablation. Conclusions: The system exhibits large potential for helping real-time tissue diagnosis in surgery, by providing histological tissue information with a large FOV and high resolution, label-free. By guiding high-energy fs laser pulses, the system is even able to remove suspicious tissue areas, as has been shown for thin tissue sections in this study.

Application of Indocyanine Green Fluorescence Imaging in Assisting Biopsy of Musculoskeletal Tumors.

(1) Background: Biopsies are the gold standard for the diagnosis of musculoskeletal tumors. In this study, we aimed to explore whether indocyanine green near-infrared fluorescence imaging can assist in the biopsy of bone and soft tissue tumors and improve the success rate of biopsy. (2) Method: We recruited patients with clinically considered bone and soft tissue tumors and planned biopsies. In the test group, indocyanine green (0.3 mg/kg) was injected. After identifying the lesion, a near-infrared fluorescence camera system was used to verify the ex vivo specimens of the biopsy in real time. If the biopsy specimens were not developed, we assumed that we failed to acquire lesions, so the needle track and needle position were adjusted for the supplementary biopsy, and then real-time imaging was performed again. Finally, we conducted a pathological examination. In the control group, normal biopsy was performed. (3) Results: The total diagnosis rate of musculoskeletal tumors in the test group was 94.92% (56/59) and that in the control group was 82.36% (42/51). In the test group, 14 cases were not developed, as seen from real-time fluorescence in the core biopsy, and then underwent the supplementary biopsy after changing the puncture direction and the location of the needle channel immediately, of which 7 cases showed new fluorescence. (4) Conclusions: Using the near-infrared fluorescence real-time development technique to assist the biopsy of musculoskeletal tumors may improve the accuracy of core biopsy and help to avoid missed diagnoses, especially for some selected tumors.

Dynamic real-time optical microscopy of oral mucosal lesions using confocal laser endomicroscopy.

OBJECTIVES: Confocal laser endomicroscopy (CLE) is a novel non-invasive point-of-care optical biopsy technology that enables real-time in vivo microscopic visualisation of cellular and tissue architecture. In this study, we assessed the diagnostic accuracy of a hand-held fluorescence single-fibre distal-scanning CLE (fsdCLE) platform for diagnosing oral epithelial dysplasia (OED) and oral squamous cell carcinoma (OSCC). MATERIALS AND METHODS: Forty-seven patients presenting with 63 distinct oral mucosal lesions were subjected to optical biopsy using a miniaturised fsdCLE system (ViewnVivo®, Optiscan Imaging Ltd) and topical exogenous acriflavine hydrochloride contrast agent before undergoing tissue biopsy and histopathological consensus review by four pathologists. CLE images were captured in vivo in real-time during clinical examination and assessed on-the-fly for the presence of cellular and architectural features of OED/OSCC offering an instantaneous diagnosis. Predicted optical diagnoses were compared to definitive consensus tissue histopathology. Sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV) and accuracy were calculated for the presence/absence of dysplasia/malignancy on optical biopsy. Percentage agreement, Fleiss' kappa, and intraclass correlation coefficient (ICC) were calculated for each assessment stage during the consensus histopathology process. RESULTS: Diagnostic accuracy was extremely high at 88.9%. Other metrics were sensitivity 86.8%, specificity 92%, PPV 94.3% and NPV 82.1%. One hundred percent of carcinoma cases were detected accurately using CLE in the clinic. CONCLUSION: fsdCLE is a highly accurate, easy-to-use, rapid and slide-free point-of-care in vivo optical technology for diagnosing OED/OSCC and discriminating between dysplastic and non-dysplastic pathology. It demonstrates near-perfect agreement with traditional consensus histopathology without the need for physical tissue biopsy.

Fatty Acids and Bilirubin as Intrinsic Autofluorescence Serum Biomarkers of Drug Action in a Rat Model of Liver Ischemia and Reperfusion.

The autofluorescence of specific fatty acids, retinoids, and bilirubin in crude serum can reflect changes in liver functional engagement in maintaining systemic metabolic homeostasis. The role of these fluorophores as intrinsic biomarkers of pharmacological actions has been investigated here in rats administered with obeticholic acid (OCA), a Farnesoid-X Receptor (FXR) agonist, proven to counteract the increase of serum bilirubin in hepatic ischemia/reperfusion (I/R) injury. Fluorescence spectroscopy has been applied to an assay serum collected from rats submitted to liver I/R (60/60 min ± OCA administration). The I/R group showed changes in the amplitude and profiles of emission spectra excited at 310 or 366 nm, indicating remarkable alterations in the retinoid and fluorescing fatty acid balance, with a particular increase in arachidonic acid. The I/R group also showed an increase in bilirubin AF, detected in the excitation spectra recorded at 570 nm. OCA greatly reversed the effects observed in the I/R group, confirmed by the biochemical analysis of bilirubin and fatty acids. These results are consistent with a relationship between OCA anti-inflammatory effects and the acknowledged roles of fatty acids as precursors of signaling agents mediating damaging responses to harmful stimuli, supporting serum autofluorescence analysis as a possible direct, real-time, cost-effective tool for pharmacological investigations.

Towards targeted colorectal cancer biopsy based on tissue morphology assessment by compression optical coherence elastography.

Identifying the precise topography of cancer for targeted biopsy in colonoscopic examination is a challenge in current diagnostic practice. For the first time we demonstrate the use of compression optical coherence elastography (C-OCE) technology as a new functional OCT modality for differentiating between cancerous and non-cancerous tissues in colon and detecting their morphological features on the basis of measurement of tissue elastic properties. The method uses pre-determined stiffness values (Young's modulus) to distinguish between different morphological structures of normal (mucosa and submucosa), benign tumor (adenoma) and malignant tumor tissue (including cancer cells, gland-like structures, cribriform gland-like structures, stromal fibers, extracellular mucin). After analyzing in excess of fifty tissue samples, a threshold stiffness value of 520 kPa was suggested above which areas of colorectal cancer were detected invariably. A high Pearson correlation (r =0.98; p <0.05), and a negligible bias (0.22) by good agreement of the segmentation results of C-OCE and histological (reference standard) images was demonstrated, indicating the efficiency of C-OCE to identify the precise localization of colorectal cancer and the possibility to perform targeted biopsy. Furthermore, we demonstrated the ability of C-OCE to differentiate morphological subtypes of colorectal cancer - low-grade and high-grade colorectal adenocarcinomas, mucinous adenocarcinoma, and cribriform patterns. The obtained ex vivo results highlight prospects of C-OCE for high-level colon malignancy detection. The future endoscopic use of C-OCE will allow targeted biopsy sampling and simultaneous rapid analysis of the heterogeneous morphology of colon tumors.

Microendoscopy in vivo for the pathological diagnosis of cervical precancerous lesions and early cervical cancer.

BACKGROUND: Cervical cancer is an important public health problem. Conventional colposcopy is inefficient in the diagnosis of cervical lesions and massive biopsies result in trauma. There is an urgent need for a new clinical strategy to triage women with abnormal cervical screening results immediately and effectively. In this study, the high-resolution microendoscopy combined with methylene blue cell staining technology was used to perform real-time in vivo imaging of the cervix for the first time. METHODS: A total of 41 patients were enrolled in the study. All patients underwent routine colposcopy and cervical biopsy, and high-resolution images of methylene blue-stained cervical lesions were obtained in vivo using microendoscopy. The cell morphological features of benign and neoplastic cervical lesions stained with methylene blue under microendoscopy were analyzed and summarized. The microendoscopy and histopathology findings of the high-grade squamous intraepithelial lesion (HSIL) and more severe lesions were compared. RESULTS: The overall consistency of microendoscopy diagnosis with pathological diagnosis was 95.12% (39/41). Diagnostic cell morphological features of cervicitis, low-grade squamous intraepithelial lesion (LSIL), HSIL, adenocarcinoma in situ, and invasive cancer were clearly demonstrated in methylene blue stained microendoscopic images. In HSIL and more severe lesions, microendoscopic methylene blue cell staining technology can show the microscopic diagnostic features consistent with histopathology. CONCLUSIONS: This study was an initial exercise in the application of the microendoscopy imaging system combined with methylene blue cell staining technology to cervical precancerous lesions and cervical cancer screening. The results provided the basis for a novel clinical strategy for triage of women with abnormal cervical screening results using in vivo non-invasive optical diagnosis technology.

A Clinical Study to Assess Diffuse Reflectance Spectroscopy with an Auto-Calibrated, Pressure-Sensing Optical Probe in Head and Neck Cancer.

Diffuse reflectance spectroscopy (DRS) is a powerful tool for quantifying optical and physiological tissue properties such as hemoglobin oxygen saturation and vascularity. DRS is increasingly used clinically for distinguishing cancerous lesions from normal tissue. However, its widespread clinical acceptance is still limited due to uncontrolled probe-tissue interface pressure that influences reproducibility and introduces operator-dependent results. In this clinical study, we assessed and validated a pressure-sensing and automatic self-calibration DRS in patients with suspected head and neck squamous cell carcinoma (HNSCC). The clinical study enrolled nineteen patients undergoing HNSCC surgical biopsy procedures. Patients consented to evaluation of this improved DRS system during surgery. For each patient, we obtained 10 repeated measurements on one tumor site and one distant normal location. Using a Monte Carlo-based model, we extracted the hemoglobin saturation data along with total hemoglobin content and scattering properties. A total of twelve cancer tissue samples from HNSCC patients and fourteen normal tissues were analyzed. A linear mixed effects model tested for significance between repeated measurements and compared tumor versus normal tissue. These results demonstrate that cancerous tissues have a significantly lower hemoglobin saturation compared to normal controls (p < 0.001), which may be reflective of tumor hypoxia. In addition, there were minimal changes over time upon probe placement and repeated measurement, indicating that the pressure-induced changes were minimal and repeated measurements did not differ significantly from the initial value. This study demonstrates the feasibility of conducting optical spectroscopy measurements on intact lesions prior to removal during HNSCC procedures, and established that this probe provides diagnostically-relevant physiologic information that may impact further treatment.