[Possibilities of multiphoton microscopy for the diagnosis of the vulvar lichen sclerosus]. / Vozmozhnosti mul'tifotonnoi mikroskopii dlya diagnostiki skleroticheskogo likhena vul'vy.

BACKGROUND: Vulvar lichen sclerosus (VLS) is a chronic and recurrent dermatosis of an inflammatory nature with severe focal atrophy of the skin. Connective tissue changes are polymorphic and are still not taken into account in histological diagnostics due to the difficulty of interpreting routine histological methods. In this work, we use multiphoton microscopy (MPM) as a new imaging technique that provides detailed information about the organization of collagen fibers in the dermis based on a non-linear second harmonic generation (SHG) process. OBJECTIVE: To determine the degree of connective tissue damage in lichen sclerosus using standard histological techniques and to reveal the diagnostic capabilities of multiphoton microscopy. MATERIAL AND METHODS: We studied 42 biopsies with a histopathological diagnosis of VLS and 10 biopsies of normal vulvar skin. Histological, histochemical and immunohistochemical evaluation was used in comparison with MPM data. Quantitative analysis included the determination of the thickness, length of collagen fibers and the average intensity of the SHG signal. RESULTS: A comprehensive study of the skin showed 4 groups of changes that can be regarded as the degree of the dermis damage: initial, mild, moderate, severe. The affected area at the initial and mild degree has subtle changes, however, it is reliably identified by quantitative analysis of the SHG signal. So, the initial degree is characterized by thin (1.3-1.8 µm) long (56-69 µm) collagen fibers, with a moderate degree, the fibers are thickened (3.4-4.3 µm) and fragmented (22-37 µm). The affected area in moderate and severe cases undergoes homogenization, which is associated with the deposition of extremely thin (0.6-0.9 µm) short (16-28 µm) collagen fibers and the expression of type V collagen. CONCLUSION: Multiphoton microscopy in the second harmonic generation mode is a reliable method for identifying collagen fibers in tissues. The study made it possible to identify 4 degrees of the dermis damage in vulvar lichen sclerosus.

Cross-Polarization Optical Coherence Tomography for Clinical Evaluation of Dermal Lesion Degrees in Vulvar Lichen Sclerosus.

The aim of the study was to identify different degrees of dermal lesions in vulvar lichen sclerosus (VLS) using cross-polarization optical coherence tomography (CP OCT) based on attenuation coefficient to detect disease early manifestations and to monitor the effectiveness of treatment. Materials and Methods: The study included 10 patients without pathology and 39 patients with VLS diagnosed histologically. CP OCT was performed in vivo on the inner surface of the labia minora, in the main lesion area. From each scanning point, a 3.4×3.4×1.25-mm3 3D data array was obtained in 26 s. CP OCT examination results were compared with histological examination of specimens stained with Van Gieson's picrofuchsin.Quantitative analysis of OCT images was performed by measuring the attenuation coefficient in co-polarization and cross-polarization. For visual analysis, color-coded charts were developed based on OCT attenuation coefficients. Results: According to histological examination, all patients with VLS were divided into 4 groups as per dermal lesion degree: initial (8 patients); mild (7 patients); moderate (9 patients); severe (15 patients). Typical features of different degrees were interfibrillary edema up to 250 µm deep for initial degree, thickened collagen bundles without edema up to 350 µm deep for mild degree, dermis homogenization up to 700 µm deep for moderate degree, dermis homogenization and total edema up to 1200 µm deep for severe degree.Pathological processes in dermis during VLS like interfibrillary edema and collagen bundles homogenization were visualized using CP OCT method based on values of attenuation coefficient in co- and cross-polarization channels. However, CP OCT method appeared to be less sensitive to changes of collagen bundles thickness not allowing to distinguish thickened collagen bundles from normal ones with enough statistical significance. The CP OCT method was able to differentiate all degrees of dermal lesions among themselves. OCT attenuation coefficients differed from normal condition with statistical significance for all degrees of lesions, except for mild. Conclusion: For the first time, quantitative parameters for each degrees of dermis lesion in VLS, including initial degree, were determined by CP OCT method allowing to detect the disease at an early stage and to monitor the applied clinical treatment effectiveness.

Massive perivillous fibrin deposition of the placenta and pregnancy outcome: A retrospective observational study.

INTRODUCTION: Massive perivillous fibrin deposition (MPVFD) in the placenta is associated with pregnancy complications and maternal disease. The aim of the current study was to contribute with increased knowledge regarding MPVFD by comparing maternal characteristics, obstetric and perinatal outcome and recurrence rate according to the degree of MPVFD. MATERIAL AND METHODS: This retrospective observational study included 141 cases of MPVFD collected between January 2003 to December 2018 in the Stockholm region, Sweden. The extent of fibrin deposition was assessed as low (20-32%), moderate (33-50%) or severe (>50%) according to macroscopic examination. RESULTS: The study covered 48 placentas with low MPVFD, 41 with moderate and 52 with severe MPVFD. Severe MPVFD was associated with more prematurity than moderate and low MPVFDs (56.3% vs 34.2% and 34.0% respectively, p = 0.05 and p = 0.04). In cases with severe MPVFD, 72.3% of the liveborn infants were growth-restricted compared to 34.2% in the moderate group (p = 0.001) and 52.2% in the low group (p = 0.06). The incidence of intrauterine fetal death was 31.3% in the severe MPVFD group, which was significantly higher than in the low MPVFD group (8.5%, p = 0.01) and twice as much as in cases with moderate MPVFD (15.8%, p = 0.07). 105 subsequent pregnancies after an index pregnancy with MPVFD were identified. The outcome was favourable with a liveborn rate of 91-100%. DISCUSSION: The extent of fibrin in the placenta plays a role in pregnancy outcome. Cases with severe MPVFD in the placenta was associated with more prematurity, fetal growth restriction and intrauterine fetal death.

Multimodal Optical Coherence Tomography for Intraoperative Evaluation of Tumor Margins and Surgical Margins in Breast-Conserving Surgery.

The aim of the study: We compare the effectiveness of multimodal optical coherence tomography (MM OCT) in the traditional structural OCT mode and the OCT elastography (OCE) mode in addressing two clinically important tasks: (1) detecting groups of tumor cells at surgical margins during breast-сonserving surgery (BСS) in breast cancer (BC) and (2) identifying breast tumor margins. The obtained results were correlated with corresponding histological sections. Materials and Methods: The study was performed on 100 surgical margin samples (top, bottom, medial, and lateral - four samples from each patient in total) obtained from 25 patients with BC who underwent BCS (lumpectomy), and on 25 postoperative tumor samples (to determine tumor margins). With MM OCT method, we visually and numerically assessed the scattering (level and depth of OCT signal penetration) and elastic (stiffness values, or Young's modulus (kPa)) properties of the tumor and non-tumor breast tissue and the obtained values were compared with the results of postoperative histological examination. Results: In 4 surgical margin samples (out of 100), with the OCE method we identified groups of histologically confirmed tumor cells ("positive" resection margins) at the distance of about 5 mm from the visible tumor margin. The identified zones were larger than 0.5 mm with stiffness of more than 400 kPa in all these cases. However, the structural OCT could not identify these groups of tumors and they were not distinguishable from the surrounding fibrous tissue.In the areas of tumor into non-tumor tissue transition, structural OCT images detected tumor margins only if they were adjacent to adipose tissue and did not detect them if there were adjacent to non-tumor fibrous tissue. OCE images with high stiffness values (more than 400 kPa) and high contrast showed a clear tumor margin with both adipose and fibrous tissue. Conclusion: The study demonstarets the potential of MM OCT, particularly its OCE mode, as a real-time method for intraoperative tumor margin and surgical margin assessment in BCS. OCE images compared to structural OCT images visualize higher contrast between different types of breast tissue (adipose tissue, fibrous stroma, hyalinized stroma, tumor cell clusters), as well as more accurate identification of the tumor border and detection of small groups of tumor cells at surgical margins. An algorithm for intraoperative MM OCT examination of the state of the resection margin is proposed in accordance with standard clinical guidelines for achieving clean surgical margins in breast cancer patients.

Perinatal outcome of pregnancies complicated by placental chorioangioma: systematic review and meta-analysis.

OBJECTIVE: To report the perinatal outcome of singleton pregnancies complicated by placental chorioangioma diagnosed on prenatal ultrasound. METHODS: MEDLINE, EMBASE, CINAHL and ClinicalTrials.gov databases were searched for studies reporting the outcome of pregnancies complicated by placental chorioangioma. Inclusion criteria were singleton pregnancy diagnosed with placental chorioangioma on prenatal ultrasound, with no other associated structural anomaly. The primary outcome was perinatal mortality. Secondary outcomes included associated non-structural anomalies detected on prenatal ultrasound (including fetal hydrops, anemia, polyhydramnios, signs of hyperdynamic circulation and small-for-gestational-age (SGA) fetus), SGA at birth, composite neonatal morbidity and preterm birth. Outcome was assessed separately in pregnancies undergoing and those not undergoing fetal therapy. Subanalyses were performed according to the presence of hydrops and the size of the tumor in all pregnancies diagnosed with chorioangioma. Random-effects meta-analyses of proportions were used to analyze the data. RESULTS: Twenty-eight studies (161 pregnancies) were included. In pregnancies complicated by chorioangioma that did not undergo intervention, intrauterine death occurred in 8.2% (95% CI, 3.8-15.0%), while neonatal death and perinatal death occurred in 3.8% (95% CI, 1.0-8.1%) and 11.1% (95% CI, 5.0-19.4%), respectively. SGA at birth was present in 24.0% (95% CI, 13.5-36.5%) of cases, while preterm birth < 37 weeks complicated 34.1% (95% CI, 21.1-48.3%) of pregnancies. Composite neonatal morbidity occurred in 12.0% (95% CI, 4.5-22.3%) of cases. On ultrasound, signs of fetal hyperdynamic circulation were present in 21.0% (95% CI, 9.6-35.3%) of cases, while peak systolic velocity in the fetal middle cerebral artery was increased in 20.6% (95% CI, 10.9-32.3%). Subanalysis according to the size of chorioangioma, including both pregnancies that did and those that did not undergo intervention, showed a progressive increase in the occurrence of most of the outcomes explored with increasing size of the tumor. Furthermore, the prevalence of adverse perinatal outcome was high in pregnancies complicated by chorioangioma presenting with fetal hydrops. There was no randomized controlled trial comparing intervention vs expectant management in pregnancies complicated by chorioangioma with signs of fetal compromise (hydrops or hyperdynamic circulation). Overall, perinatal mortality occurred in 31.2% (95% CI, 18.1-46.1%) of fetuses undergoing in-utero therapy, and 57.3% (95% CI, 39.2-74.4%) had resolution of hydrops or hyperdynamic circulation after treatment. CONCLUSIONS: Placental chorioangioma is associated with adverse perinatal outcome. The size of the mass and presence of fetal hydrops are likely to be the main determinants of perinatal outcome in affected pregnancies. Copyright © 2019 ISUOG. Published by John Wiley & Sons Ltd.

Optical coherence angiography for pre-treatment assessment and treatment monitoring following photodynamic therapy: a basal cell carcinoma patient study.

Microvascular networks of human basal cell carcinomas (BCC) and surrounding skin were assessed with optical coherence angiography (OCA) in conjunction with photodynamic therapy (PDT). OCA images were collected and analyzed in 31 lesions pre-treatment, and immediately/24 hours/3-12 months post-treatment. Pre-treatment OCA enabled differentiation between prevalent subtypes of BCC (nodular and superficial) and nodular-with-necrotic-core BCC subtypes with a diagnostic accuracy of 78%; this can facilitate more accurate biopsy reducing sampling error and better therapy regimen selection. Post-treatment OCA images at 24 hours were 98% predictive of eventual outcome. Additional findings highlight the importance of pre-treatment necrotic core, vascular metrics associated with hypertrophic scar formation, and early microvascular changes necessary in both tumorous and peri-tumorous regions to ensure treatment success.

Accurate early prediction of tumour response to PDT using optical coherence angiography.

Prediction of tumour treatment response may play a crucial role in therapy selection and optimization of its delivery parameters. Here we use optical coherence angiography (OCA) as a minimally-invasive, label-free, real-time bioimaging method to visualize normal and pathological perfused vessels and monitor treatment response following vascular-targeted photodynamic therapy (PDT). Preclinical results are reported in a convenient experimental model (CT-26 colon tumour inoculated in murine ear), enabling controlled PDT and post-treatment OCA monitoring. To accurately predict long-term treatment outcome, a robust and simple microvascular metric is proposed. It is based on perfused vessels density (PVD) at t = 24 hours post PDT, calculated for both tumour and peri-tumour regions. Histological validation in the examined experimental cohort (n = 31 animals) enabled further insight into the excellent predictive power of the derived early-response OCA microvascular metric. The results underscore the key role of peri-tumour microvasculature in determining the long-term PDT response.

In-vivo longitudinal imaging of microvascular changes in irradiated oral mucosa of radiotherapy cancer patients using optical coherence tomography.

Mucositis is the limiting toxicity of radio(chemo)therapy of head and neck cancer. Diagnostics, prophylaxis and correction of this condition demand new accurate and objective approaches. Here we report on an in vivo longitudinal monitoring of the oral mucosa dynamics in 25 patients during the course of radiotherapy of oropharyngeal and nasopharyngeal cancer using multifunctional optical coherence tomography (OCT). A spectral domain OCT system with a specially-designed oral imaging probe was used. Microvasculature visualization was based on temporal speckle variations of the full complex signal evaluated by high-pass filtering of 3D data along the slow scan axis. Angiographic image quantification demonstrated an increase of the vascular density and total length of capillary-like-vessels before visual signs or clinical symptoms of mucositis occur. Especially significant microvascular changes compared to their initial levels occurred when grade two and three mucositis developed. Further, microvascular reaction was seen to be dose-level dependent. OCT monitoring in radiotherapy offers a non-invasive, convenient, label-free quantifiable structural and functional volumetric imaging method suitable for longitudinal human patient studies, furnishing fundamental radiobiological insights and potentially providing useful feedback data to enable adaptive radiotherapy (ART).

Photodynamic therapy monitoring with optical coherence angiography.

Photodynamic therapy (PDT) is a promising modern approach for cancer therapy with low normal tissue toxicity. This study was focused on a vascular-targeting Chlorine E6 mediated PDT. A new angiographic imaging approach known as M-mode-like optical coherence angiography (MML-OCA) was able to sensitively detect PDT-induced microvascular alterations in the mouse ear tumour model CT26. Histological analysis showed that the main mechanisms of vascular PDT was thrombosis of blood vessels and hemorrhage, which agrees with angiographic imaging by MML-OCA. Relationship between MML-OCA-detected early microvascular damage post PDT (within 24 hours) and tumour regression/regrowth was confirmed by histology. The advantages of MML-OCA such as direct image acquisition, fast processing, robust and affordable system opto-electronics, and label-free high contrast 3D visualization of the microvasculature suggest attractive possibilities of this method in practical clinical monitoring of cancer therapies with microvascular involvement.

Increased placental expression and maternal serum levels of apoptosis-inducing TRAIL in recurrent miscarriage.

INTRODUCTION: Recurrent miscarriage (RM; ≥3 consecutive pregnancy losses) occurs in 1-3% of fertile couples. No biomarkers with high predictive value of threatening miscarriage have been identified. We aimed to profile whole-genome differential gene expression in RM placental tissue, and to determine the protein levels of identified loci in maternal sera in early pregnancy. METHODS: GeneChips (Affymetrix(®)) were used for discovery and Taqman RT-qPCR assays for replication of mRNA expression in placentas from RM cases (n = 13) compared to uncomplicated pregnancies matched for gestational age (n = 23). Concentrations of soluble TRAIL (sTRAIL) and calprotectin in maternal serum in normal first trimester (n = 35) and failed pregnancies (early miscarriage, n = 18, late miscarriage, n = 4; tubal pregnancy, n = 11) were determined using ELISA. RESULTS: In RM placentas 30 differentially expressed (with nominal P-value < 0.05) transcripts were identified. Significantly increased placental mRNA expression of TNF-related apoptosis-inducing ligand (TRAIL; P = 1.4 × 10(-3); fold-change 1.68) and S100A8 (P = 7.9 × 10(-4); fold-change 2.56) encoding for inflammatory marker calprotectin (S100A8/A9) was confirmed by RT-qPCR. When compared to normal first trimester pregnancy (sTRAIL 16.1 ± 1.6 pg/ml), significantly higher maternal serum concentration of sTRAIL was detected at the RM event (33.6 ± 4.3 pg/ml, P = 0.00027), and in pregnant women, who developed an unpredicted miscarriage 2-50 days after prospective serum sampling (28.5 ± 4.4 pg/ml, P = 0.039). Women with tubal pregnancy also exhibited elevated sTRAIL (30.5 ± 3.9 pg/ml, P = 0.035). Maternal serum levels of calprotectin were neither diagnostic nor prognostic to early pregnancy failures (P > 0.05). CONCLUSIONS: The study indicated of sTRAIL as a potential predictive biomarker in maternal serum for early pregnancy complications.

[Laser hyperthermia of tumors with the use of golden nanoparticles under the control of optical coherent tomography and acoustothermometry].

The local laser hyperthermia of an experimental tumor RShM-5 of mice with the use of golden plasmin resonance nanoparticles has been carried out. The accumulation of particles in the tumor was controlled by the in vivo noninvasive method of optical coherent tomography. Using this method, the time of the maximum content of nanoparticles in the tumor was determined to be 5 h after the intravenous administration during which the laser hyperthermia was performed. The control of the tumor temperature during the hyperthermia seance showed that the application of nanoparticles provides an effective temperature elevation inside the node and a more targeted heating. The local laser hyperthermia with nanoparticles induced the inhibition of the tumor growth from day 5 to day 7 after the seance with a maximum value of 56%.

Contrasting properties of gold nanoparticles for optical coherence tomography: phantom, in vivo studies and Monte Carlo simulation.

The possibility of using silica-gold nanoshells with 150 nm silica core size and 25 nm thick gold shell as contrasting agents for optical coherence tomography (OCT) is analyzed. Experiments on agar biotissue phantoms showed that the penetration of nanoshells into the phantoms increases the intensity of the optical coherence tomography (OCT) signal and the brightness of the corresponding areas of the OCT image. In vivo experiments on rabbit skin demonstrated that the application of nanoshells onto the skin provides significant contrasting of the borders between the areas containing nanoshells and those without. This effect of nanoshells on skin in vivo is manifested by the increase in intensity of the OCT signal in superficial parts of the skin, boundary contrast between superficial and deep dermis and contrast of hair follicles and glands. The presence of nanoshells in the skin was confirmed by electron microscopy. Monte Carlo simulations of OCT images confirmed the possibility of contrasting skin-layer borders and structures by the application of gold nanoshells. The Monte Carlo simulations were performed for two skin models and exhibit effects of nanoparticles similar to those obtained in the experimental part of the study, thus proving that the effects originate exactly from the presence of nanoparticles.