In vivo assessment of bladder cancer with diffuse reflectance and fluorescence spectroscopy: A comparative study.

OBJECTIVES: The aim of this work is to assess the performance of multimodal spectroscopic approach combined with single core optical fiber for detection of bladder cancer during surgery in vivo. METHODS: Multimodal approach combines diffuse reflectance spectroscopy (DRS), fluorescence spectroscopy in the visible (405 nm excitation) and near-infrared (NIR) (690 nm excitation) ranges, and high-wavenumber Raman spectroscopy. All four spectroscopic methods were combined in a single setup. For 21 patients with suspected bladder cancer or during control cystoscopy optical spectra of bladder cancer, healthy bladder wall tissue and/or scars were measured. Classification of cancerous and healthy bladder tissue was performed using machine learning methods. RESULTS: Statistically significant differences in relative total haemoglobin content, oxygenation, scattering, and visible fluorescence intensity were found between tumor and normal tissues. The combination of DRS and visible fluorescence spectroscopy allowed detecting cancerous tissue with sensitivity and specificity of 78% and 91%, respectively. The addition of features extracted from NIR fluorescence and Raman spectra did not improve the quality of classification. CONCLUSIONS: This study demonstrates that multimodal spectroscopic approach allows increasing sensitivity and specificity of bladder cancer detection in vivo. The developed approach does not require special probes and can be used with single-core optical fibers applied for laser surgery.

Predictive potential of ACE phenotyping in extrapulmonary sarcoidosis.

Elevated ACE expression in tissues (reflected by blood ACE levels) is associated with increased risk of cardiovascular diseases and is also a marker for granulomatous diseases. We developed a new approach for characterization of ACE status in the blood-ACE phenotyping and established normal values of ACE levels 50-150% of control pooled plasma. ACE phenotyping was performed in citrated plasma of 120 patients with known interstitial lung diseases. In the 1st set of 100 patients we found 22 patients with ACE levels > 150%; ACE phenotyping also objectively identified the presence of ACE inhibitors in the plasma of 15 patients. After excluding these patients and patient with ACE mutation that increases ACE shedding, 17 patients were identified as a suspicious for systemic sarcoidosis based on elevation of blood ACE (> 150% of mean). A new parameter that we have established-ACE immunoreactivity (with mAb 9B9)-allowed us to detect 22 patients with decreased values (< 80%) of this parameter, which may indicate the presence of ACE in the blood that originates from macrophages/dendritic cells of granulomas. In the remaining 20 patients, this new parameter (mAbs binding/activity ratio) was calculated using 3 mAbs (9B9, 3A5 and i1A8-having overlapping epitopes), and 8 patients were identified as having decreases in this parameter, thus increasing dramatically the sensitivity for detection of patients with systemic sarcoidosis. Whole body PET scan confirmed extrapulmonary granulomas in some patients with lower immunoreactivity towards anti-ACE mAbs. ACE phenotyping has novel potential to noninvasively detect patients with systemic sarcoidosis.

The Association of PLAUR Genotype and Soluble suPAR Serum Level with COVID-19-Related Lung Damage Severity.

Uncovering the risk factors for acute respiratory disease coronavirus 2019 (COVID-19) severity may help to provide a valuable tool for early patient stratification and proper treatment implementation, improving the patient outcome and lowering the burden on the healthcare system. Here we report the results of a single-center retrospective cohort study on 151 severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-infected symptomatic hospitalized adult patients. We assessed the association of several blood test measurements, soluble urokinase receptor (uPAR) serum level and specific single nucleotide polymorphisms of ACE (I/D), NOS3 (rs2070744, rs1799983), SERPINE1 (rs1799768), PLAU (rs2227564) and PLAUR (rs344781, rs2302524) genes, with the disease severity classified by the percentage of lung involvement on computerized tomography scans. Our findings reveal that the T/C genotype of PLAUR rs2302524 was independently associated with a less severe lung damage (odds ratio 0.258 [0.071-0.811]). Along with high C-reactive protein, fibrinogen and soluble uPAR serum levels turned out to be independently associated with more severe lung damage in COVID-19 patients. The identified factors may be further employed as predictors of a possibly severe COVID-19 clinical course.

Prediction of reproductive function recovery after microsurgical varicocelectomy in men from infertile couples: Clinical and laboratory predictors.

The objective was to reveal predictors for fertility recovery after varicocelectomy in subfertile men. This retrospective study recruited 93 men with clinical varicocele and pathozoospermia who underwent microsurgical varicocelectomy. Stepwise discriminant analysis was performed to identify predictors of spontaneous pregnancy (SP) after surgery. 'Clinically significant improvement' (CSI) following varicocelectomy was defined as an increase in total progressively motile sperm count (TPMSC) by at least 12.5 million (calculated from WHO-2010 reference values). 52% of patients showed CSI, and 28% reported SP after surgery. Patients who reported SP (group II), compared to that remained infertile (group I), were younger (27.3 ± 2.9 versus 30.2 ± 4.2 years; p < .01), had less infertility period (24.1 ± 14.0 versus 44.4 ± 32.9 months; p < .05) and had initially higher TPMSC (median (25% -75%) = 34 (11-67) versus 9.5 (0-33) mln; p < .05). The stepwise discriminant analysis showed that male age (coefficient value = -0.157), total sperm motility (0.024) and postoperative increase in TPMSC (0.010) were the significant predictors of SP. The predictive ability, sensitivity and specificity of the discriminant function were 84%, 87%, 76% respectively. This algorithm can be recommended after varicocelectomy in decision-making on natural conception or the ART protocols usage.

Skin dehydration monitoring with optical spectroscopy allows assessment of water content in the organism: Thermal and physical loads, diuretic therapy.

This study investigates the relationship between body hydration levels and skin hydration using spatially resolved diffuse reflectance spectroscopy. The research involved monitoring skin dehydration and rehydration under various conditions, including thermal and physical loads on healthy volunteers, and diuretic therapy in patients with edema syndrome. Findings indicate a correlation between body mass reduction and skin hydration: a 1% loss in body mass corresponds to a 10% decrease in skin hydration. During thermal stress, water absorption at 970 nm decreased monotonically without recovery. Physical activity resulted in approximately 10% changes in skin water content within 20 min, followed by rehydration. Patients with edema syndrome exhibited the most substantial decrease in water absorption amplitude, at nearly 30%, during diuretic treatment. These results support optical spectroscopy as a non-invasive tool for assessing body hydration, with implications for developing portable hydration monitoring devices for clinical and sports applications.

Predictors of microsurgical varicocelectomy efficacy in male infertility treatment: critical assessment and systematization.

In this review, we tried to systematize all the evidence (from PubMed [MEDLINE], Scopus, Cochrane Library, EBSCO, Embase, and Google Scholar) from 1993 to 2021 on the predictors of microsurgical varicocelectomy efficacy in male infertility treatment. Regarding the outcomes of varicocele repair, we considered semen improvement and pregnancy and analyzed them separately. Based on the 2011 Oxford CEBM Levels of Evidence, we assigned a score to each trial that studied the role of the predictor. We systematized the studied predictors based on the total points, which were, in turn, calculated based on the number and quality of studies that confirmed or rejected the studied predictor as significant, into three levels of significance: predictors of high, moderate, and low clinical significance. Preoperative total motile sperm count (TMSC) coupled with sperm concentration can be a significant predictor of semen improvement and pregnancy after varicocelectomy. In addition, for semen improvement alone, scrotal Doppler ultrasound (DUS) parameters, sperm DNA fragmentation index (DFI), and bilateral varicocelectomy are reliable predictors of microsurgical varicocelectomy efficacy.

Blood ACE Phenotyping for Personalized Medicine: Revelation of Patients with Conformationally Altered ACE.

Background: The angiotensin-converting enzyme (ACE) metabolizes a number of important peptides participating in blood pressure regulation and vascular remodeling. Elevated blood ACE is a marker for granulomatous diseases and elevated ACE expression in tissues is associated with increased risk of cardiovascular diseases. Objective and Methodology: We applied a novel approach -ACE phenotyping-to find a reason for conformationally impaired ACE in the blood of one particular donor. Similar conformationally altered ACEs were detected previously in 2-4% of the healthy population and in up to 20% of patients with uremia, and were characterized by significant increase in the rate of angiotensin I hydrolysis. Principal findings: This donor has (1) significantly increased level of endogenous ACE inhibitor in plasma with MW less than 1000; (2) increased activity toward angiotensin I; (3) M71V mutation in ABCG2 (membrane transporter for more than 200 compounds, including bilirubin). We hypothesize that this patient may also have the decreased level of free bilirubin in plasma, which normally binds to the N domain of ACE. Analysis of the local conformation of ACE in plasma of patients with Gilbert and Crigler-Najjar syndromes allowed us to speculate that binding of mAbs 1G12 and 6A12 to plasma ACE could be a natural sensor for estimation of free bilirubin level in plasma. Totally, 235 human plasma/sera samples were screened for conformational changes in soluble ACE. Conclusions/Significance: ACE phenotyping of plasma samples allows us to identify individuals with conformationally altered ACE. This type of screening has clinical significance because this conformationally altered ACE could not only result in the enhancement of the level of angiotensin II but could also serve as an indicator of free bilirubin levels.

Urinary ACE Phenotyping as a Research and Diagnostic Tool: Identification of Sex-Dependent ACE Immunoreactivity.

BACKGROUND: Angiotensin-converting enzyme (ACE) is highly expressed in renal proximal tubules, but ACE activity/levels in the urine are at least 100-fold lower than in the blood. Decreased proximal tubular ACE has been associated with renal tubular damage in both animal models and clinical studies. Because ACE is shed into urine primarily from proximal tubule epithelial cells, its urinary ACE measurement may be useful as an index of tubular damage. OBJECTIVE AND METHODOLOGY: We applied our novel approach-ACE phenotyping-to characterize urinary ACE in volunteer subjects. ACE phenotyping includes (1) determination of ACE activity using two substrates (ZPHL and HHL); (2) calculation of the ratio of hydrolysis of the two substrates (ZPHL/HHL ratio); (3) quantification of ACE immunoreactive protein levels; and (4) fine mapping of local ACE conformation with mAbs to ACE. PRINCIPAL FINDINGS: In normal volunteers, urinary ACE activity was 140-fold less than in corresponding plasma/serum samples and did not differ between males and females. However, urinary ACE immunoreactivity (normalized binding of 25 mAbs to different epitopes) was strongly sex-dependent for the several mAbs tested, an observation likely explained by differences in tissue ACE glycosylation/sialylation between males and females. Urinary ACE phenotyping also allowed the identification of ACE outliers. In addition, daily variability of urinary ACE has potential utility as a feedback marker for dieting individuals pursuing weight loss. CONCLUSIONS/SIGNIFICANCE: Urinary ACE phenotyping is a promising new approach with potential clinical significance to advance precision medicine screening techniques.

Monitoring the skin structure during edema in vivo with spatially resolved diffuse reflectance spectroscopy.

Significance: Edema occurs in the course of various skin diseases. It manifests itself in changes in water concentrations in skin layers: dermis and hypodermis and their thicknesses. In medicine and cosmetology, objective tools are required to assess the skin's physiological parameters. The dynamics of edema and the skin of healthy volunteers were studied using spatially resolved diffuse reflectance spectroscopy (DRS) in conjunction with ultrasound (US). Aim: In this work, we have developed a method based on DRS with a spatial resolution (SR DRS), allowing us to simultaneously assess water content in the dermis, dermal thickness, and hypodermal thickness. Approach: An experimental investigation of histamine included edema using SR DRS under the control of US was conducted. An approach for skin parameter determination was studied and confirmed using Monte-Carlo simulation of diffuse reflectance spectra for a three-layered system with the varying dermis and hypodermis parameters. Results: It was shown that an interfiber distance of 1 mm yields a minimal relative error of water content determination in the dermis equal to 9.3%. The lowest error of hypodermal thickness estimation was achieved with the interfiber distance of 10 mm. Dermal thickness for a group of volunteers (7 participants, 21 measurement sites) was determined using SR DRS technique with an 8.3% error using machine learning approaches, taking measurements at multiple interfiber distances into account. Hypodermis thickness was determined with root mean squared error of 0.56 mm for the same group. Conclusions: This study demonstrates that measurement of the skin diffuse reflectance response at multiple distances makes it possible to determine the main parameters of the skin and will serve as the basis for the development and verification of an approach that works in a wide range of skin structure parameters.

Initial Assessment of Boiling Histotripsy for Mechanical Ablation of Ex Vivo Human Prostate Tissue.

Boiling histotripsy (BH) is a focused ultrasound technology that uses millisecond-long pulses with shock fronts to induce mechanical tissue ablation. The pulsing scheme and mechanisms of BH differ from those of cavitation cloud histotripsy, which was previously developed for benign prostatic hyperplasia. The goal of the work described here was to evaluate the feasibility of using BH to ablate fresh ex vivo human prostate tissue as a proof of principle for developing BH for prostate applications. Fresh human prostate samples (N = 24) were obtained via rapid autopsy (<24 h after death, institutional review board exempt). Samples were analyzed using shear wave elastography to ensure that mechanical properties of autopsy tissue were clinically representative. Samples were exposed to BH using 10- or 1-ms pulses with 1% duty cycle under real-time B-mode and Doppler imaging. Volumetric lesions were created by sonicating 1-4 rectangular planes spaced 1 mm apart, containing a grid of foci spaced 1-2 mm apart. Tissue then was evaluated grossly and histologically, and the lesion content was analyzed using transmission electron microscopy and scanning electron microscopy. Observed shear wave elastography characterization of ex vivo prostate tissue (37.9 ± 22.2 kPa) was within the typical range observed clinically. During BH, hyperechoic regions were visualized at the focus on B-mode, and BH-induced bubbles were also detected using power Doppler. As treatment progressed, hypoechoic regions of tissue appeared, suggesting successful tissue fractionation. BH treatment was twofold faster using shorter pulses (1 ms vs. 10 ms). Histological analysis revealed lesions containing completely homogenized cell debris, consistent with histotripsy-induced mechanical ablation. It was therefore determined that BH is feasible in fresh ex vivo human prostate tissue producing desired mechanical ablation. The study supports further work aimed at translating BH technology as a clinical option for prostate ablation.

Ultrasound-Triggerable Coatings for Foley Catheter Balloons for Local Release of Anti-Inflammatory Drugs during Bladder Neck Dilation.

Bladder neck contracture (BNC) is a complication of the surgical treatment of benign and malignant prostate conditions and is associated with the partial or complete blockage of urination. Correction of this condition usually requires repeated surgical intervention, which does not guarantee recovery. Balloon dilation is a minimally invasive alternative to the surgical dissection of tissues; however, it significantly reduces the patient's quality of life. Additional local anti-inflammatory treatment may reduce the number of procedures requested and increase the attractiveness of this therapeutic strategy. Here, we report about an ultrathin biocompatible coating based on polylactic acid for Foley catheter balloons that can provide localized release of Prednol-L in the range of 56-99 µg in the BNC zone under conventional diagnostic ultrasound exposure. Note that the exposure of a transrectal probe with a conventional gray-scale ultrasound regimen with and without shear wave elastography (SWE) was comparably effective for Prednol-L release from the coating surface of a Foley catheter balloon. This strategy does not require additional manipulations by clinicians. The trigger for the drug release is the ultrasound exposure, which is applied for visualization of the balloon's location during the dilation process. In vivo experiments demonstrated the absence of negative effects of the usage of a coated Foley catheter for balloon dilation of the bladder neck and urethra.

Phenotyping Angiotensin-Converting Enzyme in Blood: A Necessary Approach for Precision Medicine.

BACKGROUND: Angiotensin-converting enzyme (ACE) metabolizes a number of important peptides participating in blood pressure regulation and vascular remodeling. Elevated ACE expression in tissues (which is generally reflected by ACE in blood) is associated with increased risk of cardiovascular diseases. Elevated ACE in blood is also a marker for granulomatous diseases. METHODS: We applied our novel approach-ACE phenotyping-to characterize serum ACE in 300 unrelated patients and to establish normal values for ACE levels. ACE phenotyping includes (a) determination of ACE activity with 2 substrates (Z-Phe-His-Leu [ZPHL] and Hip-His-Leu [HHL]), (b) calculation of a ratio for hydrolysis of ZPHL and HHL, and (c) quantification of ACE immunoreactive protein levels and ACE conformation with a set of monoclonal antibodies (mAbs) to ACE. RESULTS: Only a combination of ACE activity determination with 2 substrates and quantification of the amount of ACE immunoreactive protein with mAbs 1G12 and 9B9 allows for the unequivocal detection of the presence of ACE inhibitors in the blood. After excluding such subjects, we were able to establish normal values of ACE in healthy populations: 50%-150% from control pooled serum. This ACE phenotyping approach in screening format with special attention to outliers can also identify patients with various mutations in ACE and may help to identify the as yet unknown ACE secretase or other mechanistic details of precise regulation of ACE expression. CONCLUSIONS: ACE phenotyping is a promising new approach with potential clinical significance to advance precision medicine screening techniques by establishing different risk groups based on ACE phenotype.

Single-Fiber Diffuse Reflectance Spectroscopy and Spatial Frequency Domain Imaging in Surgery Guidance: A Study on Optical Phantoms.

Diffuse reflectance spectroscopy (DRS) and imaging are increasingly being used in surgical guidance for tumor margin detection during endoscopic operations. However, the accuracy of the boundary detection with optical techniques may depend on the acquisition parameters, and its evaluation is in high demand. In this work, using optical phantoms with homogeneous and heterogeneous distribution of chromophores mimicking normal and pathological bladder tissues, the accuracy of tumor margin detection using single-fiber diffuse reflectance spectroscopy and spatial frequency domain imaging was evaluated. We also showed how the diffuse reflectance response obtained at different spatial frequencies with the spatial frequency domain imaging technique could be used not only to quantitatively map absorption and scattering coefficients of normal tissues and tumor-like heterogeneities but also to estimate the tumor depth localization. The demonstrated results could be helpful for proper analysis of the DRS data measured in vivo and for translation of optical techniques for tumor margin detection to clinics.

Novel ACE mutations mimicking sarcoidosis by increasing blood ACE levels.

An elevated blood angiotensin I-converting enzyme (ACE) supports diagnosis of sarcoidosis and Gaucher disease. However, some ACE mutations increase ACE shedding, and patients with these mutations are therefore at risk of being incorrectly diagnosed with sarcoidosis because of elevated serum ACE levels. We applied a novel approach called "ACE phenotyping" to identify possible ACE mutations in 3 pulmonary clinic patients that had suspected sarcoidosis based on elevated blood ACE levels. Conformational fingerprinting of ACE indicated that these mutations may be localized in the stalk region of the protein and these were confirmed by whole exome sequencing. Index patient 1 (IP1) had a mutation (P1199L) that had been previously identified, while the other 2 patients had novel ACE mutations. IP2 had 2 mutations, T887M and N1196K (eliminating a putative glycosylation site), while IP3 had a stop codon mutation Q1124X (eliminating the transmembrane anchor). We also performed a comprehensive analysis of the existing database of all ACE mutations to estimate the proportion of mutations increasing ACE shedding. The frequency of ACE mutations resulting in increased blood ACE levels may be much higher than previously estimated. ACE phenotyping, together with whole exome sequencing, is a diagnostic approach that could prevent unnecessary invasive and/or costly diagnostic procedures, or potentially harmful treatment for patients misdiagnosed on the basis of elevated blood ACE levels.

Scrotal Tissues: The Perfect Material for Urogenital Reconstruction.

Correction of male urogenital pathology of different severity is complicated by the anatomical aspects of the penis. Skin texture, internal structures, and perfusion dynamics of the urogenital area make it a difficult area to reconstruct. We provide our experience with axial scrotal flaps for correction of penile defects of different severity and believe that these local flaps offer sufficient tissue characteristics for proper restoration of this complex region. METHODS: Forty-eight patients were divided into 3 groups depending on penile defect genesis and severity. Axial scrotal flap reconstruction was used for the correction of defects in all cases, when necessary in combination with other flaps. RESULTS: Axial scrotal flaps for total and subtotal penile reconstruction serve as valuable material for reconstruction of the urogenital area, and are to be combined with other flaps for restoration of bulk tissues. Localized defects of the penis and urethra reconstructed by axial scrotal flaps provide excellent aesthetic results with minimal scaring, stable perfusion dynamics, and high satisfaction rate. CONCLUSIONS: Scrotal tissues provide an excellent reconstruction material for penile defects because of their highly similar tissue structure as that of the penis. Scrotal axial flaps do not provide excess bulking in the postoperative period and are recommended for reconstruction of urethral and localized penile defects. Multistage surgery is recommended in cases of severe tissue damage, in combination with other flaps (inguinal, thoracodorsal, and radial).

Pericapillary Edema Assessment by Means of the Nailfold Capillaroscopy and Laser Scanning Microscopy.

Edema, i.e., fluid accumulation in the interstitial space, accompanies numerous pathological states of the human organism, including heart failure (HF), inflammatory response, and lymphedema. Nevertheless, techniques for quantitative assessment of the edema's severity and dynamics are absent in clinical practice, and the analysis is mainly limited to physical examination. This fact stimulates the development of novel methods for fast and reliable diagnostics of fluid retention in tissues. In this work, we focused on the possibilities of two microscopic techniques, nailfold video capillaroscopy (NVC) and confocal laser scanning microscopy (CLSM), in the assessment of the short-term and long-term cutaneous edema. We showed that for the patients with HF, morphological parameters obtained by NVC-namely, the apical diameter of capillaries and the size of the perivascular zone-indicate long-term edema. On the other hand, for healthy volunteers, the application of two models of short-term edema, venous occlusion, and histamine treatment of the skin, did not reveal notable changes in the capillary parameters. However, a significant reduction of the NVC image sharpness was observed in this case, which was suggested to be due to water accumulation in the epidermis. To verify these findings, we made use of CLSM, which provides the skin structure with cellular resolution. It was observed that for the histamine-treated skin, the areas of the dermal papillae become hyporefractive, leading to the loss of contrast and the lower visibility of capillaries. Similar effect was observed for patients undergoing infusion therapy. Collectively, our results reveal the parameters can be used for pericapillary edema assessment using the NVC and CLSM, and paves the way for their application in a clinical set-up.

Tissue ACE phenotyping in prostate cancer.

Epithelial cells of prostate express significant level of ACE and, as a result, seminal fluid has 50-fold more ACE than plasma. The substitution of highly specialized prostate epithelial cells by tumor cells results in dramatic decrease in ACE production in prostate tissues. We performed detailed characterization of ACE status in prostate tissues from patients with benign prostate hyperplasia (BPH) and prostate cancer (PC) using new approach- ACE phenotyping, that includes evaluation of: 1) ACE activity with two substrates (HHL and ZPHL); 2) the ratio of the rates of their hydrolysis (ZPHL/HHL ratio); 3) the ratio of immunoreactive ACE protein to ACE activity; 4) the pattern of mAbs binding to different epitopes on ACE - ACE conformational fingerprint - to reveal conformational changes in prostate ACE due to prostate pathology. ACE activity dramatically decreased and the ratio of immunoreactive ACE protein to ACE activity increased in PC tissues. The catalytic parameter, ZPHL/HHL ratio, increased in prostate tissues from all patients with PC, but was did not change for most |BPH patients. Nevertheless, prostate tissues of several patients diagnosed with BPH based on histology, also demonstrated decreased ACE activity and increased immunoreactive ACE protein/ACE activity and ZPHL/HHL ratios, that could be considered as more early indicators of prostate cancer development than routine histology. Thus, ACE phenotyping of prostate biopsies has a potential to be an effective approach for early diagnostics of prostate cancer or at least for differential diagnostics of BPH and PC.