Spatial Distribution of Macrophage and Lymphocyte Subtypes within Tumor Microenvironment to Predict Recurrence of Non-Muscle-Invasive Papillary Urothelial Carcinoma after BCG Immunotherapy.

Non-muscle-invasive papillary urothelial carcinoma (NMIPUC) of the urinary bladder is the most common type of bladder cancer. Intravesical Bacille Calmette-Guerin (BCG) immunotherapy is applied in patients with a high risk of recurrence and progression of NMIPUC to muscle-invasive disease. However, the tumor relapses in about 30% of patients despite the treatment, raising the need for better risk stratification. We explored the potential of spatial distributions of immune cell subtypes (CD20, CD11c, CD163, ICOS, and CD8) within the tumor microenvironment to predict NMIPUC recurrence following BCG immunotherapy. Based on analyses of digital whole-slide images, we assessed the densities of the immune cells in the epithelial-stromal interface zone compartments and their distribution, represented by an epithelial-stromal interface density ratio (IDR). While the densities of any cell type did not predict recurrence, a higher IDR of CD11c (HR: 0.0012, p-value = 0.0002), CD8 (HR: 0.0379, p-value = 0.005), and ICOS (HR: 0.0768, p-value = 0.0388) was associated with longer recurrence-free survival (RFS) based on the univariate Cox regression. The history of positive repeated TUR (re-TUR) (HR: 4.93, p-value = 0.0001) and T1 tumor stage (HR: 2.04, p-value = 0.0159) were associated with shorter RFS, while G3 tumor grade according to the 1973 WHO classification showed borderline significance (HR: 1.83, p-value = 0.0522). In a multivariate analysis, the two models with a concordance index exceeding 0.7 included the CD11c IDR in combination with either a history of positive re-TUR or tumor stage. We conclude that the CD11c IDR is the most informative predictor of NMIPUC recurrence after BCG immunotherapy. Our findings highlight the importance of assessment of the spatial distribution of immune cells in the tumor microenvironment.

Prediction of Non-Muscle Invasive Papillary Urothelial Carcinoma Relapse from Hematoxylin-Eosin Images Using Deep Multiple Instance Learning in Patients Treated with Bacille Calmette-Guérin Immunotherapy.

The limited reproducibility of the grading of non-muscle invasive papillary urothelial carcinoma (NMIPUC) necessitates the search for more robust image-based predictive factors. In a cohort of 157 NMIPUC patients treated with Bacille Calmette-Guérin (BCG) immunotherapy, we explored the multiple instance learning (MIL)-based classification approach for the prediction of 2-year and 5-year relapse-free survival and the multiple instance survival learning (MISL) framework for survival regression. We used features extracted from image patches sampled from whole slide images of hematoxylin-eosin-stained transurethral resection (TUR) NPMIPUC specimens and tested several patch sampling and feature extraction network variations to optimize the model performance. We selected the model showing the best patient survival stratification for further testing in the context of clinical and pathological variables. MISL with the multiresolution patch sampling technique achieved the best patient risk stratification (concordance index = 0.574, p = 0.010), followed by a 2-year MIL classification. The best-selected model revealed an independent prognostic value in the context of other clinical and pathologic variables (tumor stage, grade, and presence of tumor on the repeated TUR) with statistically significant patient risk stratification. Our findings suggest that MISL-based predictions can improve NMIPUC patient risk stratification, while validation studies are needed to test the generalizability of our models.

A Prospective Study on the Impact of Clinical Factors and Adjusted Triple D System for Success Rate of ESWL.

Objective: Our study aimed to evaluate the success rate of ESWL and identify relevant treatment-specific factors affecting treatment outcomes, as well as to assess the accuracy of the updated Triple D scoring system and compare it with older systems. Material and Methods: A prospective study of 71 patients who received ESWL treatment for renal stones that were 5-15 mm in size was completed. The patient having no residual stones or residual stones lesser than 4 mm after ESWL was identified as a treatment success. Univariate and multivariate logistic regression and ROC curves were used to identify important factors for treatment outcomes. Results: Successful treatment was achieved for 66.2% of patients. The stone volume (SV), mean stone density (MD), and delivered power to the stone volume unit ratio (SMLI/SV) were defined as the most critical factors influencing ESWL success. An updated Triple D score system with a, SMLI/SV ratio could be an alternative to older systems and reach an even higher accuracy. A limitation of this study is the limited sample size due to the COVID-19 pandemic. Conclusions: Our results show that the three factors that most influence the success of ESWL are the stone size, mean stone density, and SMLI/SV ratio. Based on this, we present a simple updated triple D score system to predict ESWL success, which could be implemented in future clinical practice.

CD8+ Cell Density Gradient across the Tumor Epithelium-Stromal Interface of Non-Muscle Invasive Papillary Urothelial Carcinoma Predicts Recurrence-Free Survival after BCG Immunotherapy.

BACKGROUND: Bacille Calmette-Guerin (BCG) immunotherapy is the first-line treatment in patients with high-risk non-muscle invasive papillary urothelial carcinoma (NMIPUC), the most common type of bladder cancer. The therapy outcomes are variable and may depend on the immune response within the tumor microenvironment. In our study, we explored the prognostic value of CD8+ cell density gradient indicators across the tumor epithelium-stroma interface of NMIPUC. METHODS: Clinical and pathologic data were retrospectively collected from 157 NMIPUC patients treated with BCG immunotherapy after transurethral resection. Whole-slide digital image analysis of CD8 immunohistochemistry slides was used for tissue segmentation, CD8+ cell quantification, and the assessment of CD8+ cell densities within the epithelium-stroma interface. Subsequently, the gradient indicators (center of mass and immunodrop) were computed to represent the density gradient across the interface. RESULTS: By univariable analysis of the clinicopathologic factors, including the history of previous NMIPUC, poor tumor differentiation, and pT1 stage, were associated with shorter RFS (p < 0.05). In CD8+ analyses, only the gradient indicators but not the absolute CD8+ densities were predictive for RFS (p < 0.05). The best-performing cross-validated model included previous episodes of NMIPUC (HR = 4.4492, p = 0.0063), poor differentiation (HR = 2.3672, p = 0.0457), and immunodrop (HR = 5.5072, p = 0.0455). CONCLUSIONS: We found that gradient indicators of CD8+ cell densities across the tumor epithelium-stroma interface, along with routine clinical and pathology data, improve the prediction of RFS in NMIPUC.

A Novel Infrared Spectroscopy Method for Analysis of Stone Dust for Establishing Final Composition of Urolithiasis.

Background: The introduction of the holmium laser for lithotripsy and minimally invasive techniques in endoscopy increased the popularity of stone dusting techniques. Retrieving stone pieces for an analysis increases the economic burden of surgery and operative time. Novel methods are needed for the analysis of convenient urolithiasis composition. Objective: This study aims to assess the efficacy of the stone dust Fourier transform infrared spectroscopy coupled with attenuated total reflection (FTIR ATR) method for accurate stone composition determination from the dust specimens compared with simultaneously retrieved standard stone fragments. Design setting and participants: From July 2021 to March 2022, a total of 75 patients who received endoscopic treatment for urolithiasis were included in this study. Outcome measurements and statistical analysis: The accuracy of the FTIR ATR method was assessed via estimates of sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV). The results were compared between samples of stone dust and the final stone composition. Results and limitations: Total or partial biochemical composition agreement was observed in 92.7% of cases and total agreement in 82.4% of cases when stone dust was compared with stone fragments. The highest accuracy rates were obtained for uric acid stones: sensitivity 100%, specificity 98.3%, PPV 90.9%, and NPV 100%. Identification of other types of stones was also of high accuracy, reaching up to 83.3% sensitivity and 100% specificity. Conclusions: The application of FTIR ATR spectroscopy for a stone dust analysis allows obtaining easy and cost-effective final composition of urolithiasis without a stone fragment analysis. This technique was shown to be feasible, and there is substantial potential for clinical practice. Patient summary: This study investigates a novel method that determines accurate stone composition without acquiring the pieces of stone during surgery. The results have shown that stone dust Fourier transform infrared spectroscopy coupled with attenuated total reflection provides accurate stone composition.

Differentiation of Urothelial Carcinoma and Normal Bladder Tissues by Means of Fiber-Based ATR IR Spectroscopy.

Surgical treatment is widely applied curative approach for bladder cancer. White light cystoscopy (WLC) is currently used for intraoperative diagnostics of malignant lesions but has relatively high false-negative rate. Here we represent an application of label free fiber-based attenuated total reflection infrared spectroscopy (ATR IR) for freshly resected human bladder tissue examination for 54 patients. Defined molecular spectral markers allow to identify normal and urothelial carcinoma tissues. While methods of statistical analysis (Hierarchical cluster analysis (HCA) and Principal component analysis (PCA)) used for spectral data treatment allow to discriminate tissue types with 91% sensitivity and 96-98% specificity. In the present study the described method was applied for tissue examination under ex vivo conditions. However, after method validation the equipment could be translated from laboratory studies to in situ or even in vivo studies in operating room.

Sulforaphane has an additive anticancer effect to FOLFOX in highly metastatic human colon carcinoma cells.

Colorectal cancer (CRC) is one of the most common malignancies worldwide. Patients with CRC may need chemotherapy (CTx) in a neoadjuvant, adjuvant or palliative setting through the course of the disease. Unfortunately, its effect is limited by chemoresistance and chemotoxicity. Novel more effective and non­toxic CTx regimens are needed to further improve CRC treatment outcomes. Thus, the present study was designed to test the hypothesis that non­toxic sulforaphane (SF) is effective against CRC and has additive effects in combination with conventional 5­fluorouracil, oxaliplatin and folinic acid (FOLFOX) CTx in vitro. Highly metastatic human colon cancer cells, CX­1, and fibroblasts were treated with FOLFOX ± SF. Cell viability was assessed using an MTT assay. The level of apoptosis and the expression of apoptotic proteins were measured by TUNEL assay and quantitative PCR analysis. Aldehyde dehydrogenase isoform 1 (ALDH1) and multidrug resistance protein 2 (MRP2) levels were evaluated. The ability of cells to form spheroids was measured in three­dimensional cell culture. SF alone and in combination with FOLFOX effectively decreased the viability of the CX­1 cells, promoted apoptosis within the CX­1 cells, prevented cellular spheroid formation and decreased ALDH1 activity. However, SF promoted MRP2 expression and protein levels. In conclusion, SF together with conventional FOLFOX has additive anticancer effects against highly metastatic human CRC in vitro.

Evaluation of Renal Function after Partial Nephrectomy and Detection of Clinically Significant Acute Kidney Injury.

Background and Objectives: Consequences of partial nephrectomy (PN), intraoperative hypotension (IOH) and postoperative neutrophil to lymphocyte ratio (NLR) may cause postoperative acute kidney injury (AKI) and in long-term-chronic kidney disease (CKD). Our study aimed to identify the AKI incidence after PN, to find clinically significant postoperative AKI and renal dysfunction, and to determine the predictor factors. Materials and Methods: A prospective observational study consisted of 91 patients who received PN with warm ischemia, and estimated preoperative glomerular filtration rate (eGFR) ≥ 60 mL/min and without abnormal albuminuria. Results: 38 (41.8%) patients experienced postoperative AKI. Twenty-one (24.1%) patients had CKD upstage after 1 year follow-up. Sixty-seven percent of CKD upstage patients had AKI 48 h after surgery and 11% after 2 months. All 15 (16.5%) patients with CKD had postoperative AKI. With IOH, OR 1.07, 95% CI 1.03−1.10 and p < 0.001, postoperative NLR after 48 h (OR 1.50, 95% CI 1.19−1.88, p < 0.001) was the major risk factor of AKI. In multivariate logistic regression analysis, the kidney's resected part volume (OR 1.08, 95% CI 1.03−1.14, p < 0.001) and IOH (OR 1.10, 95% CI 1.04−1.15, p < 0.001) were retained as statistically significant prognostic factors for detecting postoperative renal dysfunction. The independent risk factor for clinically significant postoperative AKI was only IOH (OR, 1.06; p < 0.001). Only AKI with the CKD upstage group has a statistically significant effect (p < 0.0001) on eGFR 6 and 12 months after surgery. Conclusions: The presence of AKI after PN is not rare. IOH and NLR are associated with postoperative AKI. The most important predictive factor of postoperative AKI is an NLR of over 3.5. IOH is an independent risk factor for clinically significant postoperative AKI and together with kidney resected part volume effects postoperative renal dysfunction. Only clinically significant postoperative AKI influences the reduction of postoperative eGFR after 6 and 12 months.

Toward a SERS Diagnostic Tool for Discrimination between Cancerous and Normal Bladder Tissues via Analysis of the Extracellular Fluid.

Vibrational spectroscopy provides the possibility for sensitive and precise detection of chemical changes in biomolecules due to development of cancers. In this work, label-free near-infrared surface enhanced Raman spectroscopy (SERS) was applied for the differentiation between cancerous and normal human bladder tissues via analysis of the extracellular fluid of the tissue. Specific cancer-related SERS marker bands were identified by using a 1064 nm excitation wavelength. The prominent spectral marker band was found to be located near 1052 cm-1 and was assigned to the C-C, C-O, and C-N stretching vibrations of lactic acid and/or cysteine molecules. The correct identification of 80% of samples is achieved with even limited data set and could be further improved. The further development of such a detection method could be implemented in clinical practice for the aid of surgeons in determining of boundaries of malignant tumors during the surgery.

Factors influencing extracorporeal shock wave lithotripsy efficiency for optimal patient selection.

INTRODUCTION: Current literature suggests various predictors related to the stone and patient, which could influence stone fragmentation and clearance rates. AIM: To establish clinical characteristics of stone disease for patients undergoing extracorporeal shockwave lithotripsy (ESWL) which may predict the success of the procedure. MATERIAL AND METHODS: One hundred and nine patients with renal stone disease diagnosed by non-contrast computed tomography (NCCT) who underwent ESWL between January 2015 and December 2019 were included in the study. Endpoints: patient being stone free (SF) or when < 4 mm fragments were detected. Age, gender, location, skin-to-stone distance, maximum stone length, stone volume, stone surface area, mean stone Hounsfield units (HU) and highest HU score were explored in uni- and multivariate regression analysis. RESULTS: Stone size revealed the highest prognostic power for ESWL failure, where OR for stone volume and stone surface area were 1.06 (1.03-1.10) and 1.04 (1.02-1.06), respectively (all p < 0.01) while a tendency was observed for skin-to-stone distance 1.02 (1.00-1.03). The amount of energy applied during the procedure to one cubic millimeter of stone volume (SMLI/stone volume) was predictive for treatment success (OR = 0.60, 95% CI: 0.41-0.87, p < 0.01). Stone volume (OR = 1.06, 95% CI: 1.00-1.14, p = 0.01) and stone surface area (OR = 1.03, 95% CI: 1.01-1.06, p = 0.02) remained as statistically significant prognostic factors for treatment failure. CONCLUSIONS: Both greater stone volume and stone surface area, as well as lower power delivered per stone volume unit during the ESWL procedure, were found to be significant factors and could be useful to predict treatment failure.

Complications after male adjustable suburethral sling implantation.

INTRODUCTION: Argus suburethral sling implantation is a minimally invasive operation with the possibility to adjust the tension of the sling at any time after the procedure, which provides good treatment results for male stress urinary incontinence (SUI). AIM: To determine the predictive factors, the incidence, severity and timing of the onset of complications after Argus sling implantation for males with post-operative SUI. MATERIAL AND METHODS: A total of 41 patients who underwent Argus sling implantation due to post-operative SUI were included. Median follow-up was 12 months. All complications were captured and graded according to severity and classified by timing of onset. Logistic regression analysis was performed to identify predictors of the most common side effects. RESULTS: Overall 22 (54%) of 41 males have experienced 31 complications. Three (7%) patients have experienced only intra-operative, 16 (39%) patients only post-operative and 3 (7%) patients both intra-operative and post-operative complications. The most common intra-operative complications were bladder perforation (12%) and external iliac vein injury (5%), while post-operative complications were acute urinary retention (29%), infection (10%) and perineal pain (7%). Previous radiotherapy has significantly increased the risk of intra-operative complications, while a non-significant tendency was observed for younger age, previous androgen deprivation therapy and grade 3 SUI. In terms of severity, most post-operative complications were classified as grade 3 according to the modified Clavien-Dindo system. CONCLUSIONS: Argus sling implantation provides a tolerable complication rate, where acute urinary retention was the most common side effect. Previous radiotherapy significantly increases the risk of serious intra-operative complications.

Successful laparoscopic living donor nephrectomy: first experience in Lithuania.

BACKGROUND: The aim of this paper is to share the initial results of LLDN in high-volume university centre that is performing laparoscopic nephrectomies for other indications. MATERIALS AND METHODS: During 2017, four LLDNs were performed. The transperitoneal approach was used in all cases and the kidney was removed using a suprapubic incision. All donors and recipients were prospectively analysed within six-month follow-up. The patients' clinical, laboratory, and operation-related data were collected from direct interviews with them and from medical records. All patients signed written informed consent. RESULTS: One male and three females donated their left kidneys by using the  LLDN technique. The mean age was 58 ±  9 years; two of them with a history of previous cholecystectomy. All donated kidneys had a single renal artery and renal vein. Pre-operative average eGFR was 94.2 ±  7.1 ml/min/1.73  m2, immediately after LLDN 57.5 ± 10.3 ml/min/1.73 m2, after one month 56.0 ± 9.1 ml/min/1.73 m2. There were no intraoperative complications; surgery duration was 223.75 ± 21.74 min, the cold ischemia time was 77.5 ± 28.77 min, and the warm ischemia time 6.37 ± 3.14 min. There was one postoperative donor complication, one case of acute kidney injury, and one case of prolonged postoperative abdominal pain. The only recipient complication was one case of acute kidney rejection; there were no cases of delayed graft function. CONCLUSIONS: Our initial experience confirms that LLDN is an approach that is easy to learn, especially in a high-volume university hospital with expertise in performing laparoscopic nephrectomies for other indications.

Sulforaphane decreases kidney injury after transplantation in rats: role of mitochondrial damage.

BACKGROUND: Sulforaphane is a naturally occuring antioxidative and anti-inflammatory isothiocyanat. In this study, its impact on experimental kidney transplantation was evaluated. MATERIAL AND METHODS: Male Brown Norway rats (n=112) were used as experimental animals. Donor kidneys were harvested and stored for 12 hours in HTK-solution at 4°C. D,L-Sulforaphane (4.4 mg/kg BW; 0.2ml) or normal saline (0.2 ml) was given i.v. to the recipients 24 and 1 hour before, and 6 hours after transplantation. Recipients were nephrectomized bilaterally and subsequently transplantation was performed. After 6 and 48 hours, biopsies were taken and processed for light and electron microscopy. Graft function was monitored using serum values of creatinine and BUN after 6 and 24 hours. Quantitative real-time PCR was used to detect differences in SOD2-gene expression after 6 hours and apoptotic activity was detected after 6 hours using propidium iodide flow cytometry. RESULTS: Recipient preconditioning improved reperfusion damage index from 12.8±1.6 in controls to 8.8±1.8 (p<0.001). Serum levels of creatinine and BUN decreased from 4.29±0.25 mg/dl and 119±23 mg/dl in controls to 3.65±0.7 mg/dl and 81±19 mg/dl (p<0.05). The number of severely injured tubules decreased (p<0.05). Apoptotic activity was increased in SFN-treated rats. Mitochondrial microstructure was better preserved after SFN, while SOD 2 gene expression increased (p<0.05). CONCLUSIONS: SFN ameliorates ischemia/reperfusion injury after KTx, most likely through anti-oxidative effects.

The use of high-dose melatonin in liver resection is safe: first clinical experience.

Experimental data suggest that melatonin decreases inflammatory changes after major liver resection, thus positively influencing the postoperative course. To assess the safety of a preoperative single dose of melatonin in patients undergoing major liver resection, a randomized controlled double-blind pilot clinical trial with two parallel study arms was designed at the Department of General and Transplantation Surgery, Ruprecht-Karls-University, Heidelberg. A total of 307 patients, who were referred for liver surgery, were screened. One hundred and thirteen patients, for whom a major liver resection (≥3 segments) was scheduled, were eligible. Sixty-three eligible patients refused to participate, and therefore, 50 patients were randomized. A preoperative single dose of melatonin (50 mg/kg BW) dissolved in 250 mL of milk was administered through the gastric tube after the intubation for general anesthesia. Controls were given the same amount of microcrystalline cellulose. Primary endpoint was safety. Secondary endpoints were postoperative complications. Melatonin was effectively absorbed with serum concentrations of 1142.8 ± 7.2 ng/mL (mean ± S.E.M.) versus 0.3 ± 7.8 ng/mL in controls (P < 0.0001). Melatonin treatment resulted in lower postoperative transaminases over the study period (P = 0.6). There was no serious adverse event in patients after melatonin treatment. A total of three infectious complications occurred in either group. A total of eight noninfectious complications occurred in five control patients, whereas three noninfectious complications occurred in three patients receiving preoperative melatonin (P = 0.3). There was a trend toward shorter ICU stay and total hospital stay after melatonin treatment. Therefore, a single preoperative enteral dose of melatonin is effectively absorbed and is safe and well tolerated in patients undergoing major liver surgery.