Bleeding disorder of unknown cause (BDUC) is a diagnosis of exclusion after evaluation of plasma coagulation and platelet function. Patients with BDUC (n = 375) recorded in the Vienna Bleeding Biobank were analyzed in comparison with healthy controls (HCs; n = 100) in this case-control study. Plasmin generation (PG) parameters were analyzed using calibrated fluorescence detection in citrated plasma. Turbidimetric plasma clot formation/lysis of 293 (78%) patients with BDUC and confocal microscopy of clots from representative patients with BDUC (n = 6) and HCs (n = 9) were assessed. In the PG analysis, patients with BDUC exhibited lower velocity and peak plasmin levels but a higher endogenous plasmin potential than HCs. Peak plasmin levels correlated with maximum clot absorbance but not with clot lysis time. Clot absorbance is an indicator of clot fiber density. Confocal microscopy analysis revealed a tendency towards thicker fibers in clots of patients with BDUC, which negatively correlated with peak plasmin (r = −0.561; P = .030). Peak plasmin correlated weakly with factor XIII, but not with other fibrinolytic factors (alpha2-antiplasmin, thrombin activatable fibrinolysis inhibitor, or plasminogen activator inhibitor 1) or bleeding severity. A model comprising fibrinogen and parameters of PG yielded high predictive power in discriminating between patients with BDUC and HCs across a fivefold stratified cross validation (80% of data; mean area under the curve [AUC], 0.847). The model generalized well to unseen data (20% of data; AUC, 0.856). Overall, patients with BDUC counterintuitively exhibited reduced peak plasmin levels, potentially related to altered clot structure.

Substaging of T1 urothelial cancer is associated with tumor progression and its reporting is recommended by international guidelines. However, it has not been integrated in risk stratification tools and there is no agreement on the best method to use for its reporting. We aimed to investigate the applicability, interobserver variability, and prognostic value of histological landmark based and micrometric (aggregate linear length of invasive carcinoma (ALLICA), microscopic vs. extensive system, Rete Oncologica Lombarda (ROL) system) substaging methods. A total of 79 patients with the primary diagnosis of T1 urothelial cancer treated with conventional transurethral resection and adjuvant BCG therapy between 2000 and 2020 at the Medical University of Vienna were included. The anatomical and metrical substaging systems were evaluated using agreement rate, Cohen’s kappa, Kendall’s tau, and Spearman rank correlation. Prognostic value for high-grade recurrence or T2 progression was evaluated in uni- and multivariable analysis. Applicability and reproducibility were good to moderate and varied between substaging methods. Obstacles are mainly due to fragmentation of samples. Anatomical substaging was associated with progression in univariable and multivariable analysis. In our cohort, we could only identify anatomical landmark–based substaging to be prognostic for T2 progression. A major obstacle for proper pathological assessment is fragmentation of samples due to operational procedure. Avoiding such fragmentation might improve reproducibility and significance of pathological T1 substaging of urothelial cancer.

Rapid microscopic analysis of tissue is an essential diagnostic tool in oncological surgery. The gold standard for intraoperative histological tissue evaluation is frozen sections. However, frozen sections are prone to a variety of artefacts and require skilled staff and specialized lab equipment. A potential method for rapid intraoperative tissue evaluation that does not require fixation, freezing, or sectioning of the tissue is ex vivo fluorescence confocal microscopy (FCM). The visualization of the structurally important extracellular matrix (ECM) in conventional ex vivo FCM lags behind the standards of conventional histology. The objective of this study was to find a stain that would improve the depiction of the ECM to resemble FFPE H&E sections as closely as possible. Eleven different tissue stains were tested on 122 tissue samples submitted to the Department of Pathology at the Medical University of Vienna. This study was conducted on the RS-G4 Upright (Caliber I.D. Rochester, NY, USA, distributed in Europe by MAVIG GmbH, Munich, Germany). Fast Green FCF (FGFCF) in combination with acridine orange as a nuclear stain improved the visibility of the structural details of the ECM. Morphological details in FCM were equivalent or even superior to frozen sections in most analyzed categories. The addition of FGFCF to the conventional staining protocol improves the assessment of the ECM and analysis of fibrosis. The rapid staining protocol is compatible with an application in intraoperative microscopy.

BACKGROUND: Arterial hypertension (HTN) and type 2 diabetes (T2DM) are contributors to chronic kidney disease leading to glomerulomegaly and podocyte loss. Enlarged glomeruli and podocyte depletion are associated with kidney disease progression. This retrospective study aimed to investigate morphometric changes in patients with HTN and the contribution of coexisting T2DM in HTN. METHODS: Glomerular and podocyte structure was estimated stereologically in unaffected areas of tumor nephrectomies in 99 patients. Morphometric features between subjects with HTN (n=47), HTN+T2DM (n=32), and controls without HTN and T2DM (n=20) were compared by ANOVA. Linear regression models evaluated the effect of morphometric parameters on renal compensation after nephrectomy (change of estimated glomerular filtration rate from pre-nephrectomy to 12 months post-nephrectomy). RESULTS: In total, 36% of the HTN and 50% of the patients with T2DM exhibited dipstick-positive proteinuria. Glomerular volume in HTN+T2DM was similar compared with controls (2.7±0.8 versus 2.3±0.8×10 ⁶ µm³; P =0.16) and to HTN only (2.7±0.9×10 ⁶ µm³; P =0.95). Podocyte density was reduced in HTN+T2DM versus controls (217±66 versus 279±75 per 10 ⁶ µm³; P =0.02) and not different to HTN only (233±87 per 10 ⁶ µm³; P =0.67). Podocyte nuclear volume was larger in HTN+T2DM versus controls (230±28 versus 201±32 µm³; P =0.004) and similar to HTN (221±33 µm³; P =0.39). Larger glomerular volume ( P =0.009), reduced podocyte density ( P =0.003), and nuclear hypertrophy ( P =0.01) were associated with impaired compensation. CONCLUSION: The patterns of podocyte depletion and nuclear hypertrophy were independently observed in both HTN and HTN+T2DM versus controls. Coexisting T2DM exhibited no additional contribution to glomerular and podocyte alterations.

Rapid microscopic analysis of tissue is an essential diagnostic tool in oncological surgery. The gold standard for intraoperative histological tissue evaluation are frozen sections. However, frozen sections are prone to a variety of artefacts and require skilled staff and specialized lab equipment. A potential method for rapid intraoperative tissue evaluation that does not require fixation, freezing nor sectioning of the tissue, is ex vivo fluorescence confocal microscopy (FCM). The visualization of the structurally important extracellular matrix (ECM) in conventional ex vivo FCM lags behind the standards of conventional histology. The objective of this study was to find a stain that will improve the depiction of the ECM to resemble FFPE H&E sections as closely as possible.