Whole-mount Sections Research Articles

The Status of Liver Margin in Perihilar Cholangiocarcinoma: A Multicenter Study.

This study aimed to investigate the prevalence and distribution of carcinoma in the liver margin (LM) of resected perihilar cholangiocarcinoma (pCCA) and establish a method for LM examination. LM is the largest margin in resected pCCA with undefined status and assessment method. 227 pCCA cases underwent major hepatectomy were divided into a discovery cohort (n=101) assessed using serial whole-mount digital large sections (WDLS) combined with small sections, and a control cohort (n=126) assessed using only small sections. The LM R1 resection rate was 38.6% (39/101) in the discovery cohort and 5.6% (7/126) in the control cohort. WDLS identified more LM R1 cases compared to the small section in the discovery cohort (38.6% vs. 5.9%, P<0.001). R0 patients in the discovery cohort had better overall survival and recurrence-free survival than those in the control cohort (both P<0.05). Additionally, 95% of carcinoma was found within 20mm of the proximal ductal margin (DM). A proximal DM distance of<5mm was an independent risk factor for LM R1 resection. Patients with which are more likely to experience R1 compared to those with ≥ 5mm (P<0.001). Positive LM was the significant cause for R1 resection of pCCA and the utilization of WDLS improved the diagnostic accuracy of LM. An examination methodology was established, highlighting the necessity of examining LM within a 20mm radius around the proximal DM, especially in patients with a proximal DM of<5mm.

A comparison of whole-mount and conventional sections for pathological mesorectal extension and circumferential resection margin assessment after total mesorectal excision

IntroductionThe objective of the study is to compare 2 techniques for histological handling of rectal cancer specimens, namely whole-mount in a large block vs conventional sampling using small blocks, for mesorectal pathological assessment of circumferential resection margin status and depth of tumor invasion into the mesorectal fat. MethodsThis is a prospective study including 27 total mesorectal excision specimens of rectal cancer from patients treated for primary rectal carcinoma between 2020 and 2022 in a specialized multidisciplinary Colorectal Unit. For each total mesorectal excision specimen, 2 contiguous representative tumoral slices were selected and comparatively analyzed with whole-mount and small blocks macroscopic dissection techniques, enabling comparison between them in the same surgical specimen. The agreement between the 2 techniques to assess the distance of the tumor from the circumferential resection margin as well as the depth of tumor invasion was evaluated with the Student’s t-test for paired samples, Pearson’s correlation coefficient, and the Bland-Altman method comparison analysis. ResultsComplete mesorectal excision was observed in 8% of cases. Circumferential resection margin involvement was observed in only one case (4 %). The whole-mount and small block techniques obtained similar results when we assessed the distance to the circumferential resection margin (t-test P = 0.8, r = 0.92) and the depth of mesorectal infiltration (t-test P = 0.6, r = 0.95). ConclusionsBoth gross dissection techniques (whole-mount vs multiple small cassettes) are equivalent and reliable to assess the distance to circumferential resection margin and the depth of mesorectal infiltration in the mesorectal fat in rectal cancer staging.

Clinical High-Resolution Imaging of the Inner Ear by Using Magnetic Resonance Imaging (MRI) and Cone Beam Computed Tomography (CBCT).

Imaging of the delicate inner ear morphology has become more and more precise owing to the rapid progress in magnetic resonance imaging (MRI). However, in clinical practice, the interpretation of imaging findings is hampered by a limited knowledge of anatomical details which are frequently obscured by artifacts. Corresponding review articles are as rare in journals as they are in reference books. This shortness prompted us to perform a direct comparison of imaging with anatomical whole-mount sections as a reference. It was the intention of this paper to compare the microscopic anatomy of a human inner ear as shown on anatomical whole-mount sections with high-resolution MRI and cone beam computed tomography (CBCT). Both are available in clinical routine and depict the structures with maximum spatial resolution. It was also a goal of this work to clarify if structures that were observed on MRI in a regular manner correlate with factual inner ear anatomy or correspond with artifacts typical for imaging. A fresh human anatomical specimen was examined on a clinical 3-Tesla MRI scanner using a dedicated surface coil. The same specimen was then studied with CBCT. In each imaging modality, high-resolution 3D data sets which enabled multiplanar reformatting were created. In the second step, anatomical whole-mount sections of the specimen were cut and stained. This process enabled a direct comparison of imaging with anatomical conditions. Clinical MRI was able to depict the inner ear with remarkable anatomical precision. Strongly T2-weighted imaging protocols are exquisitely capable of showing the fluid-filled components of the inner ear. The macular organs, ampullar crests and cochlear aqueduct were clearly visible. Truncation artifacts are prone to be confused with the delicate membrane separating the endolymphatic from the perilymphatic compartment. However, it was not possible to directly depict this borderline. With the maximum resolution of magnetic resonance tomography, commonly used in everyday clinical practice, even the smallest details of the inner ear structures can be reliably displayed. However, it is important to distinguish between truncation artifacts and true anatomical structures. Therefore, this study can be useful as a reference for image analysis.

Index tumor location affected early biochemical recurrence after radical prostatectomy in patients with negative surgical margin: a retrospective study

BackgroundIndex tumors are the most aggressive tumors of the prostate. However, their clinical significance remains unclear. This study aimed to assess the incidence of index tumor location according to the zonal origin and whether these locations affect the prognosis after radical prostatectomy in patients with negative surgical margins.MethodsThis single-centered, retrospective study evaluated 1,109 consecutive patients who underwent radical prostatectomies. An index tumor was defined as the largest tumor in the prostate gland. We detected these locations based on McNeal's zonal origin using whole-mount sections. Biochemical recurrence (BCR) free survival curves were generated using the Kaplan–Meier method. Univariate and multivariate analyses using the Cox proportional hazards model were performed to determine the predictive factors for early BCR (within 1-year).ResultsA total of 621 patients with negative surgical margins who did not receive adjuvant therapy were included in this study. The index tumor were located in the transitional zone in 191 patients (30.8%), the peripheral zone in 399 patients (64.3%), and the central zone in 31 patients (5.0%). In total, 22 of 621 patients (3.5%) experienced early BCR and 70 patients (11.2%) experienced overall BCR at a median follow-up of 61.7 months. According to the index tumor location, the early BCR-free rates were 99.5%, 95.7 %, and 83.3% in the transitional, peripheral, and central zones, respectively. On multivariate analysis, the index tumor in the central zone was an independent predictor of early BCR with negative surgical margins following radical prostatectomy, followed by prostatectomy pathological grade, index tumor in the peripheral zone, and high prostate-specific antigen level.ConclusionsWe assessed the significance of index tumor location in patients with negative surgical margins following radical prostatectomy. Index tumors located in the central zone, although infrequent, were the strongest predictive factors for early BCR. Our results may allow urologists and patients to reconsider the therapeutic strategies for prostate cancer.

(057) Destabilising Evidence: A Micro-Anatomical Investigation of Penile Suspensory Ligaments (PSL) Using Epoxy Sheet Plastination and Confocal Microscopy

Abstract Introduction Precise anatomy of the PSL remains controversial. It is routinely described as having 3 components: The fundiform (FL), Suspensory (SL) and Arcuate Ligaments (AL). Proponents of penile enlargement surgery (PES) ensure that division of the “superficial” PSL (FL, SL), allows desired distraction between phallus and pubic symphysis (PS) without risking instability. Seemingly in support of this rationale, many publications imply the FL and SL provide less structural integrity as they connect only superficial fascias (Origin: Scarpa’s fascia or linea alba; Insertion: Penile Dartos or Buck’s fascia). AL is reported to be the primary support structure as it connects the corporal tunica albuginea (TA) directly to the bone and preserved in PES given its deeper location. Despite this, significant sexual dysfunction (SD) from PSL deficiency after PES has been reported. An improved understanding of this anatomy is paramount for quality surgical care in both pre-operative counselling and rectification of adverse functional outcomes. Objective (1) To present initial findings of the in-situ PSL anatomy (2) To propose a new anatomical classification system Methods Data from six male cadavers (46-87 years; mean 75) were used in present study. There was no history of genitourinary operations and cause of death were either respiratory or vascular. Following deep-freeze and 0.9mm whole-mount sectioning, specimens were impregnated with epoxy-resin mixture (E12/E1/AE10/AE30). Without specific labelling, this process allows endogenous autofluorescence of collagen/elastin/myofilaments when excited using 488nm-confocal-laser microscope. Select areas were examined using stereoscopic microscopy and scanned at high-resolution. This technique ensures preservation of anatomy in-situ and allowed enhanced examination at both macroscopic and microscopic levels (Figure 1). Results Proposed classification divides PSL by anatomical location: Prepubic, Subpubic, Retropubic. While corresponding to the same distinct structures, it avoids the misnomers and discrepancies that are embedded in existing nomenclature. Findings are summarised in Table 1. The AL does not appear to the PSL apparatus. It fuses with the interosseus fibrocartilaginous lamina deep to it and attaches laterally to the inferior pubic rami. Inferiorly, its semicircular free edge rounds off the apex of the pubic arch but has no corporal attachment. Conclusions Findings of the present study do not support an anatomical rationale for PES, without risking instability. Contrary to previous reports, the subpubic PSL (or suspensory ligament) is the only component that directly connects TA with PS. The purported role of AL appears unfounded. Furthermore, perpendicular fibrous orientation of the subpubic and retropubic PSL suggests a dynamic support mechanism, not dissimilar to the knee’s cruciate ligaments. To the author’s knowledge, this is the first description of the musculotendinous origin of the retropubic PSL, whose fibres extend anteriorly from the urogenital diaphragm. Further studies are required to better characterise the retropubic PSL insertion and may lead to new insights into the origin and architecture of deep penile fascia. Based on present findings, the TA likely consists of multiple fibrous components originating from the musculotendinous aponeuroses of superficial perineal musculature. Disclosure No.

A comparison of whole-mount and conventional sections for pathological mesorectal extension and circumferential resection margin assessment after total mesorectal excision

IntroductionThe objective of the study is to compare 2 techniques for histological handling of rectal cancer specimens, namely whole-mount in a large block vs conventional sampling using small blocks, for mesorectal pathological assessment of circumferential resection margin status and depth of tumor invasion into the mesorectal fat. MethodsThis is a prospective study including 27 total mesorectal excision specimens of rectal cancer from patients treated for primary rectal carcinoma between 2020 and 2022 in a specialized multidisciplinary Colorectal Unit. For each total mesorectal excision specimen, 2 contiguous representative tumoral slices were selected and comparatively analyzed with whole-mount and small blocks macroscopic dissection techniques, enabling comparison between them in the same surgical specimen. The agreement between the 2 techniques to assess the distance of the tumor from the circumferential resection margin as well as the depth of tumor invasion was evaluated with the Student’s t-test for paired samples, Pearson’s correlation coefficient, and the Bland-Altman method comparison analysis. ResultsComplete mesorectal excision was observed in 8% of cases. Circumferential resection margin involvement was observed in only one case (4 %). The whole-mount and small block techniques obtained similar results when we assessed the distance to the circumferential resection margin (t-test P = 0.8, r = 0.92) and the depth of mesorectal infiltration (t-test P = 0.6, r = 0.95). ConclusionsBoth gross dissection techniques (whole-mount vs multiple small cassettes) are equivalent and reliable to assess the distance to circumferential resection margin and the depth of mesorectal infiltration in the mesorectal fat in rectal cancer staging.

Full resolution reconstruction of whole-mount sections from digitized individual tissue fragments

Whole-mount sectioning is a technique in histopathology where a full slice of tissue, such as a transversal cross-section of a prostate specimen, is prepared on a large microscope slide without further sectioning into smaller fragments. Although this technique can offer improved correlation with pre-operative imaging and is paramount for multimodal research, it is not commonly employed due to its technical difficulty, associated cost and cumbersome integration in (digital) pathology workflows. In this work, we present a computational tool named PythoStitcher which reconstructs artificial whole-mount sections from digitized tissue fragments, thereby bringing the benefits of whole-mount sections to pathology labs currently unable to employ this technique. Our proposed algorithm consists of a multi-step approach where it (i) automatically determines how fragments need to be reassembled, (ii) iteratively optimizes the stitch using a genetic algorithm and (iii) efficiently reconstructs the final artificial whole-mount section on full resolution (0.25 µm/pixel). PythoStitcher was validated on a total of 198 cases spanning five datasets with a varying number of tissue fragments originating from different organs from multiple centers. PythoStitcher successfully reconstructed the whole-mount section in 86–100% of cases for a given dataset with a residual registration mismatch of 0.65–2.76 mm on automatically selected landmarks. It is expected that our algorithm can aid pathology labs unable to employ whole-mount sectioning through faster clinical case evaluation and improved radiology-pathology correlation workflows.

A Stereological Approach to Quantify Immunohistochemical Staining in Zebrafish Larvae.

Immunohistochemistry (IHC) is a powerful research tool to localize specific antigens in whole-mount or tissue sections of embryos with labeled antibodies based on antigen-antibody interactions. Stereological methods are nowadays an essential tool to quantify cells or other types of structures in an unbiased and reproducible manner. In this chapter, a general protocol for a stereological estimation of the relative volume density of each structural component (Vv), which can be applied to any organ/structural component, will be described.

Predictors of Extraprostatic Extension in Patients with Prostate Cancer.

To identify effective factors predicting extraprostatic extension (EPE) in patients with prostate cancer (PCa). This retrospective cohort study recruited 898 consecutive patients with PCa treated with robot-assisted laparoscopic radical prostatectomy. The patients were divided into EPE and non-EPE groups based on the analysis of whole-mount histopathologic sections. Histopathological analysis (ISUP biopsy grade group) and magnetic resonance imaging (MRI) (PI-RADS v2.1 scores [1-5] and the Mehralivand EPE grade [0-3]) were used to assess the prediction of EPE. We also assessed the clinical usefulness of the prediction model based on decision-curve analysis. Of 800 included patients, 235 (29.3%) had EPE, and 565 patients (70.7%) did not (non-EPE). Multivariable logistic regression analysis showed that the biopsy ISUP grade, PI-RADS v2.1 score, and Mehralivand EPE grade were independent risk factors for EPE. In the regression assessment of the models, the best discrimination (area under the curve of 0.879) was obtained using the basic model (age, serum PSA, prostate volume at MRI, positive biopsy core, clinical T stage, and D'Amico risk group) and Mehralivand EPE grade 3. Decision-curve analysis showed that combining Mehralivand EPE grade 3 with the basic model resulted in superior net benefits for predicting EPE. Mehralivand EPE grades and PI-RADS v2.1 scores, in addition to basic clinical and demographic information, are potentially useful for predicting EPE in patients with PCa.

Q-VAT: Quantitative Vascular Analysis Tool.

As our imaging capability increase, so does our need for appropriate image quantification tools. Quantitative Vascular Analysis Tool (Q-VAT) is an open-source software, written for Fiji (ImageJ), that perform automated analysis and quantification on large two-dimensional images of whole tissue sections. Importantly, it allows separation of the vessel measurement based on diameter, allowing the macro- and microvasculature to be quantified separately. To enable analysis of entire tissue sections on regular laboratory computers, the vascular network of large samples is analyzed in a tile-wise manner, significantly reducing labor and bypassing several limitations related to manual quantification. Double or triple-stained slides can be analyzed, with a quantification of the percentage of vessels where the staining's overlap. To demonstrate the versatility, we applied Q-VAT to obtain morphological read-outs of the vasculature network in microscopy images of whole-mount immuno-stained sections of various mouse tissues.

Neuroprotective Effect of a Nutritional Supplement Containing Spearmint Extract, Forskolin, Homotaurine and Group B Vitamins in a Mouse Model of Transient Ocular Hypertension.

Glaucoma is one of the most common sight-threatening eye disorders and one of the main causes of irreversible blindness worldwide. The current therapies focusing on reducing intraocular pressure (IOP) are often insufficient to prevent the progression of the disease, so the therapeutic management of glaucoma remains a challenge. The aim of this study was to evaluate the neuroprotective, IOP-lowering independent effects of a nutritional supplement containing forskolin, homotaurine, spearmint extract and vitamins of the B group in a model of acute glaucoma developed in mice. Glaucoma was induced in adult wild-type C57BL/6J mice by transient elevation of IOP. The dietary supplement, branded as Gangliomix® (125 mg/kg/day), was administered by oral gavage for 17 days and ocular hypertension was induced on the 10th day of treatment. A histological analysis of the retinas was performed and RGC survival was evaluated with fluorogold labeling and Brn3a immunostaining on wholemount and retinal sections. Expression of alpha-spectrin, caspase-3, PARP-1 and GFAP was studied with western blotting or immunofluorescence. A significant increase in RGC survival was reported in the retina of mice treated with the dietary supplement as compared to vehicle-treated animals. The observed neuroprotection was associated with a calpain activity decrease, reduction in caspase-3 and PARP-1 activation, and prevention of GFAP upregulation. These effects were independent from the hypotensive effects of the supplement. Altogether, our data suggest that the dietary supplementation with forskolin, homotaurine, spearmint extract and vitamins of the B group supports RGC survival and may offer beneficial effects in glaucoma patients in combination with the currently used IOP-lowering therapy.

Development and characterization of an ETV1 rabbit monoclonal antibody for the immunohistochemical detection of ETV1 expression in cancer tissue specimens

BackgroundAberrant ETV1 overexpression arising from gene rearrangements or mutations occur frequently in prostate cancer, round cell sarcomas, gastrointestinal stromal tumors, gliomas, and other malignancies. The absence of specific monoclonal antibodies (mAb) has limited its detection and our understanding of its oncogenic function. MethodsAn ETV1 specific rabbit mAb (29E4) was raised using an immunogenic peptide. Key residues essential for its binding were probed by ELISA and its binding kinetics were measured by surface plasmon resonance imaging (SPRi). Its selective binding to ETV1 was assessed by immunoblots and immunofluorescence assays (IFA), and by both single and double-immuno-histochemistry (IHC) assays on prostate cancer tissue specimens. ResultsImmunoblot results showed that the mAb is highly specific and lacked cross-reactivity with other ETS factors. A minimal epitope with two phenylalanine residues at its core was found to be required for effective mAb binding. SPRi measurements revealed an equilibrium dissociation constant in the picomolar range, confirming its high affinity. ETV1 (+) tumors were detected in prostate cancer tissue microarray cases evaluated. IHC staining of whole-mounted sections revealed glands with a mosaic staining pattern of cells that are partly ETV1 (+) and interspersed with ETV1 (−) cells. Duplex IHC, using ETV1 and ERG mAbs, detected collision tumors containing glands with distinct ETV1 (+) and ERG (+) cells. ConclusionsThe selective detection of ETV1 by the 29E4 mAb in immunoblots, IFA, and IHC assays using human prostate tissue specimens reveals a potential utility for the diagnosis, the prognosis of prostate adenocarcinoma and other cancers, and the stratification of patients for treatment by ETV1 inhibitors.

Resolving the Pathogenesis of Anaplastic Wilms Tumors through Spatial Mapping of Cancer Cell Evolution.

While patients with intermediate-risk (IR) Wilms tumors now have an overall survival (OS) rate of almost 90%, those affected by high-stage tumors with diffuse anaplasia have an OS of only around 50%. We here identify key events in the pathogenesis of diffuse anaplasia by mapping cancer cell evolution over anatomic space in Wilms tumors. We spatially mapped subclonal landscapes in a retrospective cohort of 20 Wilms tumors using high-resolution copy-number profiling and TP53 mutation analysis followed by clonal deconvolution and phylogenetic reconstruction. Tumor whole-mount sections (WMS) were utilized to characterize the distribution of subclones across anatomically distinct tumor compartments. Compared with non-diffuse anaplasia Wilms tumors, tumors with diffuse anaplasia showed a significantly higher number of genetically distinct tumor cell subpopulations and more complex phylogenetic trees, including high levels of phylogenetic species richness, divergence, and irregularity. All regions with classical anaplasia showed TP53 alterations. TP53 mutations were frequently followed by saltatory evolution and parallel loss of the remaining wild-type (WT) allele in different regions. Morphologic features of anaplasia increased with copy-number aberration (CNA) burden and regressive features. Compartments demarcated by fibrous septae or necrosis/regression were frequently (73%) associated with the emergence of new clonal CNAs, although clonal sweeps were rare within these compartments. Wilms tumors with diffuse anaplasia display significantly more complex phylogenies compared with non-diffuse anaplasia Wilms tumors, including features of saltatory and parallel evolution. The subclonal landscape of individual tumors was constrained by anatomic compartments, which should be considered when sampling tissue for precision diagnostics.

MP40-11 CONTRIBUTION OF GLEASON PATTERN 4 PROSTATE CANCER TISSUE TO PRE-OPERATIVE PSA VALUES IN INTERMEDIATE RISK PROSTATE CANCER: RESULTS FROM 4 860 RP SPECIMENS

You have accessJournal of UrologyCME1 Apr 2023MP40-11 CONTRIBUTION OF GLEASON PATTERN 4 PROSTATE CANCER TISSUE TO PRE-OPERATIVE PSA VALUES IN INTERMEDIATE RISK PROSTATE CANCER: RESULTS FROM 4 860 RP SPECIMENS Fabian Falkenbach, Lukas Hohenhorst, Zhe Tian, Mykyta Kachanov, Tobias Maurer, Sophie Knipper, Markus Graefen, and Lars Budäus Fabian FalkenbachFabian Falkenbach More articles by this author , Lukas HohenhorstLukas Hohenhorst More articles by this author , Zhe TianZhe Tian More articles by this author , Mykyta KachanovMykyta Kachanov More articles by this author , Tobias MaurerTobias Maurer More articles by this author , Sophie KnipperSophie Knipper More articles by this author , Markus GraefenMarkus Graefen More articles by this author , and Lars BudäusLars Budäus More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003278.11AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Despite advances in imaging and genetic testing, blood PSA remains the cornerstone of prostate cancer (PCa) diagnosis and risk stratification. Especially in the management of intermediate-risk PCa, the PSA value has strong implications for clinical decision-making. It is known, that within this group Gleason pattern (GP) 4 drives biological aggressiveness. Aim of our study was to assess the relationship between pre-operative blood PSA values and volume of GP 4 compared to GP 3 and benign tissue in Radical Prostatectomy (RP) specimens. METHODS: Overall, 4860 patients with localized pT2pN0cM0, intermediate risk PCa without tertiary GP were assessed. Individual GPs were quantified at the whole-mount section analysis of RP specimen. For analysis, patients with biochemical recurrence after RP and a pre-operative PSA >50 ng/ml were further excluded for avoiding contamination of the cohort by occult metastasis. Moreover, patients with prior / neoadjuvant treatment, a gland size <10 cc after TUR-P or gland size >150 cc (associated risk of chronic inflammation) were excluded. Multivariable linear regression analysis addressed the relationship between volume of tissue types and PSA values. RESULTS: Of 4860 patients, in 4390 (90%) a Gleason Score of 3+4 and in 470 (10%) a Gleason Score of 4+3 was present. Median age at RP was 63 years (IQR: 58-68 years), median PSA was 6.4ng/ml (IQR: 4.8-8.7ng/ml) and median prostate volume was 30cc (IQR: 23-40cc). PSA as well as PSA-Density were associated with the quantitative GP 4 fraction above 50% (p-value: <0.001). GP 4 tissue contributed to the pre-operative PSA by 0.37 ng/ml per cc, GP 3 and benign prostate tissue combined contributed by 0.062 ng/ml per cc (p-value: <0.001). There was no significant difference in PSA contribution between GP 3 and benign tissue. CONCLUSIONS: The highest contribution to PSA prior RP originates from GP 4 PCa tumor foci. PSA expression of GP 3 and benign tissue were not significantly different. This confirms the discussed potential indolent nature of GP 3 PCa. Within intermediate risk PCa, PSA is a strong surrogate parameter for the amount of GP 4 PCa tissue. Further confirmatory studies including e.g. imaging data are needed for excluding potential selection bias. Source of Funding: None. © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e549 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Fabian Falkenbach More articles by this author Lukas Hohenhorst More articles by this author Zhe Tian More articles by this author Mykyta Kachanov More articles by this author Tobias Maurer More articles by this author Sophie Knipper More articles by this author Markus Graefen More articles by this author Lars Budäus More articles by this author Expand All Advertisement PDF downloadLoading ...

SMARCA4 and SMARCE1 in gastric cancer: Correlation with ARID1A, and microsatellite stability, and SMARCE1/ERBB2 co‐amplification

Recent studies have shown an association between certain subunits of the SWI/SNF complex with specific tumor characteristics in gastric cancer (GC). In an earlier study, we applied multiregional whole exome sequencing on multiple primary tumor samples and found alterations of the SWI/SNF complex in 78% of the cases. ERBB2, which encodes for Her2/neu, is a well-known predictive biomarker used to guide the treatment of GC in the palliative setting. SMARCE1, which encodes for a subunit of the SWI/SNF complex, is localized in close genetic proximity to ERBB2. As little is known about the significance of the SWI/SNF complex in GC biology and the potential relationship between ERBB2 and SMARCE1 upregulation, we examined the expression patterns of SMARCA4 and SMARCE1, two mutually exclusive catalytic ATPase subunits of the SWI/SNF complex, in a well characterized GC cohort. The expression of SMARCA4 and SMARCE1 was studied by immunohistochemistry in connection with clinicopathological patient characteristics in a cohort of 468 GCs. Digital droplet polymerase chain reaction was performed for amplification analysis on ERBB2 and SMARCE1. Immunohistochemical staining of whole-mount tissue sections found a diffusely "gray scale" expression of SMARCA4 in 446 (95.2%) GCs and of SMARCE1 in 463 (98.8%) GCs. The expression of SMARCA4 and SMARCE1 correlated significantly with ARID1A, p53, and microsatellite status. No correlation was found with the patient prognosis. The amplification analysis of SMARCE1 showed amplification in 4 of 34 cases. In 3 of 34 cases, SMARCE1 was co-amplified with ERBB2. We also found a co-expression of SMARCE1 and Her2/neu in a subset of patients. While the effect of a co-amplification is currently unknown, synergistic effects of SMARCE1 and Her2/neu overexpression should be explored in future studies, holding potential for an improved treatment of GC.

A pilot study of diabetic retinopathy in a porcine model of maturity onset diabetes of the young type 3 (MODY3)

Diabetic retinopathy (DR) is a leading cause of blindness in the working population. Because novel therapeutic intervention require testing, there is an urgent need for reliable animal models that faithfully replicate DR. Pig eyes have many similarities to human eyes anatomically and physiologically. Thus, attempts have been made to establish porcine models of DR by surgical, pharmaceutical or genetical induction of insulin deficiency, and dietary intervention. A previous study reported a transgenic pig model of maturity onset diabetes of the young type 3 (MODY3) developed signs of severe DR such as hemorrhage and proliferative tissue at the surface of the retina. However, the course of development of DR has not been studied in detail in this model. The purpose of this study was to investigate the early phase of DR in a MODY3. MODY3 and wild-type (WT) pigs underwent fundus photography and fluorescein angiogram (FA) before they developed cataracts. Animals were euthanized at age 1, 4, 7, and 10 months. Whole-mount retina and 10-μm thick paraffinized sections were stained with isolectin B4, and vessel density was determined by MATLAB software. At 4 and 7 months, retinal arterioles were immediately cannulated, and vasomotor action was measured by incubation with bradykinin and sodium nitroprusside. In the MODY3 pigs, fasting blood sugar levels gradually increased up to 500 mg/dL. Vascular tortuosity and yellowish spindle-shaped lesions were confirmed in MODY3 pigs at the age of 7 months; however, no microaneurysms were detected on FA. Compared with age-matched WT pigs, MODY3 pigs showed a significant decrease in blood vessel density in the intermediate and deep vascular plexus at 4 and 7 months of age and a slight decrease in capillary density in the superficial vascular plexus at 7 months of age. In MODY3 pigs, electron microscopy revealed thickening of the capillary basement membrane and leukostasis in the major blood vessels at 10 months of age. Bradykinin-induced dilation of retinal arterioles was diminished in MODY3 pigs as early as 7 months of age. Within 1 year after birth, MODY3 pigs show all typical early vascular lesions of diabetes except for microaneurysm formation. This pilot study suggests that the MODY3 pigs may serve as a suitable DR model to test effects of newly developed compounds on DR.

Potential use of high-resolution T2-weighted MRI with histopathologic findings in staging esophageal cancer

Magnetic resonance imaging (MRI) has shown promising capabilities in diagnosing local esophageal carcinoma. This study investigated the clinical value of high resolution (HR; small field of view and continuous thin section) axial T2-weighted MRI (HR-T2WI) as a noninvasive method for esophageal carcinoma tumor staging (T staging). Forty-two patients with biopsy-proven esophageal cancer were investigated using HR-T2WI. The discrepancies between the esophageal wall layers and tumor tissue were assessed for MRI T staging using a visual MRI signal intensity scale (low, intermediate, and high intensities). The computed tomography (CT) and MRI T staging was compared with whole-mount histopathological sections in all patients who underwent resection. HR-T2WI provided a thorough view of the esophageal wall and the tumor's anatomic layers. Of the 42 patients with histological tumors (HTs), there were 6 cases with tumors classified as HT-1a, 5 cases with HT-1b, 14 cases with HT-2, and 17 cases with HT-3/4, and their MRI T stages were 5 MRI-T1a, 6 MRI-T1b, 14 MRI-T2, and 17 MRI-T3/4, respectively. After analyzing the imaging presentation at different HT staginess, we found that HR-T2WI enabled a more accurate classification than was possible with CT. The difference in accuracy between CT and T2WI was statistically significant (P<0.05) in the entire sample and in HT1-2 tumors and HT3-4 tumors. HR-T2WI clearly identified normal esophageal wall layers; it had high diagnostic accuracy when evaluating tumor invasion and in MRI-T staging for esophageal carcinoma. This study established staging criteria of esophageal carcinoma using HR-T2WI and indicated that this approach could be used as a supplemental noninvasive method for the local T staging of esophageal carcinoma.

Exploring Hypoxia in Prostate Cancer With T2-weighted Magnetic Resonance Imaging Radiomics and Pimonidazole Scoring.

Radiomics involves high throughput extraction of mineable precise quantitative imaging features that serve as non-invasive prognostic or predictive biomarkers. High levels of hypoxia are associated with a poorer prognosis in prostate cancer and limit radiation therapy efficacy. Most patients with prostate cancer undergo magnetic resonance imaging (MRI) as a part of their diagnostics, and T2 imaging is the most utilised imaging method. The aim of this study was to determine whether hypoxia in prostate tumors could be identified using a radiomics model extracted from T2-weighted MR images. Eighty eight intermediate or high-risk prostate cancer patients were evaluated. Prior to radical prostatectomy, all patients received pimonidazole (PIMO). PIMO hypoxic scores were assigned in whole-mount sections from prostatectomy specimens by an experienced pathologist who was blinded to MRI. The region of interest used for radiomics analysis included the prostatic index tumor. Radiomics extraction yielded 165 features using a special evaluation version of RadiomiX [RadiomiX Research Toolbox version 20180831 (OncoRadiomics SA, Liège, Belgium)] for non-clinical use. Multivariable logistic regression with Elastic Net regularization was utilised using 10 times repeated 10-fold cross-validation to select the best model hyperparameters, optimizing for area under the receiver operating characteristic curve (AUC). The average (out of sample) performance based on the repeated cross validation using the ONESE model yielded an AUC of 0.60±0.2. Shape-based features were the most prominent in the model. The development of a radiomics hypoxia model using T2 weighted MR images, standard in the staging of prostate cancer, is possible.

Assessment of Prostate Carcinoma Aggressiveness: Relation to 68Ga-PSMA-11-PET/MRI and Gleason Score.

To test the correlation of 68Ga-PSMA-11 uptake and the expression of PSMA (prostatic specific membrane antigen) with the Gleason score, apparent diffusion coefficient (ADC) and pharmacokinetic parameters obtained from dynamic contrast agent-enhanced MRI/PET. Forty newly diagnosed, therapy naïve patients with prostatic carcinoma (PC) (mean age of 56.7, range=34-79), who were referred for 68Ga-PSMA-11-PET/MRI for primary staging and had undergone radical prostatectomy (RAPE) were included in this prospective study. Their blood samples were tested for serum levels of prostate-specific antigen (PSA) and proPSA. The patients' prostates were evaluated using whole-mount sections, which helped determine the extent and grade of the tumor; tests were performed to determine immunohistochemical PSMA expression. A correlation between PSMA expression and the accumulation of 68Ga-PSMA-11 was found using the Spearman correlation coefficient (p=0.0011). A stronger correlation was found between Gleason patterns 3 or 4 and PSMA expression (p=0.06). Furthermore, the correlation of Gleason score with the overall 68Ga-PSMA-11 accumulation within the tumor or non-tumor tissue was found to be significant (p=0.0157). A significant relation was found only with the Kep elimination rate constant, which was stronger in Gleason pattern 4 than in Gleason pattern 3. A weaker correlation was found between the accumulation of 68Ga-PSMA-11 and Ktrans in Gleason pattern 4: the most significant relation being between ADCmin and Gleason pattern 3 and 4 (p=0.0074). The total size of the tumor correlated with levels of proPSA (p<0.0001), and its extra prostatic extension correlated with levels of proPSA (p<0.0001). 68Ga-PSMA-11 correlates well with the expression of PSMA. Gleason pattern 3 and 4 had a higher correlation with 68Ga-PSMA-11 levels than did Gleason pattern 5. Either no correlation, or a weak correlation, was established with pharmacokinetics.

Early structural alterations of intrinsic cardiac ganglionated plexus in spontaneously hypertensive rats.

Persistent arterial hypertension leads to structural and functional remodeling of the heart resulting in myocardial ischemia, fibrosis, hypertrophy, and eventually heart failure. Previous studies have shown that individual neurons composing the intracardiac ganglia are hypertrophied in the failing human, dog, and rat hearts, indicating that this process involves changes in cardiac innervation. However, despite a wealth of data on changes in intrinsic cardiac ganglionated plexus (GP) in late-stage disease models, little is known about the effects of hypertension on cardiac innervation during the early onset of heart failure development. Thus, we examined the impact of early hypertension on the structural organization of the intrinsic cardiac ganglionated plexus in juvenile (8-9 weeks) and adult (12-18 weeks) spontaneously hypertensive (SH) and age-matched Wistar-Kyoto (WKY) rats. GP was studied using a combination of immunofluorescence confocal microscopy and transmission electron microscopy in whole-mount preparations and tissue sections. Here, we report intrinsic cardiac GP of SH rats to display multiple structural alterations: (i) a decrease in the intracardiac neuronal number, (ii) a marked reduction in axonal diameters and their proportion within intracardiac nerves, (iii) an increased density of myocardial nerve fibers, and (iv) neuropathic abnormalities in cardiac glial cells. These findings represent early neurological changes of the intrinsic ganglionated plexus of the heart introduced by early-onset arterial hypertension in young adult SH rats.