Benign Specimens Research Articles

Optimization of Fluorescence In Situ Hybridization Protocols in the Era of Precision Medicine.

Fluorescence in situ hybridization (FISH) is a cytogenetic assay that is widely used in both clinical and research settings to validate genetic aberrations. Simple in principle, it is based on denaturation and hybridization of a DNA probe and its complementary sequence; however, it is subject to continuous optimization. Here we share how in-house FISH can be optimized using different control tissues to visualize and ultimately validate common and novel genetic abnormalities unearthed by next-generation sequencing (NGS). Seven specific FISH probes were designed and labeled, and conditions for eight tissue types and one patient-derived tumor organoid were optimized. Formalin-fixed paraffin-embedded (FFPE) tissue slides were used for each experiment. Slides were first deparaffinized, then placed in a pretreatment solution followed by a digestion step. In-house FISH probes were then added to the tissue to be denatured and hybridized, and then washed twice. To obtain optimal results, probe concentration, pepsin incubation time, denaturation, and the two post-hybridization washes were optimized for each sample. By modifying the above conditions, all FISH experiments were optimized in separate tissue types to investigate specific genomic alterations in tumors arising in those tissues. Signals were clear and distinct, allowing for visualization of the selected probes. Following this protocol, our lab has quickly optimized 11 directly labeled in-house FISH probes to support genetic aberrations nominated by NGS, including most recent discoveries through whole-genome sequencing analyses. We describe a robust approach of how to advance in-house labeled FISH probes. By following these guidelines, reliable and reproducible FISH results can be obtained to interrogate FFPE slides from benign, tumor tissues, and patient-derived tumor organoid specimens. This is of most relevance in the era of NGS and precision oncology. © 2024 Wiley Periodicals LLC. Basic Protocol 1: Metaphase FISH optimization Support Protocol 1: In-house probe labeling and preparation Support Protocol 2: Metaphase spread preparation Basic Protocol 2: Optimization of FISH on formalin-fixed paraffin-embedded tissue.

Development and Potential Implementation of Endometrial Cancer Screening in a Poor-Prognosis Endemic Cancer Community [ID 2683642

INTRODUCTION: Endometrial cancer incidence and mortality both are increasing in the United States. Certain patient populations are disproportionately affected by this disease. Currently, there is no recommendation from any professional society to screen high-risk patients for premalignancy or cancer. Our patient population is composed of predominantly Black women, a group that experiences worse endometrial cancer outcomes than other demographics. A risk assessment nomogram may be able to be developed using data from this unique patient population that can identify women at high risk of developing endometrial disease. METHODS: Endometrial pathology specimens collected at a large safety-net hospital between the years of 2013 and 2022 were analyzed. A retrospective chart review was performed on this data, looking for the patient characteristics of race, age, body mass index (BMI), parity, hemoglobin A1C level, and tamoxifen use history at the time of specimen collection. Odds ratios were calculated for those variables individually as well as collectively to determine whether they could be used to predict risk of endometrial disease in a woman with some of all those characteristics. RESULTS: The odds ratios for age, BMI, and parity were statistically significant between benign and abnormal pathologic specimens. The area under the receiver operating characteristic curve was 0.76 when combining age, BMI, and parity. CONCLUSION: Our data suggest that at-risk patients can be reliably identified using simple characteristics. Strengths of our study include a unique patient population and large database. Future directions include refinement of the model and prospective application in the clinical setting.

Enhancing upper tract urothelial carcinoma diagnosis: Utility of cytokeratin 17 and CK20/CD44/p53 immunohistochemical panel

Upper tract urothelial carcinoma (UTUC) presents diagnostic challenges due to small biopsy specimen size, poor orientation, and technical obstacles that can yield equivocal diagnoses. This uncertainty often mandates repeated biopsies to evaluate the necessity of nephroureterectomy. Prior studies have suggested cytokeratin 17 (CK17) immunostain as an adjunctive tool for diagnosing bladder urothelial neoplasia in both urine cytology and tissue biopsy specimens. We evaluated the utility of CK17 in differentiating UTUC from benign urothelium and its ability to stratify low-grade from high-grade neoplasia. Our study involved a cohort of previously diagnosed cytology (n = 29) and tissue specimens from biopsies and resections (n = 85). We evaluated CK17 staining percentage in cytology and tissue samples and localization patterns in biopsy/resection samples. Our findings showed a statistically significant distinction (p < 0.05) between UTUC and benign tissue specimens based on full thickness localization pattern (odds ratio 8.8 [95% CI 1.53–67.4]). The percentage of CK17 staining failed to significantly differentiate neoplastic from non-neoplastic cases in cytology or tissue samples. Additionally, based on prior research showing the efficacy of CK20/CD44/p53 triple panel in bladder urothelial neoplasia, we utilized tissue microarrays to evaluate if these markers could distinguish UTUC from benign urothelium. We found that CK20/CD44/p53, individually or in combination, could not distinguish urothelial neoplasia from non-neoplasia. Full thickness CK17 urothelial localization by immunohistochemistry was highly reproducible with excellent interobserver agreement and may play a supplementary role in distinguishing upper tract urothelial neoplasia from benign urothelium.

Abstract 6825: Epigenetic dysregulation in well-differentiated pancreatic neuroendocrine tumor (PNET): An immunohistochemistry (IHC) analysis

Abstract Epigenetic dysregulation through DNA Methyltransferase 1 (DNMT1) is integral to pancreatic neuroendocrine tumorigenesis. We aim to investigate the interplay between epigenetic modifiers and pathogenetic markers in the PNET microenvironment. 12 paraffin-embedded PNET specimens and 12 benign adjacent tissue specimens, used as controls, were obtained from Cooper University Hospital Pathology Department (IRB 23-130). IHC was performed using epigenetic markers, including DNMT1, DNMT3A, DNMT3B, and 5-Hydroxymethylcytosine (5-HMC), and pathogenic markers, including Multiple Endocrine Neoplasia 1 (MEN1), Protein-Tyrosine Phosphatase (PTEN), and Ras Association Domain Family 1 (RASSF1). Image J software was utilized for IHC scoring under the guidance of a board-certified pathologist. DNMT1 was upregulated in PNET tumor specimens with a mean density score of 72.02 +/- 31 versus 16.02 +/- 17 in adjacent benign tissue (p&amp;lt;.001). Grade 2 tumors displayed increased DNMT1 expression compared to Grade 1 tumors with a mean score of 37 +/-21 versus 71 +/- 23 (p&amp;lt;.05). Decreased expression of 5-HMC was noted in tumor specimens in comparison to benign tissue with a mean score of 3.92 +/- 5.05 versus 19.37 +/- 5.05 (p&amp;lt;.05). Loss of 5-HMC was observed in higher tumor grades with a mean score of .79 +/- 2 versus 8.2+/-2 (p&amp;lt;.01). DNMT3B was upregulated in PNET tumor specimens with a mean score of 13.16 +/- 3.94 versus 7.02+/- 2.553 (p&amp;lt;.01) in adjacent benign tissue and was positively correlated to DNMT1 expression. DNMT3A was not found to have a difference in expression between tumor and benign tissue. MEN1 was downregulated in PNET, with 45% of tumor specimens displaying loss of MEN1 expression. Mean MEN1 density in tumor tissue was 4.47 +/- 5.95 versus 11.41 +/- 3.53 in adjacent benign (p&amp;lt;.01). Increased expression of DNMT1 found to negatively correlate with MEN1 expression (R2=.23). Loss of RASSF1 expression was present in 70% of tumor specimens with a mean score of 1.32 +/- 1.99 versus 9.50+/- 6.89 (p&amp;lt;.001) in adjacent benign tissue. RASSF1 expression correlated negatively with DNMT1 (R2=.21). PTEN expression showed intratumoral homogeneity to benign tissue, and its expression was not correlated with any epigenetic markers. Our work provides a framework of epigenetic regulators involved in the pathobiology of PNET, and can refine prognostic evaluation while providing therapeutic benefits for patients with PNET. Citation Format: Zena Saleh, Yazid Ghanem, Hansa Joshi, Mathew C. Moccia, Helen Toma, Yahui Li, Upsana Joneja, Tao Gao, Young Hong. Epigenetic dysregulation in well-differentiated pancreatic neuroendocrine tumor (PNET): An immunohistochemistry (IHC) analysis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6825.

Insulin-like Growth Factor II mRNA-binding Protein 3 is a Highly Sensitive Marker for Intravascular Large B-cell Lymphoma: Immunohistochemical Analysis of 152 Pathology Specimens From 88 Patients.

Intravascular large B-cell lymphoma (IVLBCL) is a rare type of aggressive extranodal large B-cell lymphoma characterized by the selective growth of lymphoma cells within the lumina of blood vessels, particularly capillaries. IVLBCL lacks mass formation, and its diagnosis can be challenging. We analyzed the utility of insulin-like growth factor II mRNA-binding protein 3 (IMP3) immunohistochemistry for IVLBCL diagnosis in various organs. Double staining with paired box 5 (PAX5) was performed for validation. Overall, 152 pathological specimens (111 positive and 41 negative for IVLBCL) obtained from 88 patients with a diagnosis of IVLBCL were stained for IMP3 and IMP3/PAX5. As negative controls, 40 pathology specimens from 38 patients with no history of IVLBCL or other B-cell lymphomas were stained for IMP3, which comprised 31 benign pathological specimens from 29 patients in whom malignancy was suspected, 7 cases of appendicitis with intravascular and/or intralymphatic lymphoid proliferations, and 2 cases of intravascular natural killer/T-cell lymphoma. All mononuclear cells with cytoplasmic staining were considered positive for IMP3 expression, but expression restricted to germinal center B cells was excluded from evaluation. All 111 IVLBCL pathological specimens were positive for IMP3 and IMP3/PAX5. In addition, 11 of the 41 specimens originally diagnosed as IVLBCL-negative showed IMP3/PAX5 double-positive cells, raising the suspicion of IVLBCL. However, of the 40 negative control samples, IMP3-positive non-germinal center B cells were detected in only 2 samples ( P = 0.0131) and no intravascular IMP3-positive B cells suspicious for IVLBCL were identified. Altogether, IMP3 immunohistochemistry is a highly sensitive marker of IVLBCL and can be a helpful adjunct for IVLBCL diagnosis.

Patterns of B-cell lymphocyte expression changes in pre- and post-malignant prostate tissue are associated with prostate cancer progression.

Inflammation characterized by the presence of T and B cells is often observed in prostate cancer, but it is unclear how T- and B-cell levels change during carcinogenesis and whether such changes influence disease progression. The study used a retrospective sample of 73 prostate cancer cases (45 whites and 28 African Americans) that underwent surgery as their primary treatment and had a benign prostate biopsy at least 1 year before diagnosis. CD3+, CD4+, and CD20+ lymphocytes were quantified by immunohistochemistry in paired pre- and post-diagnostic benign prostate biopsy and tumor surgical specimens, respectively. Clusters of similar trends of expression across two different timepoints and three distinct prostate regions-benign biopsy glands (BBG), tumor-adjacent benign glands (TAG), and malignant tumor glandular (MTG) regions-were identified using Time-series Anytime Density Peaks Clustering (TADPole). A Cox proportional hazards model was used to estimate the hazard ratio (HR) of time to biochemical recurrence associated with region-specific lymphocyte counts and regional trends. The risk of biochemical recurrence was significantly reduced in men with an elevated CD20+ count in TAG (HR = 0.81, p = 0.01) after adjusting for covariates. Four distinct patterns of expression change across the BBG-TAG-MTG regions were identified for each marker. For CD20+, men with low expression in BBG and higher expression in TAG compared to MTG had an adjusted HR of 3.06 (p = 0.03) compared to the reference group that had nominal differences in CD20+ expression across all three regions. The two CD3+ expression patterns that featured lower CD3+ expression in the BBG compared to the TAG and MTG regions had elevated HRs ranging from 3.03 to 4.82 but did not reach statistical significance. Longitudinal and spatial expression patterns of both CD3+ and CD20+ suggest that increased expression in benign glands during prostate carcinogenesis is associated with an aggressive disease course.

(261) Vasovasostomy Surgical Simulation Using En Bloc Orchiectomy Specimen

Abstract Introduction Surgical simulation with use of synthetic materials, animals, or human tissues is an essential part of surgical training. Prior vasovasostomy (VV) simulations have described use of silicone tubules, animal vasa, and vas segments from human prostatectomy specimen. Herein we describe a VV surgical simulation model using benign, en bloc human testicle and spermatic cord specimen acquired via gender-affirming simple orchiectomy. Objective To improve resident confidence in performing VV via a surgical wet lab using benign, en bloc human testicle and spermatic cord specimen acquired via gender-affirming simple orchiectomy. Methods En bloc testicle and spermatic cord specimen from benign orchiectomy procedures were stored in our institution’s tissue bank. After development of a tissue handling protocol, specimens were then transported to our surgical simulation center for use in a VV microsurgical surgical simulation exercise. Participants were asked to complete a six-throw VV anastomosis using specimen. The first three throws were “training naïve,” with no prior instruction and an 8-minute time cap. After the first three throws or 8 minutes of time, participants received 10 minutes of hands-on instruction from a fellowship-trained urologic microsurgeon, then completed the remaining three throws of their six-throw anastomosis. Performance was measured using a microsurgical skills assessment tool for each set of three stitches and compared (figure 1). Pre- exercise and post- exercise surveys (figure 2) were completed in which participants rated their confidence in performing VV. Two months later, the same participants returned for an interval skills assessment in which they were tasked with completing the six-throw anastomosis in a 16-minute time limit. Skills were again evaluated with the microsurgical skills assessment tool; pre- and post- exercise surveys were again completed. Results Four urology trainees participated in our exercise: two PGY2s, one PGY3 and one PGY4. Completed number of knots increased from the first 8-minute session to the final 8-minute session for all participants (table 1). Surgical performance as measured by the skills-assessment tool did not change as a result of the instruction intervention (table 1). Participant confidence in performing microsurgical VV improved between the pre- and post- exercise surveys (table 2). All participants felt the wet lab served as a realistic representation of real-life VV (table 3)). At the two-month reassessment time point, all participants completed a similar number of throws (+/- 1 throw) in an overall equal amount of time (16 consecutive minutes compared to 8 minutes twice) with similar surgical performance compared to initial assessment (table 4). Conclusions A microsurgical VV wet lab using en bloc benign orchiectomy specimen serves a realistic simulation of real-life VV and improves trainee confidence in performing this procedure. Real-time instruction likely provides less value for improving operative skill than does experience and repetitions alone. These findings support not only the use of human specimen for surgical training, but also the value of a gender-affirming surgery program which provides access to these benign specimens. Disclosure No.

ALDH1A1 drives prostate cancer metastases and radioresistance by interplay with AR- and RAR-dependent transcription.

Rationale: Current therapies for metastatic osseous disease frequently fail to provide a durable treatment response. To date, there are only limited therapeutic options for metastatic prostate cancer, the mechanisms that drive the survival of metastasis-initiating cells are poorly characterized, and reliable prognostic markers are missing. A high aldehyde dehydrogenase (ALDH) activity has been long considered a marker of cancer stem cells (CSC). Our study characterized a differential role of ALDH1A1 and ALDH1A3 genes as regulators of prostate cancer progression and metastatic growth. Methods: By genetic silencing of ALDH1A1 and ALDH1A3 in vitro, in xenografted zebrafish and murine models, and by comparative immunohistochemical analyses of benign, primary tumor, and metastatic specimens from patients with prostate cancer, we demonstrated that ALDH1A1 and ALDH1A3 maintain the CSC phenotype and radioresistance and regulate bone metastasis-initiating cells. We have validated ALDH1A1 and ALDH1A3 as potential biomarkers of clinical outcomes in the independent cohorts of patients with PCa. Furthermore, by RNAseq, chromatin immunoprecipitation (ChIP), and biostatistics analyses, we suggested the molecular mechanisms explaining the role of ALDH1A1 in PCa progression. Results: We found that aldehyde dehydrogenase protein ALDH1A1 positively regulates tumor cell survival in circulation, extravasation, and metastatic dissemination, whereas ALDH1A3 plays the opposite role. ALDH1A1 and ALDH1A3 are differentially expressed in metastatic tumors of patients with prostate cancer, and their expression levels oppositely correlate with clinical outcomes. Prostate cancer progression is associated with the increasing interplay of ALDH1A1 with androgen receptor (AR) and retinoid receptor (RAR) transcriptional programs. Polo-like kinase 3 (PLK3) was identified as a transcriptional target oppositely regulated by ALDH1A1 and ALDH1A3 genes in RAR and AR-dependent manner. PLK3 contributes to the control of prostate cancer cell proliferation, migration, DNA repair, and radioresistance. ALDH1A1 gain in prostate cancer bone metastases is associated with high PLK3 expression. Conclusion: This report provides the first evidence that ALDH1A1 and PLK3 could serve as biomarkers to predict metastatic dissemination and radiotherapy resistance in patients with prostate cancer and could be potential therapeutic targets to eliminate metastasis-initiating and radioresistant tumor cell populations.

Usability of serum hedgehog signalling proteins as biomarkers in canine mammary carcinomas

BackgroundThe hedgehog signalling pathway has been implicated in tumourigenesis and progression of many tumour types. This pathway has recently emerged as a therapeutic target, and inhibitors of hedgehog signalling have gained considerable attention. In dogs, the roles of hedgehog signals in several types of tumours have been investigated, but their relationship with canine mammary gland tumours (MGTs) has not been established. This study aimed to evaluate the expression of sonic hedgehog (SHH) and glioma-associated oncogene 1 (GLI-1) in the serum and mammary tumour tissues of dogs.ResultsSHH and GLI-1 protein expression levels were significantly higher in MGT tissues than in normal mammary gland tissues, as well as in malignant MGT specimens than in benign MGT specimens. Serum levels of SHH and GLI-1 were higher in MGT patients than in healthy controls (p < .001 and .001, respectively). Serum SHH level showed a statistically significant relationship with metastatic status (p = .01), and serum GLI-1 level showed a statistically significant relationship with histologic grade (p = 0.048) and metastatic status (p = 0.007). Serum hedgehog signalling protein levels were not significantly associated with breed size, sex, tumour size, or histologic type.ConclusionsHedgehog signalling protein expression in canine MGT tissue and serum differed according to the histological classification (benign and malignant) and metastatic status, indicating a relationship between the hedgehog signalling pathway and canine MGT. Thus, the hedgehog signalling pathway may serve as a new biomarker and therapeutic target in canine MGT patients.

Expression of the Calcium-Sensing Receptor on Normal and Abnormal Parathyroid and Thyroid Tissue

IntroductionCurrent imaging techniques have several limitations in detecting parathyroid glands. We have investigated the calcium-sensing receptor (CaSR) as a potential target for specifically labeling parathyroid glands for radiologic detection. For accurate imaging it is vital that a large differential expression exists between the target tissue and adjacent structures. We sought to investigate the relative abundance of the CaSR in normal and abnormal parathyroid tissue, as well as normal and abnormal thyroid. MethodsExisting clinical specimens were selected that represented a wide variety of pathologically and clinically confirmed malignant and benign thyroid and parathyroid specimens. Sections were stained for the CaSR using immunohistochemistry and scored for intensity and abundance of expression. (H score = intensity scored from 0 to 3 multiplied by the % of cells at each intensity. Range 0-300). ResultsAll parathyroid specimens expressed the CaSR to a high degree. Normal parathyroid had the highest H score (271, s.d. 25.4). Abnormal parathyroid specimens were slightly lower but still much higher than normal thyroid (H score 38.3, s.d. 23.3). Medullary thyroid cancer also expressed the CaSR significantly higher than normal thyroid (H score 182, s.d. 69.1, P < 0.001) but below parathyroid levels. Hürthle cell carcinoma expressed the CaSR to a lesser degree but higher than normal thyroid (H score 101, s.d. 46.4, P = 0.0037). ConclusionsThe CaSR is differentially expressed on parathyroid tissue making it a feasible target for parathyroid imaging. False positives might be anticipated with medullary and Hürthle cell cancers.

ACAA2 is a novel molecular indicator for cancers with neuroendocrine phenotype

BackgroundNeuroendocrine phenotype is commonly associated with therapy resistance and poor prognoses in small-cell neuroendocrine cancers (SCNCs), such as neuroendocrine prostate cancer (NEPC) and small-cell lung cancer (SCLC). Expression levels of current neuroendocrine markers exhibit high case-by-case variability, so multiple markers are used in combination to identify SCNCs. Here, we report that ACAA2 is elevated in SCNCs and is a potential molecular indicator for SCNCs.MethodsACAA2 expressions in tumour xenografts, tissue microarrays (TMAs), and patient tissues from prostate and lung cancers were analysed via immunohistochemistry. ACAA2 mRNA levels in lung and prostate cancer (PC) patients were assessed in published datasets.ResultsACAA2 protein and mRNA levels were elevated in SCNCs relative to non-SCNCs. Medium/high ACAA2 intensity was observed in 78% of NEPC PDXs samples (N = 27) relative to 33% of adeno-CRPC (N = 86), 2% of localised PC (N = 50), and 0% of benign prostate specimens (N = 101). ACAA2 was also elevated in lung cancer patient tissues with neuroendocrine phenotype. 83% of lung carcinoid tissues (N = 12) and 90% of SCLC tissues (N = 10) exhibited medium/high intensity relative to 40% of lung adenocarcinoma (N = 15).ConclusionACAA2 expression is elevated in aggressive SCNCs such as NEPC and SCLC, suggesting it is a potential molecular indicator for SCNCs.

Human parvovirus B19 infection in malignant and benign tissue specimens of different head and neck anatomical subsites

BackgroundThe role of human parvovirus B19 (B19V) infection in malignant and benign lesions such as head and neck squamous cell carcinomas (HNSCCs) and oral mucocele lesions has not been established. Herein, we examined, for the first time, the presence of B19V in HNSCCs from Iranian subjects.MethodsOne hundred and eight HNSCC specimens were analyzed for the presence of B19V using nested polymerase chain reaction (nPCR) and TaqMan quantitative PCR assays. Immunohistochemistry procedures were performed to evaluate the expression of B19V VP1/VP2 proteins, p16INK4a, and NF-κB in tumor tissues and their adjacent non-tumor tissues. In addition, 40 oral mucocele, 30 oral buccal mucosa swabs, and 30 nasopharyngeal swabs obtained from healthy adults were analyzed as controls.ResultsB19V DNA was detected in 36.1% of HNSCCs. Further, 23.3% of HNSCC specimens showed immunoreactivity against B19V VP1/VP2 proteins. There was a significant difference in the frequency of B19V DNA-positive cases between the patient and control groups (p < 0.0001). Moreover, comparing tumoral tissues and their adjacent non-tumor tissues in terms of immunoreactivity against B19V structural proteins, a significant association was found between tumor tissues and B19V infection (p < 0.0001). Finally, investigating the simultaneous presence of B19V and high-risk human papillomaviruses (HPV) DNA, we found a significant association between these two viral infections in HNSCCs (p = 0.031).ConclusionsTo sum up, B19V was frequently present in HNSCC tissues of Iranian patients but mostly absent in the adjacent non-tumor tissues as well as oral mucocele lesions, buccal, and nasopharyngeal swabs of healthy subjects. HPV possibly contributes to B19V persistence in HNSCC tissues. Additional research is required to investigate potential etiological or cofactor roles of B19V in the development of HNSCCs.

Venous Thromboembolism After Abdominal and Minimally Invasive Large Specimen Hysterectomy

To investigatethe incidence of venous thromboembolism (VTE) in patients undergoing large specimen hysterectomy for benign indications. To evaluate the impact of route of surgery and operative time in the development of VTE in this population. Retrospective cohort study (Canadian Task Force Classification II2) of targeted hysterectomy data prospectively collected from the American College of Surgeons National Surgical Quality Improvement Program involving over 500 hospitals across the United States SETTING: National Surgical Quality Improvement Program Database PATIENTS: Women aged 18 years or older undergoing hysterectomy for benign indications between 2014 and 2019. Patients were further classified into four groups according to uterine weight: <100g, 100-249g, 250g-499g, and specimens ≥500g. Current Procedural Terminology codes were used to identify cases. Variables including age, ethnicity, body mass index, smoking status, diabetes, hypertension, blood transfusion, and American Society of Anesthesiologists classification system scores were collected. Cases were stratified by route of surgery, operative time, and uterine weight. A total of 122,418 hysterectomies occurring between 2014-2019 were included in our study, of which 28,407 (23.2%) patients underwent abdominal, 75,490 (61.7%) laparoscopic, and 18,521 (15.1%) vaginal hysterectomy. The overall rate of VTE in patients with large specimen hysterectomies (≥500g) was 0.64%. After multivariable adjustment, there was no significant difference in the odds of VTE between uterine weight groups. Only 30% of the surgeries with uterine weight above 500g were performed with minimally invasive surgical routes. Patients who underwent minimally invasive hysterectomy had lower odds of VTE via laparoscopic (aOR 0.62; CI:0.48-0.81) and vaginal (aOR 0.46; CI:0.31-0.69) routes compared to laparotomy. Prolonged operative time (>120 min) was associated with an increased odds of VTE (aOR 1.86; CI:1.51-2.29). The occurrence of VTE after benign large specimen hysterectomy is rare. The odds of VTE is higher with longer operative times and lower with minimally invasive approaches even for markedly enlarged uteri.

The Future of Minimal-Access Myoma Surgery with In-Bag Contained Morcellation.

Contained electromechanical morcellation has emerged as a safety approach for laparoscopic myomatous tissue retrieval. This retrospective single-center analysis evaluated the bag deployment practicability and safety of electromechanical in-bag morcellation when used for big surgical benign specimens. The main age of patients was 39.3 years (range 21 to 71); 804 myomectomies, 242 supracervical hysterectomies, 73 total hysterectomies, and 1 retroperitoneal tumor extirpation were performed. A total of 78.7% of specimens weighed more than 250 g (n = 881) and 9% more than 1000 g. The largest specimens, weighing 2933 g, 3183 g, and 4780 g, required two bags for complete morcellation. Neither difficulties nor complications related to bag manipulation were recorded. Small bag puncture was detected in two cases, but peritoneal washing cytology was free of debris. One retroperitoneal angioleiomyomatosis and three malignancies were detected in histology (leiomyosarcoma = 2; sarcoma = 1); therefore, patients underwent radical surgery. All patients were disease-free at 3 years follow-up, but one patient presented multiple abdominal metastases of the leiomyosarcoma in the third year; she refused subsequent surgery and was lost from follow-up. This large series demonstrates that laparoscopic bag morcellation is a safe and comfortable method to remove large and giant uterine tumors. Bag manipulation takes only a few minutes, and perforations rarely occur and are easy to detect intraoperatively. This technique did not result in the spread of debris during myoma surgery, potentially avoiding the additional risk of parasitic fibroma or peritoneal sarcoma.

Intraoperative Use of Wide-Field Optical Coherence Tomography to Evaluate Tissue Microstructure in the Oral Cavity and Oropharynx

Involvement of deep margins represents a significant challenge in the treatment of oropharyngeal cancer, and given practical limitations of frozen-section analysis, a need exists for real-time, nondestructive intraoperative margin analysis. Wide-field optical coherence tomography (WF-OCT) has been evaluated as a tool for high-resolution adjunct specimen imaging in breast surgery, but its clinical application in head and neck surgery has not been explored. To evaluate the utility of WF-OCT for visualizing microstructures at margins of excised oral and oropharyngeal tissue. This nonrandomized, investigator-initiated qualitative study evaluated the feasibility of the Perimeter Medical Imaging AI Otis WF-OCT device at a single academic center. Included participants were adults undergoing primary ablative surgery of the oral cavity or oropharynx for squamous cell carcinoma in 2018 and 2019. Data were analyzed in October 2019. Patients were treated according to standard surgical care. Freshly resected specimens were imaged with high-resolution WF-OCT prior to routine pathology. Interdisciplinary interpretation was performed to interpret WF-OCT images and compare them with corresponding digitized pathology slides. No clinical decisions were made based on WF-OCT image data. Visual comparisons were performed between WF-OCT images and hematoxylin and eosin slides. A total of 69 specimens were collected and scanned from 53 patients (mean [SD] age, 59.4 [15.2] years; 35 [72.9%] men among 48 patients with demographic data) undergoing oral cavity or oropharynx surgery for squamous cell carcinoma, including 42 tonsillar tissue, 17 base of the tongue, 4 buccal tissue, 3 mandibular, and 3 other specimens. There were 41 malignant specimens (59.4%) and 28 benign specimens (40.6%). In visual comparisons of WF-OCT images and hematoxylin and eosin slides, visual differentiation among mucosa, submucosa, muscle, dysplastic, and benign tissue was possible in real time using WF-OCT images. Microarchitectural features observed in WF-OCT images could be matched with corresponding features within the permanent histology with fidelity. This qualitative study found that WF-OCT imaging was feasible for visualizing tissue microarchitecture at the surface of resected tissues and was not associated with changes in specimen integrity or surgical and pathology workflow. These findings suggest that formal clinical studies investigating use of WF-OCT for intraoperative analysis of deep margins in head and neck surgery may be warranted.

Venous Thromboembolism after Abdominal and Minimally Invasive Large Specimen Hysterectomy

<h3>Study Objective</h3> To investigate the incidence of venous thromboembolism (VTE) in patients undergoing large specimen hysterectomy for benign indications. To evaluate the impact of route of surgery and operative time in the development of VTE in this population. <h3>Design</h3> Cohort study of targeted hysterectomy data prospectively collected from the American College of Surgeons National Surgical Quality Improvement Program involving over 500 hospitals across the United States. <h3>Setting</h3> National Surgical Quality Improvement Program Database. <h3>Patients or Participants</h3> Women aged 18 years or older undergoing hysterectomy for benign indications between 2014 and 2019. Patients were further classified into four groups according to uterine weight: <100g, 100-249g, 250g-499g, and specimens >500g. <h3>Interventions</h3> Current Procedural Terminology codes were used to identify cases. Variables including age, ethnicity, body mass index (BMI), smoking status, diabetes, hypertension, blood transfusion, and American Society of Anesthesiologists (ASA) classification system scores were collected. Cases were stratified by route of surgery, operative time, and uterine weight. <h3>Measurements and Main Results</h3> A total of 122,408 patients were included in the study, of which 28,397 (23%) underwent abdominal, 75,490 (62%) laparoscopic, and 18,521 (15%) vaginal hysterectomy. The VTE rate for large specimen hysterectomy (>500g) was 0.64%. After adjusting for confounders, patients who underwent minimally invasive hysterectomy had a lower rate of VTE for the laparoscopic (aOR 0.62; CI:0.47-0.81) and vaginal (aOR 0.46; CI:0.31-0.68) routes when compared to open surgery. Total operative time was independently associated with postoperative VTE (>120min, aOR 1.85; CI:1.5-2.27). Uterine weight even for specimens >500g was not significantly associated with the rate of VTE. Interestingly, only 30% of the surgeries with uterine weight above 500g were done with a minimally invasive route. <h3>Conclusion</h3> The occurrence of VTE after benign large specimen hysterectomy is rare. The rate of VTE is higher with longer operative times and is lower with minimally invasive approaches even for markedly enlarged uteri.

An Immunohistochemical study of CD83 positive dendritic cell density in human benign and malignant uterine specimens

Introduction and Aim: Dendritic cells (DCs) are heterogenous group of accessory cells that function as specialized forms of antigen-presenting cells in human body. A thorough knowledge of the DC population of the human uterus, especially the CD83 population would be very useful for an effective understanding of the immune protection of the endometrium during different phases of menstrual cycle and in the pathological conditions of the uterus. Materials and Methods: A prospective study was conducted in a tertiary care referral medical college hospital in South India. Women in the age group of 35 to 50 years, who underwent hysterectomy were included in the study. Immunohistochemical study on the presence of mature CD83+DCs was carried out in the processed uterine specimens. The number of CD83 cells/20 high power fields in each of the specimen was calculated and reviewed by two independent reviewers. Results: About 43 hysterectomy specimens obtained from patients undergoing hysterectomy for benign and malignant conditions of uterus were included in the study. Fibroid uterus was the most common indication for hysterectomy. The minimum and maximum DCs were 12 and 720 DCs per 20 HPF respectively, with an overall mean of 109.28. The mean DCs amongst benign specimens was 1.2 DCs and in the malignant specimens was 22.65 DCs per HPF, with a P value of 0.0001. Conclusion: Mature dendritic cells are seen in abundance in malignant and pre-malignant tissues of human uterus. A reduction in number of CD83+ DCs was observed in patients with proliferative and secretory endometrium.

A study of incidentally detected carcinoma in patients operated for benign thyroid disease: Is it more common than we think?

Aim: In this prospective study we analysed the incidence of carcinoma in patients operated with preoperative diagnosis of benign thyroid conditions, and correlate it with risk factors for further management. Methods: A prospective analysis of 94 patients that presented to a tertiary care centre in south India and diagnosed to have benign thyroid disease that required surgical management from April 2016 to October 2017. The postoperative thyroid specimen was analysed and any evidence of malignancy was noted. These patients were further evaluated and need for further treatment was analysed based on risk factors like advanced age, male sex and size of the tumour according to AGES, AMES and MACIS scoring systems. Results: In our study, 11 out of 94 patients operated for benign thyroid disease were found to have malignancy. Thus, the incidence of occult carcinoma of thyroid was significantly higher than that of the previously published literature in India, 11.7% vs 2-7%. The most common preoperative diagnosis was that of multinodular goitre. Conclusion : The incidence of malignancy in benign thyroid specimens was high in our study, but there is a need for longer follow up, to study the biology and behavioural patterns of these probably low grade tumours.

Immunohistochemical study of CD1a positive dendritic cells in normal, benign and malignant lesions of the human uterus and cervix

Introduction and Aim: Dendritic cells (DCs) perform the function of antigen-presenting cells of the immune system of mammals. After demonstration of DCs in the human cervix, more focus fell on the role of DCs distributed in various benign and cancerous lesions of endometrium. In our study, we have studied the role of CD1a positive DCs in non-malignant and cancerous tissues of the uterus and human cervix. Materials and Methods: A prospective analysis was conducted on patients who had undergone hysterectomy and cervical biopsy. After a detailed menstrual cycle history, the processed specimens were subjected for hematoxylin-eosin and immunohistochemical staining. The CD1a positive cells were counted per 20 high power fields and the mean cells per high power field (HPF) was calculated. Results: Eighty specimens were studied. Forty-six uterine and 34 cervix specimens were studied. The average chronological age of our study patients was 45.7 years. Bleeding per vaginum was the most common presentation. Fibroid uterus was the most common indication for hysterectomy. The minimum and maximum DCs amongst the 80 specimens were 1 and 246 per 20 HPF and the overall mean was 72.7. The mean cervical and uterine DCs per HPF were 7.7 and 0.7 respectively (P=0.0116). The average DCs per HPF in benign group were more than that in malignant specimens (9.23 and 1.76, respectively). Conclusion: A higher concentration of DCs in the cervical tissue was observed. Malignant tissues showed a lesser number of DCs than the benign specimens.

Caesarean section operation is not associated with myometrial hypertrophy–a prospective cohort study

Maternity statistics of England in 2020 showed rise in Caesarean Section (CS) rate to 31%. Some studies correlated adverse gynaecological symptoms e.g. menstrual irregularities and pelvic pain to ‘niche’ formation at CS scar site. Niche formation was speculated to cause myometrial hypertrophy aggravating these symptoms. This was a prospective comparative histological study including 52 consecutive benign hysterectomy specimens which were categorised into 2 groups: (i) specimens with CS scar (n = 22), (ii) specimens with no CS scar (n = 30). Median (IQ range) uteri weight was 97.2grms (43.5–226) and 91.7grms (35.7–201.7) in study and control groups, respectively (p = .991). Mean (±SD) thickness of anterior myometrial wall was 18.7 mm (±3.6) and 19.4 mm (±4.5) in study and control groups, respectively (p = .58). Mean (±SD) thickness of posterior myometrial wall was 19.1 mm (±3.7) and 18.7 mm (±3.9) in study and control groups, respectively (p = .78). The assumption that CS scar causes myometrial hypertrophy was not demonstrated in this study. IMPACT STATEMENT What is already known on this subject? Maternity statistics world-wide show a continuous rise in the rate of Caesarean Section (CS) operation. The CS scar is assumed to be related to adverse clinical gynaecological symptoms such as intermenstrual bleeding, dysmenorrhoea, dyspareunia and chronic pelvic pain; however, the mechanism of this association is not clear. Further, little is known about the effects of CS scar on uterine wall morphology and function. What do the results of this study add? This study was the first prospective series in the literature to compare the uteri with scar with those without in respect of weight and myometrial wall thickness. It was not able to demonstrate the association between having CS scar and myometrial hypertrophy which was hypothesised to be the cause of adverse gynaecological symptoms. However, the microscopic examination of the CS scar revealed adenomyosis, haemorrhage and/or chronic inflammation in most cases. What are the implications of these findings for clinical practice and/or future research? The clinical implication of the histological changes shown in the CS scar requires large comparative clinical studies.