Robust Cell Detection and Segmentation for Image Cytometry Reveal Th17 Cell Heterogeneity.

Image cytometry enables quantitative cell characterization with preserved tissue architecture; thus, it has been highlighted in the advancement of multiplex immunohistochemistry (IHC) and digital image analysis in the context of immune-based biomarker monitoring associated with cancer immunotherapy. However, one of the challenges in the current image cytometry methodology is a technical limitation in the segmentation of nuclei and cellular components particularly in heterogeneously stained cancer tissue images. To improve the detection and specificity of single-cell segmentation in hematoxylin-stained images (which can be utilized for recently reported 12-biomarker chromogenic sequential multiplex IHC), we adapted a segmentation algorithm previously developed for hematoxlin and eosin-stained images, where morphological features are extracted based on Gabor-filtering, followed by stacking of image pixels into n-dimensional feature space and unsupervised clustering of individual pixels. Our proposed method showed improved sensitivity and specificity in comparison with standard segmentation methods. Replacing previously proposed methods with our method in multiplex IHC/image cytometry analysis, we observed higher detection of cell lineages including relatively rare TH 17 cells, further enabling sub-population analysis into TH 1-like and TH 2-like phenotypes based on T-bet and GATA3 expression. Interestingly, predominance of TH 2-like TH 17 cells was associated with human papilloma virus (HPV)-negative status of oropharyngeal squamous cell carcinoma of head and neck, known as a poor-prognostic subtype in comparison with HPV-positive status. Furthermore, TH 2-like TH 17 cells in HPV-negative head and neck cancer tissues were spatiotemporally correlated with CD66b+ granulocytes, presumably associated with an immunosuppressive microenvironment. Our cell segmentation method for multiplex IHC/image cytometry potentially contributes to in-depth immune profiling and spatial association, leading to further tissue-based biomarker exploration. © 2019 International Society for Advancement of Cytometry.

Effect of ibuprofen on severity of surgically-managed pediatric post-tonsillectomy hemorrhage.

OBJECTIVES: The association between ibuprofen use and severity of post-tonsillectomy hemorrhage (PTH) remains unclear. We aimed to compare PTH severity in patients who did or did not receive ibuprofen. METHODS: A retrospective cohort study of pediatric patients requiring operative control of PTH at a tertiary children's hospital between 2015 and 2019 was performed. PTH severity was assessed using pre-tonsillectomy and post-hemorrhage hemoglobin and hematocrit values, estimated intraoperative blood loss, estimated hemorrhage flow rate, and need for transfusion. Differences in hemorrhage severity markers between the two cohorts were compared. RESULTS: A total of 168 consecutive patients were included in this study. The mean age was 8.8 years, and 55.4% of patients were male. Sixty-five patients (38.7%) received ibuprofen postoperatively. There was no statistically significant difference in the mean change in hemoglobin (1.1 vs. 1.1, P = 0.85) or hematocrit (3.1 vs. 3.2, P = 0.97) between patients who received ibuprofen compared to those who did not. Similarly, there were no significant differences in need for transfusion (3.1% vs. 3.9%, P = 1.00) or occurrence of high-flow (arterial) blood loss (33.8% vs. 40.8%, P = 0.42) between the two groups. CONCLUSION: Postoperative ibuprofen use does not appear to significantly increase PTH severity, as measured by change in hemoglobin and hematocrit values, need for transfusion, or presence of high-flow blood loss. This study introduces previously unexplored markers to assess PTH severity and supports further prospective studies to determine the effect of ibuprofen on PTH severity.

Single-cell spatial architectures associated with clinical outcome in head and neck squamous cell carcinoma.

There is increasing evidence that the spatial organization of cells within the tumor-immune microenvironment (TiME) of solid tumors influences survival and response to therapy in numerous cancer types. Here, we report results and demonstrate the applicability of quantitative single-cell spatial proteomics analyses in the TiME of primary and recurrent human papillomavirus (HPV)-negative head and neck squamous cell carcinoma (HNSCC) tumors. Single-cell compositions of a nine patient, primary and recurrent (n = 18), HNSCC cohort is presented, followed by deeper investigation into the spatial architecture of the TiME and its relationship with clinical variables and progression free survival (PFS). Multiple spatial algorithms were used to quantify the spatial landscapes of immune cells within TiMEs and demonstrate that neoplastic tumor-immune cell spatial compartmentalization, rather than mixing, is associated with longer PFS. Mesenchymal (αSMA+) cellular neighborhoods describe distinct immune landscapes associated with neoplastic tumor-immune compartmentalization and improved patient outcomes. Results from this investigation are concordant with studies in other tumor types, suggesting that trends in TiME cellular heterogeneity and spatial organization may be shared across cancers and may provide prognostic value in multiple cancer types.

Opioid Prescribing in Patients Undergoing Neck Dissections With Short Hospitalizations.

OBJECTIVE: To evaluate postoperative opioid prescribing in patients undergoing neck dissections with short hospitalizations. STUDY DESIGN: Retrospective cohort study. SETTING: Tertiary academic hospital. METHODS: The study population included patients who underwent lateral neck dissections with or without an associated head and neck procedure and required hospitalization for ≤3 days from 2012 to 2019. Interventions to decrease opioid utilization, including preoperative counseling, multimodality pain management, and multidisciplinary collaboration, were implemented in September 2016. Patients were divided into 2 groups: preintervention (group 1) and postintervention (group 2). The mean quantity of opioids prescribed during hospitalization, at discharge, and in refills was calculated in morphine milligram equivalents (MME). RESULTS: A total of 407 patients were included in the analysis: 223 patients in group 1 and 184 patients in group 2 (42.3% female, 89.4% white; average age, 55.2 years [95% CI, 53.6-56.9]). The mean opioid quantity prescribed in unilateral neck dissection alone decreased from 353.9 MME (95% CI, 266.7-441.2) in group 1 to 113.3 MME (95% CI, 87.8-138.7) in group 2 (P < .001; effect size, 1.0). Statistically significant decreases in mean opioid quantity prescribed were also observed in unilateral neck dissection in combination with thyroidectomy, parotidectomy, glossectomy, or tonsillectomy. The percentage of patients requiring opioid prescription refills was not statistically different between the groups. CONCLUSION: This study demonstrates that the quantity of opioids prescribed in patients undergoing neck dissections and associated head and neck procedures with short hospitalizations can be reduced to as low as 100 to 125 MME with preoperative counseling, multimodality pain management, and multidisciplinary collaboration.

Prevalence and Risk Factors for Multifocality in Pediatric Thyroid Cancer.

Importance: Current guidelines recommend total thyroidectomy for the majority of pediatric thyroid cancer owing to an increased prevalence of multifocality. However, there is a paucity of information on the exact prevalence and risk factors for multifocal disease-knowledge that is critical to improving pediatric thyroid cancer management and outcomes. Objective: To determine the prevalence and risk factors for multifocal disease in pediatric patients with papillary thyroid carcinoma (PTC). Design, Setting, and Participants: This multicenter retrospective cohort study included patients 18 years or younger who underwent thyroidectomy for PTC from 2010 to 2020 at 3 tertiary pediatric hospitals and 2 tertiary adult and pediatric hospitals in the US. Main Outcomes and Measures: Demographic and clinical variables, including age, family history of thyroid cancer, autoimmune thyroiditis, prior radiation exposure, cancer predisposition syndrome, tumor size, tumor and nodal stage, PTC pathologic variant, and preoperative imaging, were assessed for association with presence of any multifocal, unilateral multifocal, and bilateral multifocal disease using multiple logistic regression analyses. Least absolute shrinkage and selection operator analysis was performed to develop a model of variables that may predict multifocal disease. Results: Of 212 patients, the mean age was 14.1 years, with 23 patients 10 years or younger; 173 (82%) patients were female. Any multifocal disease was present in 98 (46%) patients, with bilateral multifocal disease in 73 (34%). Bilateral multifocal disease was more accurately predicted on preoperative imaging than unilateral multifocal disease (48 of 73 [66%] patients vs 9 of 25 [36%] patients). Being 10 years or younger, T3 tumor stage, and N1b nodal stage were identified as predictors for multifocal and bilateral multifocal disease. Conclusions and Relevance: This large, multicenter cohort study demonstrated a high prevalence of multifocal disease in pediatric patients with PTC. Additionally, several potential predictors of multifocal disease, including age and advanced T and N stages, were identified. These risk factors and the high prevalence of multifocal disease should be considered when weighing the risks and benefits of surgical management options in pediatric patients with PTC.

Impact of AAO-HNS Guideline on Obtaining Polysomnography Prior to Tonsillectomy for Pediatric Sleep-Disordered Breathing.

OBJECTIVE: To assess the impact of the 2011 American Academy of Otolaryngology-Head and Neck Surgery (AAO-HNS) guideline on practice patterns in obtaining preadenotonsillectomy (AT) polysomnography (PSG) for pediatric sleep-disordered breathing (SDB). STUDY DESIGN: Retrospective cohort study. SETTING: Tertiary children's hospital. SUBJECT AND METHODS: The study population included all patients referred to our pediatric otolaryngology clinic for consideration of AT for SDB during two 12-month time periods: before (2010-2011) and after (2015-2016) publication of the 2011 AAO-HNS guideline. Demographic, insurance, comorbidity, and Pediatric Sleep Questionnaire (PSQ) variables were assessed for association with pre-AT PSG using bivariate and multivariate logistic regression analysis. RESULTS: A greater percentage of patients underwent pre-AT PSG in 2015-2016 vs 2010-2011 (30% vs 22%, P = .001). On multivariate analysis, presence of neuromuscular disorder was the only predictor associated with pre-AT PSG in 2010 to 2011 (odds ratio [OR], 3.00; 95% CI, 1.10, 8.06; P = .03). Presence of neuromuscular disorder (OR, 2.54; 95% CI, 1.51, 4.29; P < .0001), craniofacial anomaly (OR, 2.32; 95% CI, 1.20, 4.50; P = .013), or Down syndrome (OR, 3.45; 95% CI, 1.54, 7.72; P = .003) was associated with pre-AT PSG in 2015 to 2016. Positive PSQ screen was significantly associated with decreased odds of pre-AT PSG in both time periods. CONCLUSION: After publication of the 2011 AAO-HNS guideline, there was a significant but modest increase in pre-AT PSG utilization in children with SDB and high-risk comorbidities at our institution, consistent with AAO-HNS guideline recommendations. However, overall guideline adherence remains low and may reflect limitations in PSG testing capacity.

High-dimensional multiplexed immunohistochemical characterization of immune contexture in human cancers.

Biomarker assessments of tumor specimens is widely used in cancer research to audit tumor cell intrinsic as well as tumor cell extrinsic features including the diversity of immune, stromal, and mesenchymal cells. To comprehensively and quantitatively audit the tumor-immune microenvironment (TiME), we developed a novel multiplex immunohistochemistry (mIHC) platform and computational image processing workflow using a single formalin-fixed paraffin-embedded (FFPE) tissue section. Herein, we validated this platform using nine matched primary newly diagnosed and recurrent head and neck squamous cell carcinoma (HNSCC) sections sequentially subjected to immunodetection with a panel of 29 antibodies identifying malignant tumor cells, and 17 distinct leukocyte lineages and their functional states. Image cytometric analysis was applied to interpret chromogenic signals from digitally scanned and coregistered light microscopy-based images enabling identification and quantification of individual tumor cells, structural features, immune cell phenotypes and their functional state. In agreement with our previous study via a 12-plex imaging mIHC platform, myeloid-inflamed status in newly diagnosed primary tumors associated with significantly short progression free survival, independent of lymphoid-inflamed status. Spatial distribution of tumor and immune cell lineages in TiME was also examined and revealed statistically significant CD8+ T cell exclusion from tumor nests, whereas regulatory T cells and myeloid cells, when present in close proximity to tumor cells, highly associated with rapid cancer recurrence. These findings indicate presence of differential immune-spatial profiles in newly diagnosed and recurrent HNSCC, and establish the robustness of the 29-plex mIHC platform and associated analytics for quantitative analysis of single tissue sections revealing longitudinal TiME changes.