Shifts in Serum Bile Acid Profiles Associated With Barrett's Esophagus and Stages of Progression to Esophageal Adenocarcinoma.

INTRODUCTION: Reflux bile acids are believed to promote esophageal adenocarcinoma (EAC), but the role of systemic bile acids is unknown. This study aimed to assess associations between systemic bile acids and stages of Barrett's esophagus (BE) progression. METHODS: Subjects with and without BE were enrolled in this multicenter cross-sectional study. Targeted serum bile acid profiling was performed, and a subset of subjects completed a validated food frequency questionnaire. RNA sequencing was performed on BE or gastric cardia tissue to assess bile acid associations with gene expression. RESULTS: A total of 141 subjects were enrolled with serum bile acids profiled (49 non-BE; 92 BE: 44 no dysplasia, 25 indefinite/low grade dysplasia, 23 high-grade dysplasia/EAC). Lower Healthy Eating Index score, older age, higher body mass index, and no proton pump inhibitor use were associated with increased levels of multiple bile acids. Global bile acid pools were distinct between non-BE and stages of BE neoplasia ( P = 0.004). Increasing cholic acid was associated with high-grade dysplasia/EAC compared with non-BE, even after adjusting for EAC risk factors (adjusted odds ratio 2.03, 95% confidence interval 1.11-3.71) as was the combination of unconjugated primary bile acids (adjusted odds ratio 1.81, 95% confidence interval 1.04-3.13). High cholic acid levels were associated with tissue gene expression changes including increased DNA replication and reduced lymphocyte differentiation genes. DISCUSSION: Alterations in serum bile acids are independently associated with advanced neoplasia in BE and may contribute to neoplastic progression. Future studies should explore associated gut microbiome changes, proneoplastic effects of bile acids, and whether these bile acids, particularly cholic acid, represent potential biomarkers or viable therapeutic targets for advanced neoplasia in BE.

High risk features in colorectal adenomatous polyps: A multi-institutional study.

High risk features in colorectal adenomatous polyps include size >1 cm and advanced histology: high-grade dysplasia and villous architecture. We investigated whether the diagnostic rates of advanced histology in colorectal adenomatous polyps were similar among institutions across the United States, and if not, could differences be explained by patient age, polyp size, and/or CRC rate. Nine academic institutions contributed data from three pathologists who had signed out at least 100 colorectal adenomatous polyps each from 2018 to 2019 taken from patients undergoing screening colonoscopy. For each case, we recorded patient age and sex, polyp size and location, concurrent CRC, and presence or absence of HGD and villous features. A total of 2700 polyps from 1886 patients (mean age: 61 years) were collected. One hundred twenty-four (5 %) of the 2700 polyps had advanced histology, including 35 (1 %) with HGD and 101 (4 %) with villous features. The diagnostic rate of advanced histology varied by institution from 1.7 % to 9.3 % (median: 4.3 %, standard deviation [SD]: 2.5 %). The rate of HGD ranged from 0 % to 3.3 % (median: 1 %, SD: 1.2 %), while the rate of villous architecture varied from 1 % to 8 % (median: 3.7 %, SD: 2.5 %). In a multivariate analysis, the factor most strongly associated with advanced histology was polyp size >1 cm with an odds ratio (OR) of 31.82 (95 % confidence interval [CI]: 20.52-50.25, p < 0.05). Inter-institutional differences in the rate of polyps >1 cm likely explain some of the diagnostic variance, but pathologic subjectivity may be another contributing factor.

Esophageal squamous cell carcinoma with pagetoid spread: a clinicopathologic study.

Pagetoid spread in esophageal squamous epithelium associated with underlying esophageal adenocarcinoma (EAC) has been well studied. Case reports describing pagetoid spread of esophageal squamous cell carcinomas (ESCC) also exist in the literature. The latter, however, has not been systematically studied. In this study, we report seven cases of pagetoid spread associated with ESCC. The clinical, morphologic, and immunophenotypic profiles of pagetoid spread in the context of ESCC and EAC are compared. Cases of pagetoid spread of ESCC were identified through computerized search of pathology archives at five institutions. Additional cases were identified through manual review of surgical resection cases of treatment naive ESCC in Mass General Brigham (MGB) pathology archive. Clinical history was collected via chart review. Immunohistochemistry for CK7, CK20, CDX2, p53, p63, and p40 was performed on selected cases. A computerized search of pathology archives of five institutions revealed only two cases. A manual review of 76 resected untreated ESCC revealed five additional cases with unequivocal pagetoid spread of ESCC, indicating the condition was not uncommon but rarely reported. Patient age ranged from 54 to 78 years (median, 65). There were six women and one man. One case had in situ disease, five had pT1 (1 pT1a and 4 pT1b), and one had pT3 disease. One of the patients with pT1 tumor had a positive lymph node, while the remaining six patients were all N0. Four tumors were in the proximal to mid esophagus, and three in the distal esophagus. Patient survival ranged from 25 months to more than 288 months. The pagetoid tumor cells demonstrated enlarged, hyperchromatic nuclei with variable amounts of eosinophilic cytoplasm. The cytoplasm was often condensed to the perinuclear area, creating peripheral clearing. By immunohistochemistry, the pagetoid cells were positive for p40 (6/6) and p63 (7/7) and negative for CDX2 (7/7). The tumor cells showed mutant-type staining for p53 in five of seven cases. One of the patients had pagetoid tumor cells at the resection margin and subsequently had recurrent disease 2 years later. All other patients had negative resection margins and did not have local recurrence. Four cases of pagetoid spread in the context of EAC were used as a comparison group. Previously published studies were also analyzed. These tumors were all located in the distal esophagus or gastroesophageal junction. All cases were associated with underlying invasive EAC. Pagetoid spread associated with EAC often had cytoplasmic vacuoles or mucin. They were more frequently positive for CK7 than pagetoid ESCC (p = 0.01). Both ESCC and EAC may give rise to pagetoid spread of tumor cells within surface squamous epithelium. Pagetoid spread from ESCC and EAC have overlapping morphologic features. P40 and p63 immunostains can facilitate the distinction between ESCC and EAC. P53 immunostain can aid in confirmation of malignancy. Understanding their overlapping pathologic features will help pathologists avoid pitfalls and diagnose these lesions correctly on biopsy specimens.

Standardizing Randomized Controlled Trials in Celiac Disease: An International Multidisciplinary Appropriateness Study.

BACKGROUND & AIMS: There is a need to develop safe and effective pharmacologic options for the treatment of celiac disease (CeD); however, consensus on the appropriate design and configuration of randomized controlled trials (RCTs) in this population is lacking. METHODS: A 2-round modified Research and Development/University of California Los Angeles Appropriateness Method study was conducted. Eighteen gastroenterologists (adult and pediatric) and gastrointestinal pathologists voted on statements pertaining to the configuration of CeD RCTs, inclusion and exclusion criteria, gluten challenge, and trial outcomes. Two RCT designs were considered, representing the following distinct clinical scenarios for which pharmacotherapy may be used: trials incorporating a gluten challenge to simulate exposure; and trials evaluating reversal of histologic changes, despite attempted adherence to a gluten-free diet. Each statement was rated as appropriate, uncertain, or inappropriate, using a 9-point Likert scale. RESULTS: For trials evaluating prevention of relapse after gluten challenge, participants adherent to a gluten-free diet for 12 months or more with normal or near-normal-sized villi should be enrolled. Gluten challenge should be FODMAPS (fermentable oligosaccharides, disaccharides, monosaccharides, and polyols) free, and efficacy evaluated using histology with a secondary patient-reported outcome measure. For trials evaluating reversal of villus atrophy, the panel voted it appropriate to enroll participants with a baseline villus height to crypt depth ratio ≤2 and measure efficacy using a primary histologic end point. Guidance for measuring histologic, endoscopic, and patient-reported outcomes in adult and pediatric patients with CeD are provided, along with recommendations regarding the merits and limitations of different end points. CONCLUSIONS: We developed standardized recommendations for clinical trial design, eligibility criteria, outcome measures, gluten challenge, and disease evaluations for RCTs in patients with CeD.

Erythrophagocytosis is not a reproducible finding in liver biopsies, and is not associated with clinical diagnosis of hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a rare disease with high mortality. Liver involvement is common (based on elevated liver function tests) with most patients demonstrating acute hepatitis. Liver biopsies are frequently obtained in the setting of suspected HLH for the purpose of identification of erythrophagocytosis, and if present, this finding is thought to suggest or support the diagnosis of HLH. However, there are problems with this approach; in particular, we do not know whether this finding is reproducible or whether it is specific to HLH. Therefore, we conducted a multi-institutional study in which experienced liver pathologists reviewed images taken from liver biopsies from patients with normal liver, acute hepatitis, possible HLH, and clinical HLH to determine if there was agreement about the presence or absence of erythrophagocytosis, and to ascertain whether the finding corresponds to a clinical diagnosis of HLH. Twelve liver pathologists reviewed 141 images in isolation (i.e., no clinical information or diagnosis provided). These came from 32 patients (five normal, 17 acute hepatitis, six HLH, four possible HLH). The pathologists classified each image as negative, equivocal, or positive for erythrophagocytosis. Kappa was .08 (no agreement) for case-level and 0.1 for image-level (1.4% agreement, based on two images which were universally considered negative). There was no difference in the proportion of pathologists who diagnosed erythrophagocytosis among those with different diagnoses at case or image-level (p = 0.82 and p = 0.82, respectively). Thus, erythrophagocytosis is an entirely unreliable histologic parameter in liver, as it is irreproducible and not demonstrably associated with a clinical disease (namely, HLH). Unless and until more reliable guidelines can be established, pathologists should refrain from commenting on the presence or absence of erythrophagocytosis in liver biopsy.

A pan-cancer analysis implicates human NKIRAS1 as a tumor-suppressor gene.

The NF-κB family of transcription factors and the Ras family of small GTPases are important mediators of proproliferative signaling that drives tumorigenesis and carcinogenesis. The κB-Ras proteins were previously shown to inhibit both NF-κB and Ras activation through independent mechanisms, implicating them as tumor suppressors with potentially broad relevance to human cancers. In this study, we have used two mouse models to establish the relevance of the κB-Ras proteins for tumorigenesis. Additionally, we have utilized a pan-cancer bioinformatics analysis to explore the role of the κB-Ras proteins in human cancers. Surprisingly, we find that the genes encoding κB-Ras 1 (NKIRAS1) and κB-Ras 2 (NKIRAS2) are rarely down-regulated in tumor samples with oncogenic Ras mutations. Reduced expression of human NKIRAS1 alone is associated with worse prognosis in at least four cancer types and linked to a network of genes implicated in tumorigenesis. Our findings provide direct evidence that loss of NKIRAS1 in human tumors that do not carry oncogenic RAS mutations is associated with worse clinical outcomes.

Infection and inflammation stimulate expansion of a CD74+ Paneth cell subset to regulate disease progression.

Paneth cells (PCs), a specialized secretory cell type in the small intestine, are increasingly recognized as having an essential role in host responses to microbiome and environmental stresses. Whether and how commensal and pathogenic microbes modify PC composition to modulate inflammation remain unclear. Using newly developed PC-reporter mice under conventional and gnotobiotic conditions, we determined PC transcriptomic heterogeneity in response to commensal and invasive microbes at single cell level. Infection expands the pool of CD74+ PCs, whose number correlates with auto or allogeneic inflammatory disease progressions in mice. Similar correlation was found in human inflammatory disease tissues. Infection-stimulated cytokines increase production of reactive oxygen species (ROS) and expression of a PC-specific mucosal pentraxin (Mptx2) in activated PCs. A PC-specific ablation of MyD88 reduced CD74+ PC population, thus ameliorating pathogen-induced systemic disease. A similar phenotype was also observed in mice lacking Mptx2. Thus, infection stimulates expansion of a PC subset that influences disease progression.

Unravelling interobserver variability in gastrointestinal glandular neoplasia: a contemporary study of US and Korean pathologists.

AIMS: Interobserver variability in the assessment of gastric neoplasia biopsies between most Western and Eastern (predominantly represented by Japanese in the literature) pathologists has been documented. It is unknown if such variability exists between the US and Korean pathologists in the current era. METHODS: Ten gastrointestinal (GI) pathologists from the USA (n=5) and South Korea (n=5) evaluated 100 scanned images of gastric (n=50) and colorectal (n=50) neoplasia biopsies and answered multiple questionnaires. Consensus was defined as the answer chosen by the majority. Cohen's (κc) and Fleiss' kappa (κf) values were calculated between the consensus of the two groups and among the raters, respectively. RESULTS: Both groups reached a consensus in the majority of cases (74%-100%) with slight to perfect intergroup (κc=0.049-1.000) and no to substantial intragroup (κf=-0.083 to 0.660) agreements. For gastric neoplasia, Korean pathologists relied heavily on cytoarchitectural atypia, whereas the US pathologists focused on stromal invasion when diagnosing adenocarcinoma. For colorectal neoplasia, the Korean pathologists identified concurrent intramucosal carcinoma when diagnosing invasive adenocarcinoma, while the presence of desmoplasia was a prerequisite for the diagnosis of invasive adenocarcinoma for the US pathologists. CONCLUSIONS: For GI neoplasia biopsy interpretation, the diagnostic approach of Korean pathologists is similar to that of Eastern/Japanese pathologists. Consensus outperformed kappa statistics in capturing the magnitude of inter-rater and intergroup reliability, highlighting the potential benefit of consensus meetings to decrease the gap between Western and Eastern diagnostic approaches.

Challenges in Diagnosing and Reporting Cholangiocarcinoma.

Intrahepatic cholangiocarcinoma is a challenge to the practicing surgical pathologist for several reasons. It is rare in many parts of the world, and thus practical exposure may be limited. Related to the fact of its rarity is the fact that more common tumors which frequently metastasize to the liver can be morphologically indistinguishable (eg, pancreatic ductal adenocarcinoma). Immunohistochemical testing is generally non-contributory in this context. Other difficulties arise from the protean morphologic manifestations of cholangiocarcinoma (ie, small duct vs. large duct) and the existence of combined cholangiocarcinoma and hepatocellular carcinoma. These, and other issues of concern to the practicing diagnostic pathologist are discussed herein.

IFN-γR/STAT1 signaling in recipient hematopoietic antigen-presenting cells suppresses graft-versus-host disease.

The absence of IFN-γ receptor (IFN-γR) or STAT1 signaling in donor cells has been shown to result in reduced induction of acute graft-versus-host disease (GVHD). In this study, we unexpectedly observed increased activation and expansion of donor lymphocytes in both lymphohematopoietic organs and GVHD target tissues of IFN-γR/STAT1-deficient recipient mice, leading to rapid mortality following the induction of GVHD. LPS-matured, BM-derived Ifngr1-/- Stat1-/- DCs (BMDCs) were more potent allogeneic stimulators and expressed increased levels of MHC II and costimulatory molecules. Similar effects were observed in human antigen-presenting cells (APCs) with knockdown of Stat1 by CRISPR/Cas9 and treatment with a JAK1/2 inhibitor. Furthermore, we demonstrated that the absence of IFN-γR/STAT1 signaling in hematopoietic APCs impaired the presentation of exogenous antigens, while promoting the presentation of endogenous antigens. Thus, the indirect presentation of host antigens to donor lymphocytes was defective in IFN-γR/STAT1-deficient, donor-derived APCs in fully donor chimeric mice. The differential effects of IFN-γR/STAT1 signaling on endogenous and exogenous antigen presentation could provide further insight into the roles of the IFN-γ/STAT1 signaling pathway in the pathogenesis of GVHD, organ rejection, and autoimmune diseases.

The histological spectrum of ARB-induced gastritis.

AIMS: Olmesartan, an angiotensin receptor blocker (ARB) used for hypertension management, is known to cause a sprue-like enteropathy in a subset of patients. Rare cases of gastritis occurring with ARB use have also been reported, but the histological features of ARB-induced gastritis and the response to drug cessation have not been examined in a dedicated case-series. METHODS AND RESULTS: Cases of suspected ARB-induced gastritis were identified from the pathology archives of four institutions. Haematoxylin and eosin (H&E) slides from gastric biopsies were reviewed. Fifteen patients (14 female, one male) were identified. The most common presenting symptoms were diarrhoea (10) and weight loss (six). Gastric biopsies commonly showed a full-thickness active chronic gastritis with surface epithelial injury involving the antrum and body. Glandular atrophy, intra-epithelial lymphocytosis and/or subepithelial collagen thickening were also present in some cases. Duodenal involvement, including villous atrophy, intra-epithelial lymphocytosis and/or collagenous sprue, was identified in 11 of 13 cases with concurrent duodenal biopsies. Following drug cessation, symptomatic improvement occurred in all 11 cases for which follow-up data were available. Histological resolution occurred in five of eight cases with follow-up gastric biopsies, with improvement seen in the remaining three biopsies. CONCLUSION: ARB-induced gastritis typically presents as active chronic gastritis, frequently with associated surface epithelial injury. Glandular atrophy, intra-epithelial lymphocytosis and/or subepithelial collagen thickening may also be present. These gastric changes can be seen without associated duodenal injury in rare cases, and they should alert the pathologist to the possibility of ARB-induced injury. Drug cessation results in marked symptomatic and histological improvement.

T cell repertoire profiling in allografts and native tissues in recipients with COVID-19 after solid organ transplantation: Insight into T cell-mediated allograft protection from viral infection.

Introduction: The effects of the SARS-CoV-2 virus on the body, and why the effects are more severe in certain patients, remain incompletely understood. One population of special interest is transplant recipients because of their immunosuppressed state. Understanding the pathophysiology of graft dysfunction in transplant patients with the COVID-19 viral syndrome is important for prognosticating the risk to the graft as well as understanding how best to prevent and, if necessary, treat graft injury in these patients. Methods: We analyzed multiple types of solid organ transplant recipients (liver, kidney, heart or lung) at our institution who died from SARS-CoV-2 and underwent autopsy (n = 6) or whose grafts were biopsied during active SARS-CoV-2 infection (n = 8). Their serum inflammatory markers were examined together with the histological appearance, viral load, and TCR repertoire of their graft tissue and, for autopsy patients, several native tissues. Results: Histology and clinical lab results revealed a systemic inflammatory pattern that included elevated inflammatory markers and diffuse tissue damage regardless of graft rejection. Virus was detected throughout all tissues, although most abundant in lungs. The TCR repertoire was broadly similar throughout the tissues of each individual, with greater sharing of dominant clones associated with more rapid disease course. There was no difference in viral load or clonal distribution of overall, COVID-associated, or putative SARS-CoV-2-specific TCRs between allograft and native tissue. We further demonstrated that SARSCoV-2-specific TCR sequences in transplant patients lack a donor HLArestricted pattern, regardless of distribution in allograft or native tissues,suggesting that recognition of viral antigens on infiltrating recipient cells can effectively trigger host T cell anti-viral responses in both the host and graft. Discussion: Our findings suggest a systemic immune response to the SARS-CoV-2 virus in solid organ transplant patients that is not associated with rejection and consistent with a largely destructive effect of recipient HLA-restricted T cell clones that affects donor and native organs similarly.

Invasive carcinoma versus pseudoinvasion: interobserver variability in the assessment of left-sided colorectal polypectomies.

OBJECTIVES: Misplaced epithelium in adenomas can occasionally be difficult to distinguish from invasive adenocarcinoma. We evaluated interobserver variability in the assessment of left-sided colon polypectomies for pseudoinvasion versus invasive adenocarcinoma and further investigated relevant histological findings. METHODS: 28 consecutive left-sided colon polyps with the keywords "pseudoinvasion", "epithelial misplacement", "herniation", "prolapse" or "invasive adenocarcinoma" were collected from 28 patients and reviewed by eight gastrointestinal pathologists. Participants assessed stromal hemosiderin, lamina propria/eosinophils surrounding glands, desmoplasia, high grade dysplasia/intramucosal adenocarcinoma and margin status and rendered a diagnosis of pseudoinvasion, invasive adenocarcinoma, or both. RESULTS: Agreement among pathologists was substantial for desmoplasia (κ=0.70), high grade dysplasia/intramucosal adenocarcinoma (κ=0.66), invasive adenocarcinoma (κ=0.63) and adenocarcinoma at the margin (κ=0.65). There was moderate agreement for hemosiderin in stroma (κ=0.53) and prolapse/pseudoinvasion (κ=0.50). Agreement was low for lamina propria/eosinophils around glands (κ=0.12). For invasive adenocarcinoma, seven or more pathologists agreed in 24 of 28 cases (86%), and there was perfect agreement in 19/28 cases (68%). For pseudoinvasion, seven or more pathologists agreed in 19 of 28 cases (68%), and there was perfect agreement in 16/28 cases (57%). CONCLUSION: Moderate to substantial, though imperfect, agreement was achieved in the distinction of pseudoinvasion from invasive carcinoma.

Can lightning strike twice? Wild-type transthyretin cardiac amyloidosis associated with rare liver disease.

Wild-type ATTR cardiac amyloidosis (ATTRwt-CA) is not as rare as previously thought to be. Patients with infiltrative cardiac amyloidosis often present with right-sided heart failure (HF) symptomatology. Clinically significant liver disease and cirrhosis has not been reported in ATTRwt-CA. We present two cases of ATTRwt-CA with right-sided HF and abnormal liver function tests initially thought to be secondary to congestive hepatopathy but found to have rare and unrelated liver disease. These cases highlight the importance of developing a broad differential diagnosis and leveraging a multidisciplinary team approach in evaluating patients for unusual causes of cirrhosis/other chronic liver diseases when ATTR cardiac amyloidosis patients present with congestive hepatopathy.

A molecular single-cell lung atlas of lethal COVID-19.

Respiratory failure is the leading cause of death in patients with severe SARS-CoV-2 infection1,2, but the host response at the lung tissue level is poorly understood. Here we performed single-nucleus RNA sequencing of about 116,000 nuclei from the lungs of nineteen individuals who died of COVID-19 and underwent rapid autopsy and seven control individuals. Integrated analyses identified substantial alterations in cellular composition, transcriptional cell states, and cell-to-cell interactions, thereby providing insight into the biology of lethal COVID-19. The lungs from individuals with COVID-19 were highly inflamed, with dense infiltration of aberrantly activated monocyte-derived macrophages and alveolar macrophages, but had impaired T cell responses. Monocyte/macrophage-derived interleukin-1ß and epithelial cell-derived interleukin-6 were unique features of SARS-CoV-2 infection compared to other viral and bacterial causes of pneumonia. Alveolar type 2 cells adopted an inflammation-associated transient progenitor cell state and failed to undergo full transition into alveolar type 1 cells, resulting in impaired lung regeneration. Furthermore, we identified expansion of recently described CTHRC1+ pathological fibroblasts3 contributing to rapidly ensuing pulmonary fibrosis in COVID-19. Inference of protein activity and ligand-receptor interactions identified putative drug targets to disrupt deleterious circuits. This atlas enables the dissection of lethal COVID-19, may inform our understanding of long-term complications of COVID-19 survivors, and provides an important resource for therapeutic development.

Interobserver agreement and the impact of mentorship on the diagnosis of inflammatory bowel disease-associated dysplasia among subspecialist gastrointestinal pathologists.

The diagnosis of inflammatory bowel disease (IBD)-associated dysplasia is challenging, and past studies have demonstrated considerable interobserver variability in such diagnoses. This study aimed to assess interobserver agreement in IBD dysplasia diagnoses among subspecialty GI pathologists and to explore the impact of mentorship on diagnostic variability. Twelve GI pathologist mentees and 7 GI pathologist mentors reviewed 163 digitized slides. Participants rendered a diagnosis of negative for dysplasia, indefinite for dysplasia, low-grade dysplasia, or high-grade dysplasia and provided a confidence level for each case. Interobserver agreement and reliability were assessed using Cohen's and Fleiss' kappa (κ) statistics and intraclass correlation coefficient (ICC) analysis. The overall κ coefficient was 0.42 (95% CI: 0.38-0.46). The overall ICC was 0.67 (95% CI: 0.62-0.72). Κ coefficients ranged from 0.31 to 0.49 for mentor/mentee pairs and from 0.34 to 0.55 for pairs of mentees of the same mentor. The combined κ coefficient was 0.44 (95% CI: 0.39-0.48) for all mentees and 0.39 (95% CI: 0.34-0.43) for all mentors. Common features in low agreement cases included mucosal atrophy, areas of stark contrast, serrations, decreased goblet cells, absent surface epithelium, and poor orientation. Participants were confident in most diagnoses, and increased confidence levels generally correlated with higher interobserver agreement. Interobserver agreement among subspecialist GI pathologists in this curated cohort of IBD dysplasia cases was fair to moderate. Mentorship during GI pathology fellowship does not appear to be a significant factor contributing to interobserver variability, but increased experience also does not seem to improve interobserver agreement.