Development of an in vivo murine model of perineural invasion and spread of cutaneous squamous cell carcinoma of the head and neck.

Introduction: Cutaneous squamous cell carcinoma of the head and neck (cSCCHN) can metastasize by invading nerves and spread toward the central nervous system. This metastatic process is called perineural invasion (PNI) and spread (PNS). An in vivo sciatic nerve mouse model is used for cSCCHN PNI/PNS. Here we describe a complementary whisker pad model which allows for molecular studies investigating drivers of PNI/PNS in the head and neck environment. Methods: A431 cells were injected into the whisker pads of BALB/c Foxn1nu and NSG-A2 mice. Tumor progression was monitored by bioluminescence imaging and primary tumor resection was performed. PNI was detected by H&E and IHC. Tumor growth and PNI were assessed with inducible ablation of LOXL2. Results: The rate of PNI development in mice was 10%-28.6%. Tumors exhibited PNI/PNS reminiscent of the morphology seen in the human disease. Our model's utility was demonstrated with inducible ablation of LOXL2 reducing primary tumor growth and PNI. Discussion: This model consists in a feasible way to test molecular characteristics and potential therapies, offers to close a gap in the described in vivo methods for PNI/PNS of cSCCHN and has uses in concert with the established sciatic nerve model.

The TAS1R2 sweet taste receptor regulates skeletal muscle mass and fitness.

Muscle fitness and mass deteriorate under the conditions of obesity and aging for reasons yet to be fully elucidated. Herein, we describe a novel pathway linking peripheral nutrient sensing and skeletal muscle function through the sweet taste receptor TAS1R2 and the involvement of ERK2-PARP1-NAD signaling axis. Muscle-specific deletion of TAS1R2 (mKO) in mice produced elevated NAD levels due to suppressed PARP1 activity, improved mitochondrial function, increased muscle mass and strength, and prolonged running endurance. Deletion of TAS1R2 in obese or aged mice also ameliorated the decline in muscle mass and fitness arising from these conditions. Remarkably, partial loss-of-function of TAS1R2 (rs35874116) in older, obese humans recapitulated the healthier muscle phenotype displayed by mKO mice in response to exercise training. Our findings show that inhibition of the TAS1R2 signaling in skeletal muscle is a promising therapeutic approach to preserve muscle mass and function.

High incidence of inflammatory bowel disease in Northern Australia: a prospective community population-based Australian incidence study in the Mackay-Isaac-Whitsunday region.

BACKGROUND AND AIMS: To determine the incidence of inflammatory bowel disease (IBD) in the Mackay-Isaac-Whitsunday region in Northern Queensland (-21.14° S) and to allow a comparison with Southern Australian and New Zealand data (Geelong, Australia -38.14° S; Tasmania -41.43° S and -42.88° S (Launceston and Hobart) and Canterbury, New Zealand -43.46 °S). DESIGN: A prospective observational community population-based IBD study was conducted between 1 June 2017 and 31 May 2018. OUTCOME MEASURES: Primary includes the crude annual incidence rate of IBD, Crohn's disease (CD), ulcerative colitis (UC) and inflammatory bowel disease-unclassified (IBDU), while secondary includes disease phenotype and behaviour. RESULTS: Fifty-six new cases of IBD were identified. Twenty-three were CD, 30 were UC and 3 were IBDU. The crude annual incidence rate per 100 000 for IBD, CD, UC and IBDU were 32.2 (95% confidence interval (CI): 24.78-41.84), 13.23 (95% CI: 8.79-19.90), 17.25 (95% CI: 12.06-24.67) and 1.73 (95% CI: 0.56-5.35). When directly age-standardised to the World Health Organisation Standard Population Distribution, the overall CD, UC and IBDU incidence were 13.19, 17.34 and 1.85 per 100 000, with an overall age-standardised IBD incidence of 32.38. CONCLUSIONS: This is the first study to define the incidence of IBD in a Northern Australian cohort and to allow a comparison between North and Southern Australia. The IBD crude is the highest reported in Australia. Like others, we found a high and low incidence of upper gastrointestinal Crohn's disease and complicated disease at diagnosis respectively, likely reflective of the increased availability and early uptake of endoscopic procedures.

Cystic lesions of the retrorectal space.

Cysts of the retrorectal space comprise a heterogeneous group of rare lesions. Most develop from embryological remnants and include tailgut cysts, dermoid cysts, rectal duplication cysts, anal canal duplication cysts, sacrococcygeal teratomas and anterior meningocoele. Tailgut cyst is the most common cyst of developmental origin, usually presenting as a multilocular cystic mass with mucoid content and lined by multiple epithelial types. Compared with tailgut cysts, rectal duplication cysts display all layers of the large bowel wall including a well-defined muscularis propria. Retrorectal cysts of non-developmental origin are far less common and represent lesions that either infrequently involve the retrorectal space or undergo extensive cystic change. This review provides an overview of the various histological types of cystic lesions of the retrorectal space, divided into cysts of developmental origin and those of non-developmental origin. A practical pathological and multidisciplinary approach to diagnosing these lesions is presented.

Saccharin Stimulates Insulin Secretion Dependent on Sweet Taste Receptor-Induced Activation of PLC Signaling Axis.

BACKGROUND: Saccharin is a common artificial sweetener and a bona fide ligand for sweet taste receptors (STR). STR can regulate insulin secretion in beta cells, so we investigated whether saccharin can stimulate insulin secretion dependent on STR and the activation of phospholipase C (PLC) signaling. METHODS: We performed in vivo and in vitro approaches in mice and cells with loss-of-function of STR signaling and specifically assessed the involvement of a PLC signaling cascade using real-time biosensors and calcium imaging. RESULTS: We found that the ingestion of a physiological amount of saccharin can potentiate insulin secretion dependent on STR. Similar to natural sweeteners, saccharin triggers the activation of the PLC signaling cascade, leading to calcium influx and the vesicular exocytosis of insulin. The effects of saccharin also partially require transient receptor potential cation channel M5 (TRPM5) activity. CONCLUSIONS: Saccharin ingestion may transiently potentiate insulin secretion through the activation of the canonical STR signaling pathway. These physiological effects provide a framework for understanding the potential health impact of saccharin use and the contribution of STR in peripheral tissues.

Sampling and Reporting of Inflammatory Bowel Disease.

Pathologists have an important and expanding role in the diagnosis and management of inflammatory bowel disease. This role includes the initial diagnosis of the disease, assessment of the response to treatment and the identification of short-term complications such as cytomegalovirus infection and long-term complications such as dysplasia. Furthermore, the assessment of resection specimens for complication of disease is important to determining the risk of subsequent disease or inflammation within an ileal pouch. Adequate sampling of the disease at endoscopy and from the surgical resection specimen is vital to determining the ultimate information that can be provided by the pathologist. This sampling is determined by the clinical scenario. Similarly, a standardized approach to reporting and synthesizing the histologic findings will improve patient management. This is best exemplified by the increasing interest in histologic activity indices, such as the Nancy index in ulcerative colitis, and in the standardized reporting for inflammatory bowel disease dysplasia recommended by the SCENIC international consensus.

RAF1 rearrangements are common in pancreatic acinar cell carcinomas.

There is now evidence that gene fusions activating the MAPK pathway are relatively common in pancreatic acinar cell carcinoma with potentially actionable BRAF or RET fusions being found in ~30%. We sought to investigate the incidence of RAF1 fusions in pancreatic malignancies with acinar cell differentiation. FISH testing for RAF1 was undertaken on 30 tumors comprising 25 'pure' acinar cell carcinomas, 2 mixed pancreatic acinar-neuroendocrine carcinomas, 1 mixed acinar cell-low grade neuroendocrine tumor and 2 pancreatoblastomas. RAF1 rearrangements were identified in 5 cases and confirmed by DNA and RNA sequencing to represent oncogenic fusions (GATM-RAF1, GOLGA4-RAF1, PDZRN3-RAF1, HERPUD1-RAF1 and TRIM33-RAF1) and to be mutually exclusive with BRAF and RET fusions, as well as KRAS mutations. Large genome-wide copy number changes were common and included 1q gain and/or 1p loss in all five RAF1 FISH-positive acinar cell carcinomas. RAF1 expression by immunohistochemistry was found in 3 of 5 (60%) of fusion-positive cases and no FISH-negative cases. Phospho-ERK1/2 expression was found in 4 of 5 RAF1-fusion-positive cases. Expression of both RAF1 and phospho-ERK1/2 was heterogeneous and often only detected at the tumor-stroma interface, thus limiting their clinical utility. We conclude that RAF1 gene rearrangements are relatively common in pancreatic acinar cell carcinomas (14.3% to 18.5% of cases) and can be effectively identified by FISH with follow up molecular testing. The combined results of several studies now indicate that BRAF, RET or RAF1 fusions occur in between one third and one-half of these tumors but are extremely rare in other pancreatic malignancies. As these fusions are potentially actionable with currently available therapies, a strong argument can be made to perform FISH or molecular testing on all pancreatic acinar cell carcinomas.

Clinicopathological features of pyloric gland adenomas of the duodenum: a multicentre study of 57 cases.

AIMS: To determine the clinicopathological features of pyloric gland adenomas (PGA) that arise in the duodenum. METHODS AND RESULTS: Fifty-seven cases of duodenal PGA were identified and analysed from 56 patients. Clinicopathological and immunohistochemical analyses were performed. PGA tend to occur in older individuals (median age = 73.5), with a slight female predominance (25 males, 31 females). PGA arise more commonly in the proximal duodenum (68.75% in D1, 25% in D2 and 6.25% in D3) and usually present as mucosal nodules (98.2%) or plaques (1.8%), with a mean size of 14.8 mm. There is associated gastric heterotopia in 22.8% of cases. PGA showing features of high-grade dysplasia were significantly larger in size than PGA, showing only low-grade dysplasia (23.1 versus 8.7 mm; P = 0.0001) and more likely to show a tubulovillous rather than a pure tubular architecture (P = 0.025). In our series, 10 of 56 patients had intramucosal or invasive carcinoma associated with the duodenal PGA (17.9%). Three of these carcinomas showed lymph node metastasis. Following definitive treatment, local recurrence occurred in only three patients. CONCLUSIONS: Duodenal PGA tend to occur in the proximal duodenum of older individuals. Larger size and tubulovillous architecture correlates with high-grade dysplasia and associated adenocarcinoma. The low recurrence rate of these lesions would suggest that endoscopic management is appropriate, provided that the lesion can be completely resected.

RET gene rearrangements occur in a subset of pancreatic acinar cell carcinomas.

Pancreatic acinar cell carcinoma is relatively rare (1 to 2% of pancreatic malignancies) but may be under-recognized. In contrast to pancreatic ductal adenocarcinoma, most acinar cell carcinomas lack mutations in KRAS, DPC, CDKN2A or TP53, but appear to have a high incidence of gene rearrangements, with up to 20% reported to be driven by BRAF fusions. With the development of a new class of RET-specific tyrosine kinase inhibitors, which appear to have particularly strong activity against RET gene rearranged tumours, there is now considerable interest in identifying RET gene rearrangements across a wide range of cancers. RET rearrangements have been reported to occur at a very low incidence (<1%) in all pancreatic carcinomas. We postulated that given its unique molecular profile, RET gene rearrangements may be common in acinar cell carcinomas. We performed fluorescent in-situ hybridization (FISH) studies on a cohort of 40 acinar cell spectrum tumours comprising 36 pure acinar cell carcinomas, three pancreatoblastomas and one mixed acinar-pancreatic neuroendocrine tumour. RET gene rearrangements were identified in 3 (7.5%) cases and BRAF gene rearrangements in 5 (12.5%). All gene rearranged tumours were pure acinar cell carcinomas. Our findings indicate that amongst all pancreatic carcinomas, acinar carcinomas are highly enriched for potentially actionable gene rearrangements in RET or BRAF. FISH testing is inexpensive and readily available in the routine clinical setting and may have a role in the assessment of all acinar cell carcinomas-at this stage to recruit patients for clinical trials of new targeted therapies, but perhaps in the near future as part of routine care.

Spectrum of gastrointestinal tract pathology in a multicenter cohort of 43 Cowden syndrome patients.

Most patients with Cowden syndrome have lesions in the gastrointestinal tract, characterized by multiple polyps of various histologic types in the large bowel, polyps in the upper gastrointestinal tract, and esophageal glycogenic acanthosis. However, pathologists are often unaware of the distinctive polyposis phenotype of Cowden syndrome. In this multicenter study, we report the spectrum of gastrointestinal manifestations in a series of 43 Cowden syndrome patients who had at least one endoscopy. The median age at the first endoscopy was 46 years and 58% were women. In 24 of 29 (83%) tested patients, a pathogenic germline mutation in PTEN was identified. The histology from 199 endoscopy procedures (67 upper gastrointestinal endoscopy and 132 colonoscopies) was reviewed. Hamartomatous polyps of the large bowel were the most common lesions, present in 85% of patients. Hamartomatous polyps showed varied histology, including lymphoid aggregates in 55% of patients, a lipomatous component in 52%, a ganglioneuromatous component in 52%, and a fibrous-rich component in 14%. Polyps with at least two different stromal components were found in 55% of patients. Inflammatory polyps were present in 21% of patients. Conventional adenomas and serrated polyps were identified in 48% and 62% of patients, respectively. In the upper gastrointestinal tract, the most common lesions were esophageal glycogenic acanthosis (37%), gastric hamartomatous polyps (47%), and duodenal hamartomatous polyps (20%). All patients with glycogenic acanthosis who had a colonoscopy had hamartomatous polyps of the large bowel. In five patients, the diagnosis of Cowden syndrome was established after the pathology report raised suspicion for the diagnosis. Pathologists who are aware of the characteristic admixture of lesions in Cowden syndrome can play an essential role in recommending referral to genetic counseling and gene testing. Early diagnosis of Cowden syndrome is important, as these patients and their relatives are at increased risk for developing multiple cancers.

Actinomyces in Crohn's-like appendicitis.

AIMS: Appendicitis with a Crohn's-like histological appearance generally raises concern for Crohn's disease, Yersinia infection, and interval appendectomy. Actinomyces infection is a recognised cause of chronic appendicitis that can histologically mimic Crohn's disease. METHODS AND RESULTS: We report on 20 cases of appendicitis with Crohn's-like histological features that were due to Actinomyces. Most patients presented with acute or chronic abdominal pain. Imaging studies suggested a mass in five cases. Two patients had interval appendectomy. Histological features showed Crohn's-like appendicitis in 16 cases, with moderate to marked fibrosis and granulomas in seven cases. The other four cases had less consistent histological findings. None of the patients developed Crohn's disease during the follow-up interval (median, 37 months). CONCLUSIONS: Actinomyces can be associated with Crohn's-like appendicitis with marked fibrosis, transmural inflammation, lymphoid hyperplasia, and granulomas.

Australasian Gastrointestinal Pathology Society (AGPS) consensus guidelines for universal defective mismatch repair testing in colorectal carcinoma.

Lynch syndrome is the most common hereditary form of colorectal carcinoma caused by a constitutional pathogenic mutation in a DNA mismatch repair gene. Identifying Lynch syndrome is essential to initiate intensive surveillance program for the patient and affected relatives. On behalf of the Australasian Gastrointestinal Pathology Society (AGPS), we present in this manuscript consensus guidelines for Lynch syndrome screening in patients with colorectal carcinoma. The goal of this consensus document is to provide recommendations to pathologists for diagnosis of Lynch syndrome with discussion of the benefits and limitations of each test. Universal screening for defective mismatch repair is recommended, in agreement with the recent endorsement of universal testing by the National Health and Medical Research Council in Australia and the New Zealand Ministry of Health. The value of evaluating defective mismatch repair is acknowledged not only for Lynch syndrome screening but also for therapeutic decision information in patient management. AGPS advocates appropriate government funding for the molecular tests necessary for Lynch syndrome screening (BRAF mutation, MLH1 methylation testing).

Gastroenteropancreatic neuroendocrine neoplasms: selected pathology review and molecular updates.

Gastroenteropancreatic (GEP) neuroendocrine neoplasms can be broadly separated into well- and poorly differentiated categories. Tumours within each category have similarities in morphology and immunophenotype, but vary in grade, behaviour, molecular signature and responses to therapy. The aetiology of these differences is multifactorial. Site of origin, mucosal milieu and hereditary influences are some of the currently known factors. Given these differences, staging and grading systems continue to evolve, and the most recent World Health Organization classification of pancreatic neuroendocrine neoplasms reflects this by introducing a grade 3 neuroendocrine tumour category for morphologically well-differentiated tumours with an elevated Ki-67 proliferation index and/or mitotic count. This review aims to highlight current classification guidelines with discussion of unique site-specific features of selected GEP neuroendocrine neoplasms and an emphasis on practical issues related to daily reporting.

Brunner's gland cyst: a clinicopathological study of 25 cases highlighting an underappreciated lesion.

Brunner's gland cysts are rare, benign lesions of the duodenum usually seen as incidental, polypoid or cystic lesions of the duodenum. We present a series of 25 cases of Brunner's gland cyst, the largest case series to date. All the cases were identified during endoscopic assessment for unrelated causes. The mean age of the patients was 66.2 years and the sex ratio was approximately equal. Most were detected as a small (3-10 mm) polypoid lesion in the second part of the duodenum, away from the ampulla. Most of the cysts were unilocular and all were lined by undulating, cytologically bland cuboidal to columnar cells with clear cytoplasm and small, basal nuclei. No mitotic activity or proliferative activity was seen. No recurrence was noted, despite incomplete removal in many cases. We agree with the hypothesis that Brunner's gland cysts are likely to be caused by local obstruction to the draining duct of Brunner's glands.

Early Barrett esophagus-related neoplasia in segments 1 cm or longer is always associated with intestinal metaplasia.

The assumption that intestinal metaplasia is a prerequisite for intraepithelial neoplasia/dysplasia and adenocarcinoma in the distal esophagus has been challenged by observations of adenocarcinoma without associated intestinal metaplasia. This study describes our experience of intestinal metaplasia in association with early Barrett neoplasia in distal esophagus and gastroesophageal junction. We reviewed the first endoscopic mucosal resection of 139 patients with biopsy-proven neoplasia. In index endoscopic mucosal resection, 110/139 (79%) cases showed intestinal metaplasia. Seven had intestinal metaplasia on prior biopsy specimens and three had intestinal metaplasia in subsequent specimens, totaling 120/139 (86%) patients showing intestinal metaplasia at some point supporting the theory of sampling error for absence of intestinal metaplasia in some cases. Those without intestinal metaplasia (13%) were enriched for higher stage disease (T1a Stolte m2 or above) supporting the assertion of obliteration of intestinal metaplasia by the advancing carcinoma. All cases of intraepithelial neoplasia and T1a Stolte m1 carcinomas had intestinal metaplasia (42/42). The average density of columnar-lined mucosa showing goblet cells was significantly less in shorter segments compared to those ≥3 cm (0.31 vs 0.51, P=0.0304). Cases where segments measured less than 1 cm were seen in a higher proportion of females and also tended to lack intestinal metaplasia. We conclude that early Barrett neoplasia is always associated with intestinal metaplasia; absence of intestinal metaplasia is attributable to sampling error or obliteration of residual intestinal metaplasia by neoplasia and those with segments less than 1 cm show atypical features for Barrett-related disease (absent intestinal metaplasia and female gender), supporting that gastroesophageal junction adenocarcinomas are heterogeneous.

Histopathological findings of extra-ileal manifestations at initial diagnosis of Crohn's disease-related ileitis.

Crohn's disease is a chronic inflammatory disorder that can affect any part of the gastrointestinal tract. Our objective was to review the histological findings in index biopsies from the terminal ileum and other gastro-intestinal tract sites of Crohn's disease patients prior any treatment and to compare them with the findings from patients with non-specific ileitis. A total of 111 consecutive Crohn's disease cases (55 females, median age 27 years) with extra-ileal biopsies were retrospectively selected. Upper gastrointestinal inflammatory changes were found in 68 % of gastric biopsies, 60 % of oesophageal biopsies and 43 % of duodenal biopsies with no significant difference in frequency between paediatric and adult cases. Crohn's colitis was more common in paediatric cases than adult cases (85 % versus 57 %). Granuloma in at least one extra-ileal site was observed in 40 %, more frequently in paediatric cases than in adults (78 vs 27 %). Compared with Crohn's disease cases, the group of 151 non-specific ileitis cases (75 females, median age 52 years) were more likely to have normal upper and lower gastrointestinal biopsies and to show less frequent crypt architectural changes in the terminal ileum. In summary, Crohn's disease ileitis is often associated with inflammation elsewhere in the gastrointestinal tract while non-specific ileitis was infrequently associated with inflammation elsewhere for both paediatric and adult patients. These findings support the role of systematic biopsies in multiple gastrointestinal sites to help distinguishing Crohn's ileitis from non-specific ileitis in paediatric and adult population.

Expression profiling of cutaneous squamous cell carcinoma with perineural invasion implicates the p53 pathway in the process.

Squamous cell carcinoma (SCC) is the second most common cancer worldwide and accounts for approximately 30% of all keratinocyte cancers. The vast majority of cutaneous SCCs of the head and neck (cSCCHN) are readily curable with surgery and/or radiotherapy unless high-risk features are present. Perineural invasion (PNI) is recognized as one of these high-risk features. The molecular changes during clinical PNI in cSCCHN have not been previously investigated. In this study, we assessed the global gene expression differences between cSCCHN with or without incidental or clinical PNI. The results of the analysis showed signatures of gene expression representative of activation of p53 in tumors with PNI compared to tumors without, amongst other alterations. Immunohistochemical staining of p53 showed cSCCHN with clinical PNI to be more likely to exhibit a diffuse over-expression pattern, with no tumors showing normal p53 staining. DNA sequencing of cSCCHN samples with clinical PNI showed no difference in mutation number or position with samples without PNI, however a significant difference was observed in regulators of p53 degradation, stability and activity. Our results therefore suggest that cSCCHN with clinical PNI may be more likely to contain alterations in the p53 pathway, compared to cSCCHN without PNI.

Pathology of Perineural Spread.

The perineural space is a compartment located between the nerve axons, supporting cells and tissues, and the epineural fibrous sheath. Tumor cells invade this space in response to a complex interplay of trophic factors in the local microenviroment. This attraction of tumor cells to nerves is referred to as neurotropism. The perineural space provides a conduit for tumor spread beyond the primary site of tumor occurrence. Perineural tumor growth is of two types: perineural invasion, affecting small unnamed nerves; and perineural spread, affecting larger, named nerves and presenting with clinical symptoms related to the involved nerve. Both forms of perineural tumor growth represent an adverse prognostic feature and are an essential element of the histopathologic reporting of malignancies of the head and neck region. Perineural spread is associated with decreased overall survival. Endoneurial invasion frequently accompanies perineural spread. The epineurium is more resistant to invasion and represents an important barrier to tumor spread. Immunohistochemical stains such as broad-spectrum keratin can aid in defining the proximal extent of perineural tumor spread.