Epithelial Biopsies Research Articles

Loss of Cirbp expression is correlated with the malignant progression and poor prognosis in nasopharyngeal carcinoma.

Purpose: The correlation of cold-inducible RNA-binding protein (Cirbp) expression with clinicopathological features including patient prognosis in nasopharyngeal carcinoma (NPC) was investigated.Methods: The expression of Cirbp in NPC cell lines and tissue specimens was examined by qRT-PCR or immunohistochemistry (IHC).Results: Immunohistochemistry (IHC) results showed that high Cirbp expression was detected in 61 of 61 non-cancerous nasopharyngeal squamous epithelial biopsies, whereas the significantly reduced expression of Cirbp was observed in NPC specimens. In addition, IHC assay for Cirbp protein illustrated that the cells of 177 NPC samples and nasopharyngeal squamous epithlial cells displayed strong signals in nuclei and faint signals in cytoplasm, whereas Cirbp protein is mainly detected in the cell’s cytoplasm in many other cancers. More importantly, TNM classification displayed that the low expression of Cirbp was more frequently observed in T3-T4, N2-N3, M1 and III-IV NPC biopsies, and undifferentiated carcinoma (UDC) than T1-T2, N0-N1, M0 and I-II tumors, and differentiated nonkeratinizing carcinoma (DNKC), suggesting that Cirbp loss is a key molecular event in advanced cases of NPC. Kaplan–Meier survival analysis indicated that NPC patients showing lower Cirbp expression had a significantly shorter overall survival time than those with high Cirbp expression. Multivariate analysis suggested that the level of Cirbp expression was an independent prognostic indicator for NPC survival. Finally, we revealed a significant positive association between Cirbp expression and E-cadherin, and a notable negative correlation between Cirbp expression and Ki67 labeling index in NPC biopsies.Conclusion: Collectively, these findings demonstrate that loss of Cirbp expression is correlated with malignant progression and poor prognosis in NPC.

Flow based single cell analysis of the immune landscape distinguishes Barrett's esophagus from adjacent normal tissue.

Barrett’s esophagus (BE) is metaplasia of the squamous epithelium to a specialized columnar epithelium. BE progresses through low- and high-grade dysplasia before developing into esophageal adenocarcinoma. The BE microenvironment is not well defined. We compare 12 human clinical BE and adjacent normal squamous epithelium biopsies using single cell immunophenotyping by flow cytometry. A cassette of 19 epithelial and immune cell markers was used to detect differences between cellular compartments in normal and BE tissues. We found that the BE microenvironment has an immunological landscape distinct from adjacent normal epithelium. BE has an increased percentage of epithelial cells with a concomitant decrease in the percentage of immune cells, accompanied by a shift in the immune landscape from a predominantly T cell rich microenvironment in normal tissue to a B cell rich landscape in BE tissue. Hierarchical clustering separates BE and normal samples into two discrete groups based upon our 19-marker panel, but also reveals unexpected, shared phenotypes for three patients. Our results suggest that flow based single cell analysis may have the potential for revealing clinically relevant differences between BE and normal adjacent tissue, and that surface immunophenotypes could identify specific subpopulations from dysplastic tissue for further investigation.

Effects of clay mineral supplementation on particle-associated and epimural microbiota, and gene expression in the rumen of cows fed high-concentrate diet

The increased concentrate amounts in cow diets may initiate changes in both particle-associated (PaM) and epimural microbiota (EpM) with the potential for promoting the establishment of pathogens. Clay minerals have shown promising potentials in binding harmful microorganisms and metabolites due to their high adsorption capacity. This study evaluated the effects of a clay-mineral based product (CM) on PaM, EpM, fermentation parameters, and epithelial gene expression in cows fed a high-concentrate diet. Eight rumen-cannulated non-lactating Holstein cows received a concentrate mix supplemented with CM or not (CON) in a change-over design with an initial 100% roughage diet phase (RD, 1 week), followed by intermittent 65%-high-concentrate-diet phases (HC1, HC2; 1 and 2 week duration, respectively), interrupted by 1 week roughage only. Rumen samples for short-chain fatty acids, ammonia, and lactate quantification, as well as PaM, and epithelial biopsies for EpM examination and epithelial gene expression were collected via the cannula once during each feeding phase. Phylogenetic distance analysis of Illumina MiSeq sequencing of the 16S rRNA gene region V345 showed a clear clustering of RD microbiota compared to HC in PaM, showing the impact of the high-concentrate diet on the bacterial community. In the EpM this effect was less pronounced, due to higher variability in RD. In the PaM, a decrease (P < 0.01) of community diversity occurred with the onset of HC feeding, while in the EpM there was an increase in diversity (P < 0.05). In the PaM, CM increased the relative abundance of genus Butyrivibrio (P < 0.01), a commensal bacterium of the rumen, which was, with 6.4%, the second most abundant genus. There, the CM supplementation decreased the genera Lactobacillus, Fusobacterium, and Treponema (P = 0.05), which are potentially either lactate producing or opportunistic pathogens. In the EpM, CM decreased the relative abundance of Succiniclasticum genus (P < 0.01), a possible endotoxin producer, and increased bacteria that are associated with a normobiotic rumen, such as Campylobacter (P = 0.06). Barrier function genes were upregulated in HC2 and nutrient transport genes downregulated in HC1 (P < 0.05); however, there was little effect on pro-inflammatory genes at the epithelium. The CM showed a significant decreasing effect on the cellular metabolism genes HMGCS1 (P = 0.04). Our results suggest that CM supplementation can increase the relative abundance of commensal microbiota and decrease bacteria that could negatively impact the rumen milieu and health during high-concentrate feeding.

Patent Highlights December 2018–January 2019

Patent Highlights December 2018–January 2019

Eosinophilic esophagitis has a sub-epithelial regulatory adaptive immune response

Abstract Introduction Eosinophilic esophagitis (EoE) is food-triggered immune esophageal disease characterized by dysphagia and food impaction. The pathophysiology of EoE is poorly understood. Prior eosinophilic esophagitis studies showed abundant IgG4 with IgG4+ plasma cells in esophageal subepithelial tissue. Methods Seven esophagectomies and one deep biopsy from patients with active EoE were compared. Seven uninvolved esophageal areas from surgical resections for tumor and autopsy were used as controls. Histology, immunostains, RNA in situ hybridization, and RNA-seq findings were compared with EoE epithelial biopsy RNA-seq. Results The superficial lamina propria (LP) of EoE patients has dense fibrosis, tertiary lymphoid tissue with abundant B cells, IgG4 plasma cells, and Foxp3+ regulatory T cells. RNA-seq confirms this, with abundant B cell related transcripts, IGHG4, organized lymphoid tissue marker CCL21, and regulatory T cell markers. IL-10 in situ hybridization is present in mononuclear cells, in and near lymphoid aggregate germinal centers. Edema and degranulating mast cells are absent. By RNAseq, the LP of both eosinophilic esophagitis patients and controls have abundant thrombospondin-1, which could activate TGFβ. Nuclear immunostaining for phosphoSMAD2 and phosphoSMAD3 (suggesting active TGFβ), is present the epithelium and the superficial lamina propria. Conclusion The EoE LP immune response has abundant IgG4 plasma cells, regulatory T cells, and a predominantly immunoregulatory transcriptome. Some LP immunoregulatory features are present even in controls. Further work is needed to determine whether the immunoregulatory infiltrate contributes to the dysphagia-inducing subepithelial fibrosis.

Transplantation of cultured olfactory mucosal cells rescues optic nerve axons in a rat glaucoma model

PurposeTo determine whether transplantation of olfactory mucosal cells (OMCs) is able to rescue the loss of optic nerve axons after the intraocular pressure (IOP) is elevated in rats. MethodsThe IOP was raised by injection of magnetic microspheres into the anterior chamber of the eye. OMCs cultured from the adult olfactory mucosa were transplanted into the region of the optic disc. ResultsWe demonstrated that although the raised IOP returned to its normal level at six weeks, there was an irreversible 58% loss of optic nerve axons in the control group. However, the loss of the axons was reduced to 23% in the group with the transplanted OMCs. The Pattern Electroretinograms (pERG) showed that the decrement of the voltage amplitudes in association with the raised IOP was significantly alleviated in the group with transplantation of OMC. ConclusionsTransplantation of OMCs is able to rescue loss of optic nerve axons induced by raised IOP in the rats. The pERG recording suggested that the functional activities of the axons are also protected. Translational relevanceThe results demonstrated the ability of the transplanted OMCs to protect against the loss of the optic nerve axons and the loss of function caused by raised IOPs. The findings provide a basis for future human clinical trials by autografting OMCs from autologous nasal epithelial biopsies to treat or delay glaucoma diseases.

Chapter 4 - Anatomy of the olfactory mucosa

The classic notion that humans are microsmatic animals was born from comparative anatomy studies showing the reduction in the size of both the olfactory bulbs and the limbic brain relative to the whole brain. However, the human olfactory system contains a number of neurons comparable to that of most other mammals, and humans have exquisite olfactory abilities. Major advances in molecular and genetic research have resulted in the identification of extremely large gene families that express receptors for sensing odors. Such advances have led to a renaissance of studies focused on both human and nonhuman aspects of olfactory physiology and function. Evidence that olfactory dysfunction is among the earliest signs of a number of neurodegenerative and neuropsychiatric disorders has led to considerable interest in the use of olfactory epithelial biopsies for potentially identifying such disorders. Moreover, the unique features of the olfactory ensheathing cells have made the olfactory mucosa a promising and unexpected source of cells for treating spinal cord injuries and other neural injuries in which cell guidance is critical. The olfactory system of humans and other primates differs in many ways from that of other species. In this chapter we provide an overview of the anatomy of not only the human olfactory mucosa but of mucosae from a range of mammals from which more detailed information is available. Basic information regarding the general organization of the olfactory mucosa, including its receptor cells and the large number of other cell types critical for their maintenance and function, is provided. Cross-species comparisons are made when appropriate. The polemic issue of the human vomeronasal organ in both the adult and fetus is discussed, along with recent findings regarding olfactory subsystems within the nose of a number of mammals (e.g., the septal organ and Grüneberg ganglion).

Immunohistochemical detection of E7 human papillomavirus protein in pre‑malignant and malignant lesions of the uterine cervix.

Human papillomavirus (HPV) E7 protein expression is caused by HPV viral DNA integration into human cellular DNA, and is a prerequisite for the development and progression of cervical cancer. The present study aimed to evaluate the role of E7 protein as a biomarker for identification of transformed cervical epithelial cells during the early stages of cervical cancer. Specific monoclonal antibodies to the E7protein of high‑risk HPVs were generated and characterized for applications in immunocytochemistry and immunohistochemistry using cervical epithelial cells or biopsy tissue slides. The specificity and feasibility for detecting precancerous cells in cervical exfoliated epithelial cells was demonstrated. In addition, antibody staining of cervix biopsies indicated the pathological grades of cervical cancer and precancerous lesions. The results of the present study demonstrated the potential benefit of using E7 protein as a novel and specific clinical diagnostic marker to distinguish transient HPV infections from malignant and pre‑malignant lesions.

EARLY DIAGNOSIS AND TREATMENT IN PATIENT WITH A PRIMARY CILIARY DYSKINESIA (KARTAGENER SYNDROME): CASE REPORT

Primary ciliar dyskinesia is а rare orphan disease known for its multiple and variable symptoms caused by the marked genetic heterogenity beyond. As per the abundant symptoms in pediatric patients, a frequent inflammatory diseases of both upper and lower respiratory tract segments are the key points. A Sieberth-Kartagener Syndrome is a classical form of the primary ciliary dyskinesia covered such symptoms as the reversal placement of internal organs, chronic bronchoectases, nasal cavity hypoplasia and/or sinusitis. According to some foreign research, the age mediana value estimated in Eastern and Western European countries for a “Primary Ciliary Dyskinesia” diagnosis is equal to about 5 years. A lack of early diagnosis is nothing but a direct consequence of the poor level of awareness, so common for a primary health care system. This itself leads to increased rates of patients disability. This report deals with clinical peculiarities, diagnostics and treatment details observed and administered in a primary ciliary dyskinesia (Kartagener Syndrome) patient. Noteworthy, both mother’s tough obstetric-gynecological profile and a harsh course of this particular pregnancy were indeed taken into account. Up until 11 months, this patient was subjected to a non-clinical treatment on numerous respiratory infection occasions. A “Kartagener Syndrome” diagnosis was estimated in our clinic on the basis of laboratory and instrumental tests data. Thus, the latter prove a complete reversal of internal organs placement. Further, this diagnosis was re-confirmed by histomorphological patterns revealed in the nasal epithelial biopsy study. A difficulty to come up with the Kartagener Syndrome diagnosis at the pre-hospital treatment stage is in a focus of this specific case report. Noteworthy, we have succeed with the early diagnosis of Kartagener Syndrome and then with a following efficient therapy conducted in our clinic.

The Expression of AQP1 IS Modified in Lung of Patients With Idiopathic Pulmonary Fibrosis: Addressing a Possible New Target.

Activation of the epithelial-mesenchymal transition process (EMT) by which alveolar cells in human lung tissue undergo differentiation giving rise to a mesenchymal phenotype (fibroblast/miofibroblasts) has been well recognized as a key element in the origin of idiopathic pulmonary fibrosis (IPF). Here we analyzed expression of AQP1 in lung biopsies of patients diagnosed with IPF, and compared it to biopsies derived from patients with diverse lung pneumonies, such as hypersensitivity pneumonitis, sarcoidosis or normal lungs. Immunostaining for AQP1 showed a clear increment of AQP1 localized in the alveolar epithelium in biopsies from IPF patients alone. Moreover, to examine the possible participation of AQP1 in the pathophysiology of IPF, we evaluated its role in the pro-fibrotic transformation induced by transforming growth factor (TGF-β) in vitro. Human alveolar epithelial cells (A549), and fibroblasts derived from an IPF patient (LL29), or fibroblasts from healthy normal lung tissue (MRC-5), were treated with TGF-β, and levels of expression of AQP1, as well as those of E-cadherin, vimentin, α-SMA and collagen were analyzed by RT-qPCR, western blot and immunohistochemistry. An increase of AQP1 mRNA and protein after TGF-β treatment (4–72h) was observed either in A549 or IPF fibroblast-LL29 but not in MRC-5 fibroblasts. A gradual reduction of E-cadherin, and increased expression of vimentin, with no changes in α-SMA levels were observed in A549. Whereas in LL29 and MRC-5, TGF-β1 elicited a large production of collagen and α-SMA that was significantly greater in IPF fibroblast-LL29. Changes observed are consistent with activation of EMT by TGF-β, but whether modifications in AQP1 expression are responsible or independent events occurring at the same time is still unknown. Our results suggest that AQP1 plays a role in the pro-fibrotic TGF-β action and contributes to the etiology and pathophysiology of IPF. Understanding AQP1's role will help us comprehend the fate of this disease.

Using machine learning and RNA-seq to increase the accuracy and decrease the invasiveness of diagnosing eosinophilic esophagitis.

Abstract Eosinophilic esophagitis (EoE) is an esophageal disease characterized by dysphagia and food impaction triggered by food or drug hypersensitivity with a prevalence of ~1/1,000 people. A conclusive diagnosis of EoE requires endoscopy and multiple biopsies, followed by eosinophil counts in multiple high-power fields. This procedure is painful and expensive, and due to the focal nature of the lesions, misdiagnoses are possible if biopsies do not sample enough diseased tissue. RNA sequencing allows the entire transcriptome within a biological sample to be assayed, but due to the vastness of the data, bioinformatics tools are required for meaningful interpretation. We have trained different machine learning classification algorithms and characterized their performance. We show that accurate classification is generalizable to biopsies that were not controlled for depth and were sequenced using different protocols and machines. Furthermore, we assess whether EoE status can be predicted using buccal epithelial tissue biopsies, which can be collected non-invasively but lack histological change. Together, these results show that machine-learning algorithms based on RNA-seq data could enable more accurate diagnosis of EoE using less invasive and lower-cost biopsy protocols.

333 - Differential Effects of PPI Therapy on Hedgehog Pathway, SOX9, and P63 Gene Expression in Squamous Epithelial Biopsies from Gerd Patients with or without Barrett's Esophagus: Implications for Reversibility of Esophageal Epithelial Molecular Reprogramming

333 - Differential Effects of PPI Therapy on Hedgehog Pathway, SOX9, and P63 Gene Expression in Squamous Epithelial Biopsies from Gerd Patients with or without Barrett's Esophagus: Implications for Reversibility of Esophageal Epithelial Molecular Reprogramming

Alterations of Vascular Endothelial Growth Factor A Expression and Microvessel Density Associated With Polypropylene Vaginal Mesh.

Vaginal mesh exposure is the most common complication from mesh use in prolapse and incontinence surgery. Angiogenesis is an essential component of tissue healing, and defective angiogenesis plays a role in chronic wounds. We hypothesized that patients with exposures will have impaired angiogenesis as evidenced by altered tissue vascular endothelial growth factor A (VEGFA) expression and microvessel density. The study objective was to compare angiogenesis in women with vaginal mesh exposures, those with vaginal mesh without exposures, and in women who were mesh-naive. Patients undergoing polypropylene mesh removal and patients without mesh undergoing urinary incontinence or prolapse surgery were recruited. Full-thickness vaginal epithelial biopsies were obtained. The relative abundance of VEGFA RNA was measured with reverse transcription-polymerase chain reaction. The VEGFA and CD31 immunohistochemistry were also performed. Ninety-two subjects were enrolled and biopsied. Mean age (SD) was 57.2 (12.8) years, 16 (17.4%) were smokers, and 68 (73.9%) were postmenopausal.The VEGFA RNA expression did not differ between subjects with mesh exposure and with mesh but no exposure (P = 0.89). However, compared with subjects with no mesh, vaginal VEGFA expression was decreased in subjects with any implanted mesh (relative expression, 0.72; P = 0.02). Microvessel density was increased in subjects with mesh exposure compared with subjects with no mesh (P < 0.01). The VEGFA expression by immunohistochemistry was significantly lower in postmenopausal subjects without estrogen treatment compared with premenopausal and postmenopausal subjects being treated with estrogen (P = 0.02). The presence of polypropylene mesh and hormonal status are associated with evidence of altered angiogenesis.

NSAID use and somatic exomic mutations in Barrett\u2019s esophagus

BackgroundUse of aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs) has been shown to protect against tetraploidy, aneuploidy, and chromosomal alterations in the metaplastic condition Barrett’s esophagus (BE) and to lower the incidence and mortality of esophageal adenocarcinoma (EA). The esophagus is exposed to both intrinsic and extrinsic mutagens resulting from gastric reflux, chronic inflammation, and exposure to environmental carcinogens such as those found in cigarettes. Here we test the hypothesis that NSAID use inhibits accumulation of point mutations/indels during somatic genomic evolution in BE.MethodsWhole exome sequences were generated from 82 purified epithelial biopsies and paired blood samples from a cross-sectional study of 41 NSAID users and 41 non-users matched by sex, age, smoking, and continuous time using or not using NSAIDs.ResultsNSAID use reduced overall frequency of point mutations across the spectrum of mutation types, lowered the frequency of mutations even when adjusted for both TP53 mutation and smoking status, and decreased the prevalence of clones with high variant allele frequency. Never smokers who consistently used NSAIDs had fewer point mutations in signature 17, which is commonly found in EA. NSAID users had, on average, a 50% reduction in functional gene mutations in nine cancer-associated pathways and also had less diversity in pathway mutational burden compared to non-users.ConclusionsThese results indicate NSAID use functions to limit overall mutations on which selection can act and supports a model in which specific mutant cell populations survive or expand better in the absence of NSAIDs.

The Role of Wide Area Transepithelial Sampling in Screening for Esophageal Adenocarcinoma

Barrett's esophagus (BE) requires surveillance to monitor for dysplasia and esophageal adenocarcinoma. New methods have been developed to detect dysplasia and esophageal adenocarcinoma. One of these methods called the wide area trans epithelial sampling 3D brush biopsy (WATS 3D) is a computer-assisted brush-biopsy technique which uses an abrasive sampling instrument to detect dysplasia or esophageal adenocarcinoma. The aim of our case is to demonstrate the clinical utility of this tool in Barrett's surveillance. Patient is a 66 year old with a past medical history of lifelong heartburn and regurgitation. He was on Esomeprazole 20mg daily and denied family history of colon or esophageal cancer. The patient had a screening endoscopy for surveillance of BE three months ago, which showed a 3-5 cm area of full circumferential salmon colored mucosa suspicious for BE. Biopsies revealed squamocolumnar mucosa with intestinal metaplasia and focal high-grade glandular dysplasia without malignancy. Repeat endoscopy confirmed a Prague C5M5 lesion (Fig. 1). Repeat biopsies read by 2 different pathologists confirmed the diagnosis of high grade dysplasia (Fig. 2). Wide area trans epithelial sampling 3D brush biopsy was also performed at that time. The WATS pathology report stated that patient has at least high-grade dysplasia in his cytopathology and was suggestive of esophageal adenocarcinoma. Patient was successfully managed with 2 RFA ablation sessions with obliteration of the BE and replacement of the Barrett's mucosa with a neosquamous epithelium. Esophageal adenocarcinoma has a high mortality rate and a poor prognosis. As such screening for BE is critical in preventing progression. As illustrated in our case, the WATS brush biopsy addresses the sampling error inherent in random forceps biopsy testing of the esophagus. In just a few minutes, gastroenterologists can easily obtain a wide area, full-thickness transepithelial tissue sample for computer-assisted 3D laboratory analysis. The case was discussed with experts in BE and felt there was a small foci of adenocarcinoma in the setting of high grade dysplasia. Without evidence of invasion, lesion was safe to be treated with RFA. This foci of esophageal adenocarcinoma is likely missed with the traditional sampling methods. Overall, institutions should consider either adding WATS 3D as an adjunct to standard forceps biopsy or using it as primary tool for detecting dysplasia.Figure: Narrow band image of a C5M5 area of Barrett's seen on endoscopy.Figure: Few foci of highly dysplastic epithelium, seen singly and within apparent small vascular spaces, strongly suggestive of adenocarcinoma, in a background of Barrett's metaplasia.

Sessile Serrated Adenoma of the Appendix in an Asymptomatic Patient

A 63 year-old Hispanic male with history of hypertension and nephrolithiasis presented for an averagerisk colorectal cancer screening colonoscopy. During the study he was found with a large, raised submucosal lesion in cecum, specifically arising at the appendiceal orifice with a “volcano sign” appearance as seen in mucocele of the appendix (Image 1). Epithelial biopsy of this lesion was taken and reported a serrated adenomatous tissue. An abdominopelvic CT described a distended appendix with intraluminal fluid density. Occluding its orifice, there was a polypoid density measuring 2.0 cm x 2.3 cm x 1.7 cm with Hounsfield units compatible with soft tissue density (Image 2). There was no adjacent lymphadenopathy or focal inflammatory changes. An appendiceal mucocele as well as an obstructing adenocarcinoma were considered in the differential diagnosis. The patient underwent a laparoscopic right hemicolectomy. Pathology report confirmed a vermiform appendix with a sessile serrated adenoma occupying the entire length of the appendix, unremarkable margins of excision and lymph nodes (Image 3). Sessile serrated adenomas are a relatively recent described entity most commonly occurring in the right colon and rarely in the appendix. The term serrated adenomas originate from its histological findings of adenomas with saw tooth-like dysplastic epithelium found in more than 50% of basal crypts. It has been recognized that these lesions portraits microsatellites instability, which is a molecular pathway different from conventional adenomas and a risk factor for colorectal cancer. From 20 to 30% of colorectal cancers arise through the serrated polyp pathway. In the first small case series reported by Rubio in 2004, most serrated and villous adenomas of the appendix were diagnosed as incidental findings during appendectomy and autopsy, and appeared to be highly aggressive lesions in comparison to those arising from the colorectum. Except for rare case reports and a few case series, the true incidence of appendiceal serrated polyps is unknown. This case report describes a sessile serrated adenoma located in the appendix diagnosed by a screening colonoscopy and successfully treated by surgical resection.Figure: Endoscopic findings of a raised submucosal lesion arising at appendiceal orifice.Figure: The appendix is distended by intraluminal fluid density. No periappendiceal inflammatory changes. Occluding its orifice, there is a polypoid density which measures approximate 2 cm transversely x 2.3 cm AP x approximately 1.7 cm vertically. It shows Hounsfield unit measurements compatible with soft tissue density. No adjacent significant lymphadenopathy noted.Figure: The sections of the appendix in H&E stain show serrated mucosa throughout the appendix with irregular crypt branching, dilation and serration of crypt bases. These changes are limited to the mucosa with no invasion into the submucosa.

Clinicopathologic significance of in vivo antinuclear autoantibodies in oral mucosal biopsies

Although antinuclear autoantibody (ANA) staining of oral biopsy specimens is indicative of chronic ulcerative stomatitis, it is not known whether this staining is characteristic of other autoimmune diseases. Our study was undertaken to characterize the various in vivo ANA patterns detected in the oral mucosa by direct immunofluorescence to describe the associated hematoxylin and eosin findings, and determine whether patients with these findings had a coexisting systemic connective tissue disease. This was a retrospective analysis of oral biopsy specimens submitted from 2013 to 2016. In vivo ANA staining was present in 72 of the 2019 cases examined. Immunoglobulin G was the most common immunoreactant (71 of 72 cases), and speckled nuclear staining was the most frequent in vivo ANA pattern (52 of 72). In most cases, hematoxylin and eosin staining of biopsy specimens showed mucositis (24 of 34). Detailed clinical information was available for 10 patients, and all of them had an autoimmune disease. We found similar prevalence of ANA staining with direct immunofluorescence in oral epithelial biopsy specimens as reported for those of skin. In vivo ANA in the oral epithelium may indicate the presence of an immune-mediated disease. Patients who show ANA deposits in oral mucosal biopsy specimens should be investigated for systemic connective tissue disease as well as for chronic ulcerative stomatitis.

Subsquamous intestinal metaplasia is common in treatment-naïve Barrett’s esophagus

Barrett's intestinal metaplasia may extend beneath normal squamous epithelium at the squamocolumnar junction (SCJ) and therefore escape surveillance biopsy sampling. The prevalence of subsquamous intestinal metaplasia (SSIM) in patients undergoing Barrett's esophagus (BE) surveillance is unknown. Our aim was to examine the prevalence and distribution of SSIM proximal to the SCJ in patients undergoing BE surveillance. We enrolled consecutive patients with biopsy specimen-proven BE. Biopsy specimens were obtained from the squamous epithelium at 5mm and 10mm above the SCJ. The primary outcomes were the proportion of patients with SSIM at each level. We further assessed factors associated with SSIM. We examined 515 squamous epithelial biopsy specimens from 106 BE patients (95% men; mean age, 66 years) with a mean Barrett's length of 3.0 cm. SSIM was present in 39% at 5mm (95% CI, 29.4-48.6) and 21% (95% CI, 11.7-32.1) at 10mm proximal to the SCJ. Among all biopsy specimens, 13% (95% CI, 10.6-16.6) contained SSIM: 17% (95% CI, 13-21.6) of biopsy samples at 5mm and 8% (95% CI, 4.3-12.2) at 10mm proximal to the SCJ. SSIM was more common in the anterior/right lateral position compared with the posterior/left lateral position (21% vs 11%, P= .001). None of the biopsy specimens showed dysplasia. Length of BE or duration of reflux symptoms were not associated with the presence of SSIM. This cross-sectional study found a surprisingly high proportion of SSIM in treatment-naïve patients proximal to the SCJ. These findings raise questions regarding BE management and the prevalence of SSIM in normal-appearing esophagus.

Abstract 2052: Chemotherapy sensitivity, cytokine IL-8 level, and genetic expression mRNA IL-8 in gastric cancer treatment response prediction

Abstract Aims: To determine chemotherapy sensitivity in primary gastric cancer cell isolated from Thai gastric cancer patients against anticancer drugs. To search for the correlation of interleukin-6 and interleukin-8 levels in primary gastric cancer cell culture in comparison with tissue mRNA IL-8 gene expression and drug response or resistance. Method: The control background of corpus normal epithelial and gastric adenocarcinoma biopsy tissues were treated for primary cell culture with single cell isolation technique. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was performed to ex vivo examine the chemotherapy sensitivity to three anticancer drug, cisplatin, oxaliplatin, 5-Fluorouracil , and Irinotecan. Biopsy tissues from tumor site were collected before chemotherapy treatment in 63 patient who received neo-adjuvant chemotherapy by FOLFOX IV regimen due to advance stage III and IV in both resectable and un-resectable cases. The IL-8 mRNA gene expression was reported by relative quantitation real time RT-PCR. ELISA technique was performed to investigate the cytokine level from primary cultures taken from 41 gastric cancer patients who undergone biopsy from Maharaj Nakorn Chiang Mai Hospital during year 2014 to 2016. The clinical data base was parallel collected and analysed in correlation with clinical response and ex-vivo response. Results: Each patient has different sensitivity pattern to the standard anticancer drugs. There are no significantly direct correlations or difference between cytokine IL-8 levels or mRNA IL-8 expression, and the chemotherapy sensitivity to the actual drugs that are being used for treatment of gastric adenocarcinoma in individual patients. However, the chemotherapy resistance to all drugs in primary gastric cancer cell culture has trend to increase after the third cycle of chemotherapy treatment when compared to prior cell treatment with fluctuation of cytokine level. Different cytotoxic drug gives the same response result for chemrsensitivity test in this population. The author observed the down regulation of tissue mRNA IL-8 expression and cytokine level in the good responders. Conclusion: The results provide an additional basis of information for primary chemotherapy selection in the individualised treatment for advance gastric adenocarcinoma patients. Therefore, the author suggest to select the less toxicity cytotoxic drug for being primary chemotherapy of choice. Cytokine gene level and mRNA IL-8 gene expression show individually significantly change after chemotherapy treatment in good responder. This result may lead us to find some new targeted therapy or additional treatment for its suppression. Citation Format: Sirikan Limpakan (Yamada). Chemotherapy sensitivity, cytokine IL-8 level, and genetic expression mRNA IL-8 in gastric cancer treatment response prediction [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2052. doi:10.1158/1538-7445.AM2017-2052

Regional and temporal heterogeneity of epithelial ovarian cancer tumor biopsies: implications for therapeutic strategies.

Stage III/IV epithelial ovarian cancer (EOC) is a systemic disease. The clonal relationship among different tumor lesions at diagnosis (spatial heterogeneity) and how tumor clonal architecture evolves over time (temporal heterogeneity) have not yet been defined. Such knowledge would help to develop new target-based strategies, as biomarkers which can adjudge the success of therapeutic intervention should be independent of spatial and temporal heterogeneity.The work described in this paper addresses spatial and temporal heterogeneity in a cohort of 71 tumor biopsies using targeted NGS technology. These samples were taken from twelve high grade serous (HGS) and seven non HSG-EOC, both at the time of primary surgery when the tumor was naïve to chemotherapy and after chemotherapy.Matched tumor lesions growing in the ovary or at other anatomical sites show very different mutational landscapes with branched tumor evolution. Mutations in ATM, ATR, TGFB3, VCAM1 and COL3A1 genes were shared across all lesions. BRCA1 and BRCA2 genes were frequently mutated in synchronous lesions of non HGS-EOC. Relapsed disease seems to originate from resistant clones originally present at the time of primary surgery rather than from resistance acquired de novo during platinum based therapy. Overall the work suggests that EOC continues to evolve. More detailed mapping of genetic lesions is necessary to improve therapeutic strategies.