Human Esophageal Epithelium Research Articles

A Zebrafish Model for Studies on Esophageal Epithelial Biology.

Mammalian esophagus exhibits a remarkable change in epithelial structure during the transition from embryo to adult. However, the molecular mechanisms of esophageal epithelial development are not well understood. Zebrafish (Danio rerio), a common model organism for vertebrate development and gene function, has not previously been characterized as a model system for esophageal epithelial development. In this study, we characterized a piece of non-keratinized stratified squamous epithelium similar to human esophageal epithelium in the upper digestive tract of developing zebrafish. Under the microscope, this piece was detectable at 5dpf and became stratified at 7dpf. Expression of esophageal epithelial marker genes (Krt5, P63, Sox2 and Pax9) was detected by immunohistochemistry and in situ hybridization. Knockdown of P63, a gene known to be critical for esophageal epithelium, disrupted the development of this epithelium. With this model system, we found that Pax9 knockdown resulted in loss or disorganization of the squamous epithelium, as well as down-regulation of the differentiation markers Krt4 and Krt5. In summary, we characterized a region of stratified squamous epithelium in the zebrafish upper digestive tract which can be used for functional studies of candidate genes involved in esophageal epithelial biology.

PTH-178 In vitroandex vivomodels of topical alginate protection of human oesophageal epithelium in gastro-oesophageal reflux disease

Introduction Up to 40% of patients with gastro-oesophageal reflux disease remain symptomatic despite PPI therapy. In addition to acid the refluxate contains pepsin and bile acids, which may be important in symptom perception. A topical therapy could protect against all components of the refluxate. A reliable model for testing could be used to screen compounds for clinical evaluation. We aimed to evaluate and compare the topical protective effect of an alginate solution in a cell culture and human biopsy model. Method Primary human oesophageal epithelial cells were established into multilayer stratified squamous epithelia using an air-liquid interface technique. After placing in Ussing chambers the apical surface was “protected” with 200 µl of either alginate, viscous control, or was unprotected. After 15 min washing in neutral solution the tissue was exposed to pH 3 + bile acid solution for 30 min and TER change was calculated. Distal human oesophageal biopsies were taken, placed in Ussing chambers and “protected” as above. After 15 min or 60 min washing in neutral solution, change in TER on 30 min exposure to pH 1 + bile acid + pepsin solution was measured. Thickness of a fluorescent-labelled alginate coating on the biopsy luminal surface was measured after 15 min (5 biopsies) and 60 min (5 biopsies) washing in pH 7.4 solution. Results Acid-induced change in TER was significantly less in alginate-protected epithelium than in epithelium protected with viscous control or unprotected tissue. This was seen in both the cell culture (–1.2 ± 5.1%, n = 5 vs. –38.4 ± 6.2%, n = 4 vs. –41.9 ± 8.1%, n = 3) and human biopsy models (−2.7 ± 1.8% vs. −15.26 ± 3.7% vs. −21.1 ± 4.4%, all n = 12) after 15 min washing. Statistically significant protection was still seen after 60 min washing (–8.3 ± 2.2% vs. –25.1 ± 4.5% vs. –26.4 ± 5.3%, n = 12). After 15 min washing the mean alginate layer thickness was 5.7 ± 0.3 µm, and was 4.3 ± 0.3 µm after 60 min washing. Conclusion Alginate topical protection of human oesophageal epithelium against acid-injury is demonstrated in both cell culture and human biopsy models. Epithelial adherence persists for up to 60 min. Topical alginate mucosal protection may be promising as an add-on/alternative therapy for gastro-oesophageal reflux disease. Since the cell culture model was able to reproduce ex vivobiopsy results, it could be an effective tool to screen candidate products prior to clinical evaluation. Disclosure of interest F. Batista-Lima: None Declared, C. Lee: None Declared, D. Sifrim Grant/ Research Support from: Research grant from Reckitt-Benckiser, P. Woodland: None Declared.

Ochratoxin A induces DNA damage and G2 phase arrest in human esophageal epithelium Het-1A cells in vitro.

Ochratoxin A (OTA), a toxin produced by several species of Aspergillus and Penicillium, is one of the most abundant food-contaminating mycotoxins. The International Agency for Research on Cancer (IARC) has classified OTA as a possible human carcinogen. Our previous study showed that there were high levels of OTA contaminations in wheat in the areas with high incidence of esophageal cancer in north China. This finding suggests that exposure to low levels of OTA may be a critical etiological factor for esophageal cancer in these areas. However, up to now, the potential biological effects of OTA on human esophageal epithelial cells have not been fully elucidated. In the present study, we explored the cytotoxicity of OTA in human esophageal epithelium immortalized cells (Het-1A). We found that OTA could induce DNA strand breaks and chromosome aberrations in Het-1A cells. OTA-induced DNA damage was followed by G2 cell cycle arrest, and down-regulation of Cdc2 and cyclinB1 contributed to the OTA-induced G2 arrest in Het-1A cells. Additionally, OTA induced apoptosis in Het-1A cells by activating caspase-3. In conclusion, our results indicated that OTA could induce DNA damage, G2 arrest and apoptosis in Het-1A cells, which may be involved in the esophageal toxicity of OTA.

Sterigmatocystin induces G1 arrest in primary human esophageal epithelial cells but induces G2 arrest in immortalized cells: key mechanistic differences in these two models

Sterigmatocystin (ST), a mycotoxin commonly found in food and feed commodities, has been classified as a "possible human carcinogen." Our previous studies suggested that ST exposure might be a risk factor for esophageal cancer and that ST may induce DNA damage and G2 phase arrest in immortalized human esophageal epithelial cells (Het-1A). To further confirm and explore the cellular responses of ST in human esophageal epithelia, we comparatively evaluated DNA damage, cell cycle distribution and the relative mechanisms in primary cultured human esophageal epithelial cells (EPC), which represent a more representative model of the in vivo state, and Het-1A cells. In this study, we found that ST could induce DNA damage in both EPC and Het-1A cells but led to G1 phase arrest in EPC cells and G2 phase arrest in Het-1A cells. Furthermore, our results indicated that the activation of the ATM-Chk2 pathway was involved in ST-induced G1 phase arrest in EPC cells, whereas the p53-p21 pathway activation in ST-induced G2 phase arrest in Het-1A cells. Studies have demonstrated that SV40 large T-antigen (SV40LT) may disturb cell cycle progression by inactivating some of the proteins involved in the G1/S checkpoint. Het-1A is a non-cancerous epithelial cell line immortalized by SV40LT. To evaluate the possible perturbation effect of SV40LT on ST-induced cell cycle disturbance in Het-1A cells, we knocked down SV40LT of Het-1A cells with siRNA and found that under this condition, ST-induced G2 arrest was significantly attenuated, whereas the proportion of cells in the G1 phase was significantly increased. Furthermore, SV40LT-siRNA also inhibited the activation of the p53-p21 signaling pathway induced by ST. In conclusion, our data indicated that ST could induce DNA damage in both primary cultured and immortalized esophageal epithelial cells. In primary human esophageal epithelial cells, ST induced DNA damage and then triggered the ATM-Chk2 pathway, resulting in G1 phase arrest, whereas in SV40LT-immortalized human esophageal epithelial cells, SV40LT-mediated G1 checkpoint inactivation occurred, and ST-DNA damage activated p53-p21 signaling pathway, up-regulating G2/M phase regulatory proteins and finally leading to a G2 phase arrest. Thus, the SV40LT-mediated G1 checkpoint inactivation is responsible for the difference in the cell cycle arrest by ST between immortalized and primary cultured human esophageal epithelial cells.

Angiotensin IV and the human esophageal mucosa: An exploratory study in healthy subjects and gastroesophageal reflux disease patients

The human esophageal mucosa expresses various components of the renin-angiotensin system (RAS), e.g. the main effector peptide angiotensin II (AngII). The aim of this study was to investigate the esophageal presence of angiotensin III (AngIII) and angiotensin IV (AngIV) forming enzymes and the AngIV receptor (AT4R). The aim was also to study the actions of AngIV and to look for aberrations in patients with gastroesophageal reflux disease (GERD). Esophageal biopsies were collected from healthy volunteers (n: 19) and individuals with erosive reflux disease (n: 14). Gene transcripts and protein expression of aminopeptidase A, -B and -M, and the AT4R were investigated by reverse transcriptase polymerase chain reaction (rt-PCR), western blot (WB) and immunohistochemistry (IHC). The functional impact of AngIV was examined in an Ussing chamber. Aminopeptidase A, -B and -M and the AT4R were expressed in the esophageal epithelium. The AT4R was less prominent in certain areas in the mucosa of reflux patients. AngIV influenced the esophageal epithelial ion transport. The impact was lower in patients with GERD. The AT4R and formation enzymes of AngIII and AngIV are present in the human esophageal epithelium. Moreover, the present results suggest that AngIV exert regulatory impact on the epithelium and that RAS is involved in mucosal aberrations associated with GERD.

Tissue resistance in the normal and diseased esophagus

This paper presents commentaries on reflux-induced injury of human esophageal epithelium; inflammation in human reflux esophagitis; motor consequences of reflux-induced inflammation in esophageal epithelium; the microscopic morphology of esophageal squamous epithelium; intraluminal impedance in the evaluation of the esophageal mucosa; endoscopic tissue morphology of esophageal squamous epithelium; and the developmental biology of esophageal squamous epithelium.

A standardized method for measuring intercellular spaces in esophageal biopsies in patients with suspected gastroesophageal reflux disease (the intercellular space ratio)

Objective. Dilated intercellular space (DIS) in esophageal biopsies is regarded as a possible early sign of mucosal injury in gastroesophageal reflux disease (GERD). This study presents a standardized approach of intercellular space measurement. Material and methods. Distal and proximal esophageal biopsies were taken from 19 patients with suspected GERD, and examined with TEM. A grid containing 150 line-crossing points was applied upon each photomicrograph. The number of points falling on the intercellular space was divided by the total number of points of the grid, thereby creating a ratio called the intercellular space ratio (ISR). The ISR method was validated with regard to intra- and interobserver agreement, and was compared to a widely used method for measuring intercellular space diameter developed by Tobey et al. (Tobey NA, Carson JL, Alkiek RA, Orlando RC. Dilated intercellular spaces: a morphological feature of acid reflux–damaged human esophageal epithelium. Gastroenterology 1996;111(5):1200–1205). The ISR was also compared to other markers for GERD. Results. Pearson's correlation coefficients for intra- and interobserver agreement were 0.91 (p < 0.001) and 0.82 (p < 0.001), respectively. The Pearson's correlation coefficient between the ISR and the intercellular space diameter according to Tobey et al., measured in the same micrographs, was 0.32 (p < 0.001). The proximal ISR correlated significantly with the distal ISR (Spearman's rho = 0.57, p = 0.010), and with heartburn symptom score (Spearman's rho = 0.50, p = 0.028). Conclusions. The ISR showed a high intraobserver and interobserver agreement. It also displayed good external validity when compared to other markers for gastroesophageal reflux. A rather poor correlation was however found between the ISR and the intercellular space diameter measured as described by Tobey et al.

The renin–angiotensin system in the esophageal mucosa of healthy subjects and patients with reflux disease

Objective. Components of the renin–angiotensin system (RAS) were recently discovered in the esophagus, which could be of interest in relation to gastroesophageal reflux disease (GERD). The present study was undertaken to confirm and further investigate the expression of RAS in healthy and refluxed exposed human esophageal mucosae. Methods.Esophageal biopsies were obtained from healthy subjects (n = 34) and individuals with erosive reflux disease (ERD, n = 28). Evaluation of general morphology and histological signs of reflux as well as investigation of gene transcript, protein expression and localization of various RAS components using RT-PCR, ELISA, western blot and immunohistochemistry were performed. Physiological effects of the AT2R were investigated in Ussing chamber experiments. Results. The study confirmed histological signs of reflux in ERD and expression of ACE, AT1R, AT2R and CatD in all examined specimens. In addition, the main effector peptide AngII, the pro-hormone AGT, the Mas receptor and the angiotensin-forming enzymes renin, CMA, CatG and NEP were present. Individuals with reflux disease had higher transcription activity of ACE and AT1R, increased protein levels of AT2R and lower levels of MasR. AT2R stimulation increased the ion currents in healthy epithelium, whereas epithelium from individuals with reflux disease exhibited no significant response. Conclusions. The study demonstrated that a local RAS is present in the human esophageal epithelium. Some RAS components were significantly altered in individuals diagnosed with ERD suggesting involvement in the pathophysiology of GERD.

STAT3 is involved in esophageal carcinogenesis through regulation of Oct-1

The transcription factor signal transducer and activator of transcription 3 (STAT3) contributes to cell proliferation, apoptosis and motility in human cancer cells. We aim to elucidate the function of STAT3 in esophageal carcinogenesis process and molecular mechanisms. We showed that hyperactivated STAT3 in esophageal carcinogenesis tissues correlated with the overexpression of octamer transcription factor-1 (Oct-1). High STAT3 phosphorylation correlated with shorter survival compared with low STAT3 phosphorylation. STAT3 and Oct-1 expression levels affected the proliferation and colony formation of Eca-109 esophageal squamous cell carcinoma cells by altering Erk and Akt activation. Nevertheless, STAT3 regulated the migration and invasion of esophageal squamous cell carcinoma cells independent of Oct-1. In conjunction with Oct-1, STAT3 inhibited apoptosis in esophageal squamous cell carcinoma cells. Constitutively activated STAT3 in normal human esophageal epithelium cells (HET-1A) elevated Oct-1 expression,and promoted proliferation and decreased apoptosis. STAT3 activated HET-1A cells to form tumors in vivo, suggesting that overactivated STAT3 is sufficient for carcinogenesis. We further confirmed the colocalization of STAT3 and Oct-1 in the nucleus and found that STAT3 regulates the transcription and expression of Oct-1 by directly targeting its promoter. Activated STAT3 also upregulated many genes associated with Oct-1. Together, our results indicate that STAT3 plays a crucial role in esophageal carcinogenesis by regulating the cell proliferation and apoptosis in conjunction with Oct-1.

Surface maturation scoring for oesophageal squamous intraepithelial neoplasia: a novel diagnostic approach inspired by first endomicroscopic 3-dimensional reconstruction

ObjectiveLoss of surface maturation and cytonuclear atypia have been regarded as the pathological ‘gold standard’ for the diagnosis of oesophageal squamous cell intraepithelial neoplasia. However, there has been no satisfactory...

Bile salts disrupt human esophageal squamous epithelial barrier function by modulating tight junction proteins

Reflux of acid and bile acids contributes to epithelial tissue injury in gastro-esophageal reflux disease. However, the influence of refluxed material on human esophageal stratified epithelial barrier function and tight junction (TJ) proteins has not been fully elucidated. Here, we investigated the influence of acid and bile acids on barrier function and TJ protein distribution using a newly developed air-liquid interface (ALI) in vitro culture model of stratified squamous epithelium based on primary human esophageal epithelial cells (HEECs). Under ALI conditions, HEECs formed distinct epithelial layers on Transwell inserts after 7 days of culture. The epithelial layers formed TJ, and the presence of claudin-1, claudin-4, and occludin were detected by immunofluorescent staining. The NP-40-insoluble fraction of these TJ proteins was significantly higher by day 7 of ALI culture. Exposure of HEECs to pH 2, and taurocholic acid (TCA) and glycocholic acid (GCA) at pH 3, but not pH 4, for 1 h decreased transepithelial electrical resistance (TEER) and increased paracellular permeability. Exposure of cell layers to GCA (pH 3) and TCA (pH 3) for 1 h also markedly reduced the insoluble fractions of claudin-1 and -4. We found that deoxycholic acid (pH 7.4 or 6, 1 h) and pepsin (pH 3, 24 h) significantly decreased TEER and increased permeability. Based on these findings, ALI-cultured HEECs represent a new in vitro model of human esophageal stratified epithelium and are suitable for studying esophageal epithelial barrier functions. Using this model, we demonstrated that acid, bile acids, and pepsin disrupt squamous epithelial barrier function partly by modulating TJ proteins. These results provide new insights into understanding the role of TJ proteins in esophagitis.

Bile Acids Down-Regulate Caveolin-1 in Esophageal Epithelial Cells through Sterol Responsive Element-Binding Protein

Bile acids are synthesized from cholesterol and are major risk factors for Barrett adenocarcinoma (BAC) of the esophagus. Caveolin-1 (Cav1), a scaffold protein of membrane caveolae, is transcriptionally regulated by cholesterol via sterol-responsive element-binding protein-1 (SREBP1). Cav1 protects squamous epithelia by controlling cell growth and stabilizing cell junctions and matrix adhesion. Cav1 is frequently down-regulated in human cancers; however, the molecular mechanisms that lead to this event are unknown. We show that the basal layer of the nonneoplastic human esophageal squamous epithelium expressed Cav1 mainly at intercellular junctions. In contrast, Cav1 was lost in 95% of tissue specimens from BAC patients (n = 100). A strong cytoplasmic expression of Cav1 correlated with poor survival in a small subgroup (n = 5) of BAC patients, and stable expression of an oncogenic Cav1 variant (Cav1-P132L) in the human BAC cell line OE19 promoted proliferation. Cav1 was also detectable in immortalized human squamous epithelial, Barrett esophagus (CPC), and squamous cell carcinoma cells (OE21), but was low in BAC cell lines (OE19, OE33). Mechanistically, bile acids down-regulated Cav1 expression by inhibition of the proteolytic cleavage of 125-kDa pre-SREBP1 from the endoplasmic reticulum/Golgi apparatus and nuclear translocation of active 68-kDa SREBP1. This block in SREBP1's posttranslational processing impaired transcriptional activation of SREBP1 response elements in the proximal human Cav1 promoter. Cav1 was also down-regulated in esophagi from C57BL/6 mice on a diet enriched with 1% (wt/wt) chenodeoxycholic acid. Mice deficient for Cav1 or the nuclear bile acid receptor farnesoid X receptor showed hyperplasia and hyperkeratosis of the basal cell layer of esophageal epithelia, respectively. These data indicate that bile acid-mediated down-regulation of Cav1 marks early changes in the squamous epithelium, which may contribute to onset of Barrett esophagus metaplasia and progression to BAC.

A statistical framework for multiparameter analysis at the single-cell level

Phenotypic characterization of individual cells provides crucial insights into intercellular heterogeneity and enables access to information that is unavailable from ensemble averaged, bulk cell analyses. Single-cell studies have attracted significant interest in recent years and spurred the development of a variety of commercially available and research-grade technologies. To quantify cell-to-cell variability of cell populations, we have developed an experimental platform for real-time measurements of oxygen consumption (OC) kinetics at the single-cell level. Unique challenges inherent to these single-cell measurements arise, and no existing data analysis methodology is available to address them. Here we present a data processing and analysis method that addresses challenges encountered with this unique type of data in order to extract biologically relevant information. We applied the method to analyze OC profiles obtained with single cells of two different cell lines derived from metaplastic and dysplastic human Barrett's esophageal epithelium. In terms of method development, three main challenges were considered for this heterogeneous dynamic system: (i) high levels of noise, (ii) the lack of a priori knowledge of single-cell dynamics, and (iii) the role of intercellular variability within and across cell types. Several strategies and solutions to address each of these three challenges are presented. The features such as slopes, intercepts, breakpoint or change-point were extracted for every OC profile and compared across individual cells and cell types. The results demonstrated that the extracted features facilitated exposition of subtle differences between individual cells and their responses to cell-cell interactions. With minor modifications, this method can be used to process and analyze data from other acquisition and experimental modalities at the single-cell level, providing a valuable statistical framework for single-cell analysis.

Oxidative stress-induced posttranslational modification of TRPV1 expressed in esophageal epithelial cells

Human esophageal epithelium is continuously exposed to physical stimuli or to gastric acid that sometimes causes inflammation of the mucosa. Transient receptor potential vanilloid 1 (TRPV1) is a nociceptive, Ca(2+)-selective ion channel activated by capsaicin, heat, and protons. It has been reported that activation of TRPV1 expressed in esophageal mucosa is involved in gastroesophageal reflux disease (GERD) or in nonerosive GERD symptoms. In this study, we examined the expression and function of TRPV1 in the human esophageal epithelial cell line Het1A, focusing in particular on the role of oxidative stress. Interleukin-8 (IL-8) secreted by Het1A cells upon stimulation by capsaicin or acid with/without 4-hydroxy-2-nonenal (HNE) was measured by ELISA. Following capsaicin stimulation, the intracellular production of reactive oxygen species (ROS) was determined using a redox-sensitive fluorogenic probe, and ROS- and HNE-modified proteins were determined by Western blotting using biotinylated cysteine and anti-HNE antibody, respectively. HNE modification of TRPV1 proteins was further investigated by immunoprecipitation after treatment with synthetic HNE. Capsaicin and acid induced IL-8 production in Het1A cells, and this production was diminished by antagonists of TRPV1. Capsaicin also significantly increased the production of intracellular ROS and ROS- or HNE-modified proteins in Het1A cells. Moreover, IL-8 production in capsaicin-stimulated Het1A cells was enhanced by synthetic HNE treatment. Immunoprecipitation studies revealed that TRPV1 was modified by HNE in synthetic HNE-stimulated Het1A cells. We concluded that TRPV1 functions in chemokine production in esophageal epithelial cells, and this function may be regulated by ROS via posttranslational modification of TRPV1.

Esophageal Epithelial ASIC3 is Associated With Increase in Severity of Symptoms in Patients With Gastro-Esophageal Reflux Disease (GERD)

Introduction Immunoreactive nerves and epithelial markers have been studied in the context of acid induced oesophageal hypersensitivity. Increase in immunoreactive TRPV1 nerves are associated with presence of oesophagitis12 as well as acid exposure time.3 ASIC3 was identified in epithelium of rat oesophagus and ASIC inhibitor (Amitidine) prevented oesophageal hyperaemic response in rats to CO2 infusion (mimicking acid).4 We aim to identify ASIC 3 in human oesophageal epithelium and to determine if it is a suitable biomarker of symptom severity in patients with gastro-oesophageal reflux disease (GORD). Methods 36 oesophageal mucosal biopsy samples from patients (age 19–59 years and 17 females) with or without GORD undergoing gastroscopy were obtained and their clinical symptoms were assessed using visual analogue scale (VAS) adapted from validated questionnaire ReQUEST. The symptoms assessed were; general feeling, acid complaints, upper abdominal complaints, lower abdominal complaints, nausea, sleep disturbances. Samples were paraffin fixed and analysed using immunohistochemistry for PGP 9.5 and ASIC3. Results Epithelial ASIC3 was identified in the mucosal biopsies (14 controls, 18 non-erosive reflux disease, 4 erosive oesophagitis). ASIC3 could be classified based on distribution and morphology into 4 groups; basal layer (stratum basale), stratum spinosum perinuclear, stratum spinosum membrane bound and superficial luminal layer staining. Patients who scored higher on VAS for all 6 symptoms also demonstrated higher expression of all 4 ASIC3 features studied (median 65% (35–87) vs 57% (34–79), p=0.018). In particular, for acid complaints, sleep disturbance and lower GI complaints, this association was more significant (median 64% (46–81) vs 54% (38–69), p=0.002). Nerve fibres as stained by PGP 9.5 in human oesophageal epithelium were absent. Conclusion Presence of specific epithelial ASIC3 features in oesophageal epithelium was associated with increase in severity of common GORD symptoms. Further quantitative studies are required to study correlation of ASIC3 as an objective biomarker of symptom severity.

Variability in the Muscle Composition (Skeletal vs Smooth) of Rat Esophagus: Implication for the Mechanism of Vagally Mediated Lower Esophageal Sphincter Relaxation

G A A b st ra ct s in human esophageal epithelium as detected by PGP9.5 was absent.Epithelial ASIC3 was identified in all mucosal biopsies (14 controls, 18 non-erosive reflux disease, 4 erosive esophagitis). ASIC3 could be classified based on distribution and characteristic sub-cellular morphology into 4 groups; basal layer (stratum basale), stratum spinosum perinuclear, stratum spinosum membrane-bound, and superficial luminal layer staining. Patients with higher VAS symptom scores had greater expression of all 4 ASIC3 features studied (p= 0.018) with strongest associations between acid complaints and sleep disturbance (p=0.002). Conclusions: Presence of specific epithelial ASIC3 morphological features in esophageal epithelium was associated with increase in severity of common GERD symptoms.

Abstract 381: Light-absorbing nanoparticles induce cell death in esophageal cancer cells

Abstract Background Light-absorbing nanoparticles that are nontoxic to biological tissue possess the potential for targeted killing of cancer cells while causing minimal collateral damage to adjacent healthy tissue. Gold nanoparticles with near infrared (nIR)-absorbing light properties are ideal for this application. Aim: This study is to investigate the photo-thermal ablation effect of nIR-absorbing gold nanoparticles coated with a natural polymer (chitosan) on esophageal cancer cells as well as benign cells. Methods: Three esophageal cell lines used are human squamous esophageal epithelium cell line (Het-1A), Barrett's epithelium cell line (BAR-T), and the esophageal adenocarcinoma cell line (OE-19). Furthermore, the uptake of gold nanoparticles in the benign esophageal tissue and the response to laser exposure was also valuated in a rat esophagoduodenal anastomosis (EDA) model. Three cell lines were treated with gold nanoparticles (OD=25) then exposed to laser at a wavelength of 817 nm and power density of 3W/cm2 for 3 min. The esophageal mucosa of rats was sprayed with gold nanoparticles 24 hours befor laser application. After treatment, the rats were exposed to laser at a wavelength of 817 nm and power density of 4.15W/cm2 for 3 min. The loading of gold nanoparticles was detected by silver staining. Results: The most OE-19 cells loaded with gold nanoparticles and were completely ablated after nIR light exposure. However, most Het-1A cells and BAR-T cells didn't load on any gold nanoparticles, and they remained viable after nIR light exposure. Normal esophageal epithelium and Barrett's mateplastic epithelium show low affinity to gold nanoparticles, however, the focal region with more loaded gold nanoparticles was ablated after nIR light exposure. Conclusion: 1) Chitosan coated gold nanoparticles undergo targeted uptake by OE-19 cells, with a much higher affinity compared to benign cells. 2) Photo-thermo-ablation by nIR light exposure causes a directed cancer cell-killing effect based on preferential gold nanoparticle uptake. 3) Further study is ongoing to investigate methods of enhancing peak uptake of gold nanoparticles in the EDA rat model, to provide an optimal model for laser ablation of esophageal cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 381. doi:10.1158/1538-7445.AM2011-381

Oesophageal epithelial ASIC3 is associated with increase in severity of symptoms in patients with gastro-oesophageal reflux disease (GORD)

IntroductionImmunoreactive nerves and epithelial markers have been studied in the context of acid induced oesophageal hypersensitivity. Increase in immunoreactive TRPV1 nerves are associated with presence of oesophagitis1 2 as well...

A novel tumor suppressor gene ECRG4 interacts directly with TMPRSS11A (ECRG1) to inhibit cancer cell growth in esophageal carcinoma

BackgroundThe esophageal carcinoma related gene 4 (ECRG4) was initially identified and cloned from human normal esophageal epithelium in our laboratory (GenBank accession no.AF325503). ECRG4 has been described as a novel tumor suppressor gene associated with prognosis in esophageal squamous cell carcinoma (ESCC).MethodsIn this study, binding affinity assay in vitro and co-immunoprecipitation experiment in vivo were utilized to verify the physical interaction between ECRG4 and transmembrane protease, serine 11A (TMPRSS11A, also known as ECRG1, GenBank accession no. AF 071882). Then, p21 protein expression, cell cycle and cell proliferation regulations were examined after ECRG4 and ECRG1 co-transfection in ESCC cells.ResultsWe revealed for the first time that ECRG4 interacted directly with ECRG1 to inhibit cancer cell proliferation and induce cell cycle G1 phase block in ESCC. Binding affinity and co-immunoprecipitation assays demonstrated that ECRG4 interacted directly with ECRG1 in ESCC cells. Furthermore, the ECRG4 and ECRG1 co-expression remarkably upregulatd p21 protein level by Western blot (P < 0.001), induced cell cycle G1 phase block by flow cytometric analysis (P < 0.001) and suppressed cell proliferation by MTT and BrdU assay (both P < 0.01) in ESCC cells.ConclusionsECRG4 interacts directly with ECRG1 to upregulate p21 protein expression, induce cell cycle G1 phase block and inhibit cancer cells proliferation in ESCC.

Aberrant expression of epidermal growth factor receptor and its interaction with protein kinase C δ in inflammation associated neoplastic transformation of human esophageal epithelium in high risk populations

Esophageal cancer is the second most common cancer among Indian males and is mostly associated with tobacco smoking and alcohol consumption. Epidermal growth factor receptor (EGFR) is a member of Type I tyrosine kinases. Its activation causes the docking of various proteins in its cytosolic tail. In the present study we have analyzed the expression pattern of EGFR, protein kinase C δ (PKCδ), tumor necrosis factor-α (TNF-α), nuclear factor κB (NFκB) and the interactions between EGFR and PKCδ in various pathological conditions. Human esophageal biopsies were obtained from 93 patients with a past history of smoking and alcohol consumption: 20 showed normal mucosa, 40 with dysplasia and 33 squamous cell carcinoma (SCC). These pathological conditions were analyzed immunohistochemically for the presence of EGFR expression and then subsequently analyzed using immunoblot and immunoprecipitation. A statistically significant difference of EGFR overexpression was found between low- and high-grade dysplasia and carcinoma (χ² = 3.3, χ² = 3.42: P = 0.07, 0.33). A statistical significance was observed between dysplasia and SCC and in all histopathological types (χ² = 4, χ² = 4.9; P < 0.05, P = 0.18 and χ² = 26.3, 26.6; P < 0.001). EGFR tyrosine phosphorylation and its association with PKCδ was significantly higher in all histopathological types with χ² = 7.965; P < 0.05 and 4.0830; P = 0.2530. Altogether, our findings reveal that the activation of EGFR and its subsequent interaction with PKCδ under inflammatory conditions might positively be attributed to the transformation of normal esophageal epithelia to SCC, which could explain ongoing inflammation in normal mucosa in a population prone to smoking and alcoholism.