Artificial intelligence for the real-time classification of intrapapillary capillary loop patterns in the endoscopic diagnosis of early oesophageal squamous cell carcinoma: A proof-of-concept study.

Background: Intrapapillary capillary loops (IPCLs) represent an endoscopically visible feature of early squamous cell neoplasia (ESCN) which correlate with invasion depth - an important factor in the success of curative endoscopic therapy. IPCLs visualised on magnification endoscopy with Narrow Band Imaging (ME-NBI) can be used to train convolutional neural networks (CNNs) to detect the presence and classify staging of ESCN lesions. Methods: A total of 7046 sequential high-definition ME-NBI images from 17 patients (10 ESCN, 7 normal) were used to train a CNN. IPCL patterns were classified by three expert endoscopists according to the Japanese Endoscopic Society classification. Normal IPCLs were defined as type A, abnormal as B1-3. Matched histology was obtained for all imaged areas. Results: This CNN differentiates abnormal from normal IPCL patterns with 93.7% accuracy (86.2% to 98.3%) and sensitivity and specificity for classifying abnormal IPCL patterns of 89.3% (78.1% to 100%) and 98% (92% to 99.7%), respectively. Our CNN operates in real time with diagnostic prediction times between 26.17 ms and 37.48 ms. Conclusion: Our novel and proof-of-concept application of computer-aided endoscopic diagnosis shows that a CNN can accurately classify IPCL patterns as normal or abnormal. This system could be used as an in vivo, real-time clinical decision support tool for endoscopists assessing and directing local therapy of ESCN.

Impaired motility in Barrett's esophagus: A study using high-resolution manometry with physiologic challenge.

BACKGROUND: Esophageal dysmotility may predispose to Barrett's esophagus (BE). We hypothesized that high-resolution manometry (HRM) performed with additional physiologic challenge would better delineate dysmotility in BE. METHODS: Included patients had typical reflux symptoms and underwent endoscopy, HRM with single water swallows and adjunctive testing with solids and rapid drink challenge (RDC) before ambulatory pH-impedance monitoring. BE and endoscopy-negative reflux disease (ENRD) subjects were compared against functional heartburn patient-controls (FHC). Primary outcome was incidence of HRM contractile abnormalities with standard and adjunctive swallows. Secondary outcomes included clearance measures and symptom association on pH-impedance. KEY RESULTS: Seventy-eight patients (BE 25, ENRD 27, FHC 26) were included. Water swallow contractility was reduced in both BE (median DCI 87 mm Hg/cm/s) and ENRD (442 mm Hg/cm/s) compared to FHC (602 mm Hg/cm/s; P < .001 and .04, respectively). With the challenge of solid swallows and RDC, these parameters improved in ENRD (solids = 1732 mm Hg/cm/s), becoming similar to FHC (1242 mm Hg/cm/s; P = .93), whereas abnormalities persisted in BE (818 mm Hg/cm/s; P < .01 c.f. FHC). In BE and ENRD, reflux events (67 vs 57 events/24 hour) and symptom frequency were similar; yet symptom correlation was significantly better in ENRD compared to BE, which was comparable to FHC (symptom index 30% vs 4% vs 0%, respectively). Furthermore, bolus clearance and exposure times were more pronounced in BE (P < .01). CONCLUSIONS & INFERENCES: Reduced contractile effectiveness persisted in BE with the more representative esophageal challenge of swallowing solids and free drinking; while in ENRD and FHC peristalsis usually improved, demonstrating peristaltic reserve. Furthermore, symptom association and refluxate clearance were reduced in BE. These factors may underlie BE pathogenesis.

Validation of an updated evidence-based protocol for proactive gastrostomy tube insertion in patients with head and neck cancer.

BACKGROUND/OBJECTIVES: Evidence-based practice guidelines are available to assist in the decision making for nutrition interventions in patients with head and neck cancer. Re-assessment of guideline recommendations is important with changing demographics, new treatment regimens, advancing radiotherapy techniques, such as helical intensity-modulated radiotherapy, and the emergence of new literature. The aim of this study was to validate the updated high-risk category definition in our local hospital protocol for the swallowing and nutrition management of patients with head and neck cancer to determine the ongoing predictive ability for identifying proactive gastrostomy requirement in a new cohort. SUBJECTS/METHODS: Patients attending a major tertiary hospital for head and neck cancer treatment from 2010 to 2011 were included (n=270). Data were collected on patient demographics (age and gender), clinical factors (tumour site, staging and treatment), nutrition outcome measures (weight, enteral feeding) and protocol adherence. Sensitivity and specificity were calculated and compared with the original validation study. RESULTS: Proactive gastrostomy tubes were inserted in 86 patients. Overall protocol adherence was 93%. Sensitivity improved to 72% (increase of 18%) and specificity improved to 96% (increase of 3%) compared with the original validation study where patients received three-dimensional (3-D) conformal radiotherapy. CONCLUSIONS: The results of this study confirm that the updated high-risk category in the protocol for the swallowing and nutrition management of patients with head and neck cancer remains valid to predict proactive gastrostomy in a mixed population receiving helical intensity-modulated radiotherapy and 3-D conformal radiotherapy. The protocol has an improved sensitivity and specificity and hence remains just as relevant for advanced techniques of radiation treatment delivery.

New radiotherapy techniques do not reduce the need for nutrition intervention in patients with head and neck cancer.

BACKGROUND/OBJECTIVES: Since 2007, our institution has used validated guidelines for the insertion of proactive gastrostomy feeding tubes in patients with head and neck cancer. Helical intensity-modulated radiotherapy (H-IMRT) delivered by Tomotherapy, is an advanced radiotherapy technique introduced at our centre in 2010. This form of therapy reduces long-term treatment-related toxicity to normal tissues. The aim of this study is to compare weight change and need for tube feeding following H-IMRT (n=53) with patients that would have previously been treated with three-dimensional conformal radiotherapy (n=134). SUBJECTS/METHODS: Patients with head and neck cancer assessed as high nutritional risk with recommendation for proactive gastrostomy were identified from cohorts from 2007 to 2008 and 2010 to 2011. Retrospective data were collected on clinical factors, weight change from baseline to completion of treatment, incidence of severe weight loss (⩾ 10%) and tube feeding. Statistical analyses to compare outcomes between the two treatments included χ(2)-test, Fisher's exact and two-sample Wilcoxon tests (P<0.05). RESULTS: The H-IMRT cohort had higher proportions of patients with definitive chemoradiotherapy (P=0.032) and more advanced N stage (P<0.001). Nutrition outcomes were not significantly different between H-IMRT and conformal radiotherapy, respectively: need for proactive gastrostomy (n=49, 92% versus n=115, 86%, P=0.213), median percentage weight change (-7.2% versus -7.3%, P=0.573) and severe weight loss incidence (28% versus 27%, P=0.843). CONCLUSIONS: Both groups had median weight loss >5% and high incidences of tube feeding and severe weight loss. Nutrition intervention remains critical in this patient population, despite advances in radiotherapy techniques, and no changes to current management are recommended.

Glutathione-gated potassium efflux as a mechanism of active biofilm detachment.

Biofilm detachment often has detrimental effects such as pipe obstruction and infection, yet the detachment mechanisms underlying dispersal remain largely unknown. In this study, a stress response mechanism known as glutathione-gated potassium efflux (GGKE) was evaluated as an active detachment mechanism in the dispersal of Pseudomonas aeruginosa biofilms. N-ethylmaleimide (NEM) was used to activate potassium efflux proteins (Kef) associated with the GGKE pathway. This stress response mechanism was hypothesized to lead to altered cation concentration, which can potentially affect polymer bridging in biofilms, and ultimately cause biofilm detachment. Results showed the activation of GGKE by NEM exposure caused biofilm detachment without inducing a measurable change in viability, and detached biomass concentration and composition were dependent on NEM concentration. More detached biomass was observed with higher concentrations of NEM, with a trend of increasing polymer detachment. The detachment was likely resulting from a weakened biofilm structural integrity induced by bridge denaturing from GGKE activation. This study is important in understanding biofilm detachment from engineered systems such as membrane aerated bioreactors.

Peptide receptor radionuclide therapy with (90)Y-DOTATATE/(90)Y-DOTATOC in patients with progressive metastatic neuroendocrine tumours: assessment of response, survival and toxicity.

BACKGROUND: Peptide receptor radionuclide therapy (PRRT) is an established treatment for patients with metastatic neuroendocrine tumours (NETs), although which factors are associated with an improved overall survival (OS) remains unclear. The primary aim of this study is to determine to what extent a radiological response to (90)Y-DOTATOC/(90)Y-DOTATATE PRRT is associated with an improved OS. The association of biochemical and clinical response to OS were assessed as secondary outcome measures. METHODS: A retrospective analysis was conducted on 57 patients: radiological response was classified using RECIST criteria, biochemical response was classified using WHO criteria and clinical response was assessed subjectively. Responses were recorded as positive response (PR), stable disease (SD) or progressive disease (PD), and survival analysed. RESULTS: Radiological response was achieved in 71.5% (24.5% PR, 47% SD) and was associated with a greater OS (51 and 56 months, respectively), compared with PD (18 months). A biochemical or clinical response post PRRT were not associated with a statistically significant improvement in OS. However, when combined with radiological response a survival benefit was observed according to the number of outcomes (radiological, biochemical, clinical), in which a response was observed. Mild haematological toxicity was common, renal toxicity was rare. CONCLUSION: In patients with progressive metastatic NETs receiving (90)Y-DOTATOC/(90)Y-DOTATATE PRRT, a radiological response with either a PR or a SD post therapy confers a significant OS benefit.

A randomised controlled trial of ALA vs. Photofrin photodynamic therapy for high-grade dysplasia arising in Barrett's oesophagus.

Photofrin photodynamic therapy (PDT) is a licenced treatment for Barrett's oesophagus (BE) with high-grade dysplasia (HGD) but causes strictures and photosensitivity and complete reversal of dysplasia (CR-HGD) by 50 % at 5 years. 5-Aminolaevulinic acid (ALA) is an alternative treatment with non-randomised data suggesting 85 % CR-HGD and a low risk of side effects. We aimed to compare efficacy and side effect profile between the drugs. A single-centre randomised controlled trial was conducted. Presence of HGD was confirmed on three occasions by two specialist GI pathologists. Stratification was by length of BE and extent of dysplasia. Standard protocols for ALA and Photofrin-PDT were followed. Endoscopic follow-up with 2-cm four-quadrant biopsy was at 6 weeks, 4 months, and then annually. All adverse event data were collected. Sixty four patients were randomised, 34 ALA and 30 Photofrin-PDT. Median follow-up is 24 months. On intention-to-treat analysis, CR-HGD was 16/34 (47 %) with ALA-PDT and 12/30 (40 %) with Photofrin-PDT. The overall cancer incidence was 14 % (9/64). On sub-group log-rank analysis, for BE ≤ 6 cm, CR-HGD was significantly higher with ALA-PDT than Photofrin-PDT (χ(2) =5.39, p=0.02). Strictures and skin photosensitivity were significantly more common after treatment with Photofrin-PDT than ALA-PDT (33 vs. 9 % and 43 vs. 6 %, respectively, p<0.05). The rate of buried glands with either drug was significantly higher post-PDT (48 % of patients) than pre-PDT (20 %). ALA-PDT has a better risk profile than Photofrin-PDT. In patients with BE length ≤ 6 cm, preliminary results show ALA-PDT is associated with significantly higher CR-HGD. In longer segments of BE, neither PDT drug is sufficiently efficacious to warrant routine use.

Replication of hepatitis E virus in three-dimensional cell culture.

Hepatitis E is an acute, viral hepatitis epidemic in developing regions, but which is detected with increasing frequency in sporadic form in developed regions. Pigs and possibly some other mammals are considered reservoirs of zoonotic infection with hepatitis E virus (HEV). However, whilst the relative significance of potential transmission routes from pigs to people is still unclear, the consumption of raw or undercooked pig meat has been implicated as a source of HEV infection. The lack of information about HEV zoonotic transmission is due in part to the difficulties of in vitro propagation of HEV. The Rotating Wall Vessel (RVW) has been described as a useful tool for the culture of cell lines in a 3-dimensional (3D) configuration. The aim of this work was to develop a 3D cell culture system for HEV to facilitate studies into the viability of virions contaminating pig tissues. This study, demonstrated that HEV can replicate efficiently in the RWV in human hepatoblastoma PLC/PRF/5 cells for up to 5 months not only by real time RT-PCR but also by detection of complete virions via electron microscopy. Furthermore, the replication of HEV progeny was observed by detecting HEV RNA by RT-PCR. The progeny were able to infect fresh 3D cultures, showing that this method is able to produce infectious hepatitis E virions.

Radiofrequency ablation is effective for the treatment of high-grade dysplasia in Barrett's esophagus after failed photodynamic therapy.

Endoscopic radiofrequency ablation (RFA) is an effective treatment for high-grade dysplasia in Barrett's esophagus in ablation-naïve patients, but no studies have evaluated its use in patients in whom ablative therapy has previously failed. We describe 14 patients with residual high-grade dysplasia following aminolevulinic acid or Photofrin (porfimer sodium) photodynamic therapy (PDT). An overall complete reversal of dysplasia was achieved in 86 % with a combination of RFA and rescue endoscopic mucosal resection. The median total follow-up is 19 months. The rate of strictures was 7 % (1/14) and there was a low rate of buried glands (0.5 % follow-up biopsies). These data suggest RFA is both safe and effective for eradication of high-grade dysplasia in patients in whom PDT has failed.

A self referencing platinum nanoparticle decorated enzyme-based microbiosensor for real time measurement of physiological glucose transport.

Glucose is the central molecule in many biochemical pathways, and numerous approaches have been developed for fabricating micro biosensors designed to measure glucose concentration in/near cells and/or tissues. An inherent problem for microsensors used in physiological studies is a low signal-to-noise ratio, which is further complicated by concentration drift due to the metabolic activity of cells. A microsensor technique designed to filter extraneous electrical noise and provide direct quantification of active membrane transport is known as self-referencing. Self-referencing involves oscillation of a single microsensor via computer-controlled stepper motors within a stable gradient formed near cells/tissues (i.e., within the concentration boundary layer). The non-invasive technique provides direct measurement of trans-membrane (or trans-tissue) analyte flux. A glucose micro biosensor was fabricated using deposition of nanomaterials (platinum black, multiwalled carbon nanotubes, Nafion) and glucose oxidase on a platinum/iridium microelectrode. The highly sensitive/selective biosensor was used in the self-referencing modality for cell/tissue physiological transport studies. Detailed analysis of signal drift/noise filtering via phase sensitive detection (including a post-measurement analytical technique) are provided. Using this highly sensitive technique, physiological glucose uptake is demonstrated in a wide range of metabolic and pharmacological studies. Use of this technique is demonstrated for cancer cell physiology, bioenergetics, diabetes, and microbial biofilm physiology. This robust and versatile biosensor technique will provide much insight into biological transport in biomedical, environmental, and agricultural research applications.

Adsorption of iron cyanide complexes onto clay minerals, manganese oxide, and soil.

The adsorption characteristics of an iron cyanide complex, soluble Prussian blue KFe(III)[Fe(II)(CN)(6)], were evaluated for representative soil minerals and soil at pH 3.7, 6.4 and 9.7. Three specimen clay minerals (kaolinite, montmorillonite, and illite), two synthesized manganese oxides (birnessite and cryptomelane), and a Drummer soil from Indiana were used as the adsorbents. Surface protonation of variable charge sites increased with decreasing pH yielding positively charged sites on crystal edges and enhancing the attractive force between minerals and iron cyanide complexes. Anion adsorption on clays often is correlated to the metal content of the adsorbent, and a positive relationship was observed between iron or aluminum content and Prussian blue adsorption. Illite had high extractable iron and adsorbed more ferro-ferricyande anion, while kaolinite and montmorillonite had lower extractable iron and adsorbed less. However, less pH effect was observed on the adsorption of iron cyanide to manganese oxides. This may due to the manganese oxide mediated oxidation of ferrocyanide [Fe(II)(CN)(6)(4-)], to ferricyanide [Fe(III)(CN)(6)(3-)], which has a low affinity for manganese oxides.

Image cytometry accurately detects DNA ploidy abnormalities and predicts late relapse to high-grade dysplasia and adenocarcinoma in Barrett's oesophagus following photodynamic therapy.

BACKGROUND AND AIMS: DNA ploidy abnormalities (aneuploidy/tetraploidy) measured by flow cytometry (FC) are strong predictors of future cancer development in untreated Barrett's oesophagus, independent of histology grade. Image cytometric DNA analysis (ICDA) is an optical technique allowing visualisation of abnormal nuclei that may be undertaken on archival tissue. Our aim was to determine the accuracy of ICDA vs FC, and evaluate DNA ploidy as a prognostic biomarker after histologically successful treatment with photodynamic therapy (PDT). METHODS: Nuclei were extracted from 40 mum sections of paraffin-embedded biopsies and processed for ICDA at UCL and FC at UW using standardised protocols. Subsequently, DNA ploidy was evaluated by ICDA on a cohort of 30 patients clear of dysplasia 1 year after aminolaevulinic acid PDT for high-grade dysplasia (HGD). The results were correlated with long-term outcome. RESULTS: In the comparative study, 93% (41 out of 44) of cases were classified identically. Errors occurred in the near-diploid region by ICDA and the tetraploid region by FC. In the cohort study, there were 13 cases of late relapse (7 cancer, 6 HGD) and 17 patients who remained free of dysplasia after a mean follow-up of 44 months. Aneuploidy post-PDT was highly predictive for recurrent HGD or cancer with a hazard ratio of 8.2 (1.8-37.8) (log-rank P=0.001). CONCLUSIONS: ICDA is accurate for the detection of DNA ploidy abnormalities when compared with FC. After histologically successful PDT, patients with residual aneuploidy are significantly more likely to develop HGD or cancer than those who become diploid. DNA ploidy by ICDA is a valuable prognostic biomarker after ablative therapy.

The Malnutrition Screening Tool is a useful tool for identifying malnutrition risk in residential aged care.

BACKGROUND: The Malnutrition Screening Tool (MST) is a valid nutrition screening tool in the acute hospital setting but has not been assessed in residential aged care facilities. The aim of this secondary analysis was to determine whether the MST could be a useful nutrition screening tool when compared with a full nutrition assessment by Subjective Global Assessment (SGA) in the residential aged care setting. METHODS: Two hundred and eighty-five residents (29% male; mean age 84 +/- 9 years) from eight residential aged care facilities in Australia participated in the study. A secondary analysis of data collected during a nutrition intervention study was conducted. The MST consists of two questions related to recent weight loss and appetite. Although the MST was not specifically applied, weight loss and appetite information was available and an estimated MST score (0-5) was calculated. Nutritional status was assessed by a research assistant trained in using the SGA. RESULTS: Malnutrition prevalence was 42.8% (122 malnourished out of 285 residents). Compared to the SGA, the MST was an effective predictor of nutritional risk (sensitivity = 83.6%, specificity = 65.6%, positive predictive value = 0.65, negative predictive value = 0.84). CONCLUSIONS: The components of the MST have acceptable sensitivity and specificity, suggesting that it can play a valuable role in quickly identifying the risk of malnutrition in the residential aged care setting. Further prospective research using the MST tool against a broader array of objective and subjective nutritional parameters is required to confirm its validity as a screening tool in aged care settings.

Plant germination and growth after exposure to iron cyanide complexes.

Phytoremediation has been proposed for treatment of cyanide-contaminated soil. This study was conducted to identify plants with the highest potential for phytoremediation of iron cyanide contaminated soil. Multiple cultivars of two cyanogenic species, sorghum (Sorghum bicolor) and flax (Linum usitatissimum), and one non-cyanogenic species, switchgrass (Panicum virgatum L), were selected for evaluation. The cultivars were screened by quantifying germination and root elongation. Differences in germination emerged among the cultivars (P < 0.05), but these differences appeared to be unrelated to cyanide concentration. The presence of 1000 mg/kg Prussian blue tended to suppress root growth parameters of flax and switchgrass but did not affect sorghum similarly.

Influence of a soil enzyme on iron-cyanide complex speciation and mineral adsorption.

Cyanide is commonly found as ferrocyanide [Fe(II)(CN)(6)](-4) and in the more mobile form, ferricyanide [Fe(III)(CN)(6)](-3) in contaminated soils and sediments. Although soil minerals may influence ferrocyanide speciation, and thus mobility, the possible influence of soil enzymes has not been examined. In a series of experiments conducted under a range of soil-like conditions, laccase, a phenoloxidase enzyme derived from the fungi Trametes versicolor, was found to exert a large influence on iron-cyanide speciation and mobility. In the presence of laccase, up to 93% of ferrocyanide (36-362ppm) was oxidized to ferricyanide within 4h. No significant effect of pH (3.6 and 6.2) or initial ferrocyanide concentration on the extent or rate of oxidation was found and ferrocyanide oxidation did not occur in the absence of laccase. Relative to iron-cyanide-mineral systems without laccase, ferrocyanide adsorption to aluminum hydroxide and montmorillonite decreased in the presence of laccase and was similar to or somewhat greater than that of ferricyanide without laccase. Laccase-catalyzed conversion of ferrocyanide to ferricyanide was extensive though up to 33% of the enzyme was mineral-bound. These results demonstrate that soil enzymes can play a major role in ferrocyanide speciation and mobility. Biotic soil components must be considered as highly effective oxidation catalysts that may alter the mobility of metals and metal complexes in soil. Immobilized enzymes should also be considered for use in soil metal remediation efforts.

Bovine lymphotropic herpesvirus and non-responsive post-partum metritis in dairy herds in the UK.

Bovine lymphotropic herpesvirus (BLHV) was detected for the first time in the UK in December 2005 in a dairy herd suffering from chronic, non-responsive post-partum metritis (NPPM). A small-scale investigation was undertaken in order to determine whether this was an isolated case. Samples of vaginal exudates or vaginal swabs were collected from cows in 13 UK dairy herds with a history of post-partum metritis that had not responded to standard treatment regimes for this condition. Cows in 9/13 herds and 1/13 herds were positive for BLHV and bovine herpesvirus-4, respectively, by pan-herpesvirus polymerase chain reaction. No consistent pattern of infectious agents or nutritional/metabolic factors commonly associated with post-partum metritis was observed at the times of sampling. The detection of BLHV in association with NPPM indicates that further work is warranted to determine the impact this virus has on cattle health.

Duplicated Clock genes with unique polyglutamine domains provide evidence for nonhomologous recombination in Chinook salmon (Oncorhynchus tshawytscha).

Circadian rhythms underlie diverse life functions ranging from cellular activities to behavior. Multiple clock genes play a central role in the generation of these rhythms. We partially characterized two copies of the Clock gene from Chinook salmon (Oncorhynchus tshawytscha), OtsClock1a and OtsClock1b. The 6,460 bp OtsClock1a sequence contains 16 exons, 15 introns and encompasses three highly conserved domains indicating it is a novel member of the bHLH-PAS superfamily of transcription factors. The second copy, OtsClock1b, consists of five exons and five introns spanning 1,945 bp. A polyglutamine repeat motif (PolyQ), characteristic of a majority of CLOCK proteins, is present in both OTSCLOCK1a and OTSCLOCK1b. However, the Chinook PolyQ domains are uniquely positioned inside the gene. Interestingly, a 1,200 bp non-coding segment located downstream of the OtsClock1a PolyQ domain is absent from OtsClock1b. This insertion/deletion is 91% similar to the Salmo salar Transferrin gene. A phylogenetic analysis of 11 CLOCK proteins shows that OtsClock1a and OtsClock1b are paralogs which likely arose subsequent to the salmonid genome-wide duplication event. Ultimately, the Chinook salmon Clock genes are key components to our understanding the genetic mechanisms underlying temporally regulated life history traits in Pacific salmonids.