Partial Wave Spectroscopic Microscopy Research Articles

Early screening of colorectal cancer using feature engineering with artificial intelligence-enhanced analysis of nanoscale chromatin modifications

Colonoscopy is accurate but inefficient for colorectal cancer (CRC) prevention due to the low (~ 7 to 8%) prevalence of target lesions, advanced adenomas. We leveraged rectal mucosa to identify patients who harbor CRC field carcinogenesis by evaluating chromatin 3D architecture. Supranucleosomal disordered chromatin chains (~ 5 to 20 nm, ~1 kbp) fold into chromatin packing domains (~ 100 to 200 nm, ~ 100 to 1000 kbp). In turn, the fractal-like conformation of DNA within chromatin domains and the folding of the genome into packing domains has been shown to influence multiple facets of gene transcription, including the transcriptional plasticity of cancer cells. We deployed an optical spectroscopic nanosensing technique, chromatin-sensitive partial wave spectroscopic microscopy (csPWS), to evaluate the packing density scaling D of the chromatin chain conformation within packing domains from rectal mucosa in 256 patients with varying degrees of progression to colorectal cancer. We found average packing scaling D of chromatin domains was elevated in tumor cells, histologically normal-appearing cells 4 cm proximal to the tumor, and histologically normal-appearing rectal mucosa compared to cells from control patients (p < 0.001). Nuclear D had a robust correlation with the model of 5-year risk of CRC with r2 = 0.94. Furthermore, rectal D was evaluated as a screening biomarker for patients with advanced adenomas presenting an AUC of 0.85 and 85% sensitivity and specificity. artificial intelligence-enhanced csPWS improved diagnostic performance with AUC = 0.90. Considering the low sensitivity of existing CRC tests, including liquid biopsies, to early-stage cancers our work highlights the potential of chromatin biomarkers of field carcinogenesis in detecting early, significant precancerous colon lesions.

Early detection of lung cancer using artificial intelligence-enhanced optical nanosensing of chromatin alterations in field carcinogenesis

Supranucleosomal chromatin structure, including chromatin domain conformation, is involved in the regulation of gene expression and its dysregulation has been associated with carcinogenesis. Prior studies have shown that cells in the buccal mucosa carry a molecular signature of lung cancer among the cigarette-smoking population, the phenomenon known as field carcinogenesis or field of injury. Thus, we hypothesized that chromatin structural changes in buccal mucosa can be predictive of lung cancer. However, the small size of the chromatin chain (approximately 20 nm) folded into chromatin packing domains, themselves typically below 300 nm in diameter, preclude the detection of alterations in intradomain chromatin conformation using diffraction-limited optical microscopy. In this study, we developed an optical spectroscopic statistical nanosensing technique to detect chromatin packing domain changes in buccal mucosa as a lung cancer biomarker: chromatin-sensitive partial wave spectroscopic microscopy (csPWS). Artificial intelligence (AI) was applied to csPWS measurements of chromatin alterations to enhance diagnostic performance. Our AI-enhanced buccal csPWS nanocytology of 179 patients at two clinical sites distinguished Stage-I lung cancer versus cancer-free controls with an area under the ROC curve (AUC) of 0.92 ± 0.06 for Site 1 (in-state location) and 0.82 ± 0.11 for Site 2 (out-of-state location).

PWSpy: A Python library dedicated to the analysis of Partial Wave Spectroscopic Microscopy data.

Anthony et al., (2022). PWSpy: A Python library dedicated to the analysis of Partial Wave Spectroscopic Microscopy data.. Journal of Open Source Software, 7(72), 3957, https://doi.org/10.21105/joss.03957

Bridging Chromatin Nanoimaging and Molecular Modeling: Chromatin Packing as a Regulator of Transcriptional Heterogeneity in Carcinogenesis

The behavior of organisms is determined by both their genetic code and their capacity to explore a transcriptional landscape of thousands of genes to create new functional states. The talk will discuss the development of two synergistic platforms to study 3D chromatin structure and its regulation of transcription: The chromatin nanoimaging platform co-registers 3D electron tomography for chromatin imaging at the scale of individual DNA strands, spectroscopic photo-localization optical nanoscopy for imaging of targeted genes and transcriptional regulators, and partial wave spectroscopic microscopy for the quantification of the statistics of chromatin packing and contact probability at genomic length-scales from ∼kbp to ∼10Mbp in live cells, real-time, label-free in hundreds of cells simultaneously with single cell resolution. The computational platform enables predictive modeling of gene transcription in the context of realistic chromatin nanoscale structure from atomistic details of DNA to whole chromatin and is based on a multi-scale analysis, which includes molecular dynamics, Brownian dynamics, and molecular theory. By combining the imaging and computational platforms, we show how differential chromatin packing works across length-scales from individual DNA strands to topologically associated domains as a key regulator of intercellular transcriptional heterogeneity, transcriptional diversity, and gene-network heterogeneity, which in turn affect cells’ transcriptional access to their genomic space and have implications on cellular processes that depend on cells’ ability to adapt or respond to external stresses. Carcinogenesis provides a particularly significant testbed. We show that the regulation of the physical structure of chromatin packing allows for the predictable modulation of transcriptional heterogeneity in cancer cells during chemotherapeutic response to achieve near-complete cancer cell killing in vitro.

Preservation of cellular nano-architecture by the process of chemical fixation for nanopathology.

Transformation in chromatin organization is one of the most universal markers of carcinogenesis. Microscale chromatin alterations have been a staple of histopathological diagnosis of neoplasia, and nanoscale alterations have emerged as a promising marker for cancer prognostication and the detection of predysplastic changes. While numerous methods have been developed to detect these alterations, most methods for sample preparation remain largely validated via conventional microscopy and have not been examined with nanoscale sensitive imaging techniques. For these nanoscale sensitive techniques to become standard of care screening tools, new histological protocols must be developed that preserve nanoscale information. Partial Wave Spectroscopic (PWS) microscopy has recently emerged as a novel imaging technique sensitive to length scales ranging between 20 and 200 nanometers. As a label-free, high-throughput, and non-invasive imaging technique, PWS microscopy is an ideal tool to quantify structural information during sample preparation. Therefore, in this work we applied PWS microscopy to systematically evaluate the effects of cytological preparation on the nanoscales changes of chromatin using two live cell models: a drug-based model of Hela cells differentially treated with daunorubicin and a cell line comparison model of two cells lines with inherently distinct chromatin organizations. Notably, we show that existing cytological preparation can be modified in order to maintain clinically relevant nanoscopic differences, paving the way for the emerging field of nanopathology.

Abstract 429: Increased chromatin heterogeneity in circulating tumor cells (CTCs) is associated with high levels of HER2 expression in metastatic breast cancer (MBC)

Abstract Introduction: Molecular and genomic characterization of CTCs may help to understand MBC prognosis and predict treatment benefit. We previously reported that overexpression of HER2 in CTCs is associated with detection of clusters, more aggressive clinical features and prognosis in MBC (2018 AACR #5195). Herein, we report a new finding that a widespread variation on chromatin distribution in CTCs is associated with HER2 expression in MBC which may indicate more aggressive tumor. Methods: Whole blood sample (7.5ml/each) was collected from stage III/IV BCa patients before therapy. CTC enumeration were performed in FDA approved CELLTRACKS ANALYZERII® System (Menarini) by targeting the Epithelial Cell Adhesion Molecule antigen for capturing CTCs. After confirming CTCs were positive by CellSearch system, live CTCs enrichment was performed by using Parsortix system (ANGLE) utilizes microfluidic based 10μm separation cassettes. The captured CTCs were stained by Anti-CK-PE (specific for epithelial cells), DAPI (for nucleus), anti-CD45-APC (specific for leukocytes), and anti-HER2-FITC. CTCs chromatin packing density was scanned by Partial Wave Spectroscopic (PWS) microscopy which is a label-free spectroscopic microscopy method that resolves structures in cells and quantify the cell nucleus heterogeneity of chromatin packing density scaling between 20-350 nm, or from the kb-pair to 10 Mb-pair range. For each nuclei, the nanoscale heterogeneity of chromatin packing was analyzed as nuclear statistical parameter standard deviation Σ (sigma, RMS). Kruskal-Wallis test was used for statistics. Results: We identified 400 CTCs by CellSearch system, including 115 HER2+ CTCs and 285 HER2- CTCs. The live CTCs were sorted by Parsortix system and were proceeded with multiple staining and scanning by PWS microscopy. CTCs were classified based on morphology and correct phenotype as CK+, DAPI+ and CD45-. There were 97 acquired nuclei that were scanned and evaluated successfully, including 17 HER2+ CTCs, 12 HER2- CTCs and 80 non-CTC cells. According to the images, stronger nanoscopic variations at each pixel in internal structure represents higher chromatin distribution heterogeneity quantified by RMS. The average RMS was 0.05125 in CTC cells which is significantly higher than the non-CTC population (RMS=0.0381, p&amp;lt;0.01). Moreover, the HER2+ CTCs demonstrated the highest nuclear sigma (RMS=0.05366) among all the subgroups when compared to HER2- CTCs (RMS=0.04546, p&amp;lt;0.01). Conclusion: This is the first report of chromatin heterogeneity in CTCs, and the association between HER2 expression and high variations of chromatin distribution in MBC patients. This novel property of CTCs describing nanoscale heterogeneity of chromatin packing requires further evaluation and validation but, it may offer a novel dimension in the understanding of the metastatic process. Citation Format: Qiang Zhang, Lorenzo Gerratana, Ami N. Shah, Andrew Adam Davis, Lisa Flaum, Xiang Zhou, Youbin Zhang, Vadim Backman, Amir Behdad, Firas Wehbe, William Gradishar, Leonidas Platanias, Massimo Cristofanilli. Increased chromatin heterogeneity in circulating tumor cells (CTCs) is associated with high levels of HER2 expression in metastatic breast cancer (MBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 429.

Optical study of stress hormone-induced nanoscale structural alteration in brain using partial wave spectroscopic microscopy.

Chronic stress affects nano to microscale structures of the brain cells/tissues due to suppression of neural growths and reconnections, hence the neuronal activities. This results in depression, memory loss and even death of the brain cells. Our recently developed novel optical technique, partial wave spectroscopic microscopy has nanoscale sensitivity, and hence, can detect nanoscale changes in brain tissues due to stress. In this study, we applied this technique to quantify the stress related structural changes in the corticosterone-treated mouse model of stress. Our results show that brains from corticosterone-treated mice showed higher nanoscale structural disorder in the hippocampal region as compared to the brain from normal (vehicle) mice. The increase in structural alteration correlates with the duration of the stress. We further quantified the relative changes and the spatial localization of these changes in this mouse model and found out that the maximum changes occurred nearly symmetrically in both regions of the hippocampus. The mRNA for stress-related genes, brain-derived neurotrophic factor and tyrosine kinase-coupled receptor were also significantly reduced in the hippocampus of corticosterone-treated mice compared to that in control mice. These results indicate that chronic corticosterone treatment induces nanoscale structural alterations in mouse brain that corresponds to changes in stress-related gene expression.

Correction: Biophotonic detection of high order chromatin alterations in field carcinogenesis predicts risk of future hepatocellular carcinoma: A pilot study.

[This corrects the article DOI: 10.1371/journal.pone.0197427.].

Biophotonic detection of high order chromatin alterations in field carcinogenesis predicts risk of future hepatocellular carcinoma: A pilot study.

PurposeHepatocellular carcinoma (HCC) results from chronic inflammation/cirrhosis. Unfortunately, despite use of radiological/serological screening techniques, HCC ranks as a leading cause of cancer deaths. Our group has used alterations in high order chromatin as a marker for field carcinogenesis and hence risk for a variety of cancers (including colon, lung, prostate, ovarian, esophageal). In this study we wanted to address whether these chromatin alterations occur in HCC and if it could be used for risk stratification.Experimental designA case control study was performed in patients with cirrhosis who went on to develop HCC and patients with cirrhosis who did not develop cancer. We performed partial wave spectroscopic microscopy (PWS) which measures nanoscale alterations on formalin fixed deparaffinized liver biopsy specimens, 17 progressors and 26 non-progressors. Follow up was 2089 and 2892 days, respectively.ResultsPWS parameter disorder strength Ld were notably higher for the progressors (Ld = 1.47 ± 0.76) than the non-progressors (Ld = 1.00 ± 0.27) (p = 0.024). Overall, the Cohen’s d effect size was 0.907 (90.7%). AUROC analysis yielded an area of 0.70. There was no evidence of confounding by gender, age, BMI, smoking status and race.ConclusionsHigh order chromatin alterations, as detected by PWS, is altered in pre-malignant hepatocytes with cirrhosis and may predict future risk of HCC.

Correlating colorectal cancer risk with field carcinogenesis progression using partial wave spectroscopic microscopy.

Prior to the development of a localized cancerous tumor, diffuse molecular, and structural alterations occur throughout an organ due to genetic, environmental, and lifestyle factors. This process is known as field carcinogenesis. In this study, we used partial wave spectroscopic (PWS) microscopy to explore the progression of field carcinogenesis by measuring samples collected from 190 patients with a range of colonic history (no history, low‐risk history, and high‐risk history) and current colon health (healthy, nondiminutive adenomas (NDA; ≥5 mm and <10 mm), and advanced adenoma [AA; ≥10 mm, HGD, or >25% villous features]). The low‐risk history groups include patients with a history of NDA. The high‐risk history groups include patients with either a history of AA or colorectal cancer (CRC). PWS is a nanoscale‐sensitive imaging technique which measures the organization of intracellular structure. Previous studies have shown that PWS is sensitive to changes in the higher‐order (20–200 nm) chromatin topology that occur due to field carcinogenesis within histologically normal cells. The results of this study show that these nanoscale structural alterations are correlated with a patient's colonic history, which suggests that PWS can detect altered field carcinogenic signatures even in patients with negative colonoscopies. Furthermore, we developed a model to calculate the 5‐year risk of developing CRC for each patient group. We found that our data fit this model remarkably well (R 2 = 0.946). This correlation suggests that PWS could potentially be used to monitor CRC progression less invasively and in patients without adenomas, which opens PWS to many potential cancer care applications.

Measuring Nanoscale Chromatin Heterogeneity with Partial Wave Spectroscopic Microscopy.

Despite extensive research in the area, current understanding of the structural organization of higher-order chromatin topology (between 20 and 200nm) is limited due to a lack of proper imaging techniques at these length scales. The organization of chromatin at these scales defines the physical context (nanoenvironment) in which many important biological processes occur. Improving our understanding of the nanoenvironment is crucial because it has been shown to play a critical functional role in the regulation of chemical reactions. Recent progress in partial wave spectroscopic (PWS) microscopy enables real-time measurement of higher-order chromatin organization within label-free live cells. Specifically, PWS quantifies the nanoscale variations in mass density (heterogeneity) within the cell. These advancements have made it possible to study the functional role of chromatin topology, such as its regulation of the global transcriptional state of the cell and its role in the development of cancer. In this chapter, the importance of studying chromatin topology is explained, the theory and instrumentation of PWS are described, the measurements and analysis processes for PWS are laid out in detail, and common issues, troubleshooting steps, and validation techniques are provided.

The transformation of the nuclear nanoarchitecture in human field carcinogenesis.

Morphological alterations of the nuclear texture are a hallmark of carcinogenesis. At later stages of disease, these changes are well characterized and detectable by light microscopy. Evidence suggests that similar albeit nanoscopic alterations develop at the predysplastic stages of carcinogenesis. Using the novel optical technique partial wave spectroscopic microscopy, we identified profound changes in the nanoscale chromatin topology in microscopically normal tissue as a common event in the field carcinogenesis of many cancers. In particular, higher-order chromatin structure at supranucleosomal length scales (20–200 nm) becomes exceedingly heterogeneous, a measure we quantify using the disorder strength (Ld) of the spatial arrangement of chromatin density. Here, we review partial wave spectroscopic nanocytology clinical studies and the technology's promise as an early cancer screening technology.

Abstract B11: Label-free hyperspectral microscopy detects alterations in nanoscale cellular structure with high temporal resolution

Abstract We have previously shown that analysis of the spectral variance of a reflected light microscope image allows direct study of cellular nanoscale organization through a technique called Partial Wave Spectroscopic (PWS) microscopy. This information is produced by measuring the spectral variations in light scattering to quantify the refractive index fluctuations, which reflect the distribution of macromolecular structures, within a biological sample with length scale sensitivity on the order of 20-200nm. Previously, this approach has been used for the early detection of tumors and the structure-function study of chromatin changes in cells. Here we present a state-of-the-art system utilizing this spectroscopic principle to develop a label-free, live-cell light microscopy imaging method capable of sensing temporal changes in the nanoscale structural organization of live cells called Live-cell PWS microscopy. This system is built into a commercial epi-fluorescence live-cell microscope and data is acquired using a hyperspectral camera (IMEC), which simultaneously collects 16 spectral bands in the visible spectrum with each exposure at rates up to ~100 frames per second. In addition, the design of this instrument allows simultaneous phase contrast and hyperspectral nanoscopic measurements. Phase contrast data can be acquired simultaneously with the hyperspectral data by providing phase contrast transmission illumination within one of the cameras 16 collection bands and using a corresponding notch filter for that band on the epi-illumination source. Calculating the spectral variance of each hyperspectral data cube yields images with label-free contrast of the cells where the intensity corresponds to the heterogeneity of nanoscale macromolecular organization within each diffraction limited voxel. Using this configuration, we can simultaneously acquire both phase contrast and the nanoscale sensitive hyperspectral videos of rapidly evolving structural changes in live cells without the need of any exogenous labels. To demonstrate the unique capabilities of this system, we perform a range of experiments to induce changes in cellular ultrastructure at rapid time-scales. In this work, we will show the changes in the cellular nanoarchitecture upon (1) addition of a glucose to live HeLa cells, (2) during chemical fixation by paraformaldehyde, and (3) upon cell exposure to UV-radiation. In these experiments, we observed immediate (&amp;lt;100ms) changes in nanoscale structural organization throughout the cell, with distinct structural changes differing between the cytoplasm and nucleus. For instance, incubation of glucose results in immediate transformation of the nuclear organization, causing a rapid decrease in nanoscale heterogeneity, which then quickly returned to pretreatment levels. Comparatively in the cytoplasm, addition of glucose results in significantly increased macromolecular mass-density heterogeneity upon treatment, which remains elevated even after the nucleus had recovered to pre-treatment levels. Likewise, we also recorded the rapid changes induced by chemical fixation with 4% paraformaldehyde solution in HeLa cells. In this case, we observe a gradual homogenization of the nanoscale structure as we lose clear delineation of the nucleus from the cytoplasm after fixation. This observation can be attributed to the randomized cross-linking process of the fixative yielding a more homogenous nanoscale structural organization than the non-random structures present prior to fixation in the cells. Finally, we explore the real-time effects of UV-light exposure on cellular behavior and the chromatin nanoarchitecture. These experiments demonstrate that rapidly evolving changes in the native unlabeled nanostructure of cells can be studied with high temporal resolution making this a unique tool for studying the nanoscale cellular structure-function relationship in real-time. Citation Format: John E. Chandler, Luay M. Almassalha, Vadim Backman. Label-free hyperspectral microscopy detects alterations in nanoscale cellular structure with high temporal resolution. [abstract]. In: Proceedings of the AACR Special Conference on Engineering and Physical Sciences in Oncology; 2016 Jun 25-28; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2017;77(2 Suppl):Abstract nr B11.

Label-free imaging of the native, living cellular nanoarchitecture using partial-wave spectroscopic microscopy

The organization of chromatin is a regulator of molecular processes including transcription, replication, and DNA repair. The structures within chromatin that regulate these processes span from the nucleosomal (10-nm) to the chromosomal (>200-nm) levels, with little known about the dynamics of chromatin structure between these scales due to a lack of quantitative imaging technique in live cells. Previous work using partial-wave spectroscopic (PWS) microscopy, a quantitative imaging technique with sensitivity to macromolecular organization between 20 and 200 nm, has shown that transformation of chromatin at these length scales is a fundamental event during carcinogenesis. As the dynamics of chromatin likely play a critical regulatory role in cellular function, it is critical to develop live-cell imaging techniques that can probe the real-time temporal behavior of the chromatin nanoarchitecture. Therefore, we developed a live-cell PWS technique that allows high-throughput, label-free study of the causal relationship between nanoscale organization and molecular function in real time. In this work, we use live-cell PWS to study the change in chromatin structure due to DNA damage and expand on the link between metabolic function and the structure of higher-order chromatin. In particular, we studied the temporal changes to chromatin during UV light exposure, show that live-cell DNA-binding dyes induce damage to chromatin within seconds, and demonstrate a direct link between higher-order chromatin structure and mitochondrial membrane potential. Because biological function is tightly paired with structure, live-cell PWS is a powerful tool to study the nanoscale structure-function relationship in live cells.

Nanocytological field carcinogenesis detection to mitigate overdiagnosis of prostate cancer: a proof of concept study.

PurposeTo determine whether nano-architectural interrogation of prostate field carcinogenesis can be used to predict prognosis in patients with early stage (Gleason 6) prostate cancer (PCa), which is mostly indolent but frequently unnecessarily treated.Materials and MethodsWe previously developed partial wave spectroscopic microscopy (PWS) that enables quantification of the nanoscale intracellular architecture (20–200nm length scale) with remarkable accuracy. We adapted this technique to assess prostate needle core biopsies in a case control study from men with Gleason 6 disease who either progressed (n = 20) or remained indolent (n = 18) over a ~3 year follow up. We measured the parameter disorder strength (Ld) characterizing the spatial heterogeneity of the nanoscale cellular structure and nuclear morphology from the microscopically normal mucosa ~150 histologically normal epithelial cells.ResultsThere was a profound increase in nano-architectural disorder between progressors and non-progressors. Indeed, the Ld from future progressors was dramatically increased when compared to future non-progressors (1±0.065 versus 1.30±0.0614, respectively p = 0.002). The area under the receiver operator characteristic curve (AUC) was 0.79, yielding a sensitivity of 88% and specificity of 72% for discriminating between progressors and non-progressors. This was not confounded by demographic factors (age, smoking status, race, obesity), thus supporting the robustness of the approach.ConclusionsWe demonstrate, for the first time, that nano-architectural alterations occur in prostate cancer field carcinogenesis and can be exploited to predict prognosis of early stage PCa. This approach has promise in addressing the clinically vexing dilemma of management of Gleason 6 PCa and may provide a paradigm for dealing with the larger issue of cancer overdiagnosis.

Network signatures of nuclear and cytoplasmic density alterations in a model of pre and postmetastatic colorectal cancer

Cells contributing to the pathogenesis of cancer possess cytoplasmic and nuclear structural alterations that accompany their aberrant genetic, epigenetic, and molecular perturbations. Although it is known that architectural changes in primary and metastatic tumor cells can be quantified through variations in cellular density at the nanometer and micrometer spatial scales, the interdependent relationships among nuclear and cytoplasmic density as a function of tumorigenic potential has not been thoroughly investigated. We present a combined optical approach utilizing quantitative phase microscopy and partial wave spectroscopic microscopy to perform parallel structural characterizations of cellular architecture. Using the isogenic SW480 and SW620 cell lines as a model of pre and postmetastatic transition in colorectal cancer, we demonstrate that nuclear and cytoplasmic nanoscale disorder, micron-scale dry mass content, mean dry mass density, and shape metrics of the dry mass density histogram are uniquely correlated within and across different cellular compartments for a given cell type. The correlations of these physical parameters can be interpreted as networks whose nodal importance and level of connection independence differ according to disease stage. This work demonstrates how optically derived biophysical parameters are linked within and across different cellular compartments during the architectural orchestration of the metastatic phenotype.

Nano-Architectural Alterations in Mucus Layer Fecal Colonocytes in Field Carcinogenesis: Potential for Screening

Current fecal tests (occult blood, methylation, DNA mutations) target minute amounts of tumor products among a large amount of fecal material and thus have suboptimal performance. Our group has focused on exploiting field carcinogenesis as a modality to amplify the neoplastic signal. Specifically, we have shown that endoscopically normal rectal brushings have striking nano-architectural alterations which are detectable using a novel optical technique, partial wave spectroscopic microscopy (PWS). We therefore wished to translate this approach to a fecal assay. We examined mucus layer fecal colonocytes (MLFC) at preneoplastic and neoplastic time points (confirmed with rat colonoscopy) in the azoxymethane (AOM)-treated rat model and conducted PWS analysis to derive the nano-architectural parameter, disorder strength (Ld). We confirmed these results with studies in a genetic model (the Pirc rat). We showed that MLFC appeared microscopically normal, consistent with field carcinogenesis. Ld was elevated at an early time point (5 weeks post-AOM injection, effect size = 0.40, P = 0.024) and plateaued before adenoma formation (10 weeks post-AOM, effect size = 0.66, P = 0.001), with no dramatic increase once tumors developed. We replicated these data in the preneoplastic Pirc rat with an effect size in the MLFC that replicated the rectal brushings (increase vs. age-matched controls of 62% vs. 74%, respectively). We provide the first demonstration of a biophotonics approach to fecal assay. Furthermore, targeting the nano-architectural changes of field carcinogenesis rather than the detection of tumor products may provide a novel paradigm for colorectal cancer screening.

966 Partial Wave Spectroscopic Microscopy: A Novel Tool to Assess Perturbation in the Cellular Transcriptional Activity During Colon Carcinogenesis

with standard technique. Resection time was recorded and effort was graded using the validated NASA task load index and a 10-point visual-analog scale. Results:During submucosal dissection, the suture-pulley provided the endoscopists with direct visualization of submucosal layer. The mean time for submucosal dissection with and without the suture was 182 ±21.4 sec versus 899 ±146 sec (p,0.001). There was no significant difference in specimen diameter (34.2 vs 38.6mm, p=0.803). There was no perforation or other complication in either group. The suture-pulley method was also less demanding in all categories evaluated by the NASATLI (Table 1). Conclusion: The suture-pulley method facilitates direct visualization of the submucosal layer during ESD and significantly reduces procedure time and technical difficulty. Difficulty by NASA TLI and visual-analog scale

Spectroscopic Applications in Gastrointestinal Endoscopy

One of the major frontiers in biomedical optics has been as an adjunct to gastrointestinal endoscopy. In particular, spectroscopy of elastic light scattering has the potential of addressing many of the vexing challenges confronting endoscopists. This review discusses the principles of spectroscopy and critically evaluates performance in clinically significant scenarios. One of the best established applications is optical biopsy (in situ histological determination), and a number of techniques such as elastic scattering spectroscopy have demonstrated the ability to discriminate between neoplastic and non-neoplastic polyps. For flat dysplasia detection in Barrett's esophagus, some of the most promising spectroscopic technologies are angle-resolved low-coherence interferometry and endoscopic polarized scanning spectroscopy (the next generation light scattering spectroscopy). A new and exciting biological approach involves optical detection of field carcinogenesis. This can be exploited to reduce colonoscopic adenoma miss rate by assessing microcirculatory augmentation in the mucosa in the vicinity of the polyp using polarization-gatedspectroscopy. Furthermore, there are nano/micro-architectural correlates with diffuse field carcinogenesis throughout the colon. Indeed, technologies such as low coherence enhanced backscattering spectroscopy and partial wave spectroscopic microscopy have demonstrated that the detection of the nano-architectural alterations in the rectal mucosa can accurately sense advanced adenomas elsewhere in the colon. This may lend itself to a minimally intrusive risk stratification to identify patients who are most likely to harbor neoplasia and thus benefit from colonoscopy. Bridging these advances into the endoscopy suite requires pragmatic future development. Future studies need to focus on efficacy, cost, practicality (time required, etc), and particularly developing the paradigms that will impact upon clinical decision making.

APC Gene Testing for Familial Adenomatosis Polyposis

APC Gene Testing for Familial Adenomatosis Polyposis