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Supplemental Figure 2. IR triggers a temporary, reversible perturbation in cellular reducing potential.

Supplemental figure legends - detailed text

Ionizing radiation is the primary nonsurgical treatment modality for solid tumors. Its effectiveness is impacted by temporal constraints such as fractionation, hypoxia, and development of radioresistant clones. Biomarkers of acute radiation response are essential to developing more effective clinical algorithms. We hypothesized that acute perturbations in tumor lactate levels act as a surrogate marker of radiation response. In vitro experiments were carried out using validated human-derived cell lines from three histologies: anaplastic thyroid carcinoma (ATC), head and neck squamous cell carcinoma (HNSCC), and papillary thyroid carcinoma (PTC). Cellular metabolic activity was measured using standard biochemical assays. In vivo validation was performed using both an orthotopic and a flank derivative of a previously established ATC xenograft murine model. Irradiation of cells and tumors triggered a rapid, dose-dependent, transient decrease in lactate levels that was reversed by free radical scavengers. Acute lactate perturbations following irradiation could identify hypoxic conditions and correlated with hypoxia-induced radioresistance. Mutant TP53 cells and cells in which p53 activity was abrogated (shRNA) demonstrated a blunted lactate response to irradiation, consistent with a radioresistant phenotype. Lactate measurements therefore rapidly detected both induced (i.e., hypoxia) and intrinsic (i.e., mutTP53-driven) radioresistance. We conclude that lactate is a quantitative biomarker of acute genotoxic stress, with a temporal resolution that can inform clinical decision making. Combined with the spatial resolution of newly developed metabolic imaging platforms, this biomarker could lead to the development of truly individualized treatment strategies. Mol Cancer Ther; 14(12); 2901–8. ©2015 AACR.

Supplemental Figure 6. Loss of p53 function results in decreased radiosensitivity in HNSCC cell lines.

Supplemental Figure 1. Metabolic inhibition and exogenous ROS perturb cellular reducing potential.

Supplemental Figure 5. Acute IR effects on reducing potential and lactate can be used to detect intrinsic radioresistance driven by loss of p53 activity.

Supplemental Figure 4. Lactate accumulates in extra-cellular media as a function of cellular metabolism over time.

Supplemental Figure 3. IR effects on cellular lactate and clonogenic survival are driven by ROS.

Positive margin status after breast-conserving surgery (BCS) is a predictor of higher rates of local recurrence. Intraoperative margin assessment aims to achieve negative surgical margin status at the first operation, thus reducing the re-excision rates that are usually associated with potential surgical complications, increased medical costs, and mental pressure on patients. Microscopy with ultraviolet surface excitation (MUSE) can rapidly image tissue surfaces with subcellular resolution and sharp contrasts by utilizing the nature of the thin optical sectioning thickness of deep ultraviolet light. We have previously imaged 66 fresh human breast specimens that were topically stained with propidium iodide and eosin Y using a customized MUSE system. To achieve objective and automated assessment of MUSE images, a machine learning model is developed for binary (tumor vs. normal) classification of obtained MUSE images. Features extracted by texture analysis and pre-trained convolutional neural networks (CNN) have been investigated for sample descriptions. A sensitivity, specificity, and accuracy better than 90% have been achieved for detecting tumorous specimens. The result suggests the potential of MUSE with machine learning being utilized for intraoperative margin assessment during BCS.

Microscopy with ultraviolet surface excitation (MUSE) is increasingly studied for intraoperative assessment of tumor margins during breast-conserving surgery to reduce the re-excision rate. Here we report a two-step classification approach using texture analysis of MUSE images to automate the margin detection. A study dataset consisting of MUSE images from 66 human breast tissues was constructed for model training and validation. Features extracted using six texture analysis methods were investigated for tissue characterization, and a support vector machine was trained for binary classification of image patches within a full image based on selected feature subsets. A weighted majority voting strategy classified a sample as tumor or normal. Using the eight most predictive features ranked by the maximum relevance minimum redundancy and Laplacian scores methods has achieved a sample classification accuracy of 92.4% and 93.0%, respectively. Local binary pattern alone has achieved an accuracy of 90.3%.

Deep ultraviolet (DUV) excitation has generated high-resolution fluorescence images of fresh and unfixed breast tissues. We report that texture analysis of DUV images can potentially differentiate malignant samples from normal breast tissues.

Significance: Re-excision rates for women with invasive breast cancer undergoing breast conserving surgery (or lumpectomy) have decreased in the past decade but remain substantial. This is mainly due to the inability to assess the entire surface of an excised lumpectomy specimen efficiently and accurately during surgery. Aim: The goal of this study was to develop a deep-ultraviolet scanning fluorescence microscope (DUV-FSM) that can be used to accurately and rapidly detect cancer cells on the surface of excised breast tissue. Approach: A DUV-FSM was used to image the surfaces of 47 (31 malignant and 16 normal/benign) fresh breast tissue samples stained in propidium iodide and eosin Y solutions. A set of fluorescence images were obtained from each sample using low magnification (4×) and fully automated scanning. The images were stitched to form a color image. Three nonmedical evaluators were trained to interpret and assess the fluorescence images. Nuclear–cytoplasm ratio (N/C) was calculated and used for tissue classification. Results: DUV-FSM images a breast sample with subcellular resolution at a speed of 1.0 min/cm2. Fluorescence images show excellent visual contrast in color, tissue texture, cell density, and shape between invasive carcinomas and their normal counterparts. Visual interpretation of fluorescence images by nonmedical evaluators was able to distinguish invasive carcinoma from normal samples with high sensitivity (97.62%) and specificity (92.86%). Using N/C alone was able to differentiate patch-level invasive carcinoma from normal breast tissues with reasonable sensitivity (81.5%) and specificity (78.5%). Conclusions: DUV-FSM achieved a good balance between imaging speed and spatial resolution with excellent contrast, which allows either visual or quantitative detection of invasive cancer cells on the surfaces of a breast surgical specimen.

Breast cancer is the most commonly diagnosed cancer among women. Positive margin status after breast-conserving surgery (BCS) is a predictor of higher rates of local recurrence. Intraoperative margin detection helps to complete tumor excision at the first operation. A margin tool that is capable of imaging all six margins of large lumpectomy specimens with both high resolution and fast speed (within 20 min) is yet to be developed. Deep UV light allows simultaneous excitation of multiple fluorophores and generating surface fluorescence images. We have developed a deep UV fluorescence scanning microscope (DUV-FSM) for slide-free, high-resolution and rapid examination of tumor specimens during BCS. The DUV-FSM uses a deep UV LED for oblique back illumination of freshly excised breast tissues stained with propidium iodide and Eosin Y and motorized XY stages for mosaic scanning. Fluorescence images are captured by a color CCD camera. Both invasive lobular carcinoma (ILC) and invasive ductal carcinoma (IDC) images showed excellent contrast from that of the normal cells in color, tissue texture, and cell density and shapes. This contrast have been consistently observed in all samples (n = 20) we have imaged so far. Statistical analysis showed a significant difference (p

Background: Re-excision rates for women with invasive breast cancer undergoing breast conserving surgery (BCS or lumpectomy) have decreased in the past decade but remain substantial. This is mainly due to the inability to efficiently and accurately assess the entire surface of an excised lumpectomy specimen during surgery. To address this problem, a deep-ultraviolet scanning fluorescence microscope (DUV-FSM) imaging system was developed and evaluated to determine whether it could accurately detect cancer cells on the surface of excised breast tissue.Methods: 37 (22 malignant and 15 normal/benign) fresh breast tissue samples of variable size were stained in propidium iodide and eosin Y solutions. A set of fluorescence images were obtained from one side of each sample using low magnification (4x) and fully automated scanning. The images from each sample were stitched to form a color image. Routine histopathology was performed on each sample. Three non-medical inspectors were trained to interpret and assess the fluorescence images. Patch-level nuclear-cytoplasm ratio (N/C) was calculated and ROC analysis with the Youden index was used for tissue classification. Results: DUV-FSM images a breast sample with subcellular resolution at a speed of 1.0 minute/cm2. DUV images show excellent visual contrast in color, tissue texture, cell density and shape between invasive lobular carcinoma (ILC), invasive ductal carcinoma (IDC) and their normal counterparts. Visual interpretation of DUV images was able to distinguish invasive carcinoma from normal/benign samples with high sensitivity (95%) and specificity (94.4%). Pairwise comparison of patch-level N/C identified significant differences (pConclusions: DUV-FSM is a simple device that can rapidly image large, unprocessed breast specimens with subcellular resolution and excellent contrast which allows either visual or quantitative detection of invasive cancer cells on the surfaces of a surgical specimen. This study supports further investigation into whether this technology can be used for intraoperative assessment of surgical margins during BCS procedures.

We report a nonconcurrent dual-modality fiber-optic microendoscope (named SmartME) that integrates quantitative diffuse reflectance spectroscopy (DRS) and high-resolution fluorescence imaging (FLI) into a smartphone platform. The FLI module has a spatial resolution of ~3.5 µm, which allows the determination of the nuclear-cytoplasmic ratio (N/C) of epithelial tissues. The DRS has a spectral resolution of ~2 nm and can measure the total hemoglobin concentration (THC) and scattering properties of epithelial tissues with mean errors of 4.7% and 6.9%, respectively, which are comparable to the errors achieved with a benchtop spectrometer. Our preliminary in vivo studies from a single healthy human subject demonstrate that the SmartME can noninvasively quantify the tissue parameters of normal human oral mucosa tissues, including labial mucosa tissue, gingival tissue, and tongue dorsum tissue. The THCs of the three oral mucosa tissues are significantly different from each other (p ≤ 0.003). The reduced scattering coefficients of the gingival and labial tissues are significantly different from those of the tongue dorsum tissue (p in vivo.

Articular cartilage lacks self-healing capacity, and there is no effective therapy facilitating cartilage repair. Osteoarthritis (OA) due to cartilage defects represents large and increasing healthcare burdens worldwide. Nowadays, the generation of scaffolds to preserve bioactive factors and the biophysical environment has received increasing attention. Furthermore, improved decellularization technology has provided novel insights into OA treatment. This review provides a comparative account of different cartilage defect therapies. Furthermore, some recent effective decellularization protocols have been discussed. In particular, this review focuses on the decellularization ratio of each protocol. Moreover, these protocols were compared particularly on the basis of immunogenicity and mechanical functionality. Further, various recellularization methods have been enlisted and the reparative capacity of decellularized cartilage scaffolds is evaluated herein. The advantages and limitations of different recellularization processes have been described herein. This provides a basis for the generation of decellularized cartilage scaffolds, thereby potentially promoting the possibility of decellularization as a clinical therapeutic target.

Proteins can be post-translationally modified and this can be important in the regulation of cellular processes and function. However, little is known about whether protein phosphorylation plays a role in regulating wool fibre properties. In this study, we used a chemical labelling method combined with a high performance liquid chromatography-mass spectrometry (HPLC-MS) analysis to compare the phosphopeptides present in the wool of three Tan sheep with highly crimped wool and three Tan sheep with straighter wool. Thirty-six phosphopeptides that had differences in relative abundance between these two types of wool were identified. These peptides were derived from 28 to 33 different proteins, including two keratins (Ks) and 7 to 12 keratin-associated proteins (KAPs), with these proteins being common structural components of the wool fibre. The crimped wool had a higher relative abundance of phosphorylated K38, K72 and KAP13-x, whereas the straighter wool had a higher relative abundance of phosphorylated KAP2-1, KAP6-1, KAP4-x, KAP10-x and KAP13-y. These results confirm the phosphorylation of wool Ks and KAPs, and suggest that differential phosphorylation of Ks and KAPs may affect wool fibre crimping in Tan sheep. SIGNIFICANCE: Protein phosphorylation can alter the structural conformation and interaction of a protein, and hence affect the cellular processes that the protein undertakes. In this study, we compared the suite of phosphorylated proteins in crimped and straight wool from Chinese Tan sheep and found that some keratins and keratin-associated proteins were phosphorylated. Crimped wool had more keratin phosphorylation, while straight wool had more keratin-associated protein phosphorylation, with this suggesting that wool fibre crimping may be a regulated by phosphorylation of some wool proteins. This suggests that wool traits may be under epigenetic control and that post-translation modifications need to be considered in breeding for different wool types.

Ionizing radiation is the primary nonsurgical treatment modality for solid tumors. Its effectiveness is impacted by temporal constraints such as fractionation, hypoxia, and development of radioresistant clones. Biomarkers of acute radiation response are essential to developing more effective clinical algorithms. We hypothesized that acute perturbations in tumor lactate levels act as a surrogate marker of radiation response. In vitro experiments were carried out using validated human-derived cell lines from three histologies: anaplastic thyroid carcinoma (ATC), head and neck squamous cell carcinoma (HNSCC), and papillary thyroid carcinoma (PTC). Cellular metabolic activity was measured using standard biochemical assays. In vivo validation was performed using both an orthotopic and a flank derivative of a previously established ATC xenograft murine model. Irradiation of cells and tumors triggered a rapid, dose-dependent, transient decrease in lactate levels that was reversed by free radical scavengers. Acute lactate perturbations following irradiation could identify hypoxic conditions and correlated with hypoxia-induced radioresistance. Mutant TP53 cells and cells in which p53 activity was abrogated (shRNA) demonstrated a blunted lactate response to irradiation, consistent with a radioresistant phenotype. Lactate measurements therefore rapidly detected both induced (i.e., hypoxia) and intrinsic (i.e., mutTP53-driven) radioresistance. We conclude that lactate is a quantitative biomarker of acute genotoxic stress, with a temporal resolution that can inform clinical decision making. Combined with the spatial resolution of newly developed metabolic imaging platforms, this biomarker could lead to the development of truly individualized treatment strategies. Mol Cancer Ther; 14(12); 2901–8. ©2015 AACR.

Focusing properties of the vector vortex-bearing beams are investigated theoretically by vector diffraction theory. Simulation results show that the intensity distribution in focal region can be altered considerably by adjusting topological charge m and the numerical aperture (NA) of the focusing optical system. Focal pattern evolves from one focal spot to two overlapped intensity peaks. The two overlapped intensity peaks separate with increasing topological charge m, which leads to the focal splitting. It was found that evolution principle of focal pattern differs very remarkably with different numerical aperture of the focusing optical system. And focal split appears in focal evolution with different topological charge m in different numerical aperture of the focusing optical system. Results show that the topological charge of the vector vortex-bearing beams influences the focal intensity distribution considerably, and some novel focal patterns appear.

The logistics industry is developing rapidly; however, it is still in limited quantities and with many dynamic changes, and logistic distribution route optimization is becoming increasingly important. Since the design of a quick and effective optimization method should be given the priority in solving this problem. For this reason, the paper presents a new niche cellular genetic algorithm based on canonical cellular genetic algorithm by introducing niche technology, which can maintain the population diversity very well. This algorithm is then applied to the vehicle routing problem with time-window. And an order-reversing crossover operator is designed for solving this problem. The results show that, compared with the canonical cellular genetic algorithms, niche genetic algorithms and simple genetic algorithm, the new algorithm avoids pre-mature more effectively, and the results gained is of higher accuracy. It is of great efficiency in solving the vehicle routing problem with time-window.

Edge enhancement is a key part of image processing in order to sharpen the edge and optimize the visual quality of the input image. This paper proposes novel edge-directed enhancement spatial phase filters based on the generalized radial Hilbert transform theory. The simulated results showed that the proposed spatial phase filters are more efficient and flexible for edge-directed enhancement than conventional mathematical morphological edge enhancement algorithms.

Optical coherence tomography angiography (Angio-OCT), mainly based on the temporal dynamics of OCT scattering signals, has found a range of potential applications in clinical and scientific research. Based on the model of random phasor sums, temporal statistics of the complex-valued OCT signals are mathematically described. Statistical distributions of the amplitude differential and complex differential Angio-OCT signals are derived. The theories are validated through the flow phantom and live animal experiments. Using the model developed, the origin of the motion contrast in Angio-OCT is mathematically explained, and the implications in the improvement of motion contrast are further discussed, including threshold determination and its residual classification error, averaging method, and scanning protocol. The proposed mathematical model of Angio-OCT signals can aid in the optimal design of the system and associated algorithms., Comment: 11 pages, 11 figures