Automated Image Analysis Techniques Research Articles

Correlates and determinants of nuclear epidermal growth factor receptor (EGFR) content in an oropharyngeal cancer tissue microarray

6022 Background: We have previously reported nuclear localization of EGFR protein in oropharyngeal cancer tissue. Nuclear EGFR levels were inversely correlated with survival and response to radiotherapy. Here, we sought to identify determinants and correlates of nuclear EGFR content. Methods: We analyzed an oropharyngeal cancer tissue microarray for expression of key molecules of EGFR signaling cascade utilizing an automated image analysis technique (AQUA) scored on a scale of 0–255 which permits protein quantitation and subcellular localization. Patients with OSCC treated with radiotherapy (RT) or surgery and RT were eligible. Data were analyzed using Pearson correlations and multiple linear regression with robust standard errors. Results: Of the 95 tumors included in this study, 72 (75%) had sufficient tissue for analysis of nuclear EGFR. Nuclear EGFR levels were associated with membranous/cytoplasmic EGFR levels (rho=0.85, p<0.01), nuclear ERK2 (rho=0.34, p=0.01), tumor cyclin D1 (rho=−0.25, p=0.06) and the proliferation markers Ki-67 (rho=0.24,p=0.06) and PCNA (rho=0.38, p<0.01). Nuclear phosphorylated-Akt, PTEN, p53 levels did not correlate with nuclear EGFR level. In multivariable analysis, only PCNA retained its significant association (p=0.01). Conclusions: Our results are consistent with the preclinical model of translocation of membranous EGFR to the nucleus acting as a transcriptional factor and fostering cell proliferation. Nuclear EGFR may constitute a mechanism of resistance to current EGFR-targeted therapies. No significant financial relationships to disclose.

Segmenting Articular Cartilage Automatically Using a Voxel Classification Approach

We present a fully automatic method for articular cartilage segmentation from magnetic resonance imaging (MRI) which we use as the foundation of a quantitative cartilage assessment. We evaluate our method by comparisons to manual segmentations by a radiologist and by examining the interscan reproducibility of the volume and area estimates. Training and evaluation of the method is performed on a data set consisting of 139 scans of knees with a status ranging from healthy to severely osteoarthritic. This is, to our knowledge, the only fully automatic cartilage segmentation method that has good agreement with manual segmentations, an interscan reproducibility as good as that of a human expert, and enables the separation between healthy and osteoarthritic populations. While high-field scanners offer high-quality imaging from which the articular cartilage have been evaluated extensively using manual and automated image analysis techniques, low-field scanners on the other hand produce lower quality images but to a fraction of the cost of their high-field counterpart. For low-field MRI, there is no well-established accuracy validation for quantitative cartilage estimates, but we show that differences between healthy and osteoarthritic populations are statistically significant using our cartilage volume and surface area estimates, which suggests that low-field MRI analysis can become a useful, affordable tool in clinical studies.

Automatic content-based analysis of georeferenced image data: Detection of Beggiatoa mats in seafloor video mosaics from the HÅkon Mosby Mud Volcano

The combination of new underwater technology as remotely operating vehicles (ROVs), high-resolution video imagery, and software to compute georeferenced mosaics of the seafloor provides new opportunities for marine geological or biological studies and applications in offshore industry. Even during single surveys by ROVs or towed systems large amounts of images are compiled. While these underwater techniques are now well-engineered, there is still a lack of methods for the automatic analysis of the acquired image data. During ROV dives more than 4200 georeferenced video mosaics were compiled for the HÅkon Mosby Mud Volcano (HMMV). Mud volcanoes as HMMV are considered as significant source locations for methane characterised by unique chemoautotrophic communities as Beggiatoa mats. For the detection and quantification of the spatial distribution of Beggiatoa mats an automated image analysis technique was developed, which applies watershed transformation and relaxation-based labelling of pre-segmented regions. Comparison of the data derived by visual inspection of 2840 video images with the automated image analysis revealed similarities with a precision better than 90%. We consider this as a step towards a time-efficient and accurate analysis of seafloor images for computation of geochemical budgets and identification of habitats at the seafloor.

Effects of insertion conditions on tissue strain and vascular damage during neuroprosthetic device insertion

Long-term integration of neuroprosthetic devices is challenged by reactive responses that compromise the brain–device interface. The contribution of physical insertion parameters to immediate damage is not well described. We have developed an ex vivo preparation to capture real-time images of tissue deformation during device insertion using thick tissue slices from rat brains prepared with fluorescently labeled vasculature. Qualitative and quantitative assessments of damage were made for insertions using devices with different tip shapes inserted at different speeds. Direct damage to the vasculature included severing, rupturing and dragging, and was often observed several hundred micrometers from the insertion site. Slower insertions generally resulted in more vascular damage. Cortical surface features greatly affected insertion success; insertions attempted through pial blood vessels resulted in severe tissue compression. Automated image analysis techniques were developed to quantify tissue deformation and calculate mean effective strain. Quantitative measures demonstrated that, within the range of experimental conditions studied, faster insertion of sharp devices resulted in lower mean effective strain. Variability within each insertion condition indicates that multiple biological factors may influence insertion success. Multiple biological factors may contribute to tissue distortion, thus a wide variability was observed among insertions made under the same conditions.

A parameter investigation of shear-induced coalescence in semidilute PIB–PDMS polymer blends: effects of shear rate, shear stress volume fraction, and viscosity

In this work, drop coalescence of polymer blends under shear flow in a parallel flow apparatus was investigated by optical sectioning microscopy. In each experiment, shear rate was set at values low enough to avoid any break-up phenomena. The time evolution of the drop size distribution was determined by motorized sample scanning and iterative acquisition of stacks of images along sample depth. Drop size and location in the acquired images was found by automated image analysis techniques. A systematic experimental campaign to investigate the effects of shear rate (in the range 0.1–0.5 s−1), volume fraction (2.5–10%), and viscosity of the two phases (3–63 Pa s) at different viscosity ratio (0.1–2.3) was carried out. By comparing data from different experiments, it was found that at any strain value, the average drop size decreases monotonically with the shear stress, calculated as the product of shear rate and matrix viscosity. Furthermore, the coalescence rate slowed down with increasing viscosity ratio. Overall, these results provide an extensive set of data, which can be used as a benchmark for modeling shear-induced coalescence in polymer blends.

An MRI study of structural variations in schizophrenia using deformation field morphometry

Magnetic resonance imaging (MRI) has an important role in investigating the changes in brain structure that are associated with schizophrenia. In this study, MRI scans of patients diagnosed with schizophrenia (37 males; 19 females; 17–42 years of age) were compared with those of an age- and sex-matched group of normal subjects (37 males; 19 females; 18–40 years of age). Based on the images of the healthy control subjects, we constructed a representative average brain template. Automated image analysis techniques were used to measure differences in the regional nonlinear deformation fields between the two groups. A deformation field, which measures the spatial transformation to deform a template of brain anatomy to each individual data, was obtained as a three-dimensional displacement vector in each voxel. There was a significantly greater magnitude of the deformation fields in the superior frontal and parietal lobes as well as in the cingulate gyrus connecting both lobes of the patients with schizophrenia than in those of healthy controls, suggesting that these cerebral regions have a significantly higher structural variability in schizophrenia.

Comparing Automatic and Manual Image Processing in FLARE Assay Analysis for Colon Carcinogenesis

Measurement of the amount of oxidative damage to DNA is one tool that can be used to estimate the beneficial effect of diet on the prevention of colon carcinogenesis. The FLARE assay is a modification of the single-cell gel electrophoresis (Comet) assay, and provides a measure of the 8OHdG adduct in the cells. In this paper, we present two innovations to the existing methods of analysis. The first one is related to the FLARE assay itself. We describe automated image analysis techniques that can be expected to measure oxidative damage faster, reproducibly, with less noise, and hence achieve greater statistical power. The proposed technique is compared to an existing technique, which was more manual and thus slower. The second innovation is our statistical analysis: we exploit the shape of FLARE intensity histograms, and show statistically significant diet effects in the duodenum. Previous analyses of this data concentrated on simple summary statistics, and found only marginally statistically significant diet effects. With the new imaging method and measure of oxidative damage, we show cells in the duodenum exposed to fish oil as having more oxidative damage than cells exposed to corn oil.

Automatic analysis of cerebral asymmetry: an exploratory study of the relationship between brain torque and planum temporale asymmetry

Leftward occipital and rightward frontal lobe asymmetry (brain torque) and leftward planum temporale asymmetry have been consistently reported in postmortem and in vivo neuroimaging studies of the human brain. Here automatic image analysis techniques are applied to quantify global and local asymmetries, and investigate the relationship between brain torque and planum temporale asymmetries on T1-weighted magnetic resonance (MR) images of 30 right-handed young healthy subjects (15 male, 15 female). Previously described automatic cerebral hemisphere extraction and 3D interhemispheric reflection-based methods for studying brain asymmetry are applied with a new technique, LowD (Low Dimension), which enables automatic quantification of brain torque. LowD integrates extracted left and right cerebral hemispheres in columns orthogonal to the midsagittal plane (2D column maps), and subsequently integrates slices along the brain's anterior–posterior axis (1D slice profiles). A torque index defined as the magnitude of occipital and frontal lobe asymmetry is computed allowing exploratory investigation of relationships between this global asymmetry and local asymmetries found in the planum temporale. LowD detected significant torque in the 30 subjects with occipital and frontal components found to be highly correlated ( P < 0.02). Significant leftward planum temporale asymmetry was detected ( P < 0.05), and the torque index correlated with planum temporale asymmetry ( P < 0.001). However, torque and total brain volume were not correlated. Therefore, although components of cerebral asymmetry may be related, their magnitude is not influenced by total hemisphere volume. LowD provides increased sensitivity for detection and quantification of brain torque on an individual subject basis, and future studies will apply these techniques to investigate the relationship between cerebral asymmetry and functional laterality.

Whole-brain voxel-based statistical analysis of gray matter and white matter in temporal lobe epilepsy

Volumetric MRI studies based on manual labeling of selected anatomical structures have provided in vivo evidence that brain abnormalities associated with temporal lobe epilepsy (TLE) extend beyond the hippocampus. Voxel-based morphometry (VBM) is a fully automated image analysis technique allowing identification of regional differences in gray matter (GM) and white matter (WM) between groups of subjects without a prior region of interest. The purpose of this study was to determine whole-brain GM and WM changes in TLE and to investigate the relationship between these abnormalities and clinical parameters. We studied 85 patients with pharmacologically intractable TLE and unilateral hippocampal atrophy and 47 age- and sex-matched healthy control subjects. The seizure focus was right sided in 40 patients and left sided in 45. Student's t test statistical maps of differences between patients' and controls' GM and WM concentrations were obtained using a general linear model. A further regression against duration of epilepsy, age of onset, presence of febrile convulsions, and secondary generalized seizures was performed with the TLE population. Voxel-based morphometry revealed that GM pathology in TLE extends beyond the hippocampus involving other limbic areas such as the cingulum and the thalamus, as well as extralimbic areas, particularly the frontal lobe. White matter reduction was found only ipsilateral to the seizure focus, including the temporopolar, entorhinal, and perirhinal areas. This pattern of structural changes is suggestive of disconnection involving preferentially frontolimbic pathways in patients with pharmacologically intractable TLE.

Reliability of textural analysis of ancient plasters and mortars through automated image analysis

For the study of ancient or recent plasters and mortars, it is necessary to determinate some morphological and textural parameters to describe the aggregate fraction and its relationship with binder fraction. Textural analyses are commonly performed with simple visual comparators or, at most, with a time-consuming manual point counting. The introduction of automatic or semiautomatic image analysis techniques seems to have made this kind of determinations faster and less biased. To compare data coming from different image-processing sequences or different analytical approaches, reliability and calibration need to be tested by each operator. A critical principle setup is presented for comparison using laboratory mortars with well-known textural features; classical analysis by sieving, point-counting technique and automated image analysis were performed. This empirical strategy allowed (1) to develop a simple and easy-to-use software-based image analysis system for plaster and mortar samples; (2) to pull out the relations between samples textural features and reliability of the different approaches; (3) to provide linear relations that allow to convert apparent measured features to real features. Results also underline the need to calibrate and standardize image analysis techniques before data handling.

Quantification of depositional changes and paleo-seismic activities from laminated sediments using outcrop data

Measurements of laminations from marine and limnic sediments are commonly a time-consuming procedure. However, the resulting quantitative proxies are of importance for the interpretation of both, climate changes and paleo-seismic activities. Digital image analysis accelerates the generation and interpretation of large data sets from laminated sediments based on contrasting grey values of dark and light laminae. Statistical transformation and correlation of the grey value signals reflect high frequency cycles due to changing mean laminae thicknesses, and thus provide data monitoring climate change. Perturbations (e.g., slumping structures, seismites, and tsunamites) of the commonly continuous laminae record seismic activities and obtain proxies for paleo-earthquake frequency. Using outcrop data from (i) the Pleistocene Lisan Formation of Jordan (Dead Sea Basin) and (ii) the Carboniferous–Permian Copacabana Formation of Bolivia (Lake Titicaca), we present a two-step approach to gain high-resolution time series based on field data for both purposes from unconsolidated and lithified outcrops. Step 1 concerns the construction of a continuous digital phototransect and step 2 covers the creation of a grey density curve based on digital photos along a line transect using image analysis. The applied automated image analysis technique provides a continuous digital record of the studied sections and, therefore, serves as useful tool for the evaluation of further proxy data. Analysing the obtained grey signal of the light and dark laminae of varves using phototransects, we discuss the potential and limitations of the proposed technique.

Assessment of Image-Based Data Collection and the AASHTO Provisional Standard for Cracking on Asphalt-Surfaced Pavements

The network-level pavement management system of the Kansas Department of Transportation (KDOT) is known as the Network Optimization System (NOS). For input, KDOT manually collects data on cracking severity and extent on its network annually. In this study, pavement surface images were collected and analyzed to evaluate the impact of AASHTO provisional standard PP 44-01 for asphalt-surfaced pavement-cracking data collection on NOS. The study covered approximately 262 km (164 mi) of in-service bituminous and composite pavements in Kansas. Nearly 50,000 images of the pavement surface were obtained for manual and automated evaluation of pavement condition. Two different processes of image analysis were compared with existing results from the KDOT annual survey. Transverse and fatigue crack severity and extent were manually interpreted following KDOT crack-rating algorithms for all sections. In addition, the sections were analyzed with an automated crack identification procedure following PP 44-01. The images corresponding to approximately a 5% sample rated by KDOT were identified. Comparative data were available for a manual distress survey on 5% samples (by KDOT) and image-based manual and automated interpretation on the 5% and 100% samples. Statistical analysis shows good agreement between the 5% sample and 100% sample results from the manual survey and image-based manual interpretation, respectively, for both crack types. This implies that a 5% pavement sample is adequate for describing crack severity and extent in NOS. Severity of fatigue cracks from the images was more difficult to rate, probably because of the descriptive nature of severity levels in the NOS algorithm. Agreement between the results from the manual image analysis on both 5% and 100% pavement samples and the KDOT manual survey is acceptable. However, the automated image analysis technique following the provisional AASHTO standard tends to overestimate severity of cracking.

Automated image analysis technique for measurement of femoral component subsidence in total hip joint replacement

A new technique for the measurement of subsidence of the femoral components is proposed. The method relies on the implantation of two ball markers around the femoral stem. A single radiographic image of the hip is analysed using image-processing techniques to minimize subjectivity related to manual identification of landmarks. Dimensions of the stem are used to correct for magnification and out-of-plane rotations resulting from radiographic positioning. This technique has been applied to a specific design of implant (Exeter™). A study of the effect of radiographic positioning has been conducted using a cadaveric bone phantom. Results for the variation in the measurement of axial migration compared to the neutral position (in millimetres) were: 0.942 (10° extension); 0.347 (20° flexion); −0.435 (40° internal rotation); 0.187 (30° external rotation) for distances measured between the bone marker and the implant. Results for distances measured between the implant and the cement centralizer were: 0.107 (10° extension); −0.277 (20° flexion); 0.085 (40° internal rotation); 0.280 (30° external rotation). The variations from within a more realistic range of positions demonstrate that axial migration measurements of ca. 0.5 mm between the bone and implant, and less than 0.3 mm between the implant and the bone cement, may be expected.

Voxel-Based Morphometric Comparison of Hippocampal and Extrahippocampal Abnormalities in Patients with Left and Right Hippocampal Atrophy

We used voxel-based morphometry (VBM), an automatic whole-brain MR image analysis technique, to investigate gray matter abnormalities in patients with temporal lobe epilepsy (TLE), in whom hippocampal atrophy (HA) was demonstrated by application of the Cavalieri method of modern design stereology. VBM results (P < 0.05, corrected) indicated preferential gray matter concentration (GMC) reduction in anterior hippocampus in patients with left HA and posterior hippocampus in patients with right HA. GMC reduction was also found in right dorsal prefrontal cortex in left and right HA patients. Prefrontal atrophy may be due to epileptiform excitotoxic discharges from the reciprocally connected pathological hippocampus, and may be the underlying biological cause for executive dysfunction in patients with TLE. GMC excess in ipsilateral parahippocampal, cerebellar, and pericallosal regions was common to both left and right HA groups relative to controls, and is hypothesized to reflect diminished gray–white matter demarcation, underlying white matter atrophy, or structural displacement due to cerebrospinal fluid expansion. However, bilateral temporal lobe GMC excess was observed in left HA patients, while ipsilateral temporal lobe GMC excess was observed in right HA patients. This work demonstrates methodological consistency between automated VBM and manual stereological analysis of the hippocampus in group comparisons, indicates widespread extrahippocampal gray matter abnormalities in unilateral HA, and suggests that there may be inherent differences in the effect of TLE on temporal lobe structures depending on the side of HA.

Effect of A-L tonic capsule on DNA content in rat experimental hepatocarcinogenesis.

The effects of A-L tonic capsule on DNA content in rat experimental hepatocarcinogenesis induced by diethylnitrosamine (DENA) were observed. The experimental rats were divided into 4 groups. With exception of group D in which the rats were only administered with DENA, the rats in the groups A, B, C were previously, simultaneously and subsequently fed with A-L tonic capsule respectively while they were administered with DENA. The DNA content of all rat livers was measured using automatic image analysis technique 20 weeks after administration of DENA. The results showed that the highest and lowest DNA contents were respectively seen in the groups D and A. There was significant difference in DNA contents between the groups A or B or C and D, and also between the groups A and B or C (both P < 0.01). 4 components (4C) and > or = 5C cells were predominant in the group D, while 2C cells were the minority. The number of 2C cells in the groups A, B, C was significantly higher than that in the group D, and the number of > or = 5C cells in the groups A, B, C was markedly lower than that in the group D (P < 0.01). Also, there was very significant difference in the number of 2C and > or = 5C cells between group A and B or C (P < 0.01). It was concluded that A-L tonic capsule could effectively inhibit the increase of DNA content of hepatocytes and improve the distribution of DNA content in rat hepatocarcinogenesis, especially in group A.

A FULLY AUTOMATED MICROSCOPE BACTERIAL ENUMERATION SYSTEM FOR STUDIES OF ORAL MICROBIAL ECOLOGY

A fast and fully automated image analysis technique for the enumeration of fluorescence-labeled bacteria in oral samples was developed. This paper describes the system configuration, application strategy, automated operation, and initial validation experiments using fluorescent microspheres, bacterial cultures, in vitro grown biofilms and human dental plaque. Following a series of brief operator-controlled calibration steps, the technique automatically performs all necessary microscope operations (stage translation, focus, sampling and analysis) on slides with up to 48 wells for as many different samples. It quantifies bacteria from differential interference contrast images, images showing cells that had been labeled by immunofluorescence with monoclonal antibodies, or images with cells labeled by a fluorescent DNA stain. With all evaluated samples, close agreement between the automated system and the assessor's visual counts was observed. This novel automated image grabbing and analysis procedure is applicable to the enumeration of specific taxa in clinical samples by both immunofluorescence and fluorescent in situ hybridization.

Some fields of applications of automatic image analysis in civil engineering

This paper illustrates the use of automatic image analysis technique to investigate the morphology of cement, concrete and fibre-reinforced concrete. First the methods to be used for powders and secondly for mortar and concrete are introduced. The dispersed phases are characterized by classical morphological parameters: these also enable to accede to the hydration process. The covariances give quantitative information on the homogeneity and dispersion of the different components: gravel, air-voids and cement paste. Air-voids are characterized by granulometric distributions and their mean free paths. Rose of directions gives information on feature orientation: fibres, microcracks for fibre-reinforced concrete, etc. Finally probabilistic models can be used to simulate the microstructure of such materials.

Study of phase dispersion in concrete by image analysis

The scope of this paper is to investigate by an automatic image analysis technique the dispersion of phases in concrete. Three morphological and statistical tools were used: co-occurrence matrices, and simple and crossed-covariance. It was shown (1) that there is a repulsion between gravel, (2) that gravel and air-voids are surrounded by matrix (cement paste and sand), and (3) that the dispersion of gravel and air-voids is perfectly uniform.

Automated immunofluorescence for enumeration of selected taxa in supragingival dental plaque.

The present study investigated a recently developed automated image analysis technique for its applicability to the enumeration of selected bacteria in supragingival dental plaque. Following initial calibration, the system is capable to count fluorescence-labeled target cells in up to 48 samples without user interference. Test samples contained a characteristic mixture of planktonic bacteria, small almost planar bacterial aggregates, and large, virtually indisruptable clumps with cells from multiple species. Due to their complex composition, these samples provided a challenging validation step for the image analysis system. Automated enumeration of target bacteria was compared with visual counting of the fluorescence-labeled bacteria. Results are shown for six taxa (Actinomyces naeslundii, Fusobacterium nucleatum, Prevotella intermedia/Prev. nigrescens, Streptococcus gordonii/Strep. oralis/Strep. sanguis, Strep. sobrinus, and Veillonella dispar/ V. parvula) with characteristic differences in abundance, cell morphology and aggregation behavior. Results revealed good correspondence between the two enumeration techniques (correlation coefficients ranging from 0.77 to 0.92) provided that the portion of target bacteria exceeded 0.05% of the total bacterial cell number. This work demonstrates the applicability and usefulness of fully automated immunofluorescence to analyze such complex ecosystems as supragingival dental plaque.

Effects of frequent marijuana use on brain tissue volume and composition.

To investigate CNS effects of frequent marijuana use, brain tissue volume and composition were measured using magnetic resonance imaging (MRI) in 18 current, frequent, young adult marijuana users and 13 comparable, non-using controls. Automated image analysis techniques were used to measure global and regional brain volumes, including, for most regions, separate measures of gray and white matter. The marijuana users showed no evidence of cerebral atrophy or global or regional changes in tissue volumes. Volumes of ventricular CSF were not higher in marijuana users than controls, but were, in fact, lower. There were no clinically significant abnormalities in any subject's MRI. Sex differences were detected in several global volume measures.