Effect of age on upper and lower eyelid saccades

To study the effect of age on the metrics of upper and lower eyelid saccades, eyelid movement of two groups of 30 subjects each were measured using computed image analysis. The patients were divided on the basis of age into a younger group (20-30 years) and an older group (60-91 years). Eyelid saccade functions were fitted by the damped harmonic oscillator model. Amplitude and peak velocity were used to compare the effect of age on the saccades of the upper and lower eyelid. There was no statistically significant difference in saccade amplitude between groups for the upper eyelid (mean ± SEM; upward, young = 9.18 ± 0.32 mm, older = 8.93 ± 0.31 mm, t = 0.56, P = 0.58; downward, young = 9.11 ± 0.27 mm, older = 8.86 ± 0.32 mm, t = 0.58, P = 0.56) However, there was a clear decline in the peak velocity of the upper eyelid saccades of older subjects (upward, young = 59.06 ± 2.34 mm/s, older = 50.12 ± 1.95 mm/s, t = 2.93, P = 0.005; downward, young = 71.78 ± 1.78 mm/s, older = 60.29 ± 2.62 mm/s, t = 3.63, P = 0.0006). In contrast, for the lower eyelid there was a clear increase of saccade amplitude in the elderly group (upward, young = 2.27 ± 0.09 mm, older = 2.98 ± 0.15 mm, t = 4.33, P < 0.0001; downward, young = 2.21 ± 0.10 mm, older = 2.96 ± 0.17 mm, t = 3.85, P < 0.001). These data suggest that the aging process affects the metrics of the lid saccades in a different manner according to the eyelid. In the upper eyelid the lower tension exerted by a weak aponeurosis is reflected only on the peak velocity of the saccades. In the lower eyelid, age is accompanied by an increase in saccade amplitude which indicates that the force transmission to the lid is not affected in the elderly.

Comparison of blood neoangiogenesis and lymphatic vascularization in colorectal adenomas from patients with and without concomitant colorectal cancer

Blood and lymphatic vessel proliferation is essential for tumor growth and progression. Most colorectal carcinomas develop from adenomas (adenoma-carcinoma sequence) in a process due to accumulation of molecular genetic alterations. About 5% of adenomatous polyps are expected to become malignant, but data on the differential angiogenic patterns of these lesions in patients with and without concomitant cancer are missing. The aim of the present study is to compare the angiogenic and lymphatic patterns of adenomatous polyps from patients with and without sporadic cancer. Thirty adenomatous polyps (15 from patients with another principal malignant lesion, and 15 from patients without cancer) were submitted to immunohistochemical staining for CD105 (marker for neoangiogenesis) and D2-40 (marker for lymphatic endothelium). Microvessel density and total vascular area were determined by computer image analysis to quantify the immunostained and total areas, and to assess the number of microvessels. Adenomas from patients with carcinoma showed significantly higher values of total vascular area determined by immunostaining for CD105 (cutoff value = 4386 µm²; P = 0.019) and of lymphatic microvessel density determined by immunostaining with D2-40 (cutoff value = 11.5; P = 0.041) when compared with those from patients without cancer. The present data indicate a significant increase in blood microvascular area and in lymphatic microvascular counts in adenomas removed from patients with cancer.

The role of Ca2+/calmodulin-dependent protein kinase II and calcineurin in TNF-α-induced myocardial hypertrophy

We investigated whether Ca2+/calmodulin-dependent kinase II (CaMKII) and calcineurin (CaN) are involved in myocardial hypertrophy induced by tumor necrosis factor α (TNF-α). The cardiomyocytes of neonatal Wistar rats (1-2 days old) were cultured and stimulated by TNF-α (100 μg/L), and Ca2+ signal transduction was blocked by several antagonists, including BAPTA (4 µM), KN-93 (0.2 µM) and cyclosporin A (CsA, 0.2 µM). Protein content, protein synthesis, cardiomyocyte volumes, [Ca2+]i transients, CaMKIIδB and CaN were evaluated by the Lowry method, [³H]-leucine incorporation, a computerized image analysis system, a Till imaging system, and Western blot analysis, respectively. TNF-α induced a significant increase in protein content in a dose-dependent manner from 10 µg/L (53.56 µg protein/well) to 100 μg/L (72.18 µg protein/well), and in a time-dependent manner from 12 h (37.42 µg protein/well) to 72 h (42.81 µg protein/well). TNF-α (100 μg/L) significantly increased the amplitude of spontaneous [Ca2+]i transients, the total protein content, cell size, and [³H]-leucine incorporation in cultured cardiomyocytes, which was abolished by 4 µM BAPTA, an intracellular Ca2+ chelator. The increases in protein content, cell size and [³H]-leucine incorporation were abolished by 0.2 µM KN-93 or 0.2 µM CsA. TNF-α increased the expression of CaMKIIδB by 35.21% and that of CaN by 22.22% compared to control. These effects were abolished by 4 µM BAPTA, which itself had no effect. These results suggest that TNF-α induces increases in [Ca2+]i, CaMKIIδB and CaN and promotes cardiac hypertrophy. Therefore, we hypothesize that the Ca2+/CaMKII- and CaN-dependent signaling pathways are involved in myocardial hypertrophy induced by TNF-α.

Expression of β-catenin and c-myc during human common bile duct development: a possible role in the morphogenesis of the common bile duct

β-catenin and c-myc play important roles in the development of tissues and organs. However, little is known about their expression patterns during the development of the human common bile duct. Immunohistochemistry was used to detect β-catenin and c-myc expression in common bile duct samples from postmortem tissues of 14 premature infants and 6 spontaneously aborted fetuses. The expression of β-catenin and c-myc was also analyzed by Western blot. The samples were divided into four groups based on the stage of human fetal development: 12, 13-27, 28-37, and >37 weeks. The Image-Pro Plus v. 6.0 image analysis software was used to calculate the mean qualifying score (MQS). At fetal stages 12, 13-27, 28-37, and >37 weeks, MQS of β-catenin were 612.52±262.13, 818.38±311.73, 706.33±157.19, and 350.69±110.19, respectively. There was a significant difference in MQS among the four groups (ANOVA, P=0.0155) and between the scores at >37 and 13-27 weeks (Student-Newman-Keuls, P<0.05). At fetal stages 12, 13-27, 28-37, and >37 weeks, the MQS of c-myc were 1376.64±330.04, 1224.18±171.66, 1270.24±320.75, and 741.04±219.19, respectively. There was a significant difference in MQS among the four groups (ANOVA, P=0.0087) and between the scores at >37 and 12 weeks, >37 and 13-27 weeks, and >37 and 28-37 weeks (all P<0.05, Student-Newman-Keuls). Western blots showed that β-catenin and c-myc expression were significantly higher in fetal than in postnatal control duct tissue (P<0.05). c-myc and β-catenin are involved in the normal development of the human common bile duct.

Adopting the Vaadin Java framework to rebuild VTT’s Web Interfaces and Database Applications

With the growth in new technologies, using online tools have become an everyday lifestyle. It has a greater impact on researchers as the data obtained from various experiments needs to be analyzed and knowledge of programming has become mandatory even for pure biologists. Hence, VTT came up with a new tool, R Executables (REX) which is a web application designed to provide a graphical interface for biological data functions like Image analysis, Gene expression data analysis, plotting, disease and control studies etc., which employs R functions to provide results. REX provides a user interactive application for the biologists to directly enter the values and run the required analysis with a single click. The program processes the given data in the background and prints results rapidly. Due to growth of data and load on server, the interface has gained problems concerning time consumption, poor GUI, data storage issues, security, minimal user interactive experience and crashes with large amount of data. This thesis handles the methods by which these problems were resolved and made REX a better application for the future. The old REX was developed using Python Django and now, a new programming language, Vaadin has been implemented. Vaadin is a Java framework for developing web applications and the programming language is extremely similar to Java with new rich components. Vaadin provides better security, better speed, good and interactive interface. In this thesis, subset functionalities of REX was selected which includes IST bulk plotting and image segmentation and implemented those using Vaadin. A code of 662 lines was programmed by me which included Vaadin as the front-end handler while R language was used for back-end data retrieval, computing and plotting. The application is optimized to allow further functionalities to be migrated with ease from old REX. Future development is focused on including Hight throughput screening functions along with gene expression database handling

Partikkelikokojakaumamenetelmän testaus INCA Feature

Tässä työssä testattiin partikkelikokojakaumien analysoinnissa käytettävää kuvankäsittelyohjelmaa INCA Feature. Partikkelikokojakaumat määritettiin elektronimikroskooppikuvista INCA Feature ohjelmaa käyttäen partikkeleiden projektiokuvista päällystyspigmenttinä käytettävälle talkille ja kahdelle eri karbonaattilaadulle. Lisäksi määritettiin partikkelikokojakaumat suodatuksessa ja puhdistuksessa apuaineina käytettäville piidioksidi- ja alumiinioksidihiukkasille. Kuvankäsittelyohjelmalla määritettyjä partikkelikokojakaumia verrattiin partikkelin laskeutumisnopeuteen eli sedimentaatioon perustuvalla SediGraph 5100 analysaattorilla ja laserdiffraktioon perustuvalla Coulter LS 230 menetelmällä analysoituihin partikkelikokojakaumiin. SediGraph 5100 ja kuva-analyysiohjelma antoivat talkkipartikkelien kokojakaumalle hyvin samankaltaisen keskiarvon. Sen sijaan Coulter LS 230 laitteen antama kokojakauman keskiarvo poikkesi edellisistä. Kaikki vertailussa olleet partikkelikokojakaumamenetelmät asettivat eri näytteiden partikkelit samaan kokojärjestykseen. Kuitenkaan menetelmien tuloksia ei voida numeerisesti verrata toisiinsa, sillä kaikissa käytetyissä analyysimenetelmissä partikkelikoon mittaus perustuu partikkelin eri ominaisuuteen. Työn perusteella kaikki testatut analyysimenetelmät soveltuvat paperipigmenttien partikkelikokojakaumien määrittämiseen. Tässä työssä selvitettiin myös kuva-analyysiin tarvittava partikkelien lukumäärä, jolla analyysitulos on luotettava. Työssä todettiin, että analysoitavien partikkelien lukumäärän tulee olla vähintään 300 partikkelia. Liian suuri näytemäärä lisää kokojakauman hajontaa ja pidentää analyysiin käytettyä aikaa useaan tuntiin. Näytteenkäsittely vaatii vielä lisää tutkimuksia, sillä se on tärkein ja kriittisin vaihe SEM ja kuva-analyysiohjelmalla tehtävää partikkelikokoanalyysiä. Automaattisten mikroskooppien yleistyminen helpottaa ja nopeuttaa analyysien tekoa, jolloin menetelmän suosio tulee kasvamaan myös paperipigmenttien tutkimuksessa. Laitteiden korkea hinta ja käyttäjältä vaadittava eritysosaaminen tulevat rajaamaan käytön ainakin toistaiseksi tutkimuslaitoksiin.

Using Tomato Analyzer software to determine embryo size in x-rayed seeds

A primary interest of image analysis of X-rayed seeds is to identify whether the extent of fill in the embryo cavity is associated with to seed physiological quality. The objective of this research was to verify the accuracy of the freely available Tomato Analyzer (TA) software developed at The Ohio State University to determine the ratio of embryo size over total seed area. Seeds of pumpkin, watermelon, cucumber and cotton were X-rayed and analyzed by the software which defines seed and embryo boundaries and automatically generates numerical values to quantify that ratio. Results showed that the TA has the sensitivity to evaluate the extent of embryo growth within the cucurbits and cotton seeds and is a promising alternative for this assessment in other seed species.

Using X rays to evaluate fissures in rice seeds dried artificially

The objective of the present study was to evaluate the efficiency of X-rays in identifying fissures in artificially dried rice seeds and the relationship between damage and seed performance in the germination test. Irrigated rice seeds of the IRGA 417 and IRGA 420 cultivars were harvested with 23.3 and 24.5% water content respectively and submitted to stationary drying treatments at 32, 38, 44 and 50 °C. X-rays were taken of subsamples of 100 seeds for each treatment, using an MX-20 X-ray equipment. The X-rayed seeds were classified from 1 to 3, where 1 corresponded to seeds without fissures, 2 to seeds with non-severe fissures and 3 to seeds with severe fissures. The same X-rayed seeds were planted and on the seventh day the seedlings (normal or abnormal) and dead seeds were photographed and evaluated to verify any relationship between the fissures and physiological potential. Higher drying temperature increased the percentage of fissures in the two cultivars, which can adversely affect their germination. Seeds with fissures can be identified using X-rays.

Viability of recently harvested and stored Xylopia aromatica (Lam.) Mart. (Annonaceae) seeds

Xylopia aromatica is a native species from Brazil's "Cerrado", recommended for restoration ecology and also as a medicine. Its seeds have embryos with morphophysiological dormancy, making nursery propagation difficult. The objective of this study was to verify the efficiency of X-ray and tetrazolium tests for evaluating the viability of three seed lots, stored for different periods. All seeds were X-rayed (13 kV, 350 seconds) and samples used for tetrazolium and germination tests. In the tetrazolium test, seeds were submitted to six treatments at two temperatures (25 and 30 °C) with imbibition in distilled water and immersion in three concentrations of tetrazolium solution (0.5, 0.75 and 1%) at the two imbibition temperatures. Seeds for the germination test were placed for imbibition in distilled water and a 500 ppm Promalin® (6-Benzyladenine + GA4 + GA7) solution and later sown in sterilized sand. The embryo could not be observed with the X-ray test. However, those seeds observed with an undamaged endosperm did not differ in the percentages of seeds with firm and stained endosperms observed in the tetrazolium test for all the lots. The tetrazolium test is efficient for evaluating seed viability, principally if imbibed at 30 °C and immersed in a 0.5% solution at 30 °C.

Relationship between eggplant seed morphology and germination

Structural differences such as abnormalities, damage and free spaces in seeds may affect germination. The aim of this study was to study the relationship between eggplant seed morphology and seed germination. Ten seed lots of the eggplant cultivar Embu were evaluated by X-ray image analysis and the germination test. Seed image analysis was performed by Image Pro Plus® software and the whole seed area and free space between the embryo and endosperm were measured. The internal seed area filled by the embryo and endosperm was calculated from the difference between the whole seed and free space areas. Based on these results and visual seed analysis, seeds were classified into three categories and information on germination was obtained for each one. X-ray image analysis provides a perfect view of the internal seed parts and for seed morphology studies. An increase in seed area filled by the endosperm and embryo does not improve seed germination. Mechanical seed damage and deteriorated tissues can adversely affect seed germination.

Bioimage informatics in STED super-resolution microscopy

Optical microscopy is living its renaissance. The diffraction limit, although still physically true, plays a minor role in the achievable resolution in far-field fluorescence microscopy. Super-resolution techniques enable fluorescence microscopy at nearly molecular resolution. Modern (super-resolution) microscopy methods rely strongly on software. Software tools are needed all the way from data acquisition, data storage, image reconstruction, restoration and alignment, to quantitative image analysis and image visualization. These tools play a key role in all aspects of microscopy today – and their importance in the coming years is certainly going to increase, when microscopy little-by-little transitions from single cells into more complex and even living model systems. In this thesis, a series of bioimage informatics software tools are introduced for STED super-resolution microscopy. Tomographic reconstruction software, coupled with a novel image acquisition method STED< is shown to enable axial (3D) super-resolution imaging in a standard 2D-STED microscope. Software tools are introduced for STED super-resolution correlative imaging with transmission electron microscopes or atomic force microscopes. A novel method for automatically ranking image quality within microscope image datasets is introduced, and it is utilized to for example select the best images in a STED microscope image dataset.

Real-time monitoring of laser scribing process utilizing high-speed camera

Currently, laser scribing is growing material processing method in the industry. Benefits of laser scribing technology are studied for example for improving an efficiency of solar cells. Due high-quality requirement of the fast scribing process, it is important to monitor the process in real time for detecting possible defects during the process. However, there is a lack of studies of laser scribing real time monitoring. Commonly used monitoring methods developed for other laser processes such a laser welding, are sufficient slow and existed applications cannot be implemented in fast laser scribing monitoring. The aim of this thesis is to find a method for laser scribing monitoring with a high-speed camera and evaluate reliability and performance of the developed monitoring system with experiments. The laser used in experiments is an IPG ytterbium pulsed fiber laser with 20 W maximum average power and Scan head optics used in the laser is Scanlab’s Hurryscan 14 II with an f100 tele-centric lens. The camera was connected to laser scanner using camera adapter to follow the laser process. A powerful fully programmable industrial computer was chosen for executing image processing and analysis. Algorithms for defect analysis, which are based on particle analysis, were developed using LabVIEW system design software. The performance of the algorithms was analyzed by analyzing a non-moving image from the scribing line with resolution 960x20 pixel. As a result, the maximum analysis speed was 560 frames per second. Reliability of the algorithm was evaluated by imaging scribing path with a variable number of defects 2000 mm/s when the laser was turned off and image analysis speed was 430 frames per second. The experiment was successful and as a result, the algorithms detected all defects from the scribing path. The final monitoring experiment was performed during a laser process. However, it was challenging to get active laser illumination work with the laser scanner due physical dimensions of the laser lens and the scanner. For reliable error detection, the illumination system is needed to be replaced.

Generating Aesthetically Pleasing Images in a Virtual Environment using Particle Swarm Optimization

This research focuses on generating aesthetically pleasing images in virtual environments using the particle swarm optimization (PSO) algorithm. The PSO is a stochastic population based search algorithm that is inspired by the flocking behavior of birds. In this research, we implement swarms of cameras flying through a virtual world in search of an image that is aesthetically pleasing. Virtual world exploration using particle swarm optimization is considered to be a new research area and is of interest to both the scientific and artistic communities. Aesthetic rules such as rule of thirds, subject matter, colour similarity and horizon line are all analyzed together as a multi-objective problem to analyze and solve with rendered images. A new multi-objective PSO algorithm, the sum of ranks PSO, is introduced. It is empirically compared to other single-objective and multi-objective swarm algorithms. An advantage of the sum of ranks PSO is that it is useful for solving high-dimensional problems within the context of this research. Throughout many experiments, we show that our approach is capable of automatically producing images satisfying a variety of supplied aesthetic criteria.

Segmentation d’images intravasculaires ultrasonores

L'imagerie intravasculaire ultrasonore (IVUS) est une technologie médicale par cathéter qui produit des images de coupe des vaisseaux sanguins. Elle permet de quantifier et d'étudier la morphologie de plaques d'athérosclérose en plus de visualiser la structure des vaisseaux sanguins (lumière, intima, plaque, média et adventice) en trois dimensions. Depuis quelques années, cette méthode d'imagerie est devenue un outil de choix en recherche aussi bien qu'en clinique pour l'étude de la maladie athérosclérotique. L'imagerie IVUS est par contre affectée par des artéfacts associés aux caractéristiques des capteurs ultrasonores, par la présence de cônes d'ombre causés par les calcifications ou des artères collatérales, par des plaques dont le rendu est hétérogène ou par le chatoiement ultrasonore (speckle) sanguin. L'analyse automatisée de séquences IVUS de grande taille représente donc un défi important. Une méthode de segmentation en trois dimensions (3D) basée sur l'algorithme du fast-marching à interfaces multiples est présentée. La segmentation utilise des attributs des régions et contours des images IVUS. En effet, une nouvelle fonction de vitesse de propagation des interfaces combinant les fonctions de densité de probabilité des tons de gris des composants de la paroi vasculaire et le gradient des intensités est proposée. La segmentation est grandement automatisée puisque la lumière du vaisseau est détectée de façon entièrement automatique. Dans une procédure d'initialisation originale, un minimum d'interactions est nécessaire lorsque les contours initiaux de la paroi externe du vaisseau calculés automatiquement sont proposés à l'utilisateur pour acceptation ou correction sur un nombre limité d'images de coupe longitudinale. La segmentation a été validée à l'aide de séquences IVUS in vivo provenant d'artères fémorales provenant de différents sous-groupes d'acquisitions, c'est-à-dire pré-angioplastie par ballon, post-intervention et à un examen de contrôle 1 an suivant l'intervention. Les résultats ont été comparés avec des contours étalons tracés manuellement par différents experts en analyse d'images IVUS. Les contours de la lumière et de la paroi externe du vaisseau détectés selon la méthode du fast-marching sont en accord avec les tracés manuels des experts puisque les mesures d'aire sont similaires et les différences point-à-point entre les contours sont faibles. De plus, la segmentation par fast-marching 3D s'est effectuée en un temps grandement réduit comparativement à l'analyse manuelle. Il s'agit de la première étude rapportée dans la littérature qui évalue la performance de la segmentation sur différents types d'acquisition IVUS. En conclusion, la segmentation par fast-marching combinant les informations des distributions de tons de gris et du gradient des intensités des images est précise et efficace pour l'analyse de séquences IVUS de grandes tailles. Un outil de segmentation robuste pourrait devenir largement répandu pour la tâche ardue et fastidieuse qu'est l'analyse de ce type d'images.

Étude structurale conformationnelle des toxines de l’anthrax par cryo-microscopie et dynamique moléculaire

Les toxines de l’anthrax font partie de la famille des toxines A-B dans laquelle la moitié B se fixe à la membrane de la cellule permettant par la suite la translocation de la moitié A. Dans le cas de l’anthrax, la moitié B est représentée par le Protective Antigen (PA) et la moitié A par les deux protéines Edema Factor (EF) et Lethal Factor (LF). Après le recrutement par les récepteurs cellulaires (CMG2 et TEM8), PA s’organise en heptamère. Il peut fixer jusqu'à 3 ligands (EF et LF) avant d'être endocyté. Les modèles actuels de PA suggèrent que la baisse de pH à l’intérieur des endosomes permet un changement de conformation de la forme pré-pore vers la forme pore et que les ligands EF et LF passeraient au travers le pore pour entrer dans le cytoplasme. Cependant, le diamètre du pore est environ dix fois inférieur à celui des ligands (10 Å contre 100 Å). Un processus de folding/unfolding a été proposé mais demeure controversé. Afin d'identifier le processus de passage des facteurs EF et LF dans le cytoplasme, nous avons déterminé par cryo-microscopie électronique combinée avec l’analyse d’image les structures tridimensionnelles des complexes formés par PA et LF aux étapes prépore et pore. Par la suite, une étude complémentaire par dynamique moléculaire nous a permis de modéliser à haute résolution les différentes interactions qui ont lieu au sein du complexe. La structure 3D du complexe prépore combiné à 3 LF a été déterminée à une résolution de 14 Å. Nous avons aussi calculé une structure préliminaire du complexe pore également combiné à 3 LF Celles-ci n’ont jamais été résolues auparavant et leur connaissance permet d’envisager l’étude en profondeur du mécanisme infectieux de l’Anthrax in vivo.