Walter Blondel

Advantages and limitations of commonly used methods to assay the molecular permeability of gap junctional intercellular communication

The role of gap junctional intercellular communication (GJIC) in regulation of normal growth and differentiation is becoming increasingly recognized as a major cellular function. GJIC consists of intercellular exchange of low molecular weight molecules, and is the only means for direct contact between cytoplasms of adjacent animal cells. Disturbances of GJIC have been associated with many pathological conditions, such as carcinogenesis or hereditary illness. Reliable and accurate methods for the determination of GJIC are therefore important in cell biology studies. There are several methods used successfully in numerous laboratories to measure GJIC both in vitro and in vivo. This review comments on techniques currently used to study cell-to-cell communication, either by measuring dye transfer, as in methods like microinjection, scrape loading, gap-fluorescence recovery after photobleaching (gap-FRAP), the preloading assay, and local activation of a molecular fluorescent probe (LAMP), or by measuring electrical conductance and metabolic cooperation. As we will discuss in this review, these techniques are not equivalent but instead provide complementary information. We will focus on their main advantages and limitations. Although biological applications guide the choice of techniques we describe, we also review points that must be taken into consideration before using a methodology, such as the number of cells to analyze.

Mosaicing of Bladder Endoscopic Image Sequences: Distortion Calibration and Registration Algorithm

Cancers located on the internal wall of bladders can be detected in image sequences acquired with endoscopes. The clinical diagnosis and follow-up can be facilitated by building a unique panoramic image of the bladder with the images acquired from different viewpoints. This process, called image mosaicing, consists of two steps. In the first step, consecutive images are pairwise registered to find the local transformation matrices linking geometrically consecutive images. In the second step, all images are placed in a common and global coordinate system. In this contribution, a mutual information-based similarity measure and a stochastic gradient optimization method were implemented in the registration process. However, the images have to be preprocessed in order to register the data in a robust way. Thus, a simple correction method of the distortions affecting endoscopic images is presented. After the placement of all images in the global coordinate system, the parameters of the local transformation matrices are all adjusted to improve the visual aspect of the panoramic images. Phantoms are used to evaluate the global mosaicing accuracy and the limits of the registration algorithm. The mean distances between ground truth positions in the mosaiced image range typically in 1-3 pixels. Results given for in vivo patient data illustrate the ability of the algorithm to give coherent panoramic images in the case of bladders.

Fast construction of panoramic images for cystoscopic exploration.

Cystoscopy is used as a reference clinical examination in the detection and visualization of pathological bladder lesions. Evolution observation and analysis of these lesions is easier when panoramic images from internal bladder walls are used instead of video sequences. This work describes a fast and automatic mosaicing algorithm applied to cystoscopic video sequences, where perspective geometric transformations link successive image pairs. This mosaicing algorithm begins with a fast initialization of translation parameters computed by a cross-correlation of images, followed by an iterative optimization of transformation parameters. Finally, registered images are projected onto a global common coordinate system. A quantifying test protocol applied over a phantom yielded a mosaicing mean error lower than 4 pixels for a 1947 x 1187 pixels panoramic image. Qualitative evaluation of 10 panoramic images resulting from videos of clinical cystoscopies was performed. An analysis performed over translation values from these clinical sequences (in vivo) is used to modify the mosaicing algorithm to be able to do a dynamic selection of image pairs. Construction time of panoramic images takes some minutes. At last, algorithm limits are discussed.

Flexible calibration of structured-light systems projecting point patterns

Structured-light systems (SLSs) are widely used in active stereo vision to perform 3D modelling of a surface of interest. We propose a flexible method to calibrate SLSs projecting point patterns. The method is flexible in two respects. First, the calibration is independent of the number of points and their spatial distribution inside the pattern. Second, no positioning device is required since the projector geometry is determined in the camera coordinate system based on unknown positions of the calibration board. The projector optical center is estimated together with the 3D rays originating from the projector using a numerical optimization procedure. We study the 3D point reconstruction accuracy for two SLSs involving a laser based projector and a pico-projector, respectively, and for three point patterns. We finally illustrate the potential of our active vision system for a medical endoscopy application where a 3D cartography of the inspected organ (a large field of view surface also including image textures) can be reconstructed from a video acquisition using the laser based SLS.

Intradermal tacrolimus prevent scar hypertrophy in a rabbit ear model: a clinical, histological and spectroscopical analysis

Background: Keloids and hypertrophic scars (HSc) affect 4.5–16% of the population. Thus far, the different approaches of keloid treatment are not very efficient, with a 50% relapse rate and many ongoing researches are looking for simple, safe and more efficient therapeutic methods. Tacrolimus is an immunomodulator that could be useful in treating keloid.

Investigation of 3-D mechanical properties of blood vessels using a new in vitro tests system: results on sheep common carotid arteries

In order to investigate the three-dimensional (3-D) mechanical properties of blood vessels, a new experimental device is described allowing in vitro static and dynamic measurements on segments of arteries with high technical performances. Static tests are applied to sheep common carotid arteries. Considering a thick-walled cylindrical model of orthotropic material under large deformations, a classical 3-D approach based on strain energy density is used to calculate the resulting mechanical behavior law in radial and circumferencial directions and stresses distribution throughout the wall thickness. Results are presented with reference to unloaded and zero-stress initial state thanks to simple measurements of inner and outer circumferences. A particular ratio relating the two main stresses (circumferential and longitudinal) is calculated that put into the forth the progressive modifications in the direction of the predominant stress in the wall and the specific radial location where these changes occur. The authors observe that this point location is a function of the test conditions of the specimen, i.e., stretching length and level of pressure.

Monte Carlo modeling of multilayer phantoms with multiple fluorophores: simulation algorithm and experimental validation

This work is first a description of a statistical simulation algorithm developed for simulating the spectral absorption and emission of several fluorophores in an absorbing and diffusing multilayer model. Second, a detailed experimental validation of the simulation program is conducted on two sets of liquid and solid multilayer phantoms, containing one, two, or three fluorophores, within absorbing and scattering media. Experimental spatially resolved reflectance spectra are acquired in the wavelength band 400 to 800 nm and compared to corresponding simulated spectra. The degree of similarity between experimentation and simulation data is quantified. The results obtained underline good correlations with mean errors varying from 2 to 10%, depending on the number of layers and on the complexity of the phantoms composition.

Classification of ultraviolet irradiated mouse skin histological stages by bimodal spectroscopy: multiple excitation autofluorescence and diffuse reflectance

Histopathological analysis and in vivo optical spectroscopy were used to discriminate several histological stages of UV-irradiated mouse skin. At different times throughout the 30-week irradiation, autofluorescence (AF) and diffuse reflectance (DR) spectra were acquired in a bimodal approach. Then skin was sampled and processed to be classified, according to morphological criteria, into four histological categories: normal, and three types of hyperplasia (compensatory, atypical, and dysplastic). After extracting spectral characteristics, principal component analysis (data reduction) and the k-nearest neighbor classifying method were applied to compare diagnostic performances of monoexcitation AF (based on each of the seven excitation wavelengths: 360, 368, 390, 400, 410, 420, and 430 nm), multiexcitation AF (combining the seven excitation wavelengths), DR, and bimodal spectroscopies. Visible wavelengths are the most sensitive ones to discriminate compensatory from precancerous (atypical and dysplastic) states. Multiexcitation AF provides an average 6-percentage-point increased sensitivity compared to the best scores obtained with monoexcitation AF for all pairs of tissue categories. Bimodality results in a 4-percentage-point increase of specificity when discriminating the three types of hyperplasia. Thus, bimodal spectroscopy appears to be a promising tool to discriminate benign from precancerous stages; clinical investigations should be carried out to confirm these results.

Hybrid feature selection and SVM-based classification for mouse skin precancerous stages diagnosis from bimodal spectroscopy

This paper deals with multi-class classification of skin pre-cancerous stages based on bimodal spectroscopic features combining spatially resolved AutoFluorescence (AF) and Diffuse Reflectance (DR) measurements. A new hybrid method to extract and select features is presented. It is based on Discrete Cosine Transform (DCT) applied to AF spectra and on Mutual Information (MI) applied to DR spectra. The classification is performed by means of a multi-class SVM: the M-SVM2. Its performance is compared with the one of the One-Versus-All (OVA) decomposition method involving bi-class SVMs as base classifiers. The results of this study show that bimodality and the choice of an adequate spatial resolution allow for a significant increase in diagnostic accuracy. This accuracy can get as high as 81.7% when combining different distances in the case of bimodality.

A novel 3D surface construction approach: Application to three-dimensional endoscopic data

Video-endoscopy is the standard clinical procedure for visual exploration of internal walls of hollow organs. For the bladder, the lesion diagnosis is complex because the endoscopic images are bi-dimensional and cover only small bladder areas. 3D endoscopes, based on stereoscopic active vision principles, were recently proposed and validated. This paper presents a 3D reconstruction algorithm using 2D texture images and a few 3D points located on the internal wall surfaces provided by such endoscopes. The algorithm constructs a 3D panoramic surface using the 3D reconstruction method guided by 2D image registration. We show on realistic bladder phantoms that the algorithm is able to reconstruct 3D points and surfaces with a sub-millimetre accuracy.