Метод определения основных метрических характеристик нейроцитов в спинномозговых узлах человека

Abstract

У роботі пропонується методика отримання комплексної морфометричної характеристики нервових клітин чутливих нервових вузлів. Запропонована методика дозволяє при наявності нескладного устаткування і загальнодоступних комп’ютерних програм провести обчислення розмірів нейроцитів, кількості, відносної площі на гістологічному зрізі. Описаний метод може бути також використаний для дослідження інших елементів тканин при їх відносно однорідній структурі; В работе предлагается методика получения комплексной морфометрической характеристики нервных клеток чувствительных нервных узлов. Предложенная методика позволяет при наличии несложного оборудования и общедоступных компьютерных программ провести вычисления размеров нейроцитов, количества, относительной площади на гистологическом срезе. Описанный метод может быть также использован для исследования других элементов тканей при их относительно однородной структуре; With the development of modern digital technologies have become available resources of the electronic computing technology, with the presence of many different standard and specialized programs that allow for a new approach to solving the tasks. For example, the most modern high-light microscope equipped with software that allows you to carry out almost any morphometric study. However, these devices and their accompanying licensed programs are not always available due to their high cost. The purpose of the work. Develop a methodology for determining the basic metric characteristics dorsal root ganglion cells developed on the basis of widely available software that does not require expensive equipment. Description of the method. The described technique lies in that, initially photographed histological sections of human lumbar DRG, followed by manufacturing (in the graphic editor Adobe Photoshop) of two-dimensional spatial reconstruction of the longitudinal section. The resulting digital images are processed by outlining the contours of neurons in a standard graphic editor Paint, the operating system Microsoft Windows 7 in black, with a wide brush. Then the image opens in the editor ImageJ. Before the opening of the image program must be calibrated using the command Analyze> SetScale to define the spatial scale of the image. Active measurement results in this case are represented in the set of calibration units such as millimeters. In the next step, using the command Image> Type active image converts in 8-bit (resulting in an image in shades of gray). Then use command Image> Adjust> Threshold automatically or interactively adjustable upper and lower threshold to segment the region of interest and the background image. This removes all unwanted items, except circled circuits of neurons, resulting in a white background, which shows the black outlines of neurons. Then you need to fill the white space within the contours using command Process> Binary> FillHoles. At the next stage, convert the image into the mask by command Process> Binary> Convert to Mask and separate each element from each other at a distance of one pixel by command Process> Binary> Watershed, asking relevant parameters. Then make the setting: in the menu Analyze> SetMeasurments select Area (calculation of the total area of neurons isolated in black) and Feret’s Diameter (gives two sizes: the maximum and minimum of each individual element in black, in this case, the maximum and minimum diameter of neurocyte). Next is the calculation of the number, size and total area of regions of black color, which corresponds to the area of neurons using the Analyze> AnalyzeParticles In the window that opens, you must enter in a row Size - 100-infinity, and select points: Pixel units, Display results, Clear results, Summarize. In line Circularity leave unchanged the value (0.00-1.00), and in line Show select Outlines. Then click Ok. As a result of our actions, will open three windows: one window with the general results, which contains data on the number of neurons and their total area; the other with the size of each of the elements (the minimum - MinFeret and maximum - Feret); third window with the designation of each of the individual elements, with numbering and its contour. Results of the research. This method has been used by us for the comparative assessment of changes in quantitative characteristics of human lumbar dorsal root ganglion neurons in young and elderly people. Also, the above method may be used for research or other tissue elements at their relatively homogeneous structure. Conclusion. Thus, the above-described method allows in the presence of a minimum number of special equipment (microscope with a simple digital camera or photo adapter) and the computer with the package widely available software to perform a comprehensive morphometric study of sensory ganglia and other anatomical structures.