Quantification of cellular action fibers alignment from confocal microscopic images.

The cellular rheology has recently undergone a rapid development with particular attention to the cytoskeleton mechanical properties and its main components - actin filaments, intermediate filaments, microtubules and crosslinked proteins. However it is not clear what are the cellular structural changes that directly affect the cell mechanical properties. Thus, in this work, we aimed to quantify the structural rearrangement of these fibers that may emerge in changes in the cell mechanics. We created an image analysis platform to study smooth muscle cells from different arteries: aorta, mammary, renal, carotid and coronary and processed respectively 31, 29, 31, 30 and 35 cell image obtained by confocal microscopy. The platform was developed in Matlab (MathWorks) and it uses the Sobel operator to determine the actin fiber image orientation of the cell, labeled with phalloidin. The Sobel operator is used as a filter capable of calculating the pixel brightness gradient, point to point, in the image. The operator uses vertical and horizontal convolution kernels to calculate the magnitude and the angle of the pixel intensity gradient. The image analysis followed the sequence: (1) opens a given cells image set to be processed; (2) sets a fix threshold to eliminate noise, based on Otsu's method; (3) detect the fiber edges in the image using the Sobel operator; and (4) quantify the actin fiber orientation. Our first result is the probability distribution II(Δθ) to find a given fiber angle deviation (Δθ) from the main cell fiber orientation θ0. The II(Δθ) follows an exponential decay II(Δθ) = Aexp(-αΔθ) regarding to its θ0. We defined and determined a misalignment index α of the fibers of each artery kind: coronary αCo = (1.72 ‘+ or =’ 0.36)rad POT -1; renal αRe = (1.43 + or - 0.64)rad POT -1; aorta αAo = (1.42 + or - 0.43)rad POT -1; mammary αMa = (1.12 + or - 0.50)rad POT -1; and carotid αCa = (1.01 + or - 0.39)rad POT -1. The α of coronary and carotid are statistically different (p < 0.05) among all analyzed cells. We discussed our results correlating the misalignment index data with the experimental cell mechanical properties obtained by using Optical Magnetic Twisting Cytometry with the same group of cells.

Polymorphic-induced transformations in CaTa2O6 single-crystal fibers obtained by laser-heated pedestal growth

Calcium tantalite (CaTa2O6) single crystal fibers were obtained by the laser-heated pedestal growth method (LHPG). At room temperature, this material can present three polymorphic modifications. The rapid crystallization inherent to the LHPG method produced samples within the Pm3 space group, with some chemical disorder. In order to check for polymorphic-induced transformations, the CaTa2O6 fibers have been submitted to different thermal treatments and investigated by micro-Raman spectroscopy. For short annealing times (15 min) at 1200 °C, the cubic modification was maintained, though with an improved crystalline quality, as evidenced by the enhanced inelastic scattered intensity (by ca. 250%) and narrowing of Raman bands. The polarized Raman spectra respected very well the predicted symmetries and the selection rules for this cubic modification. On the other hand, long annealing times (24 h) at 1200 °C led to a complete (irreversible) polymorphic transformation. The Raman bands became still more intense (ca. 15 times larger than for the as-grown fibers), narrower, and several new modes appeared. Also, the spectra became unpolarized, demonstrating a polycrystalline nature of the transformed crystals. The observed Raman modes could be fully assigned to an orthorhombic modification of CaTa2O6 belonging to the Pnma space group.