Ultrastructural and morphometrical analysis of apoptosis stages in cardiomyocytes of MDX mice

Our previous study of apoptosis in mdx mouse myocardium cells demonstrated the presence of middle-sized DNA fragments (60-65 kbp) in extracts of myocardium DNA, and irregular shape of membrane enveloped nuclei in cardiomyocytes. The DNA fragmentation (DNA laddering) was observed after biomechanical stress (5 min sweeming) only. Based on these results we concluded that the majority of cardiomyocytes were at the first stage of apoptosis. The purpose of this work was to provide some morphometrical quantitive characteristics of ultrastructural properties of the nuclei and mitochondria, and to determine morphological patterns of apoptosis in cardiomyocytes of mdx and C57B1 mice. To resolve the task, we made a morphometrical analysis of the electron microscope images of nuclei and mitochondria. First of all, we divided all nuclear images into three categories: normal, semi-pathological, and pathological forms according to the extent of nuclear membrane invaginations and that of condensed chromatin spreading. The most part of C57B1 cardiomyocyte nuclei belonged to the normal form (88.9 +/- 4.3%), while the smaller part (11.1 +/- 4.3%) was regarded as semi-pathological forms. Just a reverse was observed in mdx mice: the largest part of cardiomyocytes fell into category of semi-pathological (54.6 +/- 4.4%) and pathological (31.5 +/- 4.1%) forms while, the smallest part belonged to the normal form (13.8 +/- 3.0%). 24 h after biodynamic stress, the quantity of normal nuclei of C57B1 cardiomyocytes decreased to 61 +/- 5%, the number of semi-pathological nuclei increased to 39.0 +/- 4.4% (P < 0.05). The number of pathological nuclei of mdx, cardiomyocytes fell to 15.4 +/- 3.0% (P < 0.05). It means that mdx cardiomyocytes with pathological form of their nuclei disappear because of emerging the second, destructive stage of apoptosis. To estimate the degree of ultrastructural changes in the nuclei of all three forms of cardiomyocytes we counted the square/perimeter ratio in each nucleus (circle shape factor; CSF). The value of CSF for normal nuclei of all the forms of cardiomyocytes varied between 0.65 +/- 0.02 and 0.71 +/- 0.04. In semi-pathological and pathological nuclei a significant decrease in CSF to 0.56 +/- 0.02 and 0.56 +/- 0.03 was observed, respectively (P < 0.05). The biodynamical stress did not reduce the CSF value below this level. We also counted the ratio of the square to the product of a long and a short axes (ellipse shape factor; ESF). The ESF value for normal nuclei of all forms of cardiomyocytes varied between 0.97 +/- 0.01 and 0.99 +/- 0.01. In the case of mdx mice the biodynamical stress reduced ESF to 0.95 +/- 0.01 (P < 0.05) for pathological form of nuclei. The specific density of mitochondria in mdx cardiomyocytes (0.274 +/- 0.016) was less than that in C57B1 cardiomyocytes (0.329 +/- 0.018). At the destructive stage of apoptosis, the nuclei of cardiomyocytes were round in shape, the nuclear chromatin being hypercondensed, and mitochondria swollen. The cardiomyocyte morphology was in agreement with the definition of the final stage of apoptosis as secondary necrosis. Morphometrical results show that as many as 86-90% of nuclei of mdx cardiomyocytes have abnormal structure that confirms our conclusion that mdx cardiomyocytes were at the first stage of apoptosis. The final stage of apoptosis is rarely observed by biochemical or morphological methods. It suggests the presence of some inner mechanisms regulating the initiation of the final (destructive) stage of mdx cardiomyocyte apoptosis.     

Increasing the Specific Energy Concentration of High-Current Z-Pinch Plasma by Using Transient Compression Modes

Plasma Physics Reports - The results of experiments on increasing the specific energy density of high-current Z-pinch plasma by using various compression modes are presented. The experiments were...     

Torsion lability of the O=C-N-H fragment in single dipeptide molecules of L-amino acids

On the basis of ab initio MP2 and semi-empirical PM3 quantum-chemical calculations the bistability of the nonplanar O=C-N-H fragment in the structure of simple amides and dipeptides is discussed. The influence of the nature of amino acids on the structural polymorphism of the peptide group in dipeptides is shown.     

Enzyme immunoassay for the determination of hexestrol in meat

An indirect competitive enzyme-linked immunosorbent assay (ELISA) of hexestrol (HES), an anabolic hormone forbidden for use in livestock farming, has been developed. Conditions of ELISA have been optimized by varying the concentrations of the coating conjugate (HES-ovalbumin), anti-HES antiserum, casein, and Tween 20. In the absence of Tween 20 in the reaction mixture, the detection limit (IC₁₀) equaled 0.01 ng/ml, IC₅₀ equaled 0.17 ng/ml, and the working range (IC₂₀–IC₈₀) equaled 0.03–0.86 ng/ml, while, in the presence of 0.05% Tween 20, these values equaled 0.05, 2.9, and 0.26–32.0 ng/ml, respectively. Standard deviation of the analysis results did not exceed 5.4%. If ELISA was performed in the absence of detergents, the recovery value upon HES determination in spiked beef samples ranged from 74 to 147%.     

Study of oleandomycin extraction in a methylene chloride-aqueous medium system

Influence of pH, temperature and oleandomycin concentration on distribution of the antibiotic in the methylene chloride-aqueous medium system was studied. It was shown that the oleandomycin distribution coefficient did not depend on the antibiotic concentration within the ranges of 2000 to 20,000 micrograms/ml. An equation defining relationship between the oleandomycin distribution coefficient and pH of the aqueous phase is presented.     

Study of the process of inactivating oleandomycin phosphate preparations

Ageing of oleandomycin preparations was studied. Certain biologically inactive components accumulating on storage of the preparations were isolated and investigated. A scheme for ageing of oleandomycin phosphate preparations is described. The scheme includes cleaving up of the water molecule at C11 and cleaving by the epoxide ring at the early stages and hydrolysis by the glycoside bonds at the later stages.     

Acidity and solubility of gramicidin S in water

The acid-base transformations of the gramicidin S molecule in water were studied. The protonization constants of the antibiotic amino group were calculated by the data of the potentiometric titration and the antibiotic distribution in the system of chloroform-water: K1 1.55 X 10(10), K2 1.38 X 10(6), the logarithm of the distribution coefficient of gramicidin S in the system of chloroform-water (1:1) lg alpha G 4.10. By the same data the constants of water solubility of gramicidin S base (1.02 X 10(7) mol/l), gramicidin S monohydrate (1.06 X 10(-4) mol/l) and gramicidin S dihydrochloride (2.08 X 10(-4) mol/l) were calculated.     

Accelerating the Gillespie τ-Leaping Method using graphics processing units

The Gillespie τ-Leaping Method is an approximate algorithm that is faster than the exact Direct Method (DM) due to the progression of the simulation with larger time steps. However, the procedure to compute the time leap τ is quite expensive. In this paper, we explore the acceleration of the τ-Leaping Method using Graphics Processing Unit (GPUs) for ultra-large networks (>0.5e(6) reaction channels). We have developed data structures and algorithms that take advantage of the unique hardware architecture and available libraries. Our results show that we obtain a performance gain of over 60x when compared with the best conventional implementations.