Metabolic profile of dystrophic mdx mouse muscles analyzed with in vitro magnetic resonance spectroscopy (MRS)

Duchenne muscular dystrophy (DMD) is a recessive X-linked form of muscular dystrophy characterized by progressive and irreversible degeneration of the muscles. The mdx mouse is the classical animal model for DMD, showing similar molecular and protein defects. The mdx mouse, however, does not show significant muscle weakness, and the diaphragm muscle is significantly more degenerated than skeletal muscles. In this work, (1)H magnetic resonance spectroscopy (MRS) was used to study the metabolic profile of quadriceps and diaphragm muscles from mdx and control mice. Using principal components analysis (PCA), the animals were separated into groups according to age and lineages. The classification was compared to histopathological analysis. Among the 24 metabolites identified from the nuclear MR spectra, only 19 were used by the PCA program for classification purposes. These can be important key biomarkers associated with the progression of degeneration in mdx muscles and with natural aging in control mice. Glutamate, glutamine, succinate, isoleucine, acetate, alanine and glycerol were increased in mdx samples as compared to control mice, in contrast to carnosine, taurine, glycine, methionine and creatine that were decreased. These results suggest that MRS associated with pattern recognition analysis can be a reliable tool to assess the degree of pathological and metabolic alterations in the dystrophic tissue, thereby affording the possibility of evaluation of beneficial effects of putative therapies.     

Evaluation of the ethanol intake on the Calomys callosus seminal vesicle structure

The effects of chronic alcohol ingestion on the structure of the glandular epithelium of the seminal vesicle of the rodent Calomys callosus were analyzed in 24 adult animals aged 3 months divided into three experimental groups. The control group received a solid diet and tap water, the alcoholic group received the same solid diet and ethanol P.A. diluted 20% in water (v/v) for 4 months. The abstinent group received the same liquid diet of the alcoholic one for the same period and after that the alcoholic diet was changed by water for a period of 3 months. After treatment, all animals were anesthetized, weighed and sacrificed. At the end of treatment, mean body weight did not differ between animal groups. The glandular epithelial cells of the alcoholic and abstinent groups showed atrophy and ultrastructural alterations such as the presence of altered nuclei, intense dilatation of the cisterns of the granular endoplasmic reticulum, intense digestive vacuoles and lipid droplets. Ethanol ingestion provokes marked lesions on the epithelium of the seminal vesicle probably interfering on the glandular secretion.     

Evaluation of ferritin-overexpressing brain in newly developed transgenic mice

Aberrant expression of ferritin, a major iron-binding protein, has shown to be involved in neurodegenerative diseases. In this study, we generated transgenic (Tg) mice of human ferritin heavy chain (FTH) gene and investigated the effects of ferritin overexpression in FTH-Tg brain by ¹H-MRI and ¹H-MRS. The mice displayed no apparent neurological symptoms, and no specific morphological and T₂ alterations were found in the brain by MRI, and not even by histological studies. ¹H-MRS, however, revealed that some metabolic markers were significantly altered in FTH-Tg brains compared to wild-type control brains, such as decreases in myo-inositol and glutamine, and an increase in lactate. Our present studies suggested that despite the absence of neurological, morphological, T₂, and histological signatures, brain metabolisms were significantly affected in FTH-Tg mice. This study also highlights the usefulness of ¹H-MRS in the analysis of transgenic mouse models.     

Subjective and Objective Evaluation of Voice Quality in Patients With Asthma

OBJECTIVES: To evaluate the voice quality in patients with mild-to-moderate asthma by subjective and objective methods. STUDY DESIGN: Comparative, controlled, cross-sectional study. METHODS: Patients with mild-to-moderate asthma (n=40) and age- and sex-matched healthy controls (n=40) were included. Acoustic analyses were performed by the Multi-Dimensional Voice Program (MDVP; Kay Elemetrics Corporation, Lincoln Park, NJ) and the movements of the vocal cords were examined by videolaryngostroboscopy (VLS). In addition, the duration of illness, maximum phonation time, "s/z" values, and vital capacity were evaluated. Voice Handicap Index (VHI) and GRB scales were used for subjective evaluations. RESULTS: Maximum phonation time values were significantly shorter both in male and female asthma patients compared with controls (P<0.0001). Also, average shimmer values in MDVP were higher for both sexes in the patient group compared with controls (P=0.002 and P=0.04, respectively). There was a significant difference between female patients and sex-matched controls with regard to mean noise-to-harmonic ratio values (P=0.006). Female patients with asthma had higher average jitter values compared with sex-matched controls (P<0.0001). A significant difference was noted between asthma and control groups with regard to GRB scale (P<0.0001, P<0.001, and P<0.0001, respectively). The VHI score was above the normal limit in 16 (40%), and VLS findings were abnormal in 39 (97.5%) asthmatics. CONCLUSION: In asthmatic patients, maximum phonation time, frequency, and amplitude perturbation parameters were impaired, but the vital capacity and the duration of illness did not correlate with these findings.     

Cell types in the esophageal epithelium of Anguilla anguilla (Pisces, Teleostei). Cytochemical and ultrastructural characteristics

The esophagus is an important osmoregulatory organ in euryhaline teleost fishes; however, its histochemical and ultrastructural characteristics have been studied in few species. The structure of the esophageal mucosa was analyzed in the European eel (Anguilla anguilla L.) and compared to the findings reported for other species. The histological features of the esophageal mucosa, the histochemical characteristics of mucosecretory cells in the epithelial layer and the ultrastructural features of the different cell types in this layer in animals caught in fresh water and raised in the laboratory were analyzed. The role of mucus as a protective covering and its role in ion transport an the esophagus are discussed. In addition, comments on the role of tonofilament-rich epithelial cells in ion transport are made.     

Structural characteristics of the tendinous cord–papillary muscle junction in healthy and hypertensive rats

Although the myotendinous junction (MTJ) of skeletal striated muscle is well known, more detailed studies regarding the structure of the cardiac MTJ are scarce. The objective of the present study was to investigate the morphological characteristics of the MTJ in hearts of healthy and hypertensive (SHR) female rats using histological, ultrastructural and three-dimensional (SEM) methods, as well as to evaluate the expression of vinculin by immunofluorescence. In the two groups, light microscopy showed branching tendinous cords and collagen bundles penetrating the apex of the finger-like projections of the papillary muscle. SEM analysis revealed an enlarged apex of the papillary muscle in SHR which was not observed in healthy animals. The loss of force transmission appears to be compensated by the amplified connection between the papillary muscle and valvular collagen. A large number of intercalated disks close to the fiber apex, small amounts of an amorphous intercellular substance and numerous vesicles were observed in SHR. In these animals, the expression of vinculin was more marked showing a regular distribution and a pattern of transverse striations along the sarcolemma. The presence of this protein in transverse bands suggests that vinculin surrounds myofibrils in the region of the Z band. Vinculin staining was also more marked in the region of the tendinous cord–papillary muscle junctions of SHR compared to control animals. Vinculin was quantified by electrophoresis and higher amounts of this protein were observed in SHR compared to control animals.     

Ultrastructural characteristics of the gerbil pterygoid medial muscle after experimental occlusal alteration

Ten male young adult gerbils Meriones unguiculatus weighing 55g were divided in two groups: experimental (n=5) animals submitted to exodontia of the left upper molars, and sham-operated were used as control (n=5) group. The aim of the present study was to investigate the ultrastructural effects of occlusal alteration induced by unilateral exodontia on medial pterygoid muscle. After 60 days, the animals were sacrificed by perfusion intracardially with a modified Karnovsky solution after anesthetized with overdose of urethane (3g/kg i.p.). The small samples of this muscle were fixed in 2.5% glutaraldehyde solution and post-fixed in 1% buffered osmium tetroxide solution, dehydrated in ascending concentration of ethanol, and embedded in Spurr resin. Several micrographs data showed that in this period of time, the unilateral teeth extraction was able to induce modifications on the medial pterygoid muscle fibers and capillaries at ultrastructural levels as compared to control muscles and to the ones in the contralateral side of the experimental animals. Ultrastructural changes suggest that injury was greater in the hypofunctional muscle, ipsilateral to teeth extraction.     

A study on quantitative analysis of exposure dose caused by patient depending on time and distance in nuclear medicine examination

This study evaluated possible actions that can help protect against and reduce radiation exposure by measuring the exposure dose for each type of isotope that is used frequently in nuclear medicine before performing numerical analysis of the effective half-life based on the measurement results. From July to August in 2010, the study targeted 10, 6 and 5 people who underwent an ¹⁸F-FDG (fludeoxyglucose) positron emission tomography (PET) scan, ^99mTc-HDP bone scan, and ²⁰¹Tl myocardial single-photon emission computed tomography (SPECT) scan, respectively, in the nuclear medicine department. After injecting the required medicine into the subjects, a survey meter was used to measure the dose depending on the distance from the heart and time elapsed. For the ¹⁸F-FDG PET scan, the dose decreased by approximately 66% at 90 min compared to that immediately after the injection and by 78% at a distance of 1 m compared to that at 0.3 m. In the ^99mTc-HDP bone scan, the dose decreased by approximately 71% in 200 min compared to that immediately after the injection and by approximately 78% at a distance of 1 m compared to that at 0.3 m. In the ²⁰¹Tl myocardial SPECT scan, the dose decreased by approximately 30% in 250 min compared to that immediately after the injection and by approximately 55% at a distance of 1 m compared to that at 0.3 m. In conclusion, the dose decreases by a large margin depending on the distance and time. In conclusion, this study measured the exposure doses by isotopes, distance from the heart and exposure time, and found that the doses were reduced significantly according the distance and the time.     

Structure and composition of arytenoid cartilage of the bullfrog (Lithobates catesbeianus) during maturation and aging

The aging process induces progressive and irreversible changes in the structural and functional organization of animals. The objective of this study was to evaluate the effects of aging on the structure and composition of the extracellular matrix of the arytenoid cartilage found in the larynx of male bullfrogs (Lithobates catesbeianus) kept in captivity for commercial purposes. Animals at 7, 180 and 1080 days post-metamorphosis (n = 10/age) were euthanized and the cartilage was removed and processed for structural and biochemical analysis. For the structural analyses, cartilage sections were stained with picrosirius, toluidine blue, Weigert’s resorcin-fuchsin and Von Kossa stain. The sections were also submitted to immunohistochemistry for detection of collagen types I and II. Other samples were processed for the ultrastructural and cytochemical analysis of proteoglycans. Histological sections were used to chondrocyte count. The number of positive stainings for proteoglycans was quantified by ultrastructural analysis. For quantification and analysis of glycosaminoglycans were used the dimethyl methylene blue and agarose gel electrophoresis methods. The chloramine T method was used for hydroxyproline quantification. At 7 days, basophilia was observed in the pericellular and territorial matrix, which decreased in the latter over the period studied. Collagen fibers were arranged perpendicular to the major axis of the cartilaginous plate and were thicker in older animals. Few calcification areas were observed at the periphery of the cartilage specimens in 1080-day-old animals. Type II collagen was present throughout the stroma at the different ages. Elastic fibers were found in the stroma and perichondrium and increased with age in the two regions. Proteoglycan staining significantly increased from 7 to 180 days and reduced at 1080 days. The amount of total glycosaminoglycans was higher in 180-day-old animals compared to the other ages, with marked presence of chondroitin- and dermatan-sulfate especially in this age. The content of hydroxyproline, which infers the total collagen concentration, was higher in 1080-day-old animals compared to the other ages. The results demonstrated the elastic nature of the arytenoid cartilage of L. catesbeianus and the occurrence of age-related changes in the structural organization and composition of the extracellular matrix. These changes may contribute to alter the function of the larynx in the animal during aging.     

Ultrastructural and biochemical changes of the medial pterygoid muscle induced by unilateral exodontia

The aim of the present study was to investigate the histological, biochemical and ultrastructural effects of occlusal alteration induced by unilateral exodontia on medial pterygoid muscle in guinea pigs, Cavia porcellus. Thirty (n=30) male guinea pigs (450g) were divided into two groups: experimental-animals submitted to exodontia of the left upper molars, and sham-operated were used as control. The duration of the experimental period was 60 days. Medial pterygoid muscles from ipsilateral and contralateral side were analyzed by histological (n=10), histochemical (n=10), and ultrastructural (n=10) methods. The data were submitted to statistical analysis. When the ipsilateral side was compared to the control group, it showed a significantly shorter neuromuscular spindle length (P<0.05), lower oxidative metabolic activity, and microvessel constriction, in spite of the capillary volume and surface density were not significantly different (P>0.05). In the contralateral side, the neuromuscular spindles showed significantly shorter length (P<0.05), the fibers reflected a higher oxidative capacity, the blood capillaries showed endothelial cell emitting slender sprouting along the pre-existing capillary, and significantly higher blood capillary surface density, and volume density (V(v)=89% Mann-Whitney test, P<0.05). This finding indicated a complex morphological and functional medial pterygoid muscle adaptation to occlusal alteration in this experimental model. Considering that neuromuscular spindles are responsible for the control of mandibular positioning and movements, the professional should consider if these changes interfere in the success of clinical procedures in medical field involving stomatognathic structures.     

Histological assessment of SJL/J mice treated with the antioxidants coenzyme Q10 and resveratrol

The muscular dystrophies (MDs) are genetic disorders of muscle degeneration due to mutations in genes that encode a wide variety of proteins. Dysferlinopathy are characterized by the absence of dysferlin in skeletal muscle and an autosomal recessive mode of inheritance. Both histological and ultrastructural pathology have been well established in dysferlinopathy patients and dysferlin-deficient animal models. To our knowledge the effect of antioxidant supplementation on this level has not been described previously. This article therefore focuses on the histopathology to reveal the effect of antioxidant supplementation. The study aimed to determine, at cellular level, the histopathological changes in the SJL/J mouse model following a 90 day trial with antioxidant supplementation. Markedly reduced inflammatory insult in the more affected quadriceps muscles of animals treated with high doses of CoQ10 and a combination of resveratrol/CoQ10 were observed. The outcome provides evidence that high doses of antioxidant supplementation resulted in decreased dystrophic markers and enhanced tissue integrity at cellular level.     

Bone healing process in critical-sized defects by rhBMP-2 using poloxamer gel and collagen sponge as carriers

The bone morphogenetic protein type 2, is an osteoinduction protein, but new carriers to improve its actions are being studied in recent researches. The objective of this study was to evaluate the poloxamer gel as potential carrier of the rhBMP-2 during the bone healing process and verify if the collagen sponge can improve this process. Fifty-six male Wistar rats were used, which were divided into two groups, considering two periods of time. Thus, each one of these groups was divided in four groups with seven animals each, according to the treatment and the defect filled by 4 microg rhBMP-2 in aqueous solution, 4 microg rhBMP-2 in aqueous solution+collagen sponge, 4 microg rhBMP-2 combined with poloxamer gel, and 4 microg rhBMP-2 combined with poloxamer gel+collagen sponge. After 2 and 4 weeks, respectively, animals were perfused and the hemi-mandibles removed for histological analysis. Statistical analysis of the data showed significant difference between all analyzed groups (p<0.01), and between the periods of time (p=0.0118). There was no interaction between the applied treatment and considered periods of time (p=0.642). Results showed that the rhBMP-2 used in this study was able to induce bone regeneration and had its action optimized when combined to the used carriers, being the bone repair time-dependent.     

Analysis of tracheal noise of forced human expiration according to clinical experimental data

The analysis of tracheal noise of forced expiration is one of the promising methods for the diagnosis of bronchial patency disorders, which are a sign of such widespread diseases as bronchial asthma and chronic obstructive bronchitis. The aim of this study is the verification and refinement of the acoustic model of forced expiration (Korenbaum et al., 1998) on the basis of a statistical analysis of clinical experimental data. A sample of 127 volunteers (from 18 to 74 years old) is used as an experimental statistical model: 34 persons suffering from bronchial asthma, 21 persons suffering from chronic obstructive bronchitis, 29 healthy persons, and 43 persons liable to developing the aforementioned diseases. The following parameters are analyzed: the total duration of noise of forced expiration at the trachea, the duration of wheeze of forced expiration with frequencies from 400 to 600 Hz, and the presence of narrowband high-frequency (over 600–700 Hz) spectral components at the end of forced expiration and during the whole expiration process. Reliable differences in the parameters of tracheal noise are revealed in the groups under study. The high prognostic value (a sensitivity of 89% and a specificity of 86%) of the parameters of tracheal noise of forced expiration, which were suggested a priori as the diagnostic parameters on the basis of the acoustic model (Korenbaum et al., 1998), indirectly confirms the adequacy of this model. The biomechanical-pathophysiological interpretation of the occurrence of acoustic deviations in the experimental sample groups provides an opportunity to relate the parameters of tracheal noise of forced expiration to the degree of mechanical nonuniformity of the lungs.     

Bone repair using mineral trioxide aggregate combined to a material carrier, associated or not with calcium hydroxide in bone defects

Mineral trioxide aggregate (MTA) is a powder aggregate containing mineral oxides with a good biological action and may facilitate the regeneration of the periodontal ligament and formation of bone. Calcium hydroxide demonstrates antibacterial properties, enhances tissue dissolution, and induces bone formation. The objective of this study was to evaluate the MTA in the bone healing process and verify if the calcium hydroxide P.A. can improve and accelerate this process. It was used forty male Wistar rats, which were divided into two groups, considering or not the use of calcium hydroxide P.A. solution before treatment. Thus, each one of these groups was divided in four groups with five animals each, according to the treatment and the defect filled by: animal's coagulum, monoolein gel, MTA in aqueous solution, and MTA combined with monoolein gel. After 10 days, the animals were perfused and the right hemimandibles removed for histological analysis. Statistical analysis of the data showed significant difference between all analyzed groups when it was made comparisons using or not calcium hydroxide P.A. (p<0.0001). There was found statistical difference between the groups that was inserted or not MTA, independently the calcium hydroxide application (p<0.05). Results showed that the MTA used was able to induce bone regeneration and had its action optimized when combined to calcium hydroxide P.A.     

Mössbauer Spectrometer with Novel Moving System and Resonant Detection of Gamma Rays

Temperature-dependent ⁵⁷Fe Mössbauer spectroscopy has been used to elucidate the hyperfine parameters and dynamical behavior of the metal atom in several organo-iron complexes which have one or more η⁵ P₅ ring structures as ligated groups. The spin-lattice relaxation of the (paramagnetic) one-electron oxidation products occurs on a time scale fast compared to τ_1/2 (ME) at temperatures in the range 85 < T < 320 K.     

Improved anti-Stokes energy transfer between rare earth ions in Er（0.5）Yb（9.5）：FOV oxyfluoride vitroceramics explains the strong color reversal

The widely used energy transfer theory is a foundation of luminescence,in which the rates of Stokes and antiStokes processes have the same calculation formula.An improvement on the anti-Stokes energy transfer to explain the fluorescence intensity reversal between the red and green fluorescence of Er(0.5)Yb(9.5):FOV is reported in the present article.The range of the intensity reversal Σ was measured to be 877.Dynamic processes for 16 levels were simulated.A coefficient,the improvement factor of the intensity ratio of Stokes to anti-Stokes processes in quantum Raman theory compared to classical Raman theory,is introduced to successfully describe the anti-Stokes energy transfer.A new method to calculate the distance between the rare earth ions,which is critical for the energy transfer calculation,is proposed.The validity of these important improvements is also proved by experiment.     

Depth profiling of cells and tissues by using C60 and SF5 as sputter ions

In the present study, SF₅⁺ and C₆₀⁺ were used as primary ions for sputtering and Bi₃⁺ was used as primary ions for analysis. The depth profiling procedure was utilized to make 3D images of the chemistry of single cultured cells and tissue samples of intact intestinal epithelium.The results show sputtering of organic material from cells and tissue with both SF₅⁺ and C₆₀⁺ sources. Cholesterol fragments were found in the superficial layers when sputtering with C₆₀⁺. Spectra were collected revealing the change in yield along the z-axis of the sample. 3D images of the localization of Na, K, phosphocholine and cholesterol were constructed with both ion sources for single cell cultures and the mouse intestine.Cryostate sections of mouse intestine were analysed in 2D and the results were compared with the 3D image of the intestine. The localization of cholesterol and phosphocholine was found to be similar in cryostate sections analysed in two dimensions and the sputtered, freeze-dried intestine analysed in 3D. The comparison of 2D and 3D images suggest that the phosphocholine signal faded with C₆₀⁺ sputtering. In conclusion, both C₆₀⁺ and SF₅⁺ can be used as primary ion sources for sputtering of organic material from cells and tissues. Consecutive analysis with a Bi₃⁺ source can be used to obtain image stacks that could be used for reconstruction of 3D images.     

The significance of changes in Mytella falcata (Orbigny, 1842) gill filaments chronically exposed to polluted environments

The impact of pollutants in an organism can be observed by changes in functional complexity at different levels. Bivalve gills are suitable for histopathological analysis because of their structure and function. This study aimed at examining the morphology of Mytella falcata gill filaments from three sites in the Santos estuary (São Paulo, Brazil) with different levels of environmental degradation to identify possible changes in gill structure and discuss the significance of these alterations. For this purpose, histological, histochemical and ultrastructural techniques were used. The filaments of animals from site A (less impacted site) were intact, while in sites B and C, pathological changes were observed, such as: detachment of the epithelium in the intermediate zone, morphological changes of this epithelium, inflammatory process, increase in the number of mucous cells and cell turnover processes. These results suggest that the related changes are an attempt to prevent the entrance of pollutants through gill filaments into the entire organism and that cell turnover is the final way to compensate cell injury.