Morphological characterization of rat incisor fluorotic lesions

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morphological evidences indicate that the interference of cimetidine on the peritubular components is responsible for detachment and apoptosis of Sertoli cells

Cimetidine, referred as antiandrogenic agent, has caused alterations in the seminiferous tubules, including alterations in the peritubular tissue and death of myoid cells by apoptosis. Regarding the structural and functional importance of the peritubular tissue for the maintenance of Sertoli cells (SC), we purpose to investigate the SC-basement membrane interface, focusing the morphological features of SC and their interaction with the basement membrane in the affected tubules by cimetidine. Ten animals were distributed into two groups, control (CG) and cimetidine (CmG) which received saline solution and 50 mg of cimetidine per kg of body weight, respectively, for 52 days. The testes were fixed, dehydrated and embedded for analyses under light and transmission electron microscopy. Paraffin sections were submitted to the TUNEL method; sections of testes embedded in glycol methacrylate were submitted to PAS method and stained by H&E for morphological and quantitative analyses of Sertoli Cells. In the CmG, the SC nuclei were positive to the TUNEL method and showed typical morphological alterations of cell death by apoptosis (from early to advanced stages). A significant reduction in the number of Sertoli Cells was probably due to death of these cells by apoptosis. A close relationship between SC nuclear alterations (including a high frequency of dislocated nuclei from the basal portion) and damage in the peritubular tissue was observed. The ultrastructural analysis showed a parallelism between the gradual advancement of apoptotic process in SC and detachment of the anchoring sites (hemidesmosomes) of SC plasma membrane from the lamina densa. The presence of portions of lamina densa underlying the detached hemidesmosomes indicates a continuous deposition of lamina densa, resulting in the thickening of the basal lamina. The results indicate a possible disarrangement of the SC cytoskeleton, including the focal adhesion structure. These alterations are related to SC apoptosis and probably result from disturbs induced by cimetidine on the peritubular tissue.

Morphological studies on the heart ventricle of African catfish (Clarias gariepinus)

The histological and ultrastructural characteristics of the heart ventricle in Clarias gariepinus (African catfish) has been studied by light microscopy and transmission electron microscopy. The ventricle of the heart has a saccular shape and the myocardial wall consists of an outer thin compact myocardium and an inner well-developed spongy myocardium. The myocardial layer has small myocytes, interstitial spaces and blood vessels. The myocytes are the major constituents of the ventricular wall. They are long cells, with large nuclei, and predominantly euchromatin. The sarcoplasmic reticulum of the ventricular myocytes consists of a network of tubules and subsarcolemmal cisternae oriented mainly along the longitudinal axis of the myofibrils. In contrast to the ventricular structure of other fish species described in the literature (Greer-Walker et al., 1985 Santer, 1985 Sanchez-Quintana et al., 1995, 1996), the African catfish, a freshwater sedentary fish recently introduced in neotropical climatic environments, showed a saccular ventricle that consisted of two muscle layers, a thin compact layer with large vessels and a developed spongy layer. The ultrastructure of the ventricular myocardium of C.gariepinus is similar to that of other teleosts, inclusive that of fish with other swimming habits.

Morphological and histochemical study of the myotomal muscle in pacu (Piaractus mesopotamicus Holmberg, 1887) during the initial growth phases

Morphological and histochemical methods were used to evaluate the myotomal muscle characteristics of pacu (Piaractus mesopotamicus) from hatching to 40 days old. During the larval period, the musculature consisted predominantly of white muscle. White and red muscle mass increased at 10, 20, 30 and 40 days after hatching. The larvae had round muscle fibers with a moderate degree of maturation and central nuclei. In subsequent phases, small and immature fibers were visible near larger and more differentiated fibers. Undifferentiated cells or presumptive myoblasts located in embryogenic zones were visible in the dorsal and ventral regions, and were more evident at 30 and 40 days. The red muscle fibers located in the subdermal region, had oxidative metabolism and slow contraction, whereas the more predominant white muscle fibers had glycolytic metabolism and fast contraction. Our findings indicate that during the initial phases, myotomal muscle growth in pacu occurs by both, muscle fiber hypertrophy and hyperplasia. The analysis of frequency of red and white muscle fibers shows that hyperplastic growth is intense in this period. As the growth rate in adult fish is related to the number of muscle fibers in young fish, extrinsic factors could change the muscle fiber phenotype and influence their ultimate size.

Morphological changes on the hard palatine mucosa of rats (Rattus norvegicus albinus) after chronic alcohol consumption

In order to examine the effects of alcohol on the hard palatine mucosa of rats, sixty adult female rats (Rattus norvegicus albinus) were divided into two experimental groups. The control group received solid diet (Purina rat chow) and tap water ad libitum. The alcoholic group received the same solid diet and was allowed to drink only sugar cane brandy dissolved in 30% Gay Lussac (v/v). At the end of periods of 90, 180 and 270 days of treatment, the animals at estro were sacrificed and the hard palatine mucosa were prepared for TEM and SEM methods. The basal cells of the alcoholic groups (90, 180 and 270 days of treatment) demonstrated some alterations: the intercellular spaces between these cells were higher, presented cytoplasmatic lipid droplets and autolysis. Also, the connective tissue showed intense lipid droplets accumulation in the alcoholic groups. These modifications suggested that chronic alcohol ingestion was able to modify the integrity of the cells in the rat hard palatine mucosa.

Morphological features of the epididymal epithelium of gerbil, Meriones unguiculatus

Principal cells of the ducts epididymis of the Mongolian gerbil showed ultrastructural characteristics of lining epithelium cells close related to processes of protein secretion, and transcytosis occurring between adjacent principal cells which were mainly verified in the initial segment. Principal cells also presented roles of fluid phase and adsorptive endocytoses, as well as autophagic and heterophagic lysosomal activities mainly observed in the caput epididymis. Columnar (principal) cells of the corpus epididymidis presented great number of variable vesicles and vacuoles distributed in all the cytoplasmic levels occurring a progressive coalescence pattern among them, which help to guarantee formation of cytoplasmic channels for fluid phase transport between the tubular lumen and epididymal interstitium. Clear cells were presented in the initial segment and predominately in the cauda epididymis epithelium of the gerbil and showed marked ultrastructural characteristics of endocytosis activities occurrence, perhaps directly related to the turnover of fluid phase of spermatozoa stored into the lumen of the distal tail. Other epididymal epithelium cells were verified and described such as basal, halo, apical and dark cells, but they did not presented special ultrastructural features. (c) 2007 Elsevier Ltd. All rights reserved.

Morphological alterations in the epithelium of the oral mucosa of rats (Rattus norvegicus) submitted to long-term systemic nicotine treatment

Smoking is considered to be the most albeit preventable cause of diseases and premature deaths in the history of mankind. The local action of tobacco on the oral mucosa can cause precancerous and cancerous lesions. However, there is not enough evidence to establish all the systemic effects caused by nicotine on the organism. Thus, the aim of the present study was to characterize the cellular changes of the cheek mucosa of rats submitted to long-term systemic nicotine treatment. Twenty male rats were divided into two experimental groups: a nicotine group and a control group, each consisting of 10 animals. The nicotine group was injected daily with 0.250 mg of nicotine per 100 g of body weight. All animals received a solid diet and water ad libitum. After 90 days of treatment, all animals were weighed and sacrificed. Samples of cheek mucosa were collected for light and transmission electron microscopy. The results revealed oral epithelium containing atypical cells that were characterized by atrophy, cell membrane disorganization and tissue damage. It was concluded that systemic administration of nicotine damaged the cellular integrity of the oral mucosa, impairing tissue function and predisposing the tissue to the action of different pathogenic agents and also to that of other carcinogenic substances present in tobacco. (c) 2006 Elsevier Ltd. All rights reserved.

Morphological aspects of spermatogenic cells at different stages of sexual maturation in black isogenic C57BL/6/Uni mice

In order to study the morphological changes that occur in cells of the testes of isogenic black mouse C57BL/6/Uni into three periods during spermatogenetic used 15 mice divided into 3 groups of 5 animals with 40,50 and 60 days of age. The mice were sacrificed and weighed. Testicles were weighed and measured, and histologically processed and stained with HE, PAS and Masson Massom-H and evaluated under light microscopy. It was observed that group I with 40 days of age in the seminifcrous tubules had a lumen with sparse small amount of interstitial tubular cells. In the seminiferous epithelium type A spermatogonia, intermediate and B were identified, which occupied the compartment adbasal and intermingled with these cells in spermatocytes I in Pachytene and leptotene was observed, whereas in the adluminal compartment Golgi phase spermatids we observed the presence of acrosomal granule. In group II, the cells of the seminiferous epithelium were developed and it was observed in round spermatids cephalic hood phase plus many elongated spermatids in acrosome phase and Sertoli cells. In Group III, 60 days old, it was found that seminiferous epithelium which was of the tubules had elongated spermatids in acrosome phase and maturation, with elongated nuclei and acrosomal system typical of spermiation in the presence of sperm and residual bodies near the tubular lumen. Therefore morphological evolution of germ cell testicular spermatids can be checked and recognized in its four phases: Golgi, cap, acrosome and maturation over the age of the animal.

Morphometric and morphological analysis of neuromuscular junction alterations in the denervated rat diaphragm

The morphological and structural alterations that occur in the neuromuscular junctions of the denervated rat diaphragm were studied. Fifteen adult male albino rats (Rattus norvegicus) aged about 60 days and with a mean weight of 200 g were used. Chronically denervated diaphragms were obtained and the animals were sacrificed after 4, 8 and 12 weeks of denervation. The left antimere of the diaphragm was denervated by section of the phrenic nerve and the right antimere was used as control. Each antimere was divided into three fragments: one was used for histochemical (nonspecific esterase) and morphometric study of neuromuscular junctions, and the other two were used for transmission and scanning electron microscopy (SEM) analysis. Histochemical analysis of the diaphragm neuromuscular junctions after denervation showed only small changes in junction morphology. However, these junctions became smaller and elongated and presented less visible contours with increasing time of denervation. Ultrastructural analysis of neuromuscular junctions after 12 weeks showed more or less organized junctional folds on the muscle fiber surface. The junctional cytoplasm exhibited important alterations such as mitochondrial degeneration and the presence of numerous filaments. SEM revealed the presence of deep primary synaptic grooves with peripheral excavations which housed the nerve terminal boutons and exhibited internally the secondary synaptic clefts present among the junctional folds of the sarcolemma. This study showed that some of the morphological changes demonstrated in other denervated striated skeletal muscles are not repeated at the same intensity or in the same temporal pattern in the rat diaphragm.

Morphological and morphometric study of the opossum's dorsal root ganglia neurons

The ultrastructural characteristics and the morphometric evaluation of the different types of neurons present in the dorsal root ganglia (DRG) of the South American opossum (Didelphis albiventris) were studied. Four adult male animals were used and the neurons from cervical and lumbar DRG were removed and processed for histological and transmission electron microscopy observations. The morphometric data were obtained from serial sections stained by H/E and Masson's trichrome. The number of neurons in cervical and lumbar DRG was 22 300 and 31 000, respectively. About 68% of the cervical neurons and 62.5% of the lumbar neurons presented areas up to 1300 mu m(2) and were considered as the small neurons of the DRG. The ultrastructural observations revealed two morphological types of neurons: clear large neurons and dark small neurons. The nuclei of both cell types are spherical and the chromatin is disperse and rarefected. The cytoplasm of the dark small neuron is more electron dense and shows a regular distribution of small mitochondria and many rough reticulum cisterns in the periphery. A small Golgi apparatus was close to the nucleus and many disperse neurofilaments occupy most parts of the cytoplasm. Smooth reticulum cisterns are rare and lipofucsin-like inclusions are present at some points. In the clear large neurons, the organelles are homogenously scattered through the cytoplasm. The neurofilaments are close packed forming bundles and small mitochondria and rough reticulum cisterns are disperse. Lipofucsin-like inclusions are more frequent in these cells.

Myocardial dysfunction induced by food restriction is related to morphological damage in normotensive middle-aged rats

Previous works from our laboratory have revealed that food restriction (FR) promotes discrete myocardial dysfunction in young rats. We examined the effects of FR on cardiac function, in vivo and in vitro, and ultrastructural changes in the heart of middle-aged rats. Twelve-month-old Wistar- Kyoto rats were fed a control (C) or restricted diet (daily intake reduced to 50% of the control group) for 90 days. Cardiac performance was studied by echocardiogram and in isolated left ventricular (LV) papillary muscle by isometric contraction in basal condition, after calcium chloride (5.2 mM) and beta- adrenergic stimulation with isoproterenol (10(-6) M). FR did not change left ventricular function, but increased time to peak tension, and decreased maximum rate of papillary muscle tension development. Inotropic maneuvers promoted similar effects in both groups. Ultrastructural alterations were seen in most FR rat muscle fibers and included, absence and/or disorganization of myofilaments and Z line, hyper-contracted myofibrils, polymorphic and swollen mitochondria with disorganized cristae, and a great quantity of collagen fibrils. In conclusion, cardiac muscle sensitivity to isoproterenol and elevation of extracellular calcium concentration is preserved in middle-aged FR rats. The intrinsic muscle performance depression might be related to morphological damage.

The evolution of jumping performance in anurans: morphological correlates and ecological implications

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Morphological and physiological specialization for digging in amphisbaenians, an ancient lineage of fossorial vertebrates

Amphisbaenians are legless reptiles that differ significantly from other vertebrate lineages. Most species dig underground galleries of similar diameter to that of the animal. We studied the muscle physiology and morphological attributes of digging effort in the Brazilian amphisbaenid Leposternon microcephalum (Squamata; Amphisbaenia), which burrows by compressing soil against the upper wall of the tunnel by means of upward strokes of the head. The individuals tested (<72 g) exerted forces on the soil of up to 24 N. These forces were possible because the fibres of the longissimus dorsi, the main muscle associated with burrowing, are highly pennated, thus increasing effective muscle cross-sectional area. The muscle is characterized by a metabolic transition along its length: proximal, medial and distal fibres are fast contracting and moderately oxidative, but fibres closer to the head are richer in citrate synthase and more aerobic in nature. Distal fibres, then, might be active mainly at the final step of the compression stroke, which requires more power. For animals greater than a given diameter, the work required to compress soil increases exponentially with body diameter. Leposternon microcephalum, and probably some other highly specialized amphisbaenids, are most likely constrained to small diameters and can increase muscle mass and effective muscle cross-sectional area by increasing body length, not body diameter.

Follicular diameter range based on morphological features in Synbranchus marmoratus (Bloch, 1795) (Teleostei, Synbranchiformes, Synbranchidae) from the South-central region of Brazil

The morphological characteristics of Synbranchus marmoratus female germ cells in various development stages were described in details; then measurements of ovarian follicle diameters were taken from primary and secondary growth as during these development stages the oocyte size varied considerably along the fish growth. The results were correlated to total fish length, using the individuals division in six size classes. It was possible to group oocytes by stages according to histological characteristics but not according to morphometric diameter, as there was a wide variation in diameter in each stage and overlap between different maturation stages. These data make available new information on the reproductive biology of Synbranchidae. (c) 2004 Published by Elsevier Ltd.

Ischaemia and reperfusion effects on skeletal muscle tissue: morphological and histochemical studies

This was a study on the oxidative stress due to ischaemia (I) and reperfusion (R) in skeletal muscle tissue. Using a tourniquet, groups of rats were submitted to ischaemia for 4 h, followed by different reperfusion periods. The animals were divided in four groups: control; 4 h of ischaemia (IR); 4 h of ischaemia plus 1 h reperfusion (IR-1 h); 4 h of ischaemia plus 24 h reperfusion (IR-24 h); and 4 h of ischaemia plus 72 h reperfusion (IR-72 h). At the end of the procedures, samples of soleus muscle were collected and frozen in n-hexane at -70 degrees C. Cryostat sections were submitted to haematoxylin-eosin, succinate dehydrogenase (SDH) and nicotinamide adenine dinucleotide-tetrazolium reductase (NADH-TR) stains. An additional muscle sample was processed for electron microscopy. No alterations were found in control animals. IR group showed fibres had normal aspect besides some round, acidophilic and hypertrophic fibres. There were several fibres with angular outlines and smaller diameters in this group compared with control group. NADH-TR/SDH reaction was moderately intense in most fibres. In some fibres, cytoplasm showed areas without activity and other fibres had very intense reactivity. IR-1 h group showed oedema hypercontracted fibres with disorganized myofibrils, mitochondria with focal lesions and dilated sarcoplasmic reticulum. NADH-TR/SDH reaction was moderate to weak. IR-24 h showed intense inflammatory infiltrate in the endomysium and perimysium. NADH-TR/SDH reaction was similar to IR-1 h. IR-72 h showed necrotic fibres, areas with inflammatory infiltrate, reduced muscle fibres at different stages of necrosis and phagocytosis, and many small round and basophilic fibres characterizing a regeneration process. NADH-TR/SDH reaction was weak to negative. Our results suggest that ischaemia and the subsequent 1-, 24- and 72-h reperfusions induced progressive histological damage. Although progressive, it may be reversible because there were ultrastructural signs of recovery after 72-h reperfusion. This recovery could in part be due to the low oxidative stress identified by the morphological and histochemical analysis.