Antonio Augusto Coppi Maciel Ribeiro

Telomerase activity coevolves with body mass not lifespan.

In multicellular organisms, telomerase is required to maintain telomere length in the germline but is dispensable in the soma. Mice, for example, express telomerase in somatic and germline tissues, while humans express telomerase almost exclusively in the germline. As a result, when telomeres of human somatic cells reach a critical length the cells enter irreversible growth arrest called replicative senescence. Replicative senescence is believed to be an anticancer mechanism that limits cell proliferation. The difference between mice and humans led to the hypothesis that repression of telomerase in somatic cells has evolved as a tumor‐suppressor adaptation in large, long‐lived organisms. We tested whether regulation of telomerase activity coevolves with lifespan and body mass using comparative analysis of 15 rodent species with highly diverse lifespans and body masses. Here we show that telomerase activity does not coevolve with lifespan but instead coevolves with body mass: larger rodents repress telomerase activity in somatic cells. These results suggest that large body mass presents a greater risk of cancer than long lifespan, and large animals evolve repression of telomerase activity to mitigate that risk.

Particulate Urban Air Pollution Affects the Functional Morphology of Mouse Placenta

Abstract In humans, adverse pregnancy outcomes (low birth weight, prematurity, and intrauterine growth retardation) are associated with exposure to urban air pollution. Experimental data have also shown that such exposure elicits adverse reproductive outcomes. We hypothesized that the effects of urban air pollution on pregnancy outcomes could be related to changes in functional morphology of the placenta. To test this, future dams were exposed during pregestational and gestational periods to filtered or nonfiltered air in exposure chambers. Placentas were collected from near-term pregnancies and prepared for microscopical examination. Fields of view on vertical uniform random tissue slices were analyzed using stereological methods. Volumes of placental compartments were estimated, and the labyrinth was analyzed further in terms of its maternal vascular spaces, fetal capillaries, trophoblast, and exchange surface areas. From these primary data, secondary quantities were derived: vessel calibers (expressed as diameters), trophoblast thickness (arithmetic mean), and total and mass-specific morphometric diffusive conductances for oxygen of the intervascular barrier. Two-way analysis of variance showed that both periods of exposure led to significantly smaller fetal weights. Pregestational exposure to nonfiltered air led to significant increases in fetal capillary surface area and in total and mass-specific conductances. However, the calibers of maternal blood spaces were reduced. Gestational exposure to nonfiltered air was associated with reduced volumes, calibers, and surface areas of maternal blood spaces and with greater fetal capillary surfaces and diffusive conductances. The findings indicate that urban air pollution affects placental functional morphology. Fetal weights are compromised despite attempts to improve diffusive transport across the placenta.

Distinct tumor suppressor mechanisms evolve in rodent species that differ in size and lifespan

Large, long‐lived species experience more lifetime cell divisions and hence a greater risk of spontaneous tumor formation than smaller, short‐lived species. Large, long‐lived species are thus expected to evolve more elaborate tumor suppressor systems. In previous work, we showed that telomerase activity coevolves with body mass, but not lifespan, in rodents: telomerase activity is repressed in the somatic tissues of large rodent species but remains active in small ones. Without telomerase activity, the telomeres of replicating cells become progressively shorter until, at some critical length, cells stop dividing. Our findings therefore suggested that repression of telomerase activity mitigates the increased risk of cancer in larger‐bodied species but not necessarily longer‐lived ones. These findings imply that other tumor suppressor mechanisms must mitigate increased cancer risk in long‐lived species. Here, we examined the proliferation of fibroblasts from 15 rodent species with diverse body sizes and lifespans. We show that, consistent with repressed telomerase activity, fibroblasts from large rodents undergo replicative senescence accompanied by telomere shortening and overexpression of p16Ink4a and p21Cip1/Waf1 cycline‐dependent kinase inhibitors. Interestingly, small rodents with different lifespans show a striking difference: cells from small shorter‐lived species display continuous rapid proliferation, whereas cells from small long‐lived species display continuous slow proliferation. We hypothesize that cells of small long‐lived rodents, lacking replicative senescence, have evolved alternative tumor‐suppressor mechanisms that prevent inappropriate cell division in vivo and slow cell growth in vitro. Thus, large‐bodied species and small but long‐lived species have evolved distinct tumor suppressor mechanisms.

Lymphatic drainage on healthy and neoplasic mammary glands in female dogs: Can it really be altered?

The purpose of this research was to study the mammary lymphatic drainage under a macroscopic and mesoscopic view, comparing the vascular pattern of healthy and neoplasic mammary glands injected with drawing ink alcoholic and fluorescein solutions, in 46 mongrel female dogs. The results pointed out that the thoracic gland is drained by the axillary lymph centre, but in mammary neoplasia either superficial cervical or ventral thoracic lymph centres can be involved. Cranial and caudal abdominal glands may be drained by the axillary, inguinofemoral and popliteal lymph centres. However, the popliteal drainage is specific for the healthy caudal abdominal mammary gland. The inguinal gland can be drained by both inguinofemoral and popliteal lymph centres in both neoplasic and healthy conditions. Regarding the mammary lymphatic communications, this research demonstrated that neoplasic glands present more types of anastomosis (40.9%), than healthy glands (33.33%), and an increase in contralateral anastomosis (50%) compared with healthy ones (33%). Given the data, the mammary neoplasia can change the lymphatic drainage pattern in terms of lymph centres and vascular arborization, thus forming new drainage channels and recruiting a larger number of lymph nodes. Lastly, some comments were made about the severity of a specific neoplasic mammary gland and conditions to be considered before making a decision in terms of the most adequate operative procedure, and suggestions for further investigations.

Low‐intensity treadmill exercise‐related changes in the rat stellate ganglion neurons

Stellate ganglion (SG) represents the main sympathetic input to the heart. This study aimed at investigating physical exercise–related changes in the quantitative aspects of SG neurons in treadmill‐exercised Wistar rats. By applying state‐of‐the‐art design‐based stereology, the SG volume, total number of SG neurons, mean perikaryal volume of SG neurons, and the total volume of neurons in the whole SG have been examined. Arterial pressure and heart rate were also measured at the end of the exercise period. The present study showed that a low‐intensity exercise training program caused a 12% decrease in the heart rate of trained rats. In contrast, there were no effects on systolic pressure, diastolic pressure, or mean arterial pressure. As to quantitative changes related to physical exercise, the main findings were a 21% increase in the fractional volume occupied by neurons in the SG, and an 83% increase in the mean perikaryal volume of SG neurons in treadmill‐trained rats, which shows a remarkable neuron hypertrophy. It seems reasonable to infer that neuron hypertrophy may have been the result of a functional overload imposed on the SG neurons by initial posttraining sympathetic activation. From the novel stereological data we provide, further investigations are needed to shed light on the mechanistic aspect of neuron hypertrophy: what role does neuron hypertrophy play? Could neuron hypertrophy be assigned to the functional overload induced by physical exercise?

Structure and Ultrastructure of the Celiac–Mesenteric Ganglion Complex in the Domestic Dog (Canis familiaris)

A number of neurons of the autonomic nervous system are situated in the ganglia and can be systematically divided into pre‐vertebrals, paravertebrals, intramural and para‐viscerals. The celiac–mesenteric ganglion, an important pre‐vertebral ganglion, is located together with the abdominal aorta and links the central nervous system to the peripheral system, participating in the coordination of peripheral reflexes and principally innervating the stomach, intestines, accessory glands (liver and pancreas). In addition, the celiac–mesenteric ganglion also contributes to the innervation of the spleen and has a role in gastrointestinal motility control. This study examined the structural and ultrastructural aspects of 40 celiac–mesenteric ganglia from domestic dogs. For light microscopy ganglia were included in paraplast and stained with haematoxylin–eosin, picrosirius, toluidine blue, Callejas and Massons trichrome. For examination by electron microscopy, the ganglia were submitted to cryofracture, enzyme digestion, hydrolysis and fixed in 5% glutaraldehyde in 0.1 M phosphate buffer (pH 7.4). The celiac–mesenteric ganglion was observed as a ganglionic complex composed of various ganglionic units separated by types I and III collagen fibres, predominantly unmyelinated nerve fibres and continuous capillaries. This complex is surrounded by a double‐layer capsule (internal and external). The principal ganglion cells had eccentric nuclei with two nucleoli, the nucleolemma was double and presented nuclear pores. In the cytoplasm there were vesicles of the Golgi apparatus, electron‐dense vacuoles, mitochondrias, smooth and granulated endoplasmic reticulum and free ribosomes. In conclusion, this ganglionic complex, in contrast to similar structures in the enteric nervous system, presents separate ganglionic units in a systematic arrangement related to the extrinsic and specific innervation of the target organs.

Gross Anatomic Organization of the Capybara's (Hydrochaeris hydrochaeris) Brachial Plexus

The innervation of the capybara thoracic limb was characterized. The following nerves were observed constituting the right and left brachial plexus: n. dorsalis scapulae (C4 and C5; C4, C5 and C6) which innervates the m. serratus ventralis cervicis and m. rhomboideus; n. suprascapularis (C4, C5 and C6; C5, C6 and C7) supplying the m. supraspinatus and the m. infraspinatus; cranial and caudal nn. subscapulares (C5 and C6; C5, C6 and C7) innervating the m. subscapularis; n. axillaris (C5 and C6; C6, C7 and C8) which supplies the m. triceps brachii (caput mediale); n. radialis (C6, C7, C8 and T1; C6, C7 and C8) which innervates the m. triceps brachii (caput longum and caput mediale) and the m. extensor carpi radialis, m. extensor digitorum communis, m. extensor digitorum lateralis; n. medianus joined to the n. musculocutaneus (C6, C7, C8 and T1; C6, C7 and C8) supplying the m. biceps brachii, m. flexor carpi radialis and m. coracobrachialis; n. ulnaris (C6, C7, C8 and T1; C6, C7 and C8) leading to the m. flexor carpi radialis, the m. flexor carpi ulnaris and the m. flexor digitorum superficialis; n. thoracodorsalis (C6, C7, C8 and T1; C6, C7 and C8) supplying the m. latissimus dorsi; n. thoracicus lateralis (C8, T1; C7, C8, T1) which innervates m. pectoralis profundus (caudal portion); n. thoracicus longus (C6, C7; C7, C8) which is distributed to the m. serratus ventralis thoracis. A communication between the n. radialis and n. ulnaris was observed at the left brachial plexus.

Macro- and Microstructure of the Superior Cervical Ganglion in Dogs, Cats and Horses during Maturation

The superior cervical ganglion (SCG) provides sympathetic input to the head and neck, its relation with mandible, submandibular glands, eyes (second and third order control) and pineal gland being demonstrated in laboratory animals. In addition, the SCG’s role in some neuropathies can be clearly seen in Horner’s syndrome. In spite of several studies published involving rats and mice, there is little morphological descriptive and comparative data of SCG from large mammals. Thus, we investigated the SCG’s macro- and microstructural organization in medium (dogs and cats) and large animals (horses) during a very specific period of the post-natal development, namely maturation (from young to adults). The SCG of dogs, cats and horses were spindle shaped and located deeply into the bifurcation of the common carotid artery, close to the distal vagus ganglion and more related to the internal carotid artery in dogs and horses, and to the occipital artery in cats. As to macromorphometrical data, that is ganglion length, there was a 23.6% increase from young to adult dogs, a 1.8% increase from young to adult cats and finally a 34% increase from young to adult horses. Histologically, the SCG’s microstructure was quite similar between young and adult animals and among the 3 species. The SCG was divided into distinct compartments (ganglion units) by capsular septa of connective tissue. Inside each ganglion unit the most prominent cellular elements were ganglion neurons, glial cells and small intensely fluorescent cells, comprising the ganglion’s morphological triad. Given this morphological arrangement, that is a summation of all ganglion units, SCG from dogs, cats and horses are better characterized as a ganglion complex rather than following the classical ganglion concept. During maturation (from young to adults) there was a 32.7% increase in the SCG’s connective capsule in dogs, a 25.8% increase in cats and a 33.2% increase in horses. There was an age-related increase in the neuronal profile size in the SCG from young to adult animals, that is a 1.6-fold, 1.9-fold and 1.6-fold increase in dogs, cats and horses, respectively. On the other hand, there was an age-related decrease in the nuclear profile size of SCG neurons from young to adult animals (0.9-fold, 0.7-fold and 0.8-fold in dogs, cats and horses, respectively). Ganglion connective capsule is composed of 2 or 3 layers of collagen fibres in juxtaposition and, as observed in light microscopy and independently of the animal’s age, ganglion neurons were organised in ganglionic units containing the same morphological triad seen in light microscopy.

Zinc and glutamine improve brain development in suckling mice subjected to early postnatal malnutrition.

OBJECTIVE The effect of zinc and glutamine on brain development was investigated during the lactation period in Swiss mice. METHODS Malnutrition was induced by clustering the litter size from 6-7 pups/dam (nourished control) to 12-14 pups/dam (undernourished control) following birth. Undernourished groups received daily supplementation with glutamine by subcutaneous injections starting at day 2 and continuing until day 14. Glutamine (100 mM, 40-80 microL) was used for morphological and behavioral studies. Zinc acetate was added in the drinking water (500 mg/L) to the lactating dams. Synaptophysin and myelin basic protein brain expressions were evaluated by immunoblot. Zinc serum and brain levels and hippocampal neurotransmitters were also evaluated. RESULTS Zinc with or without glutamine improved weight gain as compared to untreated, undernourished controls. In addition, zinc supplementation improved cliff avoidance and head position during swim behaviors especially on days 9 and 10. Using design-based stereological methods, we found a significant increase in the volume of CA1 neuronal cells in undernourished control mice, which was not seen in mice receiving zinc or glutamine alone or in combination. Undernourished mice given glutamine showed increased CA1 layer volume as compared with the other groups, consistent with the trend toward increased number of neurons. Brain zinc levels were increased in the nourished and undernourished-glutamine treated mice as compared to the undernourished controls on day 7. Undernourished glutamine-treated mice showed increased hippocampal gamma-aminobutyric acid and synaptophysin levels on day 14. CONCLUSION We conclude that glutamine or zinc protects against malnutrition-induced brain developmental impairments.

New alternative methods to teach surgical techniques for veterinary medicine students despite the absence of living animals. Is that an academic paradox

Due to a raised ethical mentality, veterinary schools are pursuing methods to preserve animal corpses used for surgical technique classes in an attempt to reduce the use of living animals for teaching. Generally speaking, animal and human bodies are usually preserved with 10% aqueous formalin solution especially for descriptive anatomy classes. Other possibilities include the use of glycerol, alcohol and phenol. At present, new fixatives have been developed to allow a better and longer preservation of animal corpses in order to maintain organoleptic characteristics, i.e. colour, texture, as close as possible to what students will deal with living animals. From 2004, in our college, surgical technique classes no longer use living animals for students’ training. Instead, canine corpses chemically preserved with modified Larssen (MLS) and Laskowski (LS) solutions are preferred. The purpose of this study was to investigate comparatively the biological quality of preservation of these two solutions and to evaluate students’ learning and acceptance of this new teaching method. Although these fixatives maintain body flexibility, LS solution failed to keep an ordinary tissue colouration (cadavers were intensely red) and tissue preservation was not adequate. By contrast, MLS solution, however, did not alter the colouration of cadavers which was fairly similar to that normally found in living animals. A remarkable characteristic was a very strong and unpleasant sugary odour in LS‐preserved animals and therefore the MLS solution was the elected method to preserve cadavers for surgical technique classes. The students’ feedback to the use of Larssen‐preserved cadavers was very satisfactory, i.e. 96.6% of students were in favour of the use of cadavers for surgical training and on average 91.8% (2002–2003) of students preferred the MLS solution as the chemical preserver, whereas only 8.2% elected LS solution for teaching purposes. From the students’ point of view (95.1%) the ideal class would be an initial training in MLS cadavers followed by classes with animals admitted to the Veterinary Hospital.