Estudi estructural, ultraestructural i histoquímic de les glàndules sexuals accessòries del mascle reproductor porcí (Sus domesticus)

El present treball analitza al microscopi òptic i al microscopi electrònic de transmissió les glàndules sexuals accessòries de Sus domesticus (raça Landrace - varietat anglesa) a partir de mascles reproductors porcins adults i sans. Un millor coneixement dels patrons estructural i ultraestructural normals de las glàndules sexuals accessòries permetrà diagnosticar amb facilitat quina ha estat l'estructura o la funció glandular afectada en mascles en els que s'observa una disminució de la qualitat del semen. Per altra banda, els estudis anatomopatològics s'han de complementar amb tècniques histoquímiques que generalment permeten confirmar o excloure un diagnòstic histopatològic previ. Les glàndules sexuals accessòries del mascle reproductor porcí estan molt desenvolupades i inclouen les glàndules vesiculars, la pròstata i les glàndules bulbouretrals. L'epiteli secretor de les glàndules vesiculars està format per cèl·lules columnars, cèl·lules basals i mastòcits. Les cèl·lules columnars es caracteritzen per presentar tres morfologies diferents que es consideren diferents estadis d'un mateix tipus cel·lular: les cèl·lules principals, les cèl·lules clares i les cèl·lules denses. Les cèl·lules principals secreten activament glicoproteïnes N- i O- glicosilades amb residus d'α-L-fucosa, α(16)fucosa, α-D-mannosa, α-D-glucosa, α- i -D-N-acetilgalactosamina, -D-galactosa-(13)-D-N-acetilgalactosamina, α-D-galactosa, galactosa-(14)-N-acetilglucosamina, D-N-acetilglucosamina i àcid neuramínic. Aquestes glicoproteïnes afavoreixen les interaccions entre l'espermatozoide i l'occit i regulen la permeabilitat de la membrana espermàtica. La pròstata està formada per dues porcions glandulars, el cos de la pròstata (BP) y la pròstata disseminada (DP), entre las quals s'observen diferencies estructurals, ultraestructurals, histoquímiques i funcionals. En ambdues porcions, l'epiteli secretor està constituït per cèl·lules columnars principals, denses i cèl·lules basals, i també per cèl·lules cúbiques en el BP i per cèl·lules mucoses en la DP. En ambdues porcions glandulars, se sintetitzen i secreten N- i O- glicoproteïnes neutres i àcides. Aquestes glicoproteïnes s'alliberen mitjançant un mecanisme regulat en el BP y mitjançant un mecanisme regulat i un constitutiu en la DP. Les glucoproteïnes luminals del BP contenen residus de fucosa, mannosa, α- i -D-N-acetilgalactosamina, galactosa-(14)-N-acetilglucosamina, D-N-acetilglucosamina i àcid neuramínic. En la DP les glicoproteïnes presenten, a més, -D-galactosa-(13)-D-N-acetilgalactosamina i α-D-galactosa. Les glicoproteïnes secretades en el BP i en la DP per via regulada, participen en el control de l'estabilitat del plasmalemma dels espermatozoides, eviten la resposta immune uterina i l'aglutinació dels espermatozoides i afavoreixen la seva motilitat progressiva. Les glicoproteïnes secretades per via constitutiva en la DP protegeixen i lubrifiquen la uretra pelviana. L'epiteli secretor de les glàndules bulbouretrals està format per cèl·lules piramidals principals i denses. Les cèl·lules principals sintetitzen i secreten principalment O-glicoproteïnes àcides carboxilades i sulfatades amb residus glicosídics d'N-acetilgalactosamina, -D-galactosa-(13)-D-N-acetilgalactosamina, α-D-galactosa, D-N-acetilglucosamina i àcid neuramínic. Aquests residus proporcionen resistència a la proteolisi a les O-glicoproteïnes secretades, les quals participen en la lubrificació y protecció de l'epiteli, i intervenen en el control de la permeabilitat del plasmalemma dels espermatozoides i en el transport d'ions al seu través.

Millora de la qualitat espermàtica de dosis seminals de mascles reproductors Piétrain per filtració en diverses reïnes

En aquest treball s'ha dissenyat un mètode ràpid i fiable de tinció amb fluorocroms per a l'anàlisi de la integritat i viabilitat espermàtiques a partir del marcatge de la beina mitocondrial amb MitoTracker®Green FM, de l'acrosoma amb la lectina Trypsin inhibitor from Soybean (SBTI) conjugada amb el fluorocrom Alexa Fluor®488 específic per la proacrosina i del nucli amb els fluorocroms bis-benzimida (específic per a cèl·lules viables) i iodur de propidi (específic per a cèl·lules no viables). També s'ha determinat l'efecte de la filtració de dosis seminals de mascles astentoteratonecrospèrmics en columnes de Sephadex neutre i de dosis de mascles amb baixa qualitat espermàtica per filtració en columnes de Sephadex iònic, llana de vidre i glass beads sobre la qualitat espermàtica dels diferents grups de mascles analitzats. Els resultats obtinguts han mostrat que diversos paràmetres de qualitat espermàtica milloren després de la filtració en les diferents reïnes segons la patologia que presentin.

Estudio de otros compuestos relacionados con la presencia de olor sexual no atribuible al escatol y a la 5@-Androst-16-en-3-ona en grasa dorsal de cerdo

En el presente trabajo de tesis se evalúa la posible contribución de otros compuestos al desarrollo del olor sexual mediante la aplicación de técnicas de Head Space dinámico y cromatografía de filtración de gel. El estudio se efectuó en canales de cerdos enteros previamente clasificadas con concentraciónes bajas de escatol (<0,10 µg /g) y androstenona (0,50 µg/ g) , aunque presentaban , según las respuestas de un panel sensorial, olor sexual. La selección de muestras de grasa clasificadas sin olor sexual permitió la comparación estadística de los resultados obtenidos. El análisis de los compuestos volátiles mostró una mayor abundancia de aldehidos (hexanal, heptanal), ácidos grasos de cadena corta (hexanoico, heptanico nonanoico), 1,4-diclorobenceno y estireno en las muestras de grasa con defecto organoléptico, ue puede, debido a los atributos sensoriales de estas sustancias y sus bajos umbrales de detección, favorecer el desarrollo de aromas desagradables asociados al olor sexual por los miembros de un papel sensorial. El análisis de las fracciones obtenidas por filtración de gel mediante cromatografía de gases acoplada a la espectometría de masas permitió la identificación de la 4-fenil-3-buten-2-ona en las muestras clasificadas con olor sexual y concentraciones bajas de escatol y androstenona, que junto a los resultados obtenidos en el estudio sensorial, indican una influencia de este compuesto en el olor sexual. Por una parte la 4-fenil-3-buten-2-ona presentaría una acción sinérgica con la androstenona al potenciar su detección por parte de los panelistas a concentraciones inferiores de 0,01µg/g. Además, el olor a naftalina que presenta la 4-fenil-3-buten-2-ona puede confundir a los miembros del panel debido a que es uno de los descriptores sensoriales que se asocia con la presencia de escatol en las muestras de grasa evaluadas. Así mismo, la baja transferencia a la fase vapor del escatol y, especialmente, de la androstenona determinada mediante técnicas de Head Space estático, indica que la concentración de cada uno de los compuestos en la fase vapor no se corresponde con la concentración real en las muestras de grasa . Según estos resultados, las respuestas del panel test pueden estar influenciadas por la poca volatilidad de los compuestos implicados en el olor sexual y por la presencia en la fase vapor de otros compuestos aromáticos de mayor volatilidad. La validación de métodos de análisis para la determinación de indol-escatol y androstenona-androstenoles en grasa dorsal porcina mediante cromatografía líquida en fase normal y cromatografía de gases acoplada a la espectrometría de masas, respectivamente, fue otro de los objetivos planteados del presente estudio. La simplificación del tratamiento de las muestras de grasa fue uno de los aspectos analíticos que se tuvo en mayor consideración con la finalidad de ofrecer una mayor capacidad de muestras en un intervalo de tiempo corto.

Cultiu de les cèl·lules epididimàries de "Sus domesticus": anàlisi estructural, funcional i proteòmic

En aquest treball s'han desenvolupat dos mètodes simples i ràpids pel cultiu de les cèl·lules epitelials de les tres regions de l'epidídim de Sus domesticus. Un es basa en el cultiu de fragments del túbul epididimari intactes durant 8 dies. L'altre mètode es basa en el cultiu de fragments del túbul epididimari digerits amb col·lagenasa que, després de 7 dies, donen lloc a la formació d'una monocapa de cèl·lules epitelials epididimàries que adquireixen el 90-100% de confluència després de 12-16 dies en cultiu. Aquestes cèl·lules es mantenen viables durant més de 60 dies en cultiu i no s'observa proliferació de cèl·lules no epitelials. Per determinar el nivell de conservació de les característiques epididimàries en els cultius s'ha analitzat l'estructura cel·lular, l'activitat de síntesi i secreció proteica, i el manteniment i maduració dels espermatozoides en cocultiu.

Desarrollo de métodos rápidos para el análisis de residuos en producción animal

La utilización de algunas sustancias antimicrobianas y algunos corticosteroides como promotores del crecimiento es una práctica ilegal en la UE. Una nueva aproximación para la detección y el control del suministro de estos compuestos puede ser el análisis del pelo. Esta matriz presenta ciertas ventajas frente a otras matrices de análisis, pero los métodos analíticos así como los principales mecanismos por los cuales estas sustancia se acumulan, no están del todo claros ni bien definidos. En este trabajo se desarrollan protocolos de análisis rápidos y específicos para detectar residuos de sulfametacina (SMZ), enrofloxacino (ENR) y dexametasona (DEX) en pelo, músculo e hígado de animales de producción (vacuno y porcino). Asimismo, se confirma la deposición de estos compuestos en el pelo de animales tratados y se evalúa el pelo como matriz analítica y de control de la administración de estos compuestos en producción animal.

Estudi citològic i bioquímic del fluid epididimari de "Sus domesticus"

En aquest estudi s'ha determinat que al augmentar el ritme d'extraccions de semen es produeixen canvis en el patró d'absorció i secreció del fluid epididimari, que provoquen alteracions en la maduració epididimaria dels espermatozoides i un desenvolupament anòmal de la motilitat espermàtica. La concentració de glutamat i carnitina al fluid epididimari augmenten al llarg del conducte epididimari, alhora que la concentració de myo-inositol disminueix. El contingut de myo-inositol a l'interior dels espermatozoides disminueix, mentre que el contingut de glutamat augmenta a partir del caput distal i el contingut de carnitina no varia al llarg del conducte. S'ha determinat la presència de la ruta del poliol a l'epidídim de porcí. Els resultats obtinguts indiquen que la glucosa difon de la sang cap al fluid epididimari, és convertida a sorbitol per l'aldosa reductasa, i aquest sorbitol s'acumula al fluid luminal i és convertit a fructosa per l'acció de la sorbitol deshidrogenasa.

Changes in ruminal fermentation and mineral serum level in animals kept in high temperature environments

In order to evaluate the effect of environmental temperature on ruminal fermentation and on mineral levels of growing ruminants, it was used 12 male calves (initial average weight 82.9 ± 7.7 kg, 100 days of age), were employed in a randomized block design (by weight) experiment, with repeated weight measurement and two environmental temperatures: thermoneutral (24ºC) and heat-stressed (33ºC), during 38 days. The animals exposed to 33ºC presented lower dry matter ingestion, lower T3 (triiodothyronine) serum level, higher ammoniacal nitrogen (NH3-N) level in the rumen liquid, and higher rectal and body temperatures during all the experimental period when compared to the animals kept in thermoneutral environment (24ºC). The animals kept under heat stress environment (33ºC) presented higher calcium serum level, which was the highest on 31st day and the lowest on the 38th day of the experiment; phosphorus level was the lowest during all the experimental period; sodium level was lower on the 17th, 31st and 38th experimental days. Potassium and zinc levels were lower after 24 days; copper level was lower until the 24th day; magnesium level was higher until the 17th day, if compared to the ones from the animals kept in thermoneutral environment (24ºC). The heat-stressed animals presented higher levels of ammoniacal nitrogen in the ruminal liquid and a decrease in the phosphorus, sodium, potassium and zinc serum levels. These results show the necessity of changes on feed management to ruminants in temperatures over the thermal comfort limits so that performance loss is decreased.

Occurrence and molecular characterization of Cryptosporidium spp. isolated from domestic animals in a rural area surrounding Atlantic dry forest fragments in Teodoro Sampaio municipality, State of São Paulo, Brazil

The aim of this study was to assess the occurrence of Cryptosporidium in domestic animals in rural properties surrounding rain forest fragments within the municipality of Teodoro Sampaio, southeastern Brazil. Conventional sucrose flotation method followed by molecular characterization of the parasites by sequencing PCR products amplified from SSU rRNA gene were used. Stool samples were collected from domestic animals raised as pets and livestock in all rural properties surrounding three forest fragments. Samples from cattle (197), equine (63), pigs (25), sheep (11), and dogs (28) were collected from 98 rural properties. The frequency of occurrence of Cryptosporidium within each animal species was 3.0% (6/197) among cattle and 10.7% (3/28) among dogs. Cryptosporidium was not detected in stool samples from equine, sheep, and pigs. All sequences obtained from the six samples of calves showed molecular identity with Cryptosporidium andersoni while all sequences from dog samples were similar to C. canis. The frequency of occurrence of Cryptosporidium in these domestic animal species was low. The absence of C. parvum in the present study suggests that the zoonotic cycle of cryptosporidiosis may not be relevant in the region studied. The presence of Cryptosporidium species seldom described in humans may be, otherwise, important for the wild fauna as these animals are a source of infection and dissemination of this protozoan to other animal species. The impact and magnitude of infection by C. andersoni in wild ruminants and C. canis in wild canids have to be assessed in future studies to better understand the actual importance of these species in this region.

Accelerating aging chemotherapy on aged animals: Protective effect from nutraceutical modulation

The aim of this study was to test a novel phytocompound in an experimental model of antitumor-induced immunosuppression. Five groups of mice were considered: young (Y) and aged (A) that were given intraperitoneally 10 doses of cyclophosphamide (CPX, 25mg/kg/bw) or CPX plus (150 mg/kg/bw) of the nutraceutical DTS (Denshichi-Tochiu-Sen), and control. After sacrifice, macrophage chemotaxis and serum levels of IFN-gamma, IL-2, and GM-CSF were determined. Liver and urinary bladder were examined histologically, as were the liver and kidney for redox enzymes. CPX significantly decreased macrophage chemotaxis and all cytokines (p < 0.05, A >> Y). DTS restored macrophage function and cytokine concentration (p < 0.001) and partly improved the necro-inflammatory score and substance P receptor expression in the bladder and the redox status in liver and kidney (p < 0.05). Such data suggest that DTS effectively prevents CPX-induced immune suppression and oxidative-inflammatory damage, which are particularly enhanced in aged organisms.

The diversity of the Digenea of Australian animals

The Digenea is one of five major helminth assemblages represented in Australian animals. History of the study of digeneans in Australia is reviewed briefly to show that it has never been subjected to the kind of sustained study needed to reach an understanding of it. The Australian vertebrate fauna comprises over 5500 species. These have so far been shown to harbour just over 70 families, about 306 genera and 566 species of digeneans. Digeneans occur in all classes of vertebrates in Australia but are distributed very unevenly; aquatic hosts are generally most heavily infected, but many terrestrial species are also infected. Particular weaknesses in knowledge of the fauna concern the bats, cetaceans and teleosts. Another weakness is in knowledge of life-cycles; representative life-cycles are known for only about 20 of the 70 families known in Australia. Estimates of the overall size of the fauna are dependent on an understanding of sampling strategies, the heterogeneity of distribution of the fauna, and the nature of host-specificity. These subjects are reviewed briefly and an estimate of the total fauna is made. There may be as many as 6000 species of digeneans in Australia. (C) 1998 Australian Society for Parasitology. Published by Elsevier Science Ltd.

Snakes and kittens in the grass: No evidence for preferential processing of fear-relevant animals in visual search

Ohman and colleagues provided evidence for preferential processing of pictures depicting fear-relevant animals by showing that pictures of snakes and spiders are found faster among pictures of fiowers and mushrooms than vice versa and that the speed of detecting fear-relevant animals was not affected by set size whereas the speed of detecting fiowers/mushrooms was. Experiment 1 replicated this finding. Experiment 2, however, found similar search advantages when pictures of cats and horses or of wolves and big cats were to be found among pictures of flowers and mushrooms. Moreover, Experiment 3, in a within subject comparison, failed to find faster identification of snakes and spiders than of cats and horses among flowers and mushrooms. The present findings seem to indicate that previous reports of preferential processing of pictures of snakes and spiders in a visual search task may reflect a processing advantage for animal pictures in general rather than fear-relevance.

Molecular detection of Paracoccidioides brasiliensis in road-killed wild animals

Paracoccidioides brasiliensis infections have been little studied in wild and/or domestic animals, which may represent an important indicator of the presence of the pathogen in nature. Road-killed wild animals have been used for surveillance of vectors of zoonotic pathogens and may offer new opportunities for eco-epidemiological studies of paracoccidiodomycosis (PCM). The presence of P. brasiliensis infection was evaluated by Nested-PCR in tissue samples collected from 19 road-killed animals; 3 Cavia aperea (guinea pig), 5 Cerdocyon thous (crab-eating-fox), 1 Dasypus novemcinctus (nine-banded armadillo), 1 Dasypus septemcinctus (seven-banded armadillo), 2 Didelphis albiventris (white-eared opossum), 1 Eira barbara (tayra), 2 Gallictis vittata (grison), 2 Procyon cancrivorus (raccoon) and 2 Sphiggurus spinosus (porcupine). Specific P. brasiliensis amplicons were detected in (a) several organs of the two armadillos and one guinea pig, (b) the lung and liver of the porcupine, and (c) the lungs of raccoons and grisons. P. brasiliensis infection in wild animals from endemic areas might be more common than initially postulated. Molecular techniques can be used for detecting new hosts and mapping `hot spot` areas of PCM.

Repeated stress reduces mucociliary clearance in animals with chronic allergic airway inflammation

We evaluated if repeated stress modulates mucociliary clearance and inflammatory responses in airways of guinea pigs (GP) with chronic inflammation. The GP received seven exposures of ovalbumin or saline 0.9%. After 4th inhalation, animals were submitted to repeated forced swim stressor protocol (5x/week/2 weeks). After 7th inhalation, GP were anesthetized. We measured transepithelial potential difference, ciliary beat frequency, mucociliary transport, contact angle, cough transportability and serum cortisol levels. Lungs and adrenals were removed, weighed and analyzed by morphometry. Ovalbumin-exposed animals submitted to repeated stress had a reduction in mucociliary transport, and an increase on serum cortisol, adrenals weight, mucus wettability and adhesivity, positive acid mucus area and IL-4 positive cells in airway compared to non-stressed ovalbumin-exposed animals (p < 0.05). There were no effects on eosinophilic recruitment and IL-13 positive cells. Repeated stress reduces mucociliary clearance due to mucus theological-property alterations, increasing acid mucus and its wettability and adhesivity. These effects seem to be associated with IL-4 activation. (C) 2010 Elsevier B.V. All rights reserved.

Effects of the administration of pentoxifylline and prednisolone on the evolution of portal fibrogenesis secondary to biliary obstruction-an experimental study in growing animals

Background: Many chronic liver diseases lead to progressive hepatic fibrosis, a condition that can ultimately result in loss of organ function and severe portal hypertension necessitating hepatic transplantation. Within the last few decades, studies have been conducted to demonstrate the possibility of drug modulation of hepatic fibrogenesis. Regarding biliary obstruction, it has been suggested that administration of corticosteroids could promote better late outcomes for children with biliary atresia submitted to Kasai`s portoenterostomy. Models used to test potential antifibrogenic drugs such as pentoxifylline (PTX) have not included growing animals. Methods: In this experimental study, 119 young rats (21st or 22nd days) were submitted to laparotomy and common bile duct ligation (CBDL) or to sham surgery (SHAM). Animals were allocated into 5 groups, according to surgical procedure, and administered the following solutions: (1) CBDL + distilled water, (2) SHAM + distilled water, (3) CBDL + PTX, (4) CBDL + prednisolone (PRED), and (5) CBDL + PTX + PRED (PTX + PRED). Each group was further divided into 2 subgroups according to the length of the experiment (15 or 30 days). At the end of the defined period, animals were weighed, and a hepatic fragment was collected from each one for analyses. Results: The PTX animals exhibited increased weight gain compared to animals in the PRED or PTX + PRED groups. Animals from the 3 therapeutic groups (PTX, PRED, and PTX + PRED) showed diminished collagen-filled area in portal spaces. Total portal space area was increased in the PTX group. Conclusions: Hepatic fibrosis induced by bile duct ligation in young rats could be modulated by pharmacologic interventions. Administration of PTX or PRED, or the combination of both, resulted in diminished collagen-filled areas in portal spaces. (C) 2009 Elsevier Inc. All rights reserved.

A new DTL-electrode holder for recording of electroretinograms in animals

Purpose: Contact lens electrodes (CLEs) are frequently used to register electroretinograms (ERGs) in small animals such as mice or rats. CLEs are expensive to buy or difficult to be produced individually. In addition, CLE`s have been noticed to elicit inconstant results and they carry potential to injure the cornea. Therefore, a new electrode holder was constructed based on the clinically used DTL-electrode and compared to CLEs. Material and methods: ERGs were recorded with both electrode types in nine healthy Brown-Norway rats under scotopic conditions. For low intensity responses a Naka-Rushton function was fitted and the parameters V(max), k and n were analyzed. The a-wave, b-wave and oscillatory potentials were analyzed for brighter flash intensities (1-60 scot cd s/m(2)). Repeatability was assessed for both electrode types in consecutive measurements. Results: The new electrode holder was faster in setting up than the CLE and showed lower standard deviations. No corneal alterations were observed. Slightly higher amplitudes were recorded in most of the measurements with the new electrode holder (except amplitudes induced by 60 cd s/m(2)). A Bland-Altman test showed good agreement between the DTL holder and the CLE (mean difference 35.2 mu V (Holder-CLE)). Pearson`s correlation coefficient for test-retest-reliability was r = 0.783. Conclusions: The DTL holder was superior in handling and caused far less corneal problems than the CLE and produced comparable or better electrophysiological results. The minimal production costs and the possibility of adapting the DTL holder to bigger eyes, such as for dogs or rabbits, offers with broader application prospects. (C) 2010 Elsevier B.V. All rights reserved.