A. Di Giancamillo

Influence of dietary conjugated linoleic acids and vitamin E on meat quality, and adipose tissue in rabbits

Eighty New Zealand White rabbits, age 55 days, half male and half female, were randomly assigned to one of four diets in a 2×2×2 factorial arrangement. The diets contained 0% or 0.5% CLA, and 60 or 240mg/kg α-Tocopheryl acetate. Forty-eight rabbits were slaughtered at age 92 days. Growth performances and carcass weight were higher (P<0.05) in 240mg/kg of α-tocopheryl acetate-fed rabbits. Fat and CLA isomers content of Longissimus Lumborum (LL) muscle was higher (P<0.05) in CLA-fed rabbits than control. Fatty acid composition of LL muscle was modified (P<0.05) and oxidative stability was increased (P<0.001) by both dietary treatments. CLA increased (P<0.05) triglyceride, total cholesterol and glucose levels in plasma. Adipocytes in interscapular and perirenal fat in the 240mg/kg α-tocopheryl acetate and 0.5% CLA groups were larger in size but lesser in number than in 60mg/kg α-tocopheryl acetate and no CLA rabbit (P<0.01).

The chemical code of porcine enteric neurons and the number of enteric glial cells are altered by dietary probiotics

Background  The enteric nervous system (ENS) contains chemically coded populations of neurons that serve specific functions for the control of the gastrointestinal tract. The ability of neurons to modify their chemical code in response to luminal changes has recently been discovered. It is possible that enteric neuronal plasticity may sustain the adaptability of the gut to changes in intestinal activity or injury, and that gut neurons may respond to an altered intestinal environment by changing their neuropeptide expression.

Can nutraceuticals affect the structure of intestinal mucosa? Qualitative and quantitative microanatomy in L-glutamine diet-supplemented weaning piglets

Weaning piglets were fed an L -glutamine-supplemented diet with the aim of monitoring the effects on gut mucosal turnover and barrier function, to elucidate the potential preventive or therapeutic roles of glutamine as a nutraceutical or ‘functional food’. Sixteen female weaning piglets were divided into two groups, which were fed a control diet (Ctr group: n = 8) or a Ctr + 0.5% L -glutamine diet (G group: n = 8) for 28 days. In the ileum of group G piglets the villus height (V) and crypt depth (C) were increased, and the V:C ratio was decreased (p < 0.01). The PCNA and TUNEL immunoreactivities were also tested. The number of mitotic mucosal cells (M) was increased, and that of mucosal cells with apoptotic nuclei (A) decreased in such a way that the A:M index diminished (p < 0.01). The A:M index also decreased at the level of some components of the gut-associated lymphatic tissue (GALT), thus indicating a positive effect on the gut barrier function. This trial showed that L -glutamine supplementation influenced some morphofunctional characteristics of piglet ileal mucosa. These data corroborate the nutraceutical role of glutamine as a trophic agent for mucosal repair, improvement of barrier function and gut adaptation in the swine per se and as an animal model.

Endometrial cytology and computerized morphometric analysis of epithelial nuclei: A useful tool for reproductive diagnosis in the bitch

New diagnostic approaches are required to recognize early canine hypofertility or infertility. We suggest that the identification of different cytologic types, cellular aspects, and nuclear features of the endometrial epithelial cells may be suitable for this purpose. This study was performed on the bitch (Canis familiaris) during the physiologic reproductive cycle and in uterine diseases. We also applied computerized cytomorphometry to evaluate nuclear area, perimeter, diameter, density, aspect, and roundness of endometrial epithelial cells in healthy dogs (N=35) at different stages of the reproductive cycle (before puberty, during proestrus, estrus, diestrus, and anestrus) and in bitches affected by uterine disorders (N=10). The stage of the estrous cycle was determined by vaginal cytology and progesterone evaluation and also confirmed by clinical and histologic observations. Samples for endometrial cytology were collected in vivo by uterine flushing with transcervical uterine cannulation. After uterine sampling, each dog underwent OHE or uterine stump revision. Cytologic analyses were compared with histologic examinations to verify the uterine condition. The uterine cellular population was represented by endometrial epithelial cells, erythrocytes, neutrophils, lymphocytes, eosinophils, macrophages, plasma cells, and cervical or incidental vaginal cells. Bacteria and amorphous material were observed. The proportion of different cells and nuclear features in the cytologic samples varied throughout the stages of the reproductive cycle and between normal and pathologic uterine conditions. The computer-assisted nuclear morphometry, performed in cytologic specimens by means of the six nuclear parameters chosen to evaluate the endometrial epithelial cell population, proved to be useful for determining the stage of the reproductive cycle. Furthermore, this system was demonstrated to be a valid support to diagnose and distinguish uterine disorders.

Role of autologous rabbit adipose‐derived stem cells in the early phases of the repairing process of critical bone defects

Adipose‐derived stem cells (ASCs) may represent a novel and efficient tool to promote bone regeneration. In this study, rabbit ASCs were expanded in culture and used for the regeneration of full‐thickness bone defects in the proximal epiphysis of tibia of 12 New Zealand rabbits. Defects were implanted with graft material as follows: untreated (control), empty hydroxyapatite (HA) disk, ASCs alone, and HA disk seeded with ASCs. Each isolated ASCs population was tested in vitro: they all showed a high proliferation rate, a marked clonogenic ability, and osteogenic differentiation potential. Eight weeks after implantation, macroscopic analyses of all the samples showed satisfactory filling of the lesions without any significant differences in term of stiffness between groups treated with or without cells (p > 0.05). In both the scaffold‐treated groups, a good osteointegration was radiographically observed. Even if HA was not completely reabsorbed, ASCs‐loaded HA displayed a higher scaffold resorption than the unloaded ones. Histological analyses showed that the osteogenic abilities of the scaffold‐treated defects was greater than those of scaffold‐free samples, and in particular new formed bone was more mature and more similar to native bone in presence of ASCs. These results demonstrated that autologous ASCs–HA constructs is a potential treatment for the regeneration of bone defects.

A tissue engineered osteochondral plug : an in vitro morphological evaluation

Articular cartilage lesions have a poor intrinsic healing potential. The repair tissue is often fibrous, having insufficient biomechanical properties, which could frequently lead to the development of early osteoarthritis. In the last decade, tissue engineering approaches addressed this topic in order to restore joint function with a differentiated and functional tissue. Many biomaterials and techniques have been proposed and some of them applied in clinical practice, even though several concerns have been raised on the quality of the engineered tissue and on its integration in the host joint. In this study, we focused on engineering in vitro a biphasic composite made of cellular fibrin glue and a calcium–phosphate scaffold. Biphasic composites are the latest products of tissue engineering applied to articular cartilage and they seem to allow a more efficient integration of the engineered tissue with the host. However, a firm in vitro bonding between the two components of the composite is a necessary condition to validate this model. Our study demonstrated a gross and microscopic integration of the two components and a cartilage-like quality of the newly formed matrix. Moreover, we noticed an improvement of this integration and GAGs production during the in vitro culture.

Infrared tympanic thermography as a substitute for a probe in the evaluation of ear temperature for post-mortem interval determination: A pilot study

Reported methods which have been used to measure tympanic temperatures on cadavers up to now are quite invasive. They involve the use of a probe which can perforate the tympanic membrane and frequently causes bleeding from the ear. For this reason a non traumatic method for estimating tympanic temperature should be applied. Infrared tympanic thermometry seems to be a plausible option. Reliability of infrared tympanic thermometry (ITT) has been largely assessed on living individuals but only one author up to now has assessed its applicability for post-mortem interval determination. Thus the authors set out to test the difference between ear temperatures taken with a probe vs. ITT, differences between left and right ear and reproducibility of measurements of ITT. The aim of the study was to verify whether ITT could be a plausible option for measuring ear temperature for PMI estimation. Ear temperatures were taken on 25 cadavers (15 males, 10 females). Temperatures were taken alternately by similarly trained personnel by two technical methods (Checktemp 1 thermocouple probe and First Temp Genius infrared thermometer) for a total of 93 measurements. Statistical analysis of the data was performed using SAS statistical software. The range of temperature measured was from 20 to 28 degrees C, statistical analysis revealed no differences within the two technical methods, both for right and left ear (ITT: 22.33+/-0.35 vs. probe: 23.08+/-0.25; P=0.087). The study shows the ITT method can be considered as a possible alternative to the probe for measuring ear temperative and further studies should be considered.

Changes in nitrosative stress biomarkers in swine intestine following dietary intervention with verbascoside.

In farm animals, oxidative stress can be involved in several intestinal pathological disorders, and many antioxidant molecules, especially those of plant origin, can counteract free radicals, thus stabilizing the gut environment and enhancing health. The aim of the study was to investigate whether the use of verbascoside (VB), a polyphenol plant compound, in pig feeding could modulate oxidative and/or nitrosative stress in the gut. Eighteen male piglets (Dalland) were assigned to two groups, which were fed with either a control diet (CON) or a diet supplemented with 5 mg/kg of verbascoside (VB) for 166 days. At slaughter, duodenum and jejunum specimens were collected. Immunohistochemistry and Western blot analyses were performed on the samples to evaluate free radical adducts, including acrolein (ACR), 8-hydroxydeoxyguanosine (8-OHdg) and nitrotyrosine (NT). A KRL test was also used to assess the total blood antioxidant activity, and no difference was observed. Immunohistochemistry and Western blot showed that dietary treatment decreased the levels of nitrotyrosine in enteroendocrine cell populations(P<0.05). Characterization of the enteroendocrine cell typology was then performed, and serotonin-immunoreactive cells were revealed to be directly involved in decreasing the nitrosative stress status. This preliminary study demonstrates the important role of dietary VB in decreasing stress biomarkers in swine gut, thus highlighting a possible intervention aimed at building a large prospective for antioxidant dietary supplementation in food animal species.

The effects of dietary verbascoside on blood and liver oxidative stress status induced by a high n-6 polyunsaturated fatty acids diet in piglets12

Twenty-four weaned female Hypor piglets (10.9 ± 0.1 kg mean BW) were used to evaluate the antioxidant effect of a natural extract, titrated in verbascoside, on blood and liver oxidative status in relation to a high intake of n-6 PUFA, inducing oxidative stress. Piglets were assigned to 1 of 3 experimental groups; the first group was fed a diet with 9% sunflower oil (T1) and the second received the sunflower oil diet supplemented with 5 mg of verbascoside/kg feed from Verbenaceae extract (Lippia spp.; T2). The third group was fed a control diet (CTR), in which an isoenergetic replacement of oil by starch was done. Blood samples were collected at the beginning and the end of the trial (30 d). At the end of the trial, the animals were slaughtered and the liver specimens were collected. Oxidative stress markers, including total antiradical activity, superoxide dismutase (SOD), glutathione peroxidase (GPX), and catalase (CAT) activities, were determined in blood samples. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), and γ-glutamyl transferase (GGT) plasma levels were also evaluated. Immunohistochemistry and western blot analyses were performed in liver to evaluate heat shock protein (Hsp) 70, Hsp90, and Kupffer and Ito cell activation. Liver activities of SOD, GPX, and CAT were also determined. Total antiradical activity in blood and red blood cells were affected (P < 0.01) by dietary treatments. The n-6 PUFA supplementation at a high dosage for 30 d induced oxidative stress, decreasing total antiradical activity in blood and red blood cells (CTR vs. T1 + T2; P < 0.01) and plasma CAT activity (CTR vs. T1 + T2; P = 0.088) and increasing ALT value (CTR vs. T1 + T2; P < 0.01). Also, in liver, the CAT and GPX activities tended to be lower in pigs fed n-6 PUFA diets than pigs fed a control diet (CTR vs. T1 + T2; = 0.090 and = 0.085, respectively). The liver samples presented a normal architecture and no Ito and Kupffer cell activations were observed. In liver, the SOD activity tended to be lower in the T1 group (P = 0.064) than in the CTR and T2 groups. Moreover, the level of Hsp70 was higher (P < 0.01) in the T1 group than the CTR and T2 groups. These data suggest that the dose of dietary verbascoside partially restores the antioxidant status of the liver without affecting the systemic responses to oxidative stress induced by a high-fat diet.

An in vitro tissue-engineered model for osteochondral repair

One of the main topics of regenerative medicine and tissue engineering is to address the problem of lesions involving articular cartilage. In fact, these lesions do not heal spontaneously and often lead to osteoarthritis, which causes chronic pain and worsens quality of life. Moreover, the only available treatment for osteoarthritis is symptomatic therapy and prosthetic replacement, with far from satisfactory results. A more conservative approach that restores the articular surface and function with a biologic tissue is desirable. Several strategies for regenerating articular cartilage have been proposed and applied in clinical practice but a gold standard has not yet been identified. Biphasic composites are the latest products of tissue engineering applied to articular cartilage and they seem to permit a more efficient integration of the engineered neo-tissue with the host. We present an in vitro tissue engineered model for osteochondral repair based on a composite of chondrocytes-fibrin glue gel and a calciumphosphate scaffold. This composite showed a gross integration of the two components and a cartilage-like quality of the newly formed matrix. Further studies are planned to quantify the adherence between the scaffold and the cellular fibrin glue.