Effects of opioids on proximal renal tubular cells undergoing ATP depletion.

Abstract This study investigated the effect of morphine, fentanyl, butorphanol and buprenorphine on viability and caspase-3 activity in renal proximal tubular cells exposed to opioids for 2 h before or 12 h after chemical anoxia. Cell viability decreased regardless the treatment although intracellular ATP content was elevated in morphine and fentanyl pre-treated cells at 12 h. Anoxia increased caspase activity but this effect was significantly reduced in cells treated before or after with morphine, fentanyl and in cell treated with butorphanol for 12 h. No influence of buprenorphine was detected. Morphine, fentanyl and butorphanol might have protective effects during kidney ischemia

Pain assessment in calves undergoing ring castration.

The rubber ring method for the castration of calves is quick, easy, cheap and reliable, but there are several studies indicating that it causes significant pain during and after the procedure [1]. The aim of this study was to evaluate the impact of the ring castration of 2-month-old calves on potential indicators of pain or stress. These indicators included serum cortisol concentration, leukogram, behavioural posture and activity evaluated using the UNESP-Botucatu unidimensional composite pain scale [2], local temperatures and scrotal clinical evaluation. Moreover we used the in-field measurement of Leukocyte Coping Capacity (LCC) on whole blood as a different method to evaluate suffering condition [3]. Twenty healthy calves (90 \ub1 4 kg of body weight, 2 months of age) were selected for the study and randomly assigned to either sham (n=10) or ring castration (n=10). Calves were handled in a similar manner for analogous amount of time and castrated with rubber rings using an elastrator or not castrated. Behavioural assessment, blood sampling, temperature recording and scrotal evaluation were repeated 1 hour before castration, 30 minutes after ring application and at day 3, 7 and 14 after ring application. The same timepoints were used in the sham group. Chronological changes of cortisol, leukogram, pain scale score, local temperature and LCC were analysed within each group and between groups using the SAS statistical software. P < 0.05 was considered significant. After the ring application, the scrotal sac appeared swollen at day 3, mummificated and partially detached at day 7 and at day 10 it was surgically removed. Pain scale score showed very little pain in castrated animals at any timepoints (mean score range 0,3-0,6 out of 10) and no pain in sham calves (0 out of 10). No significant differences were recorded between sham and castrated animals among timepoints in pain scores, cortisol concentrations and leukograms. LCC decreased significantly in castrated animals at day 7 compared to baseline values, thus indicating an increase in stress level. A significant difference beween groups was observed in LCC values 7 days after ring application (<0.0001). Pain indicators such as serum cortisol, behavioural observation, scrotal temperature and clinical evaluation were unaffected after ring castration. Since LCC presented a significant variation, we considered that this method might represent a more sensitive tool in case of procedures that cause mild pain

Pharmacokinetics and antinociceptive effects of tramadol and its metabolite O-desmethyltramadol following intravenous administration in sheep

Although sheep are widely used as an experimental model for various surgical procedures there is a paucity of data on the pharmacokinetics and efficacy of analgesic drugs in this species. The aims of this study were to investigate the pharmacokinetics of intravenously (IV) administered tramadol and its active metabolite O-desmethyltramadol (M1) and to assess the mechanical antinociceptive effects in sheep. In a prospective, randomized, blinded study, six healthy adult sheep were given 4 and 6\u2009mg/kg tramadol and saline IV in a cross-over design with a 2-week wash-out period. At predetermined time points blood samples were collected and physiological parameters and mechanical nociceptive threshold (MNT) values were recorded. The analytical determination of tramadol and M1 was performed using high performance liquid chromatography. Pharmacokinetic parameters fitted a two- and a non-compartmental model for tramadol and M1, respectively. Normally distributed data were analysed by a repeated mixed linear model. Plasma concentration vs. time profiles of tramadol and M1 were similar after the two doses. Tramadol and M1 plasma levels decreased rapidly in the systemic circulation, with both undetectable after 6\u2009h following drug administration. Physiological parameters did not differ between groups; MNT values were not statistically significant between groups at any time point. It was concluded that although tramadol and M1 concentrations in plasma were above the human minimum analgesic concentration after both treatments, no mechanical antinociceptive effects of tramadol were reported. Further studies are warranted to assess the analgesic efficacy of tramadol in sheep

Analgesia and/or anaesthesia during piglet castration &#8211; part II: practicability of farm protocols, resource efficiency and economic implications

Pain alleviation associated with castration of piglets is a debated welfare issue. This study compares practical aspects, resource efficiency and economic implications of two protocols involving both analgesia and anaesthesia compared to a control group: conventional castration without pain relief (CTRL); joint administration of azaperone and meloxicam (AZA-MEL), i.m.; procaine (PROC-MEL), s.c., and meloxicam, i.m. A total number of 356 male piglets (56 L), was involved. Labour, mortality during the lactation period and costs for procedures were analysed. The total amount of labour required for each single male piglet and the risk of recording at least one dead piglet during lactation in litters were significantly higher in AZA-MEL and PROC-MEL groups than in CTRL group (labour: 02:04 and 02:04 vs. 01:18 min, respectively, p <.001; mortality risk: (RR = 1.48; CI 95% = 1.02 12 2.16; p =.029). The cost estimated for the castration of each male piglet in CRTL group was 0.32 \u20ac, whereas was 3.14 \u20ac for AZA-MEL group and 3.30 \u20ac for PROC-MEL group. The results suggest that adopting analgesia and anaesthesia showed notable cost increases for farmers. This might be expected and justifiable when the management is improved to reach a higher standard quality, such as in the case of welfare-friendly surgical castration, but might be questionable when also considering the result of increased piglet mortality in the lactation period.HIGHLIGHTS To use meloxicam + azaperone or + procaine on farm during piglets castration, increases labour of workers by 76.8 and 56.5%, respectively. At the same time, also piglets mortality risk increases by 48% during lactation. Total costs for each male piglet castrated with meloxicam + azaperone or + procaine increased by 3.14 and 3.30 \u20ac, respectively

Decellularization and Delipidation Protocols of Bovine Bone and Pericardium for Bone Grafting and Guided Bone Regeneration Procedures

The combination of bone grafting materials with guided bone regeneration (GBR) membranes seems to provide promising results to restore bone defects in dental clinical practice. In the first part of this work, a novel protocol for decellularization and delipidation of bovine bone, based on multiple steps of thermal shock, washes with detergent and dehydration with alcohol, is described. This protocol is more effective in removal of cellular materials, and shows superior biocompatibility compared to other three methods tested in this study. Furthermore, histological and morphological analyses confirm the maintenance of an intact bone extracellular matrix (ECM). In vitro and in vivo experiments evidence osteoinductive and osteoconductive properties of the produced scaffold, respectively. In the second part of this study, two methods of bovine pericardium decellularization are compared. The osmotic shock-based protocol gives better results in terms of removal of cell components, biocompatibility, maintenance of native ECM structure, and host tissue reaction, in respect to the freeze/thaw method. Overall, the results of this study demonstrate the characterization of a novel protocol for the decellularization of bovine bone to be used as bone graft, and the acquisition of a method to produce a pericardium membrane suitable for GBR applications

Aortic stenting in the growing sheep causes aortic endothelial dysfunction but not hypertension: Clinical implications for coarctation repair

Stent implantation is the treatment of choice for adolescents and adults with aortic coarctation (CoAo). Despite excellent short-term results, 20%-40% of the patients develop arterial hypertension later in life, which was attributed to inappropriate response of the aortic baroreceptors to increased stiffness of the ascending aorta (ASAO), either congenital or induced by CoAo repair. In particular, it has been hypothesized that stent itself may cause or sustain hypertension. Therefore, we aimed to study the hemodynamic and structural impact following stent implantation in the normal aorta of a growing animal

Physiological pregnancy in a tiger affected by endometritis and benign mesenchymal neoplasm.

In large felids, uncomplicated endometritis and benign mesenchymal uterine tumours usually represent post-mortem findings. These pathologies generally remain clinically asymptomatic, and their impact on reproductive outcome and maternal care is unknown. A 13-year-old female tiger (Panthera tigris) was evaluated by a team of veterinarians from the University of Padova for abnormal behaviour after parturition. The animal spontaneously gave birth to a healthy male cub of 1.460 kg, but after three hours of normal parental care, she showed careless about the cub, occasional abdominal contractions, excessive self-licking and restlessness. After 10 hours of continuous observation, the tiger was anaesthetised for a complete clinical evaluation. The physical examination was unremarkable, but a vaginal bloody/mucous discharge with no obstruction of the birth canal was noted. Abdominal ultrasound revealed an enlarged left uterine horn with heterogeneous echogenicity. Blood analysis showed moderate neutrophilia and leucocytosis. Differential diagnoses included placental retention, uterine inflammation/infection, neoplasia. The tiger was transferred to the Veterinary Teaching Hospital where the explorative laparotomy and the ovario-hysterectomy were performed. The uterine horns appeared asymmetric and two nodular lesions of 1.5-cm-diameter were observed on the left horn. Histological assessment indicated the presence of a multifocal, subacute, moderate neutrophilic endometritis in both horns associated with diffuse, severe oedema. The nodules were consistent with a mesenchymal benign neoplasia. No relevant complication was observed in the following postoperative period and the cub was successfully hand raised. To the best of our knowledge, no report describes the birth of a healthy cub from a tiger affected by endometritis and neoplasia. Considering the normal maternal care observed in the previous twin-pregnancies in this animal, we cannot exclude that the uterine affection caused her abnormal behaviour in this case. Therefore, careful observation of the mother in the post-partum period is always strongly recommended

Efficacy of conventional versus innovative therapies for treating skin wounds in veterinary medicine

open16siINTRODUCTION: The skin is the largest organ of mammals. The loss of skin integrity may induce important dysfunctions or even death. For superficial wounds, the endogenous healing mechanisms in combination with traditional wound care are sufficient to achieve functional repair. In contrast, in larger wounds, like third and fourth degree burns, chronic wound or deep ulcers it is difficult to obtain the restitutio ad integrum and fibrosis and/or scar tissue develops1,2. The aim of this study was to verify the efficacy of conventional and innovative topic treatments on skin regeneration, induced experimentally in sheep. To achieve this goal different types of investigations (clinical, molecular, histological, immunohistochemical) were performed. METHODS: Six skin lesions (4x4cm) were surgically created on the back of six healthy adult sheep; every single wound was destined, in a randomized way, to one of the following treatments: Acemannan gel, Manuka Honey, hyaluronic acid, Plasma3 (ionized gas), allogeneic mesenchymal stem cells isolated from peripheral blood (PB-MSCs). The sixth wound was the placebo. Biopsies were collected with a surgical punch (0,6x0,6 cm) at time T0, T15 and T40 days. Lesions were clinically evaluated considering the presence and color of wound fluid, the state of hydration, the wound surface/surroundings and other parameters. Histological examinations considered crust formation, re-epithelization and epidermal thickness, dermis edema, extension of granulation tissue, acute and chronic inflammation. Immunohistochemistry for evaluation of inflammation, vascularization and cell proliferation was performed using CD3, CD20, MHCII, von Willebrand factor (vWF) and KI67 antibodies. Furthermore, Real time-PCR investigated genes as V ascular endothelial growth factors (VEGF), Transforming growth factor beta 1(TGFβ1), Vimentin (VIM), Collagen 1α1 (Col1α1) and hair Keratin (hKER). RESULTS: Clinically, the lesions treated with plasma healed more rapidly respect to other treatments and a reduced bacterial load was observed. At T7 wounds treated with stem cells and plasma were less macerated than lesions treated with other therapies. At T15 the wounds treated with hyaluronic acid showed a normal state of hydration while lesions treated with Manuka Honey exhibited a normal hydration from the third week only (Acemannan gel at fourth week). From the second week onwards all wounds did not show presence of fluid and exhibited a dry and clean secondary layer. All lesions, excluded wounds treated with acemannan gel, presented a red (hyaluronic acid and plasma) and dark red (Manuka Honey, PB-MSCs) granulation tissue starting from the first week. Molecular analysis showed a correspondence between clinical and molecular/histologic results. For instance, VEGF mRNA expression confirms angiogenetic events observed at histological level while TGF-β, CD3 and CD20 mRNA/protein expression indicated the presence/absence of inflammation in the used treatments. DISCUSSION & CONCLUSIONS: Innovative therapies led to surprising results regarding regeneration of mammalian skin. Indeed, on the basis of clinical analysis, wounds treated with plasma and MSC healed more rapidly. Further examinations are ongoing in order to elucidate possible mechanisms explaining these differences. REFERENCES: 1S.Y. Broeckx, S. Maes, T. Martinello, et al (2014) Equine epidermis: a source of epithelial-like stem/progenitor cells with in vitro and in vivo regenerative capacities Stem Cells Dev, pp 1134-48. 2J.H. Spaas, C. Gomiero, S.Y. Broeckx, et al (2016) Wound healing markers after autologous and allogeneic epithelial-like stem cell treatment Cytotherapy 2016 (in press). 3E. Martines, M. Zuin, R. Cavazzana, et al. (2009) A novel plasma source for sterilization of living tissues, New J. Phys. 11, 115014.openPatruno, MARCO VINCENZO; Gomiero, Chiara; Martinello, Tiziana; Perazzi, Anna; Gemignani, F; DE BENEDICTIS, GIULIA MARIA; Ferro, Silvia; Zuin, M; Martines, E; Cordaro, Luigi; Brun, Paola; Maccatrozzo, Lisa; Broeckx, Sy; Spaas, Jh; Chiers, K; Iacopetti, IlariaPatruno, MARCO VINCENZO; Gomiero, Chiara; Martinello, Tiziana; Perazzi, Anna; Gemignani, F; DE BENEDICTIS, GIULIA MARIA; Ferro, Silvia; Zuin, M; Martines, E; Cordaro, Luigi; Brun, Paola; Maccatrozzo, Lisa; Broeckx, Sy; Spaas, Jh; Chiers, K; Iacopetti, Ilari