Mini-invasive approach for removal of iliopsoas migrating grass awns with an atraumatic wound retractor.

This case series describes a novel mini coeliotomy approach using a radial, atraumatic self-retaining retractor for the retrieval of migrant plant foreign bodies from the iliopsoas muscles of six male dogs under intra-operative ultrasonographic guidance. Four dogs had a history of pulmonary disease potentially compatible with inhalation of a foreign body approximately 2-4 months before presentation. Under ultrasonographic guidance, the grass awns were identified in the iliopsoas muscle and were completely removed. In this case series, the annular ring device provided an excellent view of the surgical field for intra-abdominal manipulations. Patient follow-up at 15 days and 6 to 12 months after surgery indicated a full recovery, and no grass awn fragment residues were identified.

Characterisation and intracellular labelling of mesenchymal stromal cells derived from synovial fluid of horses and sheep.

Multipotent mesenchymal stromal cells (MSCs) derived from synovial fluid (SF) are considered to be a promising cell type for therapeutic applications in joint disease. However, despite their potential relevance for clinical and experimental studies, there is insufficient knowledge about SF-derived MSCs isolated from horses and sheep. In this study, cells were recovered from healthy SF and bone marrow (BM) of sheep, and from healthy and osteoarthritic SF of horses. Ovine SF-MSCs were used to assess the efficiency of intracellular labelling with quantum dots (QDs). Colony forming units, generation times, trilineage differentiation potential and expression of CD73, CD90 and CD105 at mRNA level were assessed. QD labelling was efficient, with >98% positive cells directly after labelling at 10 nmol/L and >95% positive cells directly after labelling at 2 nmol/L. The label decreased over 7 days of culture, with more persistence at the higher labelling concentration. No significant differences in proliferation were observed. All MSCs had trilineage differentiation potential, but adipogenesis was more distinct in equine samples and chondrogenesis was most pronounced in ovine SF-MSCs. CD73, CD90 and CD105 were expressed in equine and ovine MSCs.

Survival of bone marrow mesenchymal stem cells labelled with red fluorescent protein in an ovine model of collagenase-induced tendinitis.

OBJECTIVE: The aim of this study was to track the survival and efficacy of allogeneic bone marrow mesenchymal stem cells (BM-MSC) marked with red fluorescent protein (BM-MSCRFP) in an ovine model of collagenase-induced tendinopathy. METHODS: Bone marrow was harvested from one donor sheep and BM-MSC were isolated, cultivated and transfected with red fluorescent protein (BM-MSCRFP). Collagenase was injected into both Achilles tendons in the remaining nine sheep. After two weeks the left tendon was injected with a solution of 6 x 106 BM-MSCRFP and fibrin glue, while only fibrin glue was administered to the contra-lateral tendon in each sheep. After three, four and six weeks the tendons were harvested and evaluated for morphology, collagen I deposition, presence of CD34+ cells, and fluorescent labelled BM-MSC. RESULTS: We demonstrated that delivery of BM-MSC into tendon lesions had positive effects on the injured tendons. The BM-MSCRFP survived at three, four and six weeks after treatment, leading to better quality healing of tendons as compared to the controls, where no labelled cells were detected. Interestingly, we demonstrated high expression of CD34+ cells in tendons that had been treated with BM-MSCRFP. CLINICAL RELEVANCE: Mesenchymal stem cell allografts have a positive effect on tendon healing and local injection of BM-MSC directly into the tendon allows the homing of BM-MSC for good efficiency of engraftment.

Comparison of intra-operative analgesia provided by intravenous regional anesthesia or brachial plexus block for pancarpal arthrodesis in dogs.

The aim of this study was to compare intravenous regional anesthesia (IVRA) and brachial plexus block (BPB) for intra-operative analgesia in dogs undergoing pancarpal arthrodesis (PA). Twenty dogs scheduled for PA were intramuscularly sedated with acepromazine (0.03 mg/kg), general anesthesia was intravenously (IV) induced with thiopental (10 mg/kg) and, after intubation, maintained with isoflurane in oxygen. In 10 dogs (GIVRA) IVRA was performed on the injured limb administering 0.6 ml/kg of 0.5% lidocaine. In 10 dogs (GBPB) the BPB was performed at the axillary level with the help of a nerve stimulator and 0.3 ml/kg of a 1:1 solution of 2% lidocaine and 1% ropivacaine was injected. During surgery fentanyl (0.002 mg/kg IV) was administered if there was a 15% increase of HR and/or MAP compared to the values before surgical stimulation. All the standard cardiovascular and respiratory parameters were continuously monitored during surgery. The duration of surgery and the time of extubation were recorded. Data were compared with a 1-way ANOVA test (P<0.05). No patients required fentanyl administration during surgery. All the recorded parameters were similar in the two groups. The two techniques were similar in providing intra-operative analgesia in dogs undergoing orthopaedic surgery.

Alveolar recruiting maneuver in dogs under general anesthesia: effects on alveolar ventilation, gas exchange, and respiratory mechanics.

The aim of this study was to evaluate the effects of a recruiting maneuver (RM) on lung aeration, gas exchange, and respiratory mechanics during general anesthesia in mechanically ventilated dogs. A thoracic computed tomography (CT) scan, an arterial blood sample, and measurement of respiratory mechanics were performed 10 min before (baseline) and both 5 and 30 min after a vital capacity RM in 10 dogs under general anesthesia. The RM was performed by inflating the lung at 40 cm H(2)O for 20 s. Lung aeration was estimated by analyzing the radiographic attenuation of the CT images. Lung aeration and gas exchange improved significantly 5 min after the RM compared to baseline and returned to values similar to baseline by 30 min. Static lung compliance was not significantly affected by the RM. An RM induces a temporary improvement in lung function in healthy dogs under general anesthesia.

Ocular dirofilariosis by Dirofilaria immitis in a dog: first case report from Europe.

A five-year-old, entire female mixed-breed dog was presented with corneal oedema and episcleral hyperaemia in the left eye. The ophthalmological examination revealed the presence of a free-swimming nematode in the anterior chamber. Circulating microfilariae were not observed by a modified Knott test nor were adult antigens detected in serum by a commercial ELISA. The parasite was surgically removed from the dog's eye, but its anterior end was damaged during the surgery. Based on the morphology of the posterior end, the nematode was preliminarily identified as a male Dirofilaria immitis. The species identification was confirmed by PCR amplification and sequencing of the mitochondrial coxI and 12S rDNA genes, using a DNA barcoding approach. Although other cases of ocular dirofilariosis by D. immitis have been previously recorded in Australia and the United States, the case reported herein is the first in a dog from Europe.

Activation of PI3-kinase/Akt induced small bowel cell apoptosis during laparoscopic ischaemia-reperfusion of swine jejunum.

INTRODUCTION: This study aimed to detect small bowel cell apoptosis by evaluating the nuclear activation of transcription factors, nuclear factor-kappaB (NF-kappaB) and Akt pathways as well as detecting apoptotic cells by TUNEL assay, during ischaemia-reperfusion (I/R) injury by Arterio-Venous Occlusion (AVO) of ajejunal loop in swine, using a laparoscopic procedure. MATERIALS AND METHODS: A jejunal loop was isolated and a biopsy sample (T0) obtained. Biopsies were obtained at 30 minutes (T30I) and 60 minutes (T60I) after clamping and again during reperfusion (R), T30R and T60R. Standard H&E stains and immunohistochemical study for infiltrating polymorphonuclear leukocytes and TUNEL assays were performed. The activation of Akt and NF-kappaB in the animal model was evaluated by indirect immunofluorescence and confocal microscopy. CONCLUSIONS: Our results confirmed that I/R injury of the jejunum did not only cause epithelial damage, but also activated the molecular pathways triggering an apoptotic reaction (by pro-inflammatory cytokines) and an anti-apoptotic mechanism that can lead to the regeneration of injured tissue.

Experimental modified orthotopic piggy-back liver autotransplantation.

UNLABELLED: The classical orthotopic liver autotransplantation is a very challenging and time wasting technique; it includes the division of major hepatic vessels and choledocus, and subsequent reconnection by end to end anastomoses. The caval end to end anastomoses are the most difficult to be performed and the interposition of a prosthesis can be required. We adopted the classical orthotopic liver autotransplantation technique in 2 patients affected with diffused liver metastases from colorectal cancer, for extracorporeal neutron capture therapy (BNCT). The procedure required very long operating times and the extracorporeal circulation (ECC) set up; furthermore the vena cava reconstruction was performed by the interposition of a goretex-prosthesis. We propose a "modified orthotopic piggy-back technique" to simplify liver reconnection and shorten the operating time. MATERIALS AND METHODS: The technique was developed in the swine (25 kg body weight), under general anaesthesia. We performed the resection of the retro-hepatic vena cava with preservation of the caval flow during the anhepatic phase, by interposing a goretex-prosthesis. The reconstruction of the vena cava was then performed by a side-to-side cava-prosthesis anastomosis with lateral clamping of the prosthesis. The procedure was then completed according to the classical technique of liver transplantation. RESULTS: The mean time for VC reconstruction was 56 (+/-10)min. and the mean time for side-to-side VC-prosthesis anastomosis was 13(+/-4)min. CONCLUSIONS: The "modified orthotopic piggy-back technique" can simplify the reimplant of the liver during autotransplantation and shorten the operating time. Furthermore also the time of total extracorporeal circulation is reduced, as during the anhepatic phase and during the side-to-side cava-prosthesis anastomosis the flow in the inferior vena cava is uninterrupted.

Histology and immunohistochemistry study of ovine tendon grafted with cBMSCs and BMMNCs after collagenase-induced tendinitis.

OBJECTIVES: The aim of this study was to compare the regeneration abilities of cultured bone marrow mesenchymal cells (cBMSC) and bone marrow mononuclear cells (BMMNC) with fibrin glue, saline solution and sham control in collagenase-induced tendinitis of the Achilles tendon in sheep. METHODS: Six sheep were recruited randomly to each group: cBMSC, BMMNC, fibrin, saline and sham control. Each group received the relative treatment two weeks after inducing lesions (T(0)). After eight weeks (T(8)) of treatment, the tendons were harvested and evaluated for histomorphology, Collagen type I, III, Cartilage Oligomeric Matrix Protein (COMP) and CD34 positive cells expression. RESULTS: Histology and immunohistochemistry showed similar capabilities of cBMSC and BMMNC to restore the architecture of fibres and Extra Cellular Matrix (ECM), with a high expression of collagen type I and COMP and a very low expression of collagen type III in treated tendons. The complete architectural disruption of fibres, dramatic reduction of collagen Type I and COMP expression and increase collagen type III expression were commonly observed in tendons treated with fibrin or saline only. The presence of CD34 positive cells was appreciable in the BMMNC group while few cBMSC showed this cluster of differentiation, not expressed in tendons treated with fibrin or saline. CLINICAL SIGNIFICANCE: The data in this study show the efficacy of cBMSC and BMMNC in regenerating tendon tissue after collagenase-induced tendinitis.

Cell therapy for tendinitis, experimental and clinical report.

To compare cultured bone marrow mesenchymal cells (cBMSC), bone marrow mononucleated cells (BMMNCs), and placebo to repair collagenase-induced tissue damage in an equine model of experimental tendonitis, 6 Standardbred horses with no signs of previous SDF tendon injury have been recruited. Three weeks after collagenase treatment an average of either 5.5 x 10(6) cBMSCs or 122.3 x 10(6) BMMNCs, saline solution (placebo) or fibrin glue were injected intralesionally in random order. Horses were stall rested for 21 weeks, and tendon ultrasound scans performed before and during this period. Horses were euthanized and tendons harvested for histology and immunohistochemistry. Data observed in this study showed effectiveness of cBMSC and BMMNC in regenerating tendon tissue after collagenase -induced tendonitis. Both cBMSC and BMMNC transplantation resulted in qualitatively similar regeneration of tendon extracellular matrix in terms of type I/III collagen ratio, fiber orientation, and COMP expression. After this favourable results, 20 horses were recruited referred for spontaneous lesions of the flexor tendons or the suspensory ligament. Horses were treated with autologous graft of BMMNCs.After treatment the. the exercise program allowed was 8 weeks stall rest, 4 weeks hand walking, 4 weeks trotting, 4 weeks of gradually raising of exercise level then horses were gone back to race. US characteristics of lesions started to improve at T3. CSA-l, FPS and TLS were better in all patients, with an appreciable filling of lesions indicated by a decreasing of CSA-l and increasing of TLS. When horses started the exercise program T8 tendon architecture improved, demonstrated by their longitudinal alignment and length. At T6, and persistently in later follow-up, no lameness was evident by clinical examination. At time of writing 12 patients (60%) were go back to races, while other 8 (40%) are under controlled exercise program. Re-injury rate was assessed at 25%. All the owners judged good to excellent the outcome in term of athletic success.

Use of autologous bone marrow mononuclear cells and cultured bone marrow stromal cells in dogs with orthopaedic lesions.

The aim of the study is to evaluate the clinical application in veterinary orthopedics of bone marrow mononuclear cells (BMMNCs) and cultured bone marrow stromal cells (cBMSCs) for the treatment of some orthopaedic lesions in the dog. The authors carried out a clinical study on 14 dogs of different breed, age and size with the following lesions: 1 bone cyst of the glenoid rime; 2 nonunion of the tibia; 3 nonunion of the femur; 2 lengthening of the radius; 1 large bone defect of the distal radius;1 nonunion with carpus valgus; 4 Legg-Calvé-Perthés disease. In 9 cases the BMMCNs were used in combination with a three dimensional resorbable osteogenic scaffold the chemical composition and size of which facilitates the ingrowth of bone. In these cases the BMMNCs were suspended in an adequate amount of fibrin glue and then distribuited uniformly on a Tricalcium-Phosphate (TCP) scaffold onto which were also added some drops of thrombin. In 1 case of nonunion of the tibia and in 3 cases of Legg-Calvè-Perthés (LCP) disease the cultured BMSCs were used instead because of the small size of the dogs and of the little amount of aspirated bone marrow. X-ray examinations were performed immediately after the surgery. Clinical, ultrasounds and X-ray examinations were performed after 20 days and then every month. Until now the treated dogs have shown very good clinical and X-ray results. One of the objectives of the study was to use the BMMNCs in clinical application in orthopaedic lesions in the dog. The advantages of using the cells immediately after the bone marrow is collected, are that the surgery can be performed the same day, the cells do not need to be expanded in vitro, they preserve their osteogenic potential to form bone and promote the proper integration of the implant with the bone and lastly, the technique is easier and the costs are lower.

Surgery plus chondroprotection for canine cranial cruciate ligament (CCL) rupture: a proton-NMR study.

Rupture of the cranial cruciate ligament (CCL) is one of the most frequent causes of lameness of the rear limb in the dog. Regardless of the type of treatment, CCL rupture inevitably leads to knee osteoarthritis (OA). The purpose of this study was to evaluate the efficacy of associating surgical treatment of spontaneous rupture of the CCL with a chondroprotector, that is called 'supraadditive' because it is formulated to counteract not only chondrodegeneration, but also the oxidative and inflammatory processes of OA. The open-label controlled study used proton NMR spectroscopy to evaluate the synovial fluid of the stifle of 10 dogs with monolateral rupture of the CCL, selected for the study based on specific inclusive criteria. The dogs were assigned randomly into two groups. Five dogs received the supra-additive chondroprotector for 60 days, starting on the day after surgery. Five dogs only underwent surgical reconstruction of the CCL. The results were analysed with the ANOVA unstructured variance matrix-covariance test. The trend over time of the synovial concentration of four metabolites (lactate, alanine, acetyl groups of N-acetylated sugars on glycoproteins and alpha-anomers of glucose) was found to differ to a statistically significant extent between the two groups, suggesting that the supra-additive chondroprotector produces an intra-articular metabolic rebalance. The results support the adjuvant use of the chondroprotector in the management of CCL rupture, in view of its control of the OA changes that accompany this orthopaedic disabling condition.

Tissue engineering of bone: search for a better scaffold.

BACKGROUND: Large bone defects still represent a major problem in orthopedics. Traditional bone-repair treatments can be divided into two groups: the bone transport (Ilizarov technology) and the graft transplant (autologous or allogeneic bone grafts). Thus far, none of these strategies have proven to be always resolving. As an alternative, a tissue engineering approach has been proposed where osteogenic cells, bioceramic scaffolds, growth factors and physical forces concur to the bone defect repair. Different sources of osteoprogenitor cells have been suggested, bone marrow stromal cells (BMSC) being in most cases the first choice. METHODS AND RESULTS: In association with mineral tridimensional scaffolds, BMSC form a primary bone tissue which is highly vascularized and colonized by host hemopoietic marrow. The chemical composition of the scaffold is crucial for the osteoconductive properties and the resorbability of the material. In addition, scaffolds should have an internal structure permissive for vascular invasion. Porous bioceramics [hydroxyapatite (HA) and tricalcium phosphate] are osteoconductive and are particularly advantageous for bone tissue engineering application as they induce neither an immune nor an inflammatory response in the implanted host. Earlier, we first reported a cell-based tissue engineering procedure to treat three patients with long bone segmental defects. Cells were loaded on a 100% HA porous ceramic. These scaffolds proved to have good osteoconductive properties resulting in a good functional recovery, but they have not been resorbed after more than 5 years from the implant. In addition, due to the high density of the mineral and the relatively low porosity (50-60%), it was very difficult to monitor the patient recovery during the post-surgery time using X-rays. CONCLUSIONS: We report here some pre-clinical testing of new scaffolds. To compare these second generation ceramic scaffolds more suitable for a tissue engineering approach we had to first establish animal models and analysis procedures including the use of X-ray-computed microtomography associated with X-rays synchroton radiation.

Bladder Z-plasty for the repair of ureteric injuries. Experimental study in sheep.

There is general acceptance that the best substitute for a damaged distal ureter is an elongated bladder and that the bladder psoas hitch is the most reliable procedure. Nevertheless, it is not always sufficient to bridge a long ureteric gap. In order to obtain an equally safe but wider bladder elongation than the psoas hitch can provide, we have previously studied the effect of several Z-shaped incisions on isolated pig bladders. The aim of the present study was to verify, in vivo, the efficacy and safety of the new technique. Five sheep underwent the psoas hitch procedure and 6 the Z-plasty procedure. The results showed that Z-plasty provides an equally safe but longer bladder elongation than the psoas hitch procedure.