Could cold plasma act synergistically with allogeneic mesenchymal stem cells to improve wound skin regeneration in a large size animal model?

Skin wound healing may sometimes lead to open sores that persist for long periods and expensive hospitalization is needed. Among different kinds of therapeutic innovative approaches, mesenchymal stem cells (MSCs) and low-temperature atmospheric pressure cold plasma (ionized gas) have been recently tested to improve this regenerative process. To optimize wound healing the present study intended to combine, for the first time, these two novel approaches in a large size animal wound healing model with the aim of assessing the putative dual beneficial effects. Based on clinical, histopathological, and molecular results a synergistic action in a second intention healing wound in sheep has been observed. Experimental wounds treated with cold plasma and MSCs showed a slower but more effective healing compared to the single treatment, as observed in previous studies. The combined treatment improved the correct development of skin appendages and structural proteins of the dermis showing the potential of the dual combination as a safe and effective tool for skin regeneration in the veterinary clinical field.

Maxillomandibulocardiac reflex in a dog.

BACKGROUND: The trigeminocardiac reflex (TCR) is a brainstem reflex that may be observed in anaesthesia during surgical procedures stimulating the intracranial or peripheral portion of the trigeminal nerve. The peripheral TCR is divided into the oculocardiac reflex and the maxillomandibulocardiac reflex based on the affected sensory branches of the trigeminal nerve. In veterinary medicine the oculocardiac reflex has been described, however the maxillomandibulocardiac reflex has never been reported. CASE PRESENTATION: A 5-year-old male Epagneul Breton was presented for surgical management of an upper lip mass. During surgery, a sudden severe bradycardia and a decrease in systemic arterial blood pressure developed. The occurrence of a maxillomandibulocardiac reflex was suspected on the basis of the temporary link between surgical stimulation and haemodynamic changes. Three doses of atropine were given before starting a dopamine infusion due to lack of response. The dopamine infusion normalized heart rhythm and blood pressure. The dog recovered uneventfully and he was discharged 24 h later with a sinus rhythm and no sign of recurrence of arrhythmias. CONCLUSION: The TCR is a rare but potentially life-threatening complication of procedures involving the sensory areas innervated by the three branches of the trigeminal nerve and it may cause bradycardia with hypotension. The use of a ß1-adrenergic receptor agonist such as dopamine may be indicated in cases of a refractory response to the conventional treatment with atropine.

Endodontic treatment of canine teeth in a cynomolgus monkey ( Macaca fascicularis).

A case of bilateral superior canine tooth pulp necrosis in an 18-year-old male Macaca fascicularis is described . A skull computed tomography scan was performed to confirm the diagnostic suspicion. A successful bilateral endodontic treatment with coronotomy was performed using rotary nickel-titanium instruments. Post-operative dental radiographs confirmed the correct positioning of the sealant. A rapid and excellent clinical recovery was obtained after each surgery. This innovative approach may contribute to improving the quality of root canal treatments provided to non-human primates.

Cytology of the healthy canine and feline ocular surface: comparison between cytobrush and impression technique.

BACKGROUND: Impression cytology (IC) is a noninvasive technique in which filters are used to sample superficial layers of ocular epithelium. OBJECTIVES: The aim of this study was to compare cytology specimens obtained by IC and cytobrush from healthy canine and feline eyes. METHODS: Dogs and cats were prospectively sampled using polytetrafluorethylene filters on the right eye, and cytobrush on the left eye. Wright-Giemsa-stained specimens were evaluated by 2 observers. Cellularity, preservation, and morphology of cells and presence of goblet and inflammatory cells were scored with a 4-grade scale. Inter-observer agreement and effects of topical anesthesia were analyzed. RESULTS: In 20 canine IC samples, 10 showed good cellularity (score 2-3) and 13 good preservation. Superficial epithelial cells (SEC) were present in 13/20 of IC, while basal-intermediate cells (BIC) were seen in 14/20. In 6/20 and 7/20, goblet and inflammatory cells were noted, respectively. In 20 cats, 15 of IC showed good cellularity and 14 good preservation, and SEC were present in 16/20 of IC and BIC in 17/20. In 13/20 and 3/20 cats, goblet cells and inflammatory cells were noted, respectively. Canine cytobrush specimens appeared well preserved (9/20) and had good cellularity (8/20). In feline cytobrush specimens, good preservation and cellularity were observed in 16/20 and 14/20, respectively. In both species, all cell types were present without a clear separation. There was moderate to fair agreement about cellular morphology in IC between observers. Specimens obtained with and without anesthesia were comparable. CONCLUSION: Impression cytology allowed collection of samples with maintained cytoarchitecture, while cytoplasmatic and nuclear details were often difficult to evaluate.

Pocket technique or pocket technique combined with modified orbital rim anchorage for the replacement of a prolapsed gland of the third eyelid in dogs: 353 dogs.

The aim of this retrospective study was to evaluate the results obtained in 353 dogs (420 eyes) using two different surgical techniques for correction of a prolapsed gland of the third eyelid: the Morgan's pocket technique and a technique combining Morgan's approach with a slightly modified periosteal anchoring technique of Stanley and Kaswan. The pocket technique was used in 234 eyes and the combined technique in 186 eyes. Successful repositioning was obtained in 95% of all cases, with recurrence occurring in 5%. The recurrence rate in large breed dogs such as the English Bulldog and Boxer was lower with the combined technique than with the pocket technique.

Might the Masson trichrome stain be considered a useful method for categorizing experimental tendon lesions?

Strain injuries of tendons are the most common orthopedic injuries in athletic subjects, be they equine or human. When the tendon is suddenly damaged, an acute inflammatory phase occurs whereas its repetitive overloading may cause chronic injuries. Currently the criteria used for grading injuries are general and subjective, and therefore a reliable grading method would be an improvement. The main purpose of this study was to assess qualitatively the histological pattern of Masson trichrome stain in healthy and injured tendons; indeed, the known "paradox" of Masson staining was used to create an evaluation for the matrix of tendons, following experimental lesions and natural repair processes. A statistically significant difference of aniline-staining between healthy and lesioned tendons was observed. Overall, we think that the Masson staining might be regarded as an informative tool in discerning the collagen spatial arrangement and therefore the histological characteristics of tendons.

Effect of MLS(®) laser therapy with different dose regimes for the treatment of experimentally induced tendinopathy in sheep: pilot study.

OBJECTIVE: The aim of this preliminary study was to investigate the effect of Multiwave Locked System (MLS(®)), a particular model of low-level laser, in the acute phase of collagenase-induced tendon lesions in six adult sheep randomly assigned to two groups. BACKGROUND DATA: Tendon injuries are common among human athletes and in sport horses, require a long recovery time, and have a high risk of relapse. Many traditional treatments are not able to repair the injured tendon tissue correctly. In recent years, the use of low-level laser therapy (LLLT) produced interesting results in inflammatory modulation in different musculoskeletal disorders. METHODS: Group 1 received 10 treatments of MLS laser therapy at a fluence of 5 J/cm(2) on the left hindlimb. Group 2 received 10 treatments of MLS laser therapy at a fluence of 2.5 J/cm(2) on the left hindlimb. In every subject in both groups, the right hindlimb was considered as the control leg. RESULTS: Clinical follow-up and ultrasonography examinations were performed during the postoperative period, and histological examinations were performed at day 30 after the first application of laser therapy. In particular, results from histological examinations indicate that both treatments induced a statistically significant cell number decrease, although only in the second group did the values return to normal. Moreover, the MLS laser therapy dose of 2.5 J/cm(2) (group 2) caused a significant decrease of vessel area. CONCLUSIONS: In this study, clinical and histological evaluation demonstrated that a therapeutic dose <5 J/cm(2) furnished an anti-inflammatory effect, and induced a decrease of fibroblasts and vessel area. Overall, our results suggest that MLS laser therapy was effective in improving collagen fiber organization in the deep digital flexor tendon.

Multiple dermoid sinuses of type Vb and IIIb on the head of a Saint Bernard dog.

Dermoid sinus, a congenital malformation of neural tube development, has been reported in humans and several animal species including dogs. It is typically found in the dorsal midline and commonly occurs in the Rhodesian Ridgeback breed. A case of multiple dermoid sinuses in the fronto-occipital region is described. An 11-month-old, intact female Saint Bernard dog was presented with a 2 day history of discharge from a large irregular subcutaneous mass in the fronto-occipital region. The dog was otherwise healthy. The dog had two circular skin lesions (approximately 4 × 4 and 4 × 2 cm diameter) surrounded by multiple irregular elevated masses. The masses had multiple small openings on the skin surface with tufts of hair protruding from the apertures. The masses were surgically removed, and the diagnosis of multiple dermoid sinuses was confirmed by histological examination. Histopathological examination showed multiple, variably sized, spherical to tubular cysts expanding the dermis and subcutis. Cysts were filled with hair shafts and lamellar keratin and were lined by a stratified squamous epithelium. Sebaceous and apocrine gland adnexal structures were also observed. To the best of our knowledge, this is the first reported case of multiple dermoid sinuses of two different types in the head of a Saint Bernard dog.

Effects of in vivo applications of peripheral blood-derived mesenchymal stromal cells (PB-MSCs) and platlet-rich plasma (PRP) on experimentally injured deep digital flexor tendons of sheep.

Tendon injuries, degenerative tendinopathies, and overuse tendinitis are common in races horses. Novel therapies aim to restore tendon functionality by means of cell-based therapy, growth factor delivery, and tissue engineering approaches. This study examined the use of autologous mesenchymal stromal cells derived from peripheral blood (PB-MSCs), platelet-rich plasma (PRP) and a combination of both for ameliorating experimental lesions on deep digital flexor tendons (DDFT) of Bergamasca sheep. In particular, testing the combination of blood-derived MSCs and PRP in an experimental animal model represents one of the few studies exploring a putative synergistic action of these treatments. Effectiveness of treatments was evaluated at 30 and 120 days comparing clinical, ultrasonographic, and histological features together with immunohistochemical expression of collagen types 1 and 3, and cartilage oligomeric matrix protein (COMP). Significant differences were found between treated groups and their corresponding controls (placebo) regarding tendon morphology and extracellular matrix (ECM) composition. However, our results indicate that the combined use of PRP and MSCs did not produce an additive or synergistic regenerative response and highlighted the predominant effect of MSCs on tendon healing, enhanced tissue remodeling and improved structural organization.

Biological fate of tissue-engineered porcine valvular conduits xenotransplanted in the sheep thoracic aorta.

The ideal prosthesis to replace the diseased human aortic valve is not yet available. We have previously shown that porcine acellular aortic-valve conduits, obtained by detergent-enzymatic method, display hemodynamic performances similar to those of their native counterparts. Hence, it seemed worthwhile to ascertain whether these tissue-engineered prostheses can be successfully xenotransplanted. Porcine acellular conduits, which immunocytochemistry demonstrated to lack MHC class I and II antigens, were implanted in the thoracic aorta of 9 sheep. Two animals died just after surgery, and the other 7 sheep were sacrificed 1 or 5 months after transplantation. A rather favorable outcome of the implant was observed in 4 sheep. In these animals, aortic valves remained pliable and coaptive, and the luminal surface of the conduits was endothelized just after one month from surgery. An intense inflammatory response was present at 1 month, and, although attennuated, it persisted for 5 months, located mainly between the tunica intima and media and at the border of the implant. Vimentin-positive and smooth muscle actin-positive myofibroblasts proliferated within tunica media and adventitia, and an obvious thickening of the tunica intima was also observed. Small vessels were seen in the adventitia, and elastic fibers were well-preserved in both the aorta wall and valve leaflets. In the cases of unfavorable outcome (3 of 7 survived sheep), implants were detached from the aorta recipient and surrounded by a connective mass that almost completely obstructed their lumen. These masses were composed of a fibromyxoid background where proliferating cells, resembling those occurring in human reactive myofibroblastic lesions (proliferative fascitis), were embedded. Collectively, these rather disappointing findings indicate that acellular valve conduits, obtained by the detergent-enzymatic method, are presently not suitable for clinical applications because of the persistent inflammatory response, which conceivably triggers overgrowth mechanisms that lead to implant failure.