The clinical translation of bone tissue engineering for reconstructing large bone defects has not advanced without hurdles. The in vivo bioreactor (IVB) concept may therefore bridge between bone ...tissue engineering and reconstructive surgery by employing the patient body for prefabricating new prevascularized tissues. Ideally, IVB should minimize the need for exogenous growth factors/cells. Periosteal tissues are promising for IVB approaches to prefabricate tissue-engineered bone (TEB) flaps. However, the significance of preserving the periosteal vascular supply has not been adequately investigated. This study assessed muscle IVB with and without periosteal/pericranial grafts and flaps for prefabricating TEB flaps to reconstruct mandibular defects in sheep. The sheep (n = 14) were allocated into 4 groups: muscle IVB (M group; nM = 3), muscle + periosteal graft (MP group; nMP = 4), muscle + periosteal flap (MVP group; nMVP = 4), and control group (nControl = 3). In the first surgery, alloplastic bone blocks were implanted in the brachiocephalic muscle (M) with a periosteal graft (MP) or with a vascularized periosteal flap (MVP). After 9 wk, the prefabricated TEB flaps were transplanted to reconstruct a mandibular angle defect. In the control group, the defects were reconstructed by non-prevascularized bone blocks. Computed tomography (CT) scans were performed after 13 wk and after 23 wk at termination, followed by micro-CT (µCT) and histological analyses. Both CT and µCT analysis revealed enhanced new bone formation and decreased residual biomaterial volume in the MVP group compared with control and MP groups, while the M group showed less new bone formation and more residual biomaterial. The histological analysis showed that most of the newly formed bone emerged from defect edges, but larger areas of new bone islands were found in MP and MVP groups. The MVP group showed enhanced vascularization and higher biomaterial remodeling rates. The periosteal flaps boosted the reconstructive potential of the prefabricated TEB flaps. The regenerative potential of the periosteum was manifested after the transplantation into the mechanically stimulated bony defect microenvironment.
Background
Acute‐phase proteins (APPs) are sensitive markers of inflammation, and serum C‐reactive protein (CRP) recently has been shown to be a useful diagnostic marker in dogs with bacterial ...pneumonia (BP). In humans with community‐acquired pneumonia, APPs also have great utility as follow‐up markers aiding in the assessment of treatment response.
Objectives
The aim of our study was to investigate the applicability of APPs as markers of treatment response in dogs with BP.
Animals
Nineteen dogs diagnosed with BP and 64 healthy dogs.
Methods
The study was conducted as a prospective longitudinal observational study. Serum CRP, serum amyloid A (SAA), and haptoglobin concentrations were followed during a natural course of BP. Normalization of serum CRP was used to guide the duration of antibiotic treatment (treatment was stopped 5–7 days after CRP normalized) in 8 of 17 dogs surviving to discharge; 9 of 17 dogs were treated according to conventional recommendations.
Results
All measured APPs initially were significantly increased, but the magnitude of increase was not correlated to disease severity. C‐reactive protein and SAA concentrations decreased rapidly after initiation of antimicrobial treatment. When normalization of serum CRP was used to guide the duration of antibiotic treatment, treatment duration was significantly (P = .015) decreased without increasing the number of relapses.
Conclusions and Clinical Importance
Serum CRP and SAA reflected the recovery process well and therefore may be used as markers of treatment response. According to the results, the normalization of serum CRP may be used to guide the duration of antibiotic treatment in dogs with BP.
One to two percent of all children are born with a developmental disorder requiring pediatric hospital admissions. For many such syndromes, the molecular pathogenesis remains poorly characterized. ...Parallel developmental disorders in other species could provide complementary models for human rare diseases by uncovering new candidate genes, improving the understanding of the molecular mechanisms and opening possibilities for therapeutic trials. We performed various experiments, e.g. combined genome-wide association and next generation sequencing, to investigate the clinico-pathological features and genetic causes of three developmental syndromes in dogs, including craniomandibular osteopathy (CMO), a previously undescribed skeletal syndrome, and dental hypomineralization, for which we identified pathogenic variants in the canine SLC37A2 (truncating splicing enhancer variant), SCARF2 (truncating 2-bp deletion) and FAM20C (missense variant) genes, respectively. CMO is a clinical equivalent to an infantile cortical hyperostosis (Caffey disease), for which SLC37A2 is a new candidate gene. SLC37A2 is a poorly characterized member of a glucose-phosphate transporter family without previous disease associations. It is expressed in many tissues, including cells of the macrophage lineage, e.g. osteoclasts, and suggests a disease mechanism, in which an impaired glucose homeostasis in osteoclasts compromises their function in the developing bone, leading to hyperostosis. Mutations in SCARF2 and FAM20C have been associated with the human van den Ende-Gupta and Raine syndromes that include numerous features similar to the affected dogs. Given the growing interest in the molecular characterization and treatment of human rare diseases, our study presents three novel physiologically relevant models for further research and therapy approaches, while providing the molecular identity for the canine conditions.
Background
Chiari‐like malformation (CM) and syringomyelia (SM) are widely reported in Cavalier King Charles Spaniels and Griffon Bruxellois dogs. Increasing evidence indicates that CM and SM also ...occur in other small and toy breed dogs, such as Chihuahuas.
Objectives
To describe the presence of SM and craniocervical junction (CCJ) abnormalities in Chihuahuas and to evaluate the possible association of CCJ abnormalities with SM. To describe CM/SM‐related clinical signs and neurologic deficits and to investigate the association of CM/SM‐related clinical signs with signalment, SM, or CCJ abnormalities.
Animals
Fifty‐three client‐owned Chihuahuas.
Methods
Prospective study. Questionnaire analyses and physical and neurologic examinations were obtained before magnetic resonance and computed tomography imaging. Images were evaluated for the presence of SM, CM, and atlantooccipital overlapping. Additionally, medullary kinking, dorsal spinal cord compression, and their sum indices were calculated.
Results
Scratching was the most common CM/SM‐related clinical sign and decreased postural reaction the most common neurologic deficit in 73 and 87% of dogs, respectively. Chiari‐like malformation and SM were present in 100 and 38% of dogs, respectively. Syringomyelia was associated with the presence of CM/SM‐related clinical signs (P = 0.034), and medullary kinking and sum indices were higher in dogs with clinical signs (P = 0.016 and P = 0.007, respectively).
Conclusions and Clinical Importance
Syringomyelia and CCJ abnormalities are prevalent in Chihuahuas. Syringomyelia was an important factor for the presence of CM/SM‐related clinical signs, but many dogs suffered from similar clinical signs without being affected by SM, highlighting the clinical importance of CCJ abnormalities in Chihuahuas.
•Pre-treatment with vatinoxan (MK-467) was evaluated in sevoflurane-anaesthetised sheep receiving dexmedetomidine.•Effects on cardiopulmonary function, lung computed tomography and bronchoalveolar ...lavage were investigated.•Dexmedetomidine-induced cardiovascular effects were minimal and pulmonary effects were highly variable among sheep.•No obvious changes were observed in bronchoscopy or in bronchoalveolar lavage in either treatment group.•Vatinoxan alleviated a dexmedetomidine-induced increase in airway pressure and subclinical pulmonary oedema formation.
The effects of pre-treatment with vatinoxan (MK-467) on dexmedetomidine-induced cardiopulmonary alterations were investigated in sheep. In a crossover study design with a 20-day washout, seven sheep were anaesthetised with sevoflurane in oxygen and air. The sheep were ventilated with the pressure-limited volume-controlled mode and a positive end-expiratory pressure of 5cmH2O. Peak inspiratory pressure (PIP) was set at 25cmH2O. The sheep received either 150μg/kg vatinoxan HCl (VAT+DEX) or saline intravenously (IV) 10min before IV dexmedetomidine HCl (3μg/kg, DEX). Cardiopulmonary variables were measured before treatments (baseline), 3min after vatinoxan or saline, and 5, 15 and 25min after dexmedetomidine. Computed tomography (CT) of lung parenchyma was performed at baseline, 2min before dexmedetomidine, and 10, 20 and 30min after DEX. Bronchoalveolar lavage (BAL) was performed after the last CT scan and shortly before sheep recovered from anaesthesia. After VAT, cardiac output significantly increased from baseline. DEX alone significantly decreased partial arterial oxygen tension, total dynamic compliance and tidal volume, whereas PIP was significantly increased. With VAT+DEX, these changes were minimal. No significant changes were detected in haemodynamics from baseline after DEX. With VAT+DEX, mean arterial pressure and systemic vascular resistance were significantly decreased from baseline, although hypotension was not detected. On CT, lung density was significantly increased with DEX as compared to baseline. No visual abnormalities were detected in bronchoscopy and no differences were detected in the BAL fluid after either treatment. The pre-administration of vatinoxan alleviates dexmedetomidine-induced bronchoconstriction, oedema and hypoxaemia in sevoflurane-anaesthetised sheep.
BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is an incurable interstitial lung disease occurring mainly in West Highland White Terriers (WHWTs). The effects of IPF on survival and on exercise ...tolerance in WHWTs are unknown. OBJECTIVES: To evaluate survival, prognostic factors, and exercise tolerance in WHWTs with IPF. ANIMALS: Privately owned WHWTs; 15 with IPF and 11 healthy controls. METHODS: Prospective case‐control study conducted in 2007–2012. For survival, descriptive statistics and Kaplan–Meier (KM) survival curves with Cox proportional hazard ratios were performed. For the prognostic factor study, KM curves, Cox regression analysis, and logistic regression models were used. The 6‐minute walk test (6MWT) was used for measurement of exercise tolerance. RESULTS: The median IPF‐specific survival of deceased WHWTs (7/15) with IPF was 32 (range 2–51) months from onset of clinical signs. The risk of death from birth in WHWTs with IPF in age‐adjusted Cox model was significantly higher (hazard ratio 4.6; 95% confidence interval 1.05–19.74, P = .04) than in control WHWTs. No significant prognostic factors were identified. In 6MWT, WHWTs with IPF walked a shorter distance, median 398 m (range 273–519 m), than healthy controls, median 492 m (420–568 m), P = .05, and the partial pressure of oxygen in arterial blood in diseased dogs had a moderate positive correlation with walking distance (Kendall′s tau‐b = 0.69, P = .06). CONCLUSION AND CLINICAL IMPORTANCE: IPF had a negative impact on life expectancy, but individual survival varied considerably. 6MWT proved to be a well‐tolerated, noninvasive test to evaluate exercise tolerance.
Background: Idiopathic pulmonary fibrosis (IPF) is a chronic, interstitial lung disease primarily affecting West Highland White Terriers (WHWTs). Objective: To describe the clinicopathological and ...diagnostic imaging features in WHWTs with IPF. Animals: Twelve WHWTs with IPF and 14 healthy control WHWTs. Method: Prospective study. Clinical signs and findings of physical examination, blood and arterial blood gas analyses, radiography, high-resolution computed tomography (HRCT), bronchoscopy and bronchoalveolar lavage (BAL) of IPF dogs were obtained and compared with controls. Histopathologic changes in IPF dogs were evaluated. Results: Mean partial pressure of oxygen was significantly lower in IPF (mean ± SD, 65.5 ± 15.4 mmHg) than in controls (99.1 ± 7.8 mmHg, P<.001). The alveolar-arterial oxygen gradient was significantly higher in IPF (50.1 ± 17.3 mmHg) than in controls (17.5 ± 4.9 mmHg, P<.001). In HRCT, ground glass opacity (GGO) was detected in all IPF dogs, traction bronchiectasis in 4, and honeycombing in 1. Bronchoscopic airway changes were noted in all IPF dogs. On BAL fluid (BALF) cytology, the total cell count (TCC) was higher in IPF dogs, and the numbers but not the percentages of macrophages, neutrophils, and mast cells were increased. On histopathology, multifocal or diffuse interstitial fibrosis, type II pneumocyte hyperplasia, prominent intraalveolar macrophages, distortion of alveolar architecture, and emphysematous change were detected. Conclusion and Clinical Importance: IPF causes substantial hypoxemia. In HRCT, GGO is a consistent finding. IPF dogs have concurrent airway changes and an increase in BALF TCC.
Inherited neurodegenerative disorders are debilitating diseases that occur across different species. We have performed clinical, pathological and genetic studies to characterize a novel canine ...neurodegenerative disease present in the Lagotto Romagnolo dog breed. Affected dogs suffer from progressive cerebellar ataxia, sometimes accompanied by episodic nystagmus and behavioral changes. Histological examination revealed unique pathological changes, including profound neuronal cytoplasmic vacuolization in the nervous system, as well as spheroid formation and cytoplasmic aggregation of vacuoles in secretory epithelial tissues and mesenchymal cells. Genetic analyses uncovered a missense change, c.1288G>A; p.A430T, in the autophagy-related ATG4D gene on canine chromosome 20 with a highly significant disease association (p = 3.8 x 10-136) in a cohort of more than 2300 Lagotto Romagnolo dogs. ATG4D encodes a poorly characterized cysteine protease belonging to the macroautophagy pathway. Accordingly, our histological analyses indicated altered autophagic flux in affected tissues. The knockdown of the zebrafish homologue atg4da resulted in a widespread developmental disturbance and neurodegeneration in the central nervous system. Our study describes a previously unknown canine neurological disease with particular pathological features and implicates the ATG4D protein as an important autophagy mediator in neuronal homeostasis. The canine phenotype serves as a model to delineate the disease-causing pathological mechanism(s) and ATG4D function, and can also be used to explore treatment options. Furthermore, our results reveal a novel candidate gene for human neurodegeneration and enable the development of a genetic test for veterinary diagnostic and breeding purposes.
The production of functional human embryonic stem cell (hESC)-derived neuronal cells is critical for the application of hESCs in treating neurodegenerative disorders. To study the potential ...functionality of hESC-derived neurons, we cultured and monitored the development of hESC-derived neuronal networks on microelectrode arrays. Immunocytochemical studies revealed that these networks were positive for the neuronal marker proteins β-tubulin
III and microtubule-associated protein 2 (MAP-2). The hESC-derived neuronal networks were spontaneously active and exhibited a multitude of electrical impulse firing patterns. Synchronous bursts of electrical activity similar to those reported for hippocampal neurons and rodent embryonic stem cell-derived neuronal networks were recorded from the differentiated cultures until up to 4 months. The dependence of the observed neuronal network activity on sodium ion channels was examined using tetrodotoxin (TTX). Antagonists for the glutamate receptors NMDA D(−)-2-amino-5-phosphonopentanoic acid and AMPA/kainate 6-cyano-7-nitroquinoxaline-2,3-dione, and for GABA
A receptors (−)-bicuculline methiodide modulated the spontaneous electrical activity, indicating that pharmacologically susceptible neuronal networks with functional synapses had been generated. The findings indicate that hESC-derived neuronal cells can generate spontaneously active networks with synchronous communication
in vitro, and are therefore suitable for use in developmental and drug screening studies, as well as for regenerative medicine.
Summary
Background
Mast cells (MCs) are multi‐functional effector cells with an essential role in innate immunity and host defence, and under several pathological conditions, such as allergy. Here, ...we aimed at defining the culture conditions that would allow efficient generation of mature and functional human MCs from their progenitor cells.
Methods
Human peripheral blood‐derived CD34+ progenitor cells were cultured in vitro under serum‐free conditions with human stem cell factor for 9 weeks. Growth and differentiation of the cells into MCs were optimized by selected cytokines and a combination of hypoxic and normoxic conditions. MCs were phenotypically characterized by immunocytochemistry, their preformed mediators were quantified, and their functional ability to degranulate and release histamine was tested.
Results
On average, 20 × 106 mature MCs were generated from 0.5 × 106 progenitor cells during 9 weeks of culture, i.e. at least a 40‐fold increase in cell number was achieved. The mature MCs had oval‐shaped non‐lobular nuclei, contained histamine, heparin, tryptase, chymase, and cathepsin G in their secretory granules, and strongly expressed c‐kit (CD117) and Fc epsilon receptor I on their surface. Histamine release from the cells could be brought about by IgE–anti‐IgE cross‐linkage, compound 48/80, substance P, and anaphylatoxin C3a. The MCs remained functional for several weeks after their maturation.
Conclusion
This study describes an efficient protocol for generating mature MCs from human peripheral blood with a functional phenotype of connective tissue‐type MCs. Use of these cultured human MCs will increase our knowledge and understanding about human MC development and biology in human disease.
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