Differential association of MUC5AC and CLCA1 expression in small cartilaginous airways of RAO-affected and control horses

REASONS FOR PERFORMING STUDY: Airway mucus accumulation is associated with indoor irritant and allergen exposure in horses with recurrent airway obstruction (RAO). Epidermal growth factor receptor (EGFR) and a chloride channel (calcium activated, family member 1; CLCA1) are key signalling molecules involved in mucin gene expression. OBJECTIVES: We hypothesised that exposure to irritants and aeroallergens would lead to increased expression of the mucin gene eqMUC5AC and increased stored mucosubstance in the airways of RAO-affected horses, associated with increased neutrophils and CLCA1 and EGFR mRNA levels. METHODS: We performed quantitative RT-PCR of eqMUC5AC, CLCA1 and EGFR; volume density measurements of intraepithelial mucosubstances; and cytological differentiation of intraluminal inflammatory cells in small cartilaginous airways from cranial left and right and caudal left and right lung lobes of 5 clinically healthy and 5 RAO-affected horses that had been exposed to indoor stable environment for 5 days before euthanasia. RESULTS: Neutrophils were increased in RAO-affected horses compared to clinically healthy controls. EqMUC5AC mRNA levels were positively correlated with both CLCA1 and EGFR mRNA levels in RAO-affected horses but only with CLCA1 in controls. The relationship between eqMUC5AC and CLCA1 differed in the 2 groups of horses with RAO-affected animals overexpressing CLCA1 in relation to eqMUC5AC. CONCLUSIONS: These data implicate CLCA1 as a signalling molecule in the expression of eqMUC5AC in horses but also suggest differential regulation by CLCA1 and EGFR between horses with RAO and those with milder degrees of airway inflammation.

Magnetic resonance imaging anatomy of the normal equine larynx and pharynx

The purpose of the present study was to describe normal magnetic resonance (MR) imaging anatomy of the equine larynx and pharynx and to present the optimal protocol, sequences, and possible limitations of this examination technique. Using a 0.3 T unit, the laryngeal and pharyngeal regions was imaged in two horses. The protocol consisted of sagittal and transverse T2-weighted (T2w) fast spin echo, transverse T1-weighted (T1w) spin echo, and dorsal high-resolution T1w gradient echo (both pre- and postcontrast enhancement) sequences. Euthanasia was performed at the end of the imaging procedure. Macroscopic anatomy of the cadaver sections were compared with the MR images in transverse, midsagittal, and parasagittal planes. There was good differentiation of anatomic structures, including soft tissues. The laryngeal cartilages, hyoid apparatus, and upper airway muscle groups with their attachments could be clearly identified. However, it was not always possible to delineate individual muscles in each plane. Most useful were both T2w and T1w transverse sequences. Intravenous application of contrast medium was helpful to identify blood vessels. The MR images corresponded with the macroscopic anatomy of cadaver sections.

Intracoronary Optical Coherence Tomography and Histology of Overlapping Everolimus-Eluting Bioresorbable Vascular Scaffolds in a Porcine Coronary Artery Model

OBJECTIVES: This study sought to assess the vascular response of overlapping Absorb stents compared with overlapping newer-generation everolimus-eluting metallic platform stents (Xience V [XV]) in a porcine coronary artery model. BACKGROUND: The everolimus-eluting bioresorbable vascular scaffold (Absorb) is a novel approach to treating coronary lesions. A persistent inflammatory response, fibrin deposition, and delayed endothelialization have been reported with overlapping first-generation drug-eluting stents. METHODS: Forty-one overlapping Absorb and overlapping Xience V (XV) devices (3.0 × 12 mm) were implanted in the main coronary arteries of 17 nonatherosclerotic pigs with 10% overstretch. Implanted coronary arteries were evaluated by optical coherence tomography (OCT) at 28 days (Absorb n = 11, XV n = 7) and 90 days (Absorb n = 11, XV n = 8), with immediate histological evaluation following euthanasia at the same time points. One animal from each time point was evaluated with scanning electron microscopy alone. A total of 1,407 cross sections were analyzed by OCT and 148 cross sections analyzed histologically. RESULTS: At 28 days in the overlap, OCT analyses indicated 80.1% of Absorb struts and 99.4% of XV struts to be covered (p < 0.0001), corresponding to histological observations of struts with cellular coverage of 75.4% and 99.6%, respectively (p < 0.001). Uncovered struts were almost exclusively related to the presence of "stacked" Absorb struts, that is, with a direct overlay configuration. At 90 days, overlapping Absorb and overlapping XV struts demonstrated >99% strut coverage by OCT and histology, with no evidence of a significant inflammatory process, and comparable % volume obstructions. CONCLUSIONS: In porcine coronary arteries implanted with overlapping Absorb or overlapping XV struts, strut coverage is delayed at 28 days in overlapping Absorb, dependent on the overlay configuration of the thicker Absorb struts. At 90 days, both overlapping Absorb and overlapping XV have comparable strut coverage. The implications of increased strut thickness may have important clinical and design considerations for bioresorbable platforms.

Pulmonary Disease due to Mycobacterium tuberculosis in a Horse: Zoonotic Concerns and Limitations of Antemortem Testing.

A case of pulmonary tuberculosis caused by Mycobacterium tuberculosis was diagnosed in a horse. Clinical evaluation performed prior to euthanasia did not suggest tuberculosis, but postmortem examination provided pathological and bacteriological evidence of mycobacteriosis. In the lungs, multiple tuberculoid granulomas communicating with the bronchiolar lumen, pleural effusion, and a granulomatous lymphadenitis involving mediastinal and tracheobronchial lymph nodes were found. Serologic response to M. tuberculosis antigens was detected in the infected horse, but not in the group of 42 potentially exposed animals (18 horses, 14 alpacas, 6 donkeys, and 4 dogs) which showed no signs of disease. Diagnosis of tuberculosis in live horses remains extremely difficult. Four of 20 animal handlers at the farm were positive for tuberculous infection upon follow-up testing by interferon-gamma release assay, indicating a possibility of interspecies transmission of M. tuberculosis.

PANEL DISCUSSION: "Medical Ethics, Eugenics, and the Holocaust"

A panel discussion moderated by Dr. Thomas R. Cole, McGovern Chair in Medical Humanities and Director of the John P. McGovern Center for Humanities and Ethics at the University of Texas Health Science Center in Houston. Panelists include: Rabbi Samuel E. Karff, Rabbi Emeritus of Congregation Beth Israel and Associate Director of the John P. McGovern Center for Humanities and Ethics and Visiting Professor in the Department of Family Medicine at the University of Texas Health Science Center at the Texas Medical Center. Cardinal DiNardo, the second Archbishop of the Archdiocese of Galveston-Houston and the first cardinal archbishop from a diocese in the Southern United States. Dr. Sheldon Rubenfeld, Clinical Professor of Medicine at Baylor College of Medicine. He is Board Certified in Internal Medicine and in Endocrinology, Diabetes, and Metabolism, and is a Fellow in both the American College of Physicians and the American College of Endocrinology. Dr. Rubenfeld has taught "Healing by Killing: Medicine During the Third Reich" for three years and "Jewish Medical Ethics" for seven years at Baylor College of Medicine. He created a six-month program about Medicine and the Holocaust at Holocaust Museum Houston, including an exhibit entitled How Healing Becomes Killing: Eugenics, Euthanasia, Extermination and a series of lectures by distinguished speakers entitled "The Michael E. DeBakey Medical Ethics Lecture Series".

Nitric oxide metabolite production in the cranial cruciate ligament, synovial membrane, and articular cartilage of dogs with cranial cruciate ligament rupture

OBJECTIVE To measure concentrations of nitric oxide metabolites (nitrite-nitrate [NOt]) in cartilage, synovial membrane, and cranial cruciate ligament (CCL) in dogs and evaluate associations with osteoarthritis in dogs with CCL rupture. ANIMALS 46 dogs with CCL rupture and 54 control dogs without joint disease. PROCEDURE Tissue specimens for histologic examination and explant culture were harvested during surgery in the CCL group or immediately after euthanasia in the control group; NOt concentrations were measured in supernatant of explant cultures and compared among dogs with various degrees of osteoarthritis and between dogs with and without CCL rupture. RESULTS Osteoarthritic cartilage had significantly higher NOt concentration (1,171.6 nmol/g) than did healthy cartilage (491.0 nmol/g); NOt concentration was associated with severity of macroscopic and microscopic lesions. Synovial membrane NOt concentration did not differ between dogs with and without CCL rupture. Ruptured CCL produced less NOt than did intact ligaments. In control dogs, NOt concentrations were similar for intact ligaments (568.1 nmol/g) and articular cartilage (491.0 nmol/g). Synthesis of NOt was inhibited substantially by coincubation with inhibitors. CONCLUSIONS AND CLINICAL RELEVANCE Results suggest that NOt in canine joint tissues originates from the inducible nitric oxide synthase pathway. Nitric oxide metabolite production in cartilage was greater in dogs with osteoarthritis than in healthy dogs and was associated with lesion severity, suggesting that nitric oxide inhibitors may be considered as a treatment for osteoarthritis. The CCL produces substantial concentrations of NOt; the importance of this finding is unknown.

The effects of doxycycline on nitric oxide and stromelysin production in dogs with cranial cruciate ligament rupture

OBJECTIVE To investigate the potential of doxycycline to reduce stromelysin and inducible nitric oxide synthase (iNOS) activity in dogs with osteoarthritis (OA) secondary to spontaneous cranial cruciate ligament (CCL) rupture. STUDY DESIGN Prospective, clinical study. ANIMALS Eighty-one dogs with OA secondary to CCL rupture and 54 normal dogs. METHODS Dogs with OA secondary to CCL rupture were divided into 2 groups before surgery. The Doxy-CCl group received 3 to 4 mg/kg doxycycline orally every 24 hours for 7 to 10 days (n = 35). The CCL group received no treatment (n = 46). Synovial fluid, articular cartilage, synovial membrane, and CCL samples were collected during surgery (Doxy-CCL group and CCL group) or immediately after euthanasia from healthy dogs (control group). Synovial fluid samples were examined cytologically. Total nitric oxide (NOt) concentrations were measured in the supernatant of explant cultures of all tissue samples, and stromelysin activity was measured in the supernatant of explant cultures of cartilage. RESULTS NOt concentrations measured in cartilage were significantly lower in the Doxy-CCL group than in the CCL group, but were not different from those measured in the control group. Doxycycline treatment did not have a significant effect on cartilage stromelysin levels. CONCLUSION The findings in this study indicate that doxycycline inhibits NO production in cartilage in dogs with CCL rupture. CLINICAL RELEVANCE Doxycycline may have a role in the treatment of canine OA by inhibiting NO production.

Endogenous nitric oxide production in canine osteoarthritis: Detection in urine, serum, and synovial fluid specimens

OBJECTIVE To measure nitric oxide (NO) concentrations in serum, urine, and synovial fluid (SF) of dogs with naturally occurring cranial cruciate ligament (CCL) rupture and normal dogs, and to compare these with clinical and histologic changes of osteoarthritis (OA). STUDY DESIGN Prospective clinical study including 2 groups of animals selected from the hospital population. ANIMALS Forty-three dogs (CCL group) with OA secondary to CCL rupture; 30 healthy dogs (control group) without CCL rupture. METHODS Serum, urine, and SF were collected before and during surgery in the CCL group or immediately after euthanasia in the control group. Articular cartilage and synovial membrane tissue specimens were prepared for routine histologic examination. The stable end products of NO, total nitrite and nitrate (NOt) activity, were measured in body fluids and compared with macroscopic and histologic degrees of OA. Urinary NOt concentration was compared with urinary creatinine concentration and stated as urinary NOt:creatinine ratio (UNCR). RESULTS-SF NOt concentrations were not significantly different between the 2 groups. Serum NOt concentrations (45.6 vs 28.9 micromol/L; P =.042) and the UNCR (0.007 vs 0.004; P =.035) were significantly higher in dogs of the CCL group compared with the control population. An association between UNCR and histologic and macroscopical OA grades could be demonstrated. CONCLUSION UNCR might be a useful indicator of nitrite and nitrate production and, therefore, osteoarthritic changes in joints. CLINICAL RELEVANCE UNCR could be used as a tool to evaluate the NOt production by joint tissues over time and might therefore provide a method of evaluating the effects of drugs in the control of osteoarthritis.

The role of Acinetobacter baumannii as a nosocomial pathogen for dogs and cats in an intensive care unit.

Acinetobacter baumannii is a nosocomial pathogen associated with high morbidity and mortality in humans. Whereas infections with strains of Acinetobacter species have been reported in various situations, the importance of A baumannii as a nosocomial pathogen in veterinary hospitals has not been studied so far. In this retrospective case series, we describe 17 dogs and 2 cats from which A baumannii had been isolated during a 2 1/2-year period. In 7 dogs, A baumannii induced systemic signs of illness, whereas 12 animals showed signs of local infection. In all animals with systemic infection, and in 2 with localized infection, A baumannii contributed to the death of the animal or contributed to euthanasia; the remaining 8 dogs and both cats recovered. Molecular typing of the isolates with restriction polymorphisms of ribosomal DNA provided evidence of nosocomial spread of this pathogen and for the presence of several strains of A baumannii in the hospital environment.

Hepatic fungal infection in a young beagle with unrecognised hereditary cobalamin deficiency (Imerslund-Gräsbeck syndrome)

A 12-month-old beagle presented for anorexia, pyrexia and vomiting. The dog had been treated intermittently with antibiotics and corticosteroids for inappetence and lethargy since five months of age. Previous laboratory abnormalities included macrocytosis and neutropenia. At presentation, the dog was lethargic, febrile and thin. Laboratory examination findings included anaemia, a left shift, thrombocytopenia, hypoglycaemia and hyperbilirubinaemia. Multiple, small, hypoechoic, round hepatic lesions were observed on abdominal ultrasound. Cytological examination of hepatic fine needle aspirates revealed a fungal infection and associated pyogranulomatous inflammation. The dog's general condition deteriorated despite supportive measures and treatment with fluconazole, and owners opted for euthanasia before hypocobalaminaemia was identified. Subsequent genomic analysis revealed a CUBN:c.786delC mutation in a homozygous state, confirming hereditary cobalamin malabsorption (Imerslund-Gräsbeck syndrome). Similar to human infants, dogs with Imerslund-Gräsbeck syndrome may rarely be presented for infectious diseases, distracting focus from the underlying primary disorder.

Pulmonary Echinococcus multilocularis metastasis in a dog.

A young adult Labrador retriever dog was presented for surgical debulking of hepatic alveolar echinococcosis. Computed tomography detected hepatomegaly with multiple large cavitary masses with extension of tissue from a lesion wall into the caudal vena cava and numerous nodules in all lung lobes. Following euthanasia, histology confirmed parasitic vesicles with granulomatous reaction in all lesions, and polymerase chain reaction (PCR) established the causative agent to be Echinococcus multilocularis. This report is the first to present imaging features of pulmonary E. multilocularis granulomata in a dog.

A Nonsense Variant in COL6A1 in Landseer Dogs with Muscular Dystrophy.

A novel canine muscular dystrophy in Landseer dogs was observed. We had access to five affected dogs from two litters. The clinical signs started at a few weeks of age and the severe progressive muscle weakness led to euthanasia between 5 and 15 months of age. The pedigrees of the affected dogs suggested a monogenic autosomal recessive inheritance of the trait. Linkage and homozygosity mapping indicated two potential genome segments for the causative variant on chromosomes 10 and 31 harboring a total of 4.8 Mb of DNA or 0.2% of the canine genome. Using the illumina sequencing technology we obtained a whole genome sequence from one affected Landseer. Variants were called with respect to the dog reference genome and compared to the genetic variants of 170 control dogs from other breeds. The affected Landseer dog was homozygous for a single private non-synonymous variant in the critical intervals, a nonsense variant in the COL6A1 gene (Chr31:39,303,964G>T; COL6A1:c.289G>T; p.E97*). Genotypes at this variant showed perfect concordance with the muscular dystrophy phenotype in all five cases and more than one thousand control dogs. Variants in the human COL6A1 gene cause Bethlem myopathy or Ullrich congenital muscular dystrophy. We therefore conclude that the identified canine COL6A1 variant is most likely causative for the observed muscular dystrophy in Landseer dogs. Based on the nature of the genetic variant in Landseer dogs and their severe clinical phenotype these dogs represent a model for human Ullrich congenital muscular dystrophy.

A RAB3GAP1 SINE Insertion in Alaskan Huskies with Polyneuropathy, Ocular Abnormalities and Neuronal Vacuolation (POANV) Resembling Human Warburg Micro Syndrome 1 (WARBM1)

We observed a hereditary phenotype in Alaskan Huskies, which was characterized by polyneuropathy with ocular abnormalities and neuronal vacuolation (POANV). The affected dogs developed a progressive severe ataxia, which led to euthanasia between 8 and 16 months of age. The pedigrees were consistent with a monogenic autosomal recessive inheritance. We localized the causative genetic defect to a 4 Mb interval on chromosome 19 by a combined linkage and homozygosity mapping approach. Whole genome sequencing of one affected dog, an obligate carrier and an unrelated control revealed a 218 bp SINE insertion into exon 7 of the RAB3GAP1 gene. The SINE insertion was perfectly associated with the disease phenotype in a cohort of 43 Alaskan Huskies and it was absent from 541 control dogs of diverse other breeds. The SINE insertion induced aberrant splicing and led to a transcript with a greatly altered exon 7. RAB3GAP1 loss-of-function variants in humans cause Warburg Micro Syndrome 1 (WARBM1), which is characterized by additional developmental defects compared to canine POANV, whereas Rab3gap1 deficient mice have a much milder phenotype than either humans or dogs. Thus the RAB3GAP1 mutant Alaskan Huskies provide an interesting intermediate phenotype that may help to better understand the function of RAB3GAP1 in development. Furthermore, the identification of the presumed causative genetic variant will enable genetic testing to avoid the non-intentional breeding of affected dogs.

Evaluation of serum biochemical marker concentrations and survival time in dogs with protein-losing enteropathy

OBJECTIVE: To evaluate serum concentrations of biochemical markers and survival time in dogs with protein-losing enteropathy (PLE). DESIGN: Prospective study. ANIMALS: 29 dogs with PLE and 18 dogs with food-responsive diarrhea (FRD). PROCEDURES: Data regarding serum concentrations of various biochemical markers at the initial evaluation were available for 18 of the 29 dogs with PLE and compared with findings for dogs with FRD. Correlations between biochemical marker concentrations and survival time (interval between time of initial evaluation and death or euthanasia) for dogs with PLE were evaluated. RESULTS: Serum C-reactive protein concentration was high in 13 of 18 dogs with PLE and in 2 of 18 dogs with FRD. Serum concentration of canine pancreatic lipase immunoreactivity was high in 3 dogs with PLE but within the reference interval in all dogs with FRD. Serum α1-proteinase inhibitor concentration was less than the lower reference limit in 9 dogs with PLE and 1 dog with FRD. Compared with findings in dogs with FRD, values of those 3 variables in dogs with PLE were significantly different. Serum calprotectin (measured by radioimmunoassay and ELISA) and S100A12 concentrations were high but did not differ significantly between groups. Seventeen of the 29 dogs with PLE were euthanized owing to this disease; median survival time was 67 days (range, 2 to 2,551 days). CONCLUSIONS AND CLINICAL RELEVANCE: Serum C-reactive protein, canine pancreatic lipase immunoreactivity, and α1-proteinase inhibitor concentrations differed significantly between dogs with PLE and FRD. Most initial biomarker concentrations were not predictive of survival time in dogs with PLE.

Development of post-pump syndrome in a sheep after mitral valve stenting

A one-year-old healthy sheep received an implant stenting the mural ('posterior') leaflet of the mitral valve. The experiment was authorized by the Cantonal Ethical Committee. The surgery was performed on the open, beating heart during cardiopulmonary bypass (CPB). Management of anaesthesia was based on isoflurane with mechanical intermittent positive pressure ventilation (IPPV) of the lungs, combined with intercostal nerve blocks and intravenous fentanyl and lidocaine. Marked cardiovascular depression occurred towards the end of CPB time and required high doses of dopamine, dobutamine, lidocaine and ephedrine to allow for weaning off the CPB pump. Moreover, severe pulmonary dysfunction developed when IPPV was re-initiated after CPB. Hypoxaemia persisted throughout the recovery from general anaesthesia. Multiple organ failure developed gradually during the three postoperative days, leading to euthanasia of the animal. As described in this case, marked lung injury associated with some degree of failure of other vital organs may occur in sheep after CPB. Intraoperative cardiorespiratory complications when weaning-off may indicate the development of 'post-pump syndrome'.