Comparative transcriptomic profiling of myxomatous mitral valve disease in the cavalier King Charles spaniel.

BACKGROUND: Almost all elderly dogs develop myxomatous mitral valve disease by the end of their life, but the cavalier King Charles spaniel (CKCS) has a heightened susceptibility, frequently resulting in death at a young age and suggesting that there is a genetic component to the condition in this breed. Transcriptional profiling can reveal the impact of genetic variation through differences in gene expression levels. The aim of this study was to determine whether expression patterns were different in mitral valves showing myxomatous degeneration from CKCS dogs compared to valves from non-CKCS dogs. RESULTS: Gene expression patterns in three groups of canine valves resulted in distinct separation of normal valves, diseased valves from CKCS and diseased valves from other breeds; the latter were more similar to the normal valves than were the valves from CKCS. Gene expression patterns in diseased valves from CKCS dogs were quite different from those in the valves from other dogs, both affected and normal. Patterns in all diseased valves (from CKCS and other breeds) were also somewhat different from normal non-diseased samples. Analysis of differentially expressed genes showed enrichment in GO terms relating to cardiac development and function and to calcium signalling canonical pathway in the genes down-regulated in the diseased valves from CKCS, compared to normal valves and to diseased valves from other breeds. F2 (prothrombin) (CKCS diseased valves compared to normal) and MEF2C pathway activation (CKCS diseased valves compared to non-CKCS diseased valves) had the strongest association with the gene changes. A large number of genes that were differentially expressed in the CKCS diseased valves compared with normal valves and diseased valves from other breeds were associated with cardiomyocytes including CASQ2, TNNI3 and RYR2. CONCLUSION: Transcriptomic profiling identified gene expression changes in CKCS diseased valves that were not present in age and disease severity-matched non-CKCS valves. These genes are associated with cardiomyocytes, coagulation and extra-cellular matrix remodelling. Identification of genes that vary in the CKCS will allow exploration of genetic variation to understand the aetiology of the disease in this breed, and ultimately development of breeding strategies to eliminate this disease from the breed.

Large animal models of cardiovascular disease.

The human cardiovascular system is a complex arrangement of specialized structures with distinct functions. The molecular landscape, including the genome, transcriptome and proteome, is pivotal to the biological complexity of both normal and abnormal mammalian processes. Despite our advancing knowledge and understanding of cardiovascular disease (CVD) through the principal use of rodent models, this continues to be an increasing issue in today's world. For instance, as the ageing population increases, so does the incidence of heart valve dysfunction. This may be because of changes in molecular composition and structure of the extracellular matrix, or from the pathological process of vascular calcification in which bone-formation related factors cause ectopic mineralization. However, significant differences between mice and men exist in terms of cardiovascular anatomy, physiology and pathology. In contrast, large animal models can show considerably greater similarity to humans. Furthermore, precise and efficient genome editing techniques enable the generation of tailored models for translational research. These novel systems provide a huge potential for large animal models to investigate the regulatory factors and molecular pathways that contribute to CVD in vivo. In turn, this will help bridge the gap between basic science and clinical applications by facilitating the refinement of therapies for cardiovascular disease.

Topographical distribution and radiographic pattern of lung lesions in canine eosinophilic bronchopneumopathy.

OBJECTIVES: To evaluate the radiographic lung pattern and topographical distribution in canine eosinophilic bronchopneumopathy. MATERIALS AND METHODS: Medical records were retrospectively reviewed for dogs diagnosed with eosinophilic bronchopneumopathy. Lateral thoracic radiographs were examined for the presence of increased radiopacity, classification of pattern, topography of lung changes (cranioventral, perihilar, caudodorsal, caudoventral) and severity of pulmonary lesions. RESULTS: Forty-four cases were identified with the Labrador retriever being the most commonly affected breed; there was a mean age of 5 years and an equal gender distribution. Coughing was the most common clinical sign. Circulating eosinophilia was present in 39% of dogs, with a mean peripheral eosinophilia of 5.1×109 cells/L and a mean bronchoalveolar lavage fluid eosinophilia of 40%. Eighty percent of dogs had an abnormal lung pattern in at least one of the four lung fields; the remaining had normal thoracic radiographs. The most common patterns were a bronchial and a bronchointerstitial pattern, with 41 and 89% distribution to the caudodorsal lung field, respectively. CLINICAL SIGNIFICANCE: A bronchial and bronchointerstitial pattern are the most common radiographic lung patterns seen in canine eosinophilic bronchopneumopathy with these patterns most frequently topographically distributed to at least the caudodorsal lung field. Furthermore, within the caudodorsal lung field, a bronchointerstitial pattern predominates. This radiographic and topographical finding may allow eosinophilic bronchopneumopathy to take precedence on a differential diagnoses list before confirmatory bronchoalveolar lavage fluid sampling.

Glucocorticoid metabolism and the action of 11 beta-hydroxysteroid dehydrogenase 2 in canine congestive heart failure.

The enzyme 11-beta-hydroxysteroid dehydrogenase isoenzyme 2 (11BHSD2) is responsible for converting the active glucocorticoid cortisol to inactive cortisone and in the renal medulla protects the mineralocorticoid receptor (MR) from activation by cortisol. Derangements in 11BHSD2 activity can result in reduced conversion of cortisol to cortisone, activation of the MR by cortisol and, consequently, sodium and water retention. The objective of this study was to examine glucocorticoid metabolism in canine congestive heart failure (CHF), specifically to evaluate whether renal 11BHSD2 activity and expression were altered. Dogs were prospectively recruited into one of two phases; the first phase (n=56) utilized gas chromatography-tandem mass spectrometry to examine steroid hormone metabolites normalised to creatinine in home-caught urine samples. Total serum cortisol was also evaluated. The second phase consisted of dogs (n=18) euthanased for refractory CHF or for behavioural reasons. Tissue was collected from the renal medulla for examination by quantitative reverse transcription polymerase chain reaction, immunohistochemistry and protein immune-blotting. Heart failure did not change urinary cortisol:cortisone ratio (P=0.388), or modify renal expression (P=0.303), translation (P=0.427) or distribution of 11BHSD2 (P=0.325). However, CHF did increase excretion of 5α-tetrahydrocortisone (P=0.004), α-cortol (P=0.002) and α-cortolone (P=0.009). Congestive heart failure modifies glucocorticoid metabolism in dogs by increasing 5α-reductase and 20α-hydroxysteroid dehydrogenase activity. Differences between groups in age, sex and underlying disease processes may have influenced these results. However, 11BHSD2 does not appear to be a potential therapeutic target in canine CHF.

Colour M-mode tissue Doppler imaging in healthy cats and cats with hypertrophic cardiomyopathy.

OBJECTIVES: To determine whether decreased diastolic and systolic myocardial velocity gradient between the endocardium and the epicardium exist in the left ventricle of cats with hypertrophic cardiomyopathy. METHODS: Myocardial velocity gradient and mean myocardial velocities were measured by colour M-mode tissue Doppler imaging in the left ventricular free wall of 20 normal cats and 17 cats with hypertrophic cardiomyopathy. RESULTS: The peak myocardial velocity gradient (sec(-1)) during the first (E1) (5.71+/-1.75 versus 11.38+/-3.1, P<0.001) and second phase (E2) (3.09+/-1.53 versus 7.02+/-3.1, P=0.005) of early diastole and also the maximum early diastolic myocardial velocity gradient (Emax) (6.12+/-2.1 versus 10.76+/-3.2, P<0.001) were reduced in cats with hypertrophic cardiomyopathy compared with normal cats. Peak myocardial velocity gradient during early systole (Se) was lower in affected cats than in normal cats (6.26+/-2.08 versus 8.67+/-2.83, P=0.006). Affected cats had a lower peak mean myocardial velocities (mm/s) during the two isovolumic periods (IVRb and IVCb) compared with normal cats (2.97+/-6.76 versus 12.74+/-5.5 and 22.28+/-9.96 versus 38.65+/-10.1, P<0.001, respectively). CLINICAL SIGNIFICANCE: This study shows that hypertrophic cardiomyopathy cats have decreased myocardial velocity gradient during both diastole and systole and also altered myocardial motion during the two isovolumic periods. Myocardial velocity gradients recorded by colour M-mode tissue Doppler imaging can discriminate between the healthy and diseased myocardium.

Development and Evaluation of a Tissue-Engineered Fibrin-based Canine Mitral Valve Three-dimensional Cell Culture System.

Myxomatous mitral valve disease is the most common cardiac disease of the dog, but examination of the associated cellular and molecular events has relied on the use of cadaveric valve tissue, in which functional studies cannot be undertaken. The aim of this study was to develop a three-dimensional (3D) cell co-culture model as an experimental platform to examine disease pathogenesis. Mitral valve interstitial (VIC) and endothelial (VEC) cells were cultured from normal and diseased canine (VIC only) valves. VICs were embedded in a fibrin-based hydrogel matrix and one surface was lined with VECs. The 3D static cultures (constructs) were examined qualitatively and semiquantitatively by light microscopy, immunofluorescence microscopy and protein immunoblotting. Some constructs were manipulated and the endothelium damaged, and the response examined. The construct gross morphology and histology demonstrated native tissue-like features and comparable expression patterns of cellular (α-smooth muscle actin [SMA] and embryonic smooth muscle myosin heavy chain [SMemb]) and extracellular matrix associated markers (matrix metalloproteinase [MMP]-1 and MMP-3), reminiscent of diseased valves. There were no differences between constructs containing normal valve VICs and VECs (type 1) and those containing diseased valve VICs and normal valve VECs (type 2). Mechanical manipulation and endothelial damage (type 3) tended to decrease α-SMA and SMemb expression, suggesting reversal of VIC activation, but with retention of SMemb+ cells adjacent to the wounded endothelium consistent with response to injury. Fibrin-based 3D mitral valve constructs can be produced using primary cell cultures derived from canine mitral valves, and show a phenotype reminiscent of diseased valves. The constructs demonstrate a response to endothelial damage indicating their utility as experimental platforms.

Pulsus alternans in English cocker spaniels with dilated cardiomyopathy.

OBJECTIVES: To report the clinical findings associated with pulsus alternans in English cocker spaniels with dilated cardiomyopathy and to review the phenomenon of mechanical alternans, usually found in patients with severe left ventricular systolic dysfunction. METHODS: The case records of 10 English cocker spaniels with dilated cardiomyopathy and pulsus alternans were reviewed. All dogs had been referred in moderate to severe congestive heart failure. Pulsus alternans was clinically recognised by palpation of a regular alternation of femoral pulse amplitude and confirmed by echocardiography in all cases. Pulsus alternans was an intermittent finding in all 10 dogs. RESULTS: Phonocardiographic recordings were obtained in three dogs to document variation in the intensity of the heart sounds, with one case exhibiting absence of the second heart sound in alternate beats. M-mode echocardiography performed in all dogs showed alternation of the pattern of mitral valve diastolic motion. Doppler echocardiography showed marked alternation in stroke volume in the aortic outflow in all cases. It also showed alternation in mitral regurgitation and in the velocity of early ventricular filling. CLINICAL SIGNIFICANCE: Pulsus alternans may be more prevalent in English cocker spaniels than in other breeds, and because of its intermittent nature, its incidence may be higher than that previously reported. It has diagnostic relevance as it usually indicates severe myocardial depression. Echocardiography allows non-invasive detection of mechanical alternans and provides further insights into its pathophysiology and clinical significance.

Myxomatous Degeneration of the Canine Mitral Valve: From Gross Changes to Molecular Events.

Myxomatous mitral valve disease (MMVD) is the single most common acquired heart disease of the dog, but is also of emerging importance in human medicine, with some features of the disease shared between both species. There has been increased understanding of this disease in recent years, with most research aiming to elucidate the cellular and molecular events of disease pathogenesis. For gross and histological changes, much of our understanding is based on historical studies and there has been no comprehensive reappraisal of the pathology of MMVD. This paper reviews the gross, histological, ultrastructural, cellular and molecular changes in canine MMVD.

Pulsed tissue Doppler imaging in normal cats and cats with hypertrophic cardiomyopathy.

Myocardial motion was quantified in normal cats (n = 25) and cats with hypertrophic cardiomyopathy (HCM) (n = 23) using the pulsed tissue Doppler imaging (TDI) technique. A physiologic nonuniformity was documented in the myocardial motion of normal cats, which was detected as higher early diastolic velocities, acceleration, and deceleration in the interventricular septum compared with the left ventricular free wall (LVFW). HCM cats exhibited lower early diastolic velocities, acceleration, and deceleration and also prolonged isovolumic relaxation time compared with normal cats. These differences were detected mainly along the longitudinal axis of the heart. A cutoff value of E' in the LVFW along the longitudinal axis >7.2 cm/s discriminated normal from HCM cats with a sensitivity of 92% and a specificity of 87%. The physiologic nonuniformity of myocardial motion during diastole was lost in affected cats. Systolic impairment (decreased late-systolic velocities in most segments along the longitudinal axis and decreased early systolic acceleration in both mitral annular sites) was evident in HCM cats irrespective of the presence of left ventricular outflow tract obstruction and congestive heart failure. Postsystolic thickening was recorded in the LVFW along the longitudinal axis only in affected cats (n = 6) and was another finding indicative of systolic impairment in the HCM of this species. This study identified both diastolic and systolic impairment in cats with HCM compared with normal cats. The study also documents the normal physiologic nonhomogeneity in myocardial motion in cats and the subsequent loss of this feature in the HCM diseased state.

Prevalence of pancreatic, hepatic and renal microscopic lesions in post-mortem samples from Cavalier King Charles spaniels.

OBJECTIVES: The objective of this study was to describe the prevalence of microscopic pancreatic, hepatic and renal lesions in post-mortem samples from Cavalier King Charles spaniels. METHODS: The prevalence of microscopic lesions was determined by routine histopathology and compared to ante-mortem clinical signs. RESULTS: There was evidence of chronic pancreatitis in 51·9% of the cases, and age correlated with severity. Renal lesions were diagnosed in 52·2% of cases, most of which were inflammatory. Ante-mortem diagnosis of pancreatic and renal disease was 25 and 16·7%, respectively. Primary hepatic lesions were diagnosed in 11·1% of cases; secondary hepatic lesions were diagnosed in 64·8%. CLINICAL SIGNIFICANCE: Pancreatic and renal lesions are common in Cavalier King Charles spaniels, but they have similar rates of hepatic disease as the general population. The increasing prevalence of pancreatic lesions with age suggests that it might be a progressive condition.

Thoracic high-resolution computed tomographic findings in dogs with canine idiopathic pulmonary fibrosis.

OBJECTIVES: To identify features of canine idiopathic pulmonary fibrosis (CIPF) using high-resolution computed tomography (HRCT), and to correlate these features with clinical stages of the disease. METHODS: Ten dogs with signalment, history and clinical signs highly suggestive of CIPF were investigated. Thoracic radiography, haematology and biochemistry profiles, bronchoscopy and bronchoalveolar lavage and HRCT were performed in each case. The dogs were classified as having mild, moderate or severe stages of the disease based on their clinical presentation and radiographic signs. Features identified by HRCT were correlated with these disease stages. RESULTS: HRCT showed a spectrum of pulmonary changes and it was possible to match some of these to the stage of CIPF. CLINICAL SIGNIFICANCE: HRCT is used extensively in humans in the diagnosis and treatment planning of idiopathic pulmonary fibrosis. This preliminary study identifies the high-resolution computed tomographic features of CIPF in a small group of dogs. With further work it may be possible to use HRCT as a non-invasive tool for diagnosing, staging and therapeutically monitoring CIPF.

Efficacy and safet of pimobendan in canine heart failure caused by myxomatous mitral valve disease.

OBJECTIVES: To evaluate the clinical efficacy and safety of pimobendan by comparing it with ramipril over a six-month period in dogs with mild to moderate heart failure (HF) caused by myxomatous mitral valve disease (MMVD). METHODS: This was a prospective randomised, single-blind, parallel-group trial. Client-owned dogs (n = 43) with mild to moderate HF caused by MMVD were randomly assigned to one of two groups, which received either pimobendan (P dogs) or ramipril (R dogs) for six months. The outcome measures studied were: adverse HF outcome, defined as failure to complete the trial as a direct consequence of HF; maximum furosemide dose (mg/kg/day) administered during the study period; and any requirement for additional visits to the clinic as a direct consequence of HF. RESULTS: Treatment with pimobendan was well tolerated compared with treatment with ramipril. P dogs were 25 per cent as likely as R dogs to have an adverse HF outcome (odds ratio 4.09, 95 per cent confidence interval 1.03 to 16.3, P = 0.046). CLINICAL SIGNIFICANCE: R dogs had a higher overall score and thus may have had more advanced disease than P dogs at baseline (P = 0.04). These results should be interpreted cautiously but such a high odds ratio warrants further investigation.

Gene network and canonical pathway analysis in canine myxomatous mitral valve disease: a microarray study.

Myxomatous mitral valve disease (MMVD) is the single most common acquired heart disease of the dog and is particularly common in small pedigree breed dogs such as the Cavalier King Charles spaniel (CKCS). There are limited data on the mitral valve transcriptome and the aim of this study was to use the microarray technology in conjunction with bioinformatics platforms to analyse transcript changes in MMVD in CKCS compared to normal dogs (non-CKCS). Differentially expressed genes (n = 5397) were identified using cut-off settings of fold change, false discovery rate (FDR) and P <0.05. In total, 4002 genes were annotated to a specific transcript in the Affymetrix canine database, and after further filtering, 591 annotated canine genes were identified: 322 (55%) were up-regulated and 269 (45%) were down-regulated. Canine microRNAs (cfa-miR; n = 59) were also identified. Gene ontology and network analysis platforms identified between six and 10 significantly different biological function clusters from which the following were selected as relevant to MMVD: inflammation, cell movement, cardiovascular development, extracellular matrix organisation and epithelial-to-mesenchymal (EMT) transition. Ingenuity Pathway Analysis identified three canonical pathways relevant to MMVD: caveolar-mediated endocytosis, remodelling of epithelial adherens junctions, and endothelin-1 signalling. Considering the biological relevance to MMVD, the gene families of importance with significant difference between groups included collagens, ADAMTS peptidases, proteoglycans, matrix metalloproteinases (MMPs) and their inhibitors, basement membrane components, cathepsin S, integrins, tight junction cell adhesion proteins, cadherins, other matrix-associated proteins, and members of the serotonin (5-HT)/transforming growth factor -ß signalling pathway.

Culture and characterisation of canine mitral valve interstitial and endothelial cells.

Valve interstitial cells (VICs) have an important role in the aetiopathogenesis of myxomatous mitral valve disease (MMVD) in the dog. Furthermore, there is evidence that valve endothelial cells (VECs) also contribute to disease development. In addition to examining native valve tissue to understand MMVD, another strategy is to separately examine VIC and VEC biology under in vitro culture conditions. The aim of this study was to isolate and characterise canine mitral VICs and VECs from normal dog valves using a combination of morphology, immunohistochemistry and reverse transcription PCR (RT-PCR). Canine mitral VECs and VICs were isolated and cultured in vitro. The two cell populations exhibited different morphologies and growth patterns. VECs, but not VICs, expressed the endothelial markers, platelet endothelial cell adhesion molecule (PECAM-1 or CD31) and acetylated low density lipoprotein (Dil-Ac-LDL). Both VECs and VICs expressed vimentin and embryonic non-smooth muscle myosin heavy chain (SMemb), an activated mesenchymal cell marker. The myofibroblast marker, alpha smooth muscle actin (α-SMA), was detected at the mRNA level in both VEC and VIC cultures, but only at the protein level in VIC cultures. The morphological heterogeneity and expression of non-endothelial phenotypic markers in VEC cultures suggested that a mixture of cell types was present, which might be due to cell contamination and/or endothelial-mesenchymal transition (EndoMT). The use of a specific endothelial culture medium for primary VEC cultures enhanced the endothelial properties of the cells and reduced α-SMA and SMemb expression.

Distribution of calcitonin gene-related peptide, vasoactive intestinal polypeptide, neuropeptide Y, substance P and dopamine beta-hydroxylase immunoreactive nerve fibres in the trachea of sheep.

The distribution of nerve fibres containing calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP), neuropeptide Y (NPY), substance P (SP) and the catecholamine enzyme marker dopamine beta-hydroxylase (DBH) in the trachea of sheep was investigated by immunohistochemical methods. A semi-quantitative assessment of the extent of the nerve fibres immunoreactive for the various antigens was also made. Moderate to large numbers of CGRP-like immunoreactive (CGRP-Li) nerve fibres were present in all parts of the trachea including the epithelial layer, lamina propria, smooth muscle, closer to the mucous glands and blood vessels, and in the elastic fibre connective tissue layer. Although SP-Li nerve fibres had a similar distribution to CGRP, they were absent from the epithelial layer and only small numbers of fibres were present in other areas. Moderate numbers of VIP-Li fibres were present in the smooth muscle and close to mucous glands and blood vessels. Moderate numbers of NPY-Li fibres were present in the smooth muscle with smaller numbers close to mucous glands and blood vessels. Large numbers of DBH-Li nerve fibres were present in the smooth muscle and they had a similar distribution to NPY. The presence of both NPY and DBH in most DBH-Li nerve fibres was established by a double-straining technique, but not all the DBH-Li fibres contained NPY. The DBH/NPY-Li nerve fibres in the smooth muscle formed an extensive and dense interconnecting network and were the most common types of nerve fibres observed.

Age-related respiratory responses to substance P in normal sheep.

The in vivo effects of substance P (SP) on respiratory parameters in four different age groups of sheep were examined. Intravenous SP (10(-8) to 5 x 10(-6) mol kg-1 bodyweight) caused a dose-dependent reduction in dynamic compliance and increase in respiratory resistance in all four groups. The bronchoconstrictor response was age-related, with the greatest response occurring in the youngest age group (four to six months). In the oldest group (over four years) there was minimal bronchomotor response to SP, but a dose-dependent apnoea was present. These findings indicate that there is an age-related alteration in the respiratory response to SP in sheep.

The distribution of nerve fibres immunoreactive for vasoactive intestinal peptide, calcitonin gene-related peptide, substance P and dopamine beta-hydroxylase in the normal equine larynx.

The autonomic innervation of the mammalian respiratory system is complex, and involves a wide variety of peptide and non-peptide neurotransmitters which will have an important role in normal laryngeal function and the response to disease. This innervation has been partially described in the horse airway and lung, but there is no information on the equine larynx. This paper describes the expression and distribution of nerve fibres immunoreactive for vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP), substance P (SP) and the adrenergic enzymatic marker dopamine beta-hydroxylase (DBetaH) in the mucosa of the equine larynx. The overall relative density of nerve fibres immunoreactive for the different antigens was VIP>>CGRP>SP>>DBetaH. There were differences in the distribution of nerve fibre types, although each antigen was found in nerve fibres adjacent to blood vessels and mucous glands. VIP -like immunoreactivity (VIP -Li) was particularly extensive in association with mucous glands. SP - and CGRP -like immunoreactivity (SP -Li, CGRP -Li) were also seen close to the epithelium, with occasional nerve fibres coursing beneath and between the epithelial cells. Fragments of SP -Li and CGRP -Li fibres were also present in large nerve fibre bundles and ganglionic cell clusters, but not in the neurons themselves. The density of nerve fibres immunoreactive for DBetaH was very low and restricted to blood vessels and mucous glands. There was marked variation in the density of nerve fibres at the different sites, with the greatest density, particularly for VIP, over the arytenoid cartilage. Immunoreactive nerve fibres were less plentiful over the epiglottis, and the density of all types of nerve fibres was low over the cricoid cartilage. Overall VIP -Li nerve fibres were the most plentiful.

Changes in nasal mucosal innervation in horses with grass sickness.

REASONS FOR PERFORMING STUDY: Equine grass sickness is a dysautonomia characterised by widespread destruction of autonomic ganglia, resulting in the clinical signs of dysphagia, constipation, profuse sweating, tachycardia, rhinitis sicca and high mortality rate. Rhinitis sicca is a common finding in horses with the chronic form and we have postulated that alterations in autonomic innervation of the nasal mucosa might underlie this clinical presentation. OBJECTIVES: In this study, the expression and distribution of nerve fibres immunoreactive for calcitonin gene-related peptide (CGRP), substance P (SP), the general neuronal marker protein gene-product 9.5 (PGP 9.5; ubiquitin) and the intermediate neurofilaments (PAN-N; neurorfilaments L, M and H) in the nasal mucosa of normal horses (n = 10) and horses with EGS (n = 18; acute n = 8, subacute n = 3, chronic n = 7) was assessed. METHODS: Innervation density and distribution was investigated in the different groups using standard immunohistochemical techniques. RESULTS: Significant differences were noted when comparing the density and distribution of nerve fibres immunoreactive for PGP 95 and PAN-N, with PGP 95 consistently giving better staining in all groups and at all sites in the nasal mucosa. An apparent increase in the density of innervation was noted for acute vs. normal cases. A significant reduction in the density of innervation was noted only with PAN-N when comparing normal horses and acute cases with the chronic group (P < 0.05). CGRP and SP immunoreactive nerve fibres were typically most abundant in the epithelial and subepithelial layers, but the quality of staining and nerve fibre density was greater for SP, achieving significant difference in several comparisons. The density of innervation for SP was significantly reduced in the chronic group compared to the normal and acute groups (P < 0.01). A significant decrease was noted for CGRP only for the acute and chronic groups (P<0.05). CONCLUSIONS: These results demonstrate a reduction in the expression of the sensory neuropeptides in nasal mucosal innervation as a consequence of equine dysautonomia, and may underlie the clinical presentation of rhinitis sicca noted with this disease. POTENTIAL RELEVANCE: Nasal biopsy may be of use in antemortem diagnosis of grass sickness and identification of mucosal denervation; and might also be useful in the treatment of rhinitis in EGS cases.

Peak mean myocardial velocities and velocity gradients measured by color M-mode tissue Doppler imaging in healthy cats.

We sought to assess the feasibility of recording the myocardial velocity gradients (MVGs) and mean myocardial velocities (MMVs) measured by color M-mode tissue Doppler imaging (TDI) in the free wall of unsedated normal cats (n = 18) with a 7.4-MHz probe equipped to record TDI images. The peak MVG and MMV values during the different phases of the cardiac cycle corresponded to certain color velocity patterns occurring in the left ventricular free wall (LVFW). Biphasic shifts were recorded in the tracings of both the MVG and MMV during early diastole (E1 and E2) as well as during the isovolumic relaxation (IVR) and isovolumic contraction (IVC) phases. Stepwise regression analysis showed that age was the only significant predictor for the peak MVG values during the 2nd phase of early diastole (E2) (r = -0.79, r2 = 0.63, and P < .001). The peak late diastolic MVG values were associated positively with age (r = 0.50, r2 = 0.25, and P < .05). The peak MMV values showed a negative association with age during E2 (r = -0.71, r2 = 0.50, and P < .001) as well as during early systole (Se) (r = -0.55, r2 = 0.30, and P < .05) and late systole (SI) (r = -0.62, r2 = 0.39, and P < .01). A positive association was found between age and the peak MMV values during late diastole (r = 0.54, r2 =- 0.29, and P < .05). The MVG values showed cyclic variations consistent with wall thickness changes. The accuracy of velocity determination and the spatial resolution of the system used were validated with a phantom. To our knowledge, this study is the 1st report of the application of this technique to the myocardium of cats,providing insights into the physiology of myocardial motion. It provides reference ranges of the peak MVG and MMV values for future studies of feline myocardial diseases.

Obstructive inflammatory laryngeal disease in three cats.

Three cats with upper respiratory tract obstruction due to laryngeal inflammation are presented. Cervical radiography showed the presence of a soft tissue mass in the laryngeal region in all cases, and laryngoscopy allowed direct visualization of a mass associated with the larynx. Laryngeal samples were obtained by a combination of fine needle aspiration, cutting biopsy forceps, by ventral laryngotomy, and at post-mortem. Histopathology of the laryngeal samples showed the presence of a predominantly granulomatous inflammation, with macrophage and lymphocyte infiltration. One case was euthanased due to severe dyspnoea. The remaining two cases underwent combined medical (corticosteroid and antibiotic) and surgical (permanent tracheostomy or excision of laryngeal tissue by ventral laryngotomy) treatment. One case died of an undetermined cause 15 weeks after surgery while the other case remains clinically well 20 months after diagnosis. Recognition of the existence of granulomatous laryngitis is important as clinical signs and radiographic findings are indistinguishable from laryngeal neoplasia.