Differences in Left Ventricular Enlargement Secondary to Chronic Volume Loading Between English Springer Spaniels and Two Similar Sporting Breeds.

INTRODUCTION: English springer spaniels have larger, rounder ventricles than most other breeds. How this geometry impacts responses to volume overload remains unknown. We compared left ventricular size between English springer spaniels and two similarly sized sporting breeds (Border collies and Labrador retrievers) in naturally occurring chronic left ventricular volume loading conditions (mitral regurgitation and patent ductus arteriosus [PDA]) to examine whether differences in remodelling responses exist between these breeds. ANIMALS, MATERIALS AND METHODS: We searched records for cases of mitral regurgitation and PDA in three breeds. We recorded age, sex, presence of congestive heart failure (CHF), body weight and specific echocardiographic variables. We compared normalised measures of left ventricular size between breeds. Cases with CHF were further examined as a separate group. RESULTS: One-hundred-and-ninety-one dogs were included: 110 with degenerative mitral valve disease, 42 with mitral dysplasia and 39 with PDA. One third of all cases had CHF. All measures of left ventricular size were larger in English springer spaniels in mitral regurgitation cases (P<0.001), whereas PDA cases did not differ. English springer spaniels with PDA resulting in CHF had larger systolic dimensions and volumes than similarly affected non-English Springer Spaniel dogs (P=0.003). CONCLUSIONS: English springer spaniels have greater left ventricular dimensions when exposed to chronic mitral regurgitation, compared with Border collies and Labrador retrievers, but not when exposed to volume overload from a PDA. English springer spaniels differ in their left ventricular morphology from two other sporting breeds, supporting previous studies that they have a unique cardiac morphotype.

Echocardiographic reference intervals in healthy UK deerhounds and prevalence of preclinical dilated cardiomyopathy: a prospective, longitudinal study.

BACKGROUND: Sighthounds have high echocardiographic (ECHO) left ventricular volumes. Establishing robust breed-specific ECHO reference intervals (RI) for screening is important. End-diastolic volume index (EDVI), end-systolic volume index (ESVI) and ejection fraction (EF) reference ranges derived by Simpson's method of discs are not available for deerhounds. The influence of sex or body weight (BW) on left ventricular diameter during diastole (LVDd) and systole (LVDs) has never been reported. OBJECTIVES: Prospectively determine ECHO RI and assess prevalence of dilated cardiomyopathy (DCM) in healthy UK deerhounds. ANIMALS: Ninety-nine deerhounds. METHODS: Deerhounds scored on ECHO and ECG variables then classified as normal (NORM), equivocal (EQUIV) or affected (AFF) with DCM. Fifty-nine NORM deerhounds used to determine ECHO RI. RESULTS: Prevalence of DCM was 21.6%. There were significant differences in BW (P<0.001), LVDd (P<0.001) and LVDs (P<0.05) between female and male deerhounds. Cut-off values for EDVI (≥140.2 mL/m2: 79% sensitivity/97% specificity), ESVI (≥71.9 mL/m2: 94.7% sensitivity/94.2% specificity) and EF (≤42.1%: 84.2% sensitivity/92.8% specificity) were proposed to help diagnose DCM. The most reliable ECHO variables to identify AFF dogs were LVDs indexed to BW by allometric scaling and ESVI; one of the least reliable was sphericity index. Ventricular arrhythmias (VA) were identified in 13.6% of the population, with the highest prevalence in AFF deerhounds (42%). CONCLUSIONS: Preclinical DCM in deerhounds is common and VA may be associated with DCM. Healthy deerhounds have higher LVDd, LVDs and EDVI compared with other breeds. This study provides ECHO RIs for deerhounds; sex or BW RIs should be used when screening.

An update on canine cardiomyopathies - is it all in the genes?

Dilated cardiomyopathy is the second most common cardiac disease in dogs and causes considerable morbidity and mortality. Primary dilated cardiomyopathy is suspected to be familial, and genetic loci have been associated with the disease in a number of breeds. Because it is an adult-onset disease, usually with late onset, testing breeding dogs and bitches before breeding for a genetic mutation that could lead to dilated cardiomyopathy would be helpful to prevent disease. There is growing evidence that the genetic basis may be multigenic rather than monogenic in the majority of studied breeds. This review article describes the known genetic aspects of canine dilated cardiomyopathy and the implications of genetic tests on heart testing and the future of veterinary cardiology.

Serum cardiac troponin I concentrations in dogs with generalised seizures.

OBJECTIVES: To determine if serum cardiac troponin I concentrations - measured with both a first-generation assay and a high-sensitivity assay - were greater in dogs with generalised seizures than in controls and to identify clinical variables associated with cardiac troponin I concentration. MATERIALS AND METHODS: Prospective study of 30 dogs with recent generalised seizures and 30 healthy controls. Serum cardiac troponin I concentration was measured using two commercially available assays, and the correlation of clinical factors with concentration was examined. RESULTS: Serum concentrations of cardiac troponin I were higher in dogs that had recent seizures compared to controls when measured by both assays. The predictors most clearly associated with cardiac troponin I concentration were number of seizures and age. Both predictors were positively associated with increasing concentrations of troponin I. CLINICAL SIGNIFICANCE: Serum cardiac troponin I concentration was significantly elevated in dogs that had recent generalised seizures when compared to controls, and concentrations were higher in dogs that experienced more seizures. This association may indicate that generalised seizures are associated with damage to the myocardium.

Serum cardiac troponin I in canine syncope and seizures.

OBJECTIVE: To determine if serum cardiac troponin I (cTnI) concentration distinguishes between cardiogenic syncope and collapsing dogs presenting with either generalized epileptic seizures (both with and without cardiac disease) or vasovagal syncope. ANIMALS: Seventy-nine prospectively recruited dogs, grouped according to aetiology of collapse: generalized epileptic seizures (group E), cardiogenic syncope (group C), dogs with both epileptic seizures and cardiac disease (group B), vasovagal syncope (group V) or unclassified (group U). METHODS: Most patients had ECG (n = 78), echocardiography (n = 78) and BP measurement (n = 74) performed. Dogs with a history of intoxications, trauma, evidence of metabolic disorders or renal insufficiency (based on serum creatinine concentrations >150 µmol/L and urine specific gravity <1.030) were excluded. Serum cTnI concentrations were measured and compared between groups using non-parametric statistical methods. Multivariable regression analysis investigated factors associated with cTnI. Receiver operator characteristic curve analysis examined whether cTnI could identify cardiogenic syncope. RESULTS: Median cTnI concentrations were higher in group C than E (cTnI: 0.165 [0.02-27.41] vs. 0.03 [0.01-1.92] ng/mL; p<0.05). Regression analysis found that serum cTnI concentrations decreased with increasing time from collapse (p=0.015) and increased with increasing creatinine concentration (p=0.028). Serum cTnI diagnosed cardiogenic syncope with a sensitivity of 75% and specificity of 80%. CONCLUSIONS: Serum cTnI concentrations were significantly different between groups C and E. However, due to the overlap in cTnI concentrations between groups cTnI, measurement in an individual is not optimally discriminatory to differentiate cardiogenic syncope from collapse with generalized epileptic seizures (both with and without cardiac disease) or vasovagal syncope.

A structured, group-based diabetes self-management education (DSME) programme for people, families and whanau with type 2 diabetes (T2DM) in New Zealand: an observational study.

BACKGROUND: Group-based diabetes self-management education (DSME) programmes have been shown to be effective. A programme tailored for the unique social and ethnic environment of New Zealand (NZ) was developed using concepts from internationally developed programmes. AIM: To assess the effectiveness of a 6 week New Zealand specific DSME programme. METHODS: In this observational study people with type 2 diabetes (aged 18-80 years) from diverse cultural backgrounds were recruited from primary care. Seventeen groups of six education sessions were run. Clinical data were collected from primary care at baseline, 3, 6 and 9 months. Participants also completed a self-administered questionnaire on diabetes knowledge, and self-management behaviours. RESULTS: 107 participants, mean age 56.7±11.3 years and mean duration of diabetes 7.5±7 years (NZ European (44%), Maori (24%), Pacific (16%) and Indian (16%)), were enrolled. Confidence in self-managing diabetes, regular examination of feet, physical activity levels and smoking rates all improved. Glycaemic control improved between baseline and 6 months (HbA1C 64.9±20.0 mmol/mol to 59.9±13.9 mmol/mol (p<0.05) (baseline 8.07%±1.80, 6 months 7.62%±1.25)), but was no different to baseline at 9 months. Systolic BP reduced from 131.9±16.4 to 127.4±18.2 mmHg (p<0.05) at 6 months, but increased to baseline levels by 9 months. Diastolic BP, triglycerides and urine microalbumin:creatinine ratio were significantly reduced at 3, 6 and 9 months. CONCLUSION: A group-based DSME programme designed specifically for the NZ population was effective at improving aspects of diabetes care at 6 months. The attenuation of these improvements after 6 months suggests a refresher course at that time may be beneficial.

Suspected exercise-induced seizures in a young dog.

A 12-month-old female neutered crossbreed was referred for investigation of seizure-like episodes occurring only at intense exercise. Thorough medical, neurological and cardiac investigations were performed and excluded the most commonly known causes of seizure-like activity. The dog was fitted with an ambulatory electrocardiography device and underwent another exercise-induced seizure. The electrocardiogram during the episode revealed a sinus tachycardia at approximately 300 beats/minute. A video recording of the episode revealed generalised tonic clonic limb activity with jaw chomping and frothing at the mouth typical of seizure activity. Antiepileptic medications were not prescribed and the owner was advised not to exercise the dog intensely. The dog responded well and did not seizure after 12 months of mild-moderate off-lead exercise. As all the seizures in this case were triggered by intense physical activity, it is suggested that this may be a new form of reflex seizure activity.

The persistently variable "background" stratospheric aerosol layer and global climate change.

Recent measurements demonstrate that the "background" stratospheric aerosol layer is persistently variable rather than constant, even in the absence of major volcanic eruptions. Several independent data sets show that stratospheric aerosols have increased in abundance since 2000. Near-global satellite aerosol data imply a negative radiative forcing due to stratospheric aerosol changes over this period of about -0.1 watt per square meter, reducing the recent global warming that would otherwise have occurred. Observations from earlier periods are limited but suggest an additional negative radiative forcing of about -0.1 watt per square meter from 1960 to 1990. Climate model projections neglecting these changes would continue to overestimate the radiative forcing and global warming in coming decades if these aerosols remain present at current values or increase.

Do satellites detect trends in surface solar radiation?

Long-term variations in solar radiation at Earth's surface (S) can affect our climate, the hydrological cycle, plant photosynthesis, and solar power. Sustained decreases in S have been widely reported from about the year 1960 to 1990. Here we present an estimate of global temporal variations in S by using the longest available satellite record. We observed an overall increase in S from 1983 to 2001 at a rate of 0.16 watts per square meter (0.10%) per year; this change is a combination of a decrease until about 1990, followed by a sustained increase. The global-scale findings are consistent with recent independent satellite observations but differ in sign and magnitude from previously reported ground observations. Unlike ground stations, satellites can uniformly sample the entire globe.

Cleavage and purification of intein fusion proteins using the Streptococcus gordonii spex system.

A gram-positive bacterial expression vector using Streptococcus gordonii has been developed for expression and secretion, or surface anchoring of heterologous proteins. This system, termed Surface Protein Expression system or SPEX, has been used to express a variety of surface anchored and secreted proteins. In this study, the Mycobacterium xenopi (Mxe) GyrA intein and chitin binding domain from Bacillus circulans chitinase Al were used in conjunction with SPEX to express a fusion protein to facilitate secretion, cleavage, and purification. Streptococcus gordonii was transformed to express a secreted fusion protein consisting of a target protein with a C-terminal intein and chitin-binding domain. Two target proteins, the C-repeat region of the Streptococcus pyogenes M6 protein (M6) and the nuclease A (NucA) enzyme of Staphylococcus aureus, were expressed and tested for intein cleavage. The secreted fusion proteins were purified from culture medium by binding to chitin beads and subjected to reaction conditions to induce intein self-cleavage to release the target protein. The M6 and NucA fusion proteins were shown to bind chitin beads and elute under cleavage reaction conditions. In addition, NucA demonstrated enzyme activity both before and after intein cleavage.

Inactivation of the srtA gene in Streptococcus gordonii inhibits cell wall anchoring of surface proteins and decreases in vitro and in vivo adhesion.

The srtA gene product, SrtA, has been shown to be required for cell wall anchoring of protein A as well as virulence in the pathogenic bacterium Staphylococcus aureus. There are five major mechanisms for displaying proteins at the surface of gram-positive bacteria (P. Cossart and R. Jonquieres, Proc. Natl. Acad. Sci. USA 97:5013-5015, 2000). However, since many of the known surface proteins of gram-positive bacteria are believed to be exported and anchored via the sortase pathway, it was of interest to determine if srtA plays a similar role in other gram-positive bacteria. To that end, the srtA gene in the human oral commensal organism Streptococcus gordonii was insertionally inactivated. The srtA mutant S. gordonii exhibited a marked reduction in quantity of a specific anchored surface protein. Furthermore, the srtA mutant had reduced binding to immobilized human fibronectin and had a decreased ability to colonize the oral mucosa of mice. Taken together, these results suggest that the activity of SrtA plays an important role in the biology of nonpathogenic as well as pathogenic gram-positive cocci.

Expression and purification of histidine-tagged proteins from the gram-positive Streptococcus gordonii SPEX system.

Streptococcus gordonii (S. gordonii) has been used as a gram-positive bacterial expression vector for secreted or surface-anchored recombinant proteins. Fusion of the gram-positive bacterial N-terminal signal sequence to the target protein is all that is required for efficient export. This system is termed SPEX for Surface Protein EXpression and has been used to express proteins for a variety of uses. In this study, the SPEX system has been further developed by the construction of vectors that express polyhistidine-tagged fusion proteins. SPEX vectors were constructed with an N-terminal or C-terminal histidine tag. The C-repeat region (CRR) from Streptococcus pyogenes M6 protein and the Staphylococcus aureus nuclease A (NucA) enzyme were tested for expression. The fusion proteins were purified using metal affinity chromatography (MAC). Results show that the fusion proteins were expressed and secreted from S. gordonii with the His tag at either the N- or C-terminal position and could be purified using MAC. The M6 fusions retained immunoreactivity after expression and purification as determined by immunoblots and ELISA analyses. In addition, NucA fusions retained functional activity after MAC purification. The M6-His and NucA-His fusions were purified approximately 15- and 10-fold respectively with approximately 30% recovery of protein using MAC. This study shows that the polyhistidine tag in either the N- or C-terminal position is a viable way to purify secreted heterologous proteins from the supernatant of recombinant S. gordonii cultures. This study further illustrates the value of the SPEX system for secreted expression and purification of proteins.

Expression of active monomeric and dimeric nuclease A from the gram-positive Streptococcus gordonii surface protein expression system.

We used the surface protein expression (SPEX) system to express an anchored and a secreted form of staphylococcal nuclease A (NucA) from gram-positive bacteria. NucA is a small ( approximately 18 kDa), extracellular, monomeric enzyme from Staphylococcus aureus. A deletion of amino acids 114-119 causes monomeric NucA to form homodimers. The DNA sequence encoding either wild-type or deletion mutant NucA was cloned via homologous recombination into Streptococcus gordonii. S. gordonii strains expressing either anchored or secreted, monomeric or dimeric NucA were isolated and tested for enzymatic activity using a novel fluorescence enzyme assay. We show that active monomeric and dimeric NucA enzyme can be expressed either anchored on the cell surface or secreted into the culture medium. The activity of the dimer NucA was approximately 100-fold less than the monomer. Secreted and anchored, monomeric NucA migrated on SDS-polyacrylamide gels at approximately 18 or approximately 30 kDa, respectively. In addition, similar to S. aureus NucA, the S. gordonii recombinant NucA enzyme was dependent on CaCl(2) and was heat stable. In contrast, however, the recombinant NucA activity was maximal at pH 7.0-7.5 whereas S. aureus NucA was maximal at pH 9.0. These results show, for the first time, expression of active enzyme and polymeric protein in secreted and anchored forms using SPEX. This further demonstrates the utility of this gram-positive surface protein expression system as a potential commensal bacterial delivery system for active, therapeutic enzymes, biopharmaceuticals, or vaccines.

The value of an aide memoire in the dermatology clinic.

A randomized study was undertaken to investigate the effectiveness of a chart as an aid to memory illustrating the topical medication most frequently prescribed both in the dermatology clinic, and in general practice. One hundred subjects, half new referrals and half follow-ups, were recruited and asked to recall all of the topical preparations that had previously been prescribed for this condition. They were then asked to consult the chart, and any additional medication recognized at this time was noted, along with any clarification of formulation and strength where possible. Despite some limitations, our chart proved to be of considerable value. Thirty-eight patients could identify between three and eight additional preparations. Overall, the mean number of additional preparations recalled per patient was two. Clarification of strength and formulation was achieved by 21 patients. Eight charts were used, and provided additional information at consultation in 87% of subjects interviewed.

Localisation of the imprinted gene neuronatin, Nnat, confirms and refines the location of a second imprinting region on mouse chromosome 2.

Nine regions on six mouse autosomes are subject to imprinting and uniparental inheritance of any one of these regions results in mice with phenotypic anomalies. So far on distal Chromosome (Chr) 2 there is a unique imprinting region between 2H3 and 2H4 associated with two behavioural disorders and neonatal lethality. A maternally imprinted gene, Nnat, has been identified which is expressed in the nervous system and maps to distal Chr 2. Nnat has been excluded as a candidate for either or both the behavioural phenotypes as it lies proximal to the 2H3-2H4 imprinting region. Here we have mapped Nnat to band 2H1 which is at least 18 Mb proximal to the previously described imprinting region. It maps close to agouti, some alleles of which show differential expression according to parental origin. The localisation of Nnat to band H1 confirms and refines the map location of a second imprinting region on mouse Chr 2.

Glomerular-specific imprinting of the mouse gsalpha gene: how does this relate to hormone resistance in albright hereditary osteodystrophy?

The gene for alpha-stimulating guanine-nucleotide binding polypeptide, Gnas, has been considered as a candidate for the imprinting effects ascribed to distal mouse Chromosome (Chr) 2. Its human homologue (GNAS1) appears, from clinical and biochemical studies of patients with Albright hereditary osteodystrophy, to be paternally imprinted. GNAS1 maps to 20q13, a region that shows linkage conservation with distal mouse Chr 2. We have mapped Gnas within the imprinting region on distal Chr 2 by linkage analysis. To establish if Gnas is imprinted, we have looked for expression differences in tissues taken from mice carrying maternal duplication/paternal deficiency for distal Chr 2 (MatDp2) and its reciprocal (PatDp2). RNA in situ hybridization revealed high levels of Gnas mRNA in glomeruli of PatDp2 embryos at late gestation and lower levels in glomeruli of MatDp2 embryos. These results strongly suggest that Gnas is maternally imprinted and suggest that the mouse gene may be imprinted in a manner opposite that predicted in human.

Acetylcholine receptor aggregation at nerve-muscle contacts in mammalian cultures: induction by ventral spinal cord neurons is specific to axons.

We used a novel mammalian coculture system to study ACh receptor (AChR) redistribution and synaptic structure at nerve-muscle contacts. Ventral spinal cord (VSC) neurons were plated on cultures containing extensive myotubes but few fibroblasts. Neurite-induced redistribution of AChRs occurred within 6 hr after plating neurons and was maximal between 36-48 hr. This AChR redistribution appeared in two patterns: (1) AChR density at sites directly apposed to the neurite where neurites crossed preexisting AChR patches was sharply reduced, (2) Newly aggregated AChRs formed swaths lateral to the neurite path. VSC neurons induced more AChR aggregation than hippocampal, superior cervical ganglion and dorsal root ganglion neurons. The 43 and 58 kDa postsynaptic proteins were colocalized with AChR-enriched domains in all VSC neurite-induced aggregates whereas the colocalization of laminin was variable. Electron microscopy of regions with neurite-induced AChR aggregation showed postsynaptic membrane specializations characteristic of developing synapses and, in older cultures, features of more mature synaptic structure. Thus, the coculture system is useful for studying early stages of neuromuscular junction (NMJ) formation. Neurites in these cocultures were identified as axons or dendrites by morphological criteria and by their immunoreactivity for synaptophysin and phosphorylated heavy neurofilament subunits or for microtubule associated protein 2 (MAP2), respectively. Axons showed a 10-fold higher induction of AChR aggregation than did dendrites. Thus, at least one essential signaling molecule necessary for the induction of AChR aggregation at sites of interaction with muscle appears to be expressed in a polarized fashion in developing VSC neurons.

Thirteen genes (Cebpb, E2f1, Tcf4, Cyp24, Pck1, Acra4, Edn3, Kcnb1, Mc3r, Ntsr, Cd40, Plcg1 and Rcad) that probably lie in the distal imprinting region of mouse chromosome 2 are not monoallelically expressed.

Seven imprinted genes are currently known in the mouse but none have been identified yet in the distal imprinting region of mouse Chromosome (Chr) 2, a region which shows striking linkage conservation with human chromosome 20q13. Both maternal duplication/paternal deficiency and its reciprocal for distal Chr 2 lead to mice with abnormal body shapes and behavioural abnormalities. We have tested a number of candidate genes, that are either likely or known to lie within the distal imprinting region, for monoallelic expression. These included 3 genes (Cebpb, E2f1 and Tcf4) that express transcription factors, 2 genes (Cyp24 and Pck1) that are involved in growth, 5 genes (Acra4, Edn3, Kcnb1, Mc3r and Ntsr) where a defect could lead to neurological and probably behavioural problems, and 3 genes (Cd40, Plcg1 and Rcad) that are less obvious candidates but sequence information was available for designing primers to test their expression. On/off expression of each gene was tested by reverse transcription-polymerase chain reaction (RT-PCR) analysis of RNA extracted from tissues of mice with maternal duplication/paternal deficiency and its reciprocal for the distal region of Chr 2. None of the 13 genes is monoallelically expressed in the appropriate tissues before and shortly after birth which suggests that these genes are not imprinted later in development. This study has narrowed down the search for imprinted genes, and valuable information on which genes have been tested for on/off expression is provided. Since there is considerable evidence of conservation of imprinting between mouse and human, we would predict that the 13 genes are not imprinted in human. Five of the genes: E2f1, Tcf4, Kcnb1, Cd40 and Rcad, have not yet been mapped in human. However, because of the striking linkage conservation observed between mouse Chr 2 and human chromosome 20, we would expect these genes to map on human chromosome 20q13.

Absorption of solar radiation by clouds: observations versus models.

There has been a long history of unexplained anomalous absorption of solar radiation by clouds. Collocated satellite and surface measurements of solar radiation at five geographically diverse locations showed significant solar absorption by clouds, resulting in about 25 watts per square meter more global-mean absorption by the cloudy atmosphere than predicted by theoretical models. It has often been suggested that tropospheric aerosols could increase cloud absorption. But these aerosols are temporally and spatially heterogeneous, whereas the observed cloud absorption is remarkably invariant with respect to season and location. Although its physical cause is unknown, enhanced cloud absorption substantially alters our understanding of the atmosphere's energy budget.