Mark Dunning

Population genomics of human gene expression

Genetic variation influences gene expression, and this variation in gene expression can be efficiently mapped to specific genomic regions and variants. Here we have used gene expression profiling of Epstein-Barr virus–transformed lymphoblastoid cell lines of all 270 individuals genotyped in the HapMap Consortium to elucidate the detailed features of genetic variation underlying gene expression variation. We find that gene expression is heritable and that differentiation between populations is in agreement with earlier small-scale studies. A detailed association analysis of over 2.2 million common SNPs per population (5% frequency in HapMap) with gene expression identified at least 1,348 genes with association signals in cis and at least 180 in trans. Replication in at least one independent population was achieved for 37% of cis signals and 15% of trans signals, respectively. Our results strongly support an abundance of cis-regulatory variation in the human genome. Detection of trans effects is limited but suggests that regulatory variation may be the key primary effect contributing to phenotypic variation in humans. We also explore several methodologies that improve the current state of analysis of gene expression variation.

Superparamagnetic Iron Oxide-Labeled Schwann Cells and Olfactory Ensheathing Cells Can Be Traced In Vivo by Magnetic Resonance Imaging and Retain Functional Properties after Transplantation into the CNS

Schwann cell (SC) and olfactory ensheathing cell (OEC) transplantation has been shown experimentally to promote CNS axonal regeneration and remyelination. To advance this technique into a clinical setting it is important to be able to follow the fates of transplanted cells by noninvasive imaging. Previous studies, using complex modification processes to enable uptake of contrast agents, have shown that cells labeled in vitro with paramagnetic contrast agents transplanted into rodent CNS can be visualized using magnetic resonance imaging (MRI). Here we show that SCs and OECs efficiently internalize dextran-coated superparamagnetic iron oxide (SPIO) from the culture medium by fluid phase pinocytosis. After transplantation into focal areas of demyelination in adult rat spinal cord both transplanted SPIO-labeled SCs and OECs produce a signal reduction using T2-weighted MRI in anesthetized rats that persists for up to 4 weeks. Although signal reduction was discernable after transplantation of unlabelled cells, this is nevertheless distinguishable from that produced by transplanted labeled cells. The region of signal reduction in SPIO-labeled cell recipients correlates closely with areas of remyelination. Because the retention of functional integrity by labeled cells is paramount, we also show that SPIO-labeled SCs and OECs are able to myelinate normally after transplantation into focal areas of demyelination. These studies demonstrate the feasibility of noninvasive imaging of transplanted SCs and OECs and represent a significant step toward the clinical application of promising experimental approaches.

Increasing local levels of neuregulin (glial growth factor-2) by direct infusion into areas of demyelination does not alter remyelination in the rat CNS

Glial growth factor‐2 (GGF‐2) is a neuronally derived isoform of neuregulin shown in vitro to promote proliferation and survival of oligodendrocytes, the myelinating cells of the CNS. Enhanced remyelination has been demonstrated in vivo following systemic delivery of human recombinant GGF‐2 (rhGGF‐2) in experimental autoimmune encephalomyelitis (EAE). However, it is uncertain whether this is the result of direct effects of rhGGF‐2 on cells of the oligodendrocyte lineage or due to modulation of the immune or inflammatory response. If this enhanced remyelination was due to direct effects of rhGGF‐2 on cells of the oligodendrocyte lineage then one would expect rhGGF‐2 to induce a similar proremyelinating response in nonimmune, gliotoxin models of demyelination. Using a gliotoxin model of demyelination we were therefore able to ascertain the in vivo effect of rhGGF‐2 following local CNS delivery in a model that is not confounded by the concurrent presence of an immune‐mediated process. No significant alteration in the rate or character of remyelination was evident following local delivery as compared to controls, and indeed nor following systemic delivery in the gliotoxin model. The results of this study therefore indicate that both direct infusion and systemic delivery of rhGGF‐2 do not alter remyelination in a nonimmune, gliotoxin model of demyelination. This suggests that the proremyelinating effects of systemically delivered rhGGF‐2 in EAE are unlikely to be due to direct effects on the oligodendrocyte lineage, but may be mediated by rhGGF‐2 inducing an environment more favourable to remyelination, possibly through modulation of the immune response.

Comparison of survival after surgical or medical treatment in dogs with a congenital portosystemic shunt

OBJECTIVE To compare survival of dogs with a congenital portosystemic shunt (CPSS) that received medical or surgical treatment. DESIGN Prospective cohort study. ANIMALS 126 client-owned dogs with a single CPSS. PROCEDURES Dogs were examined at 1 of 3 referral clinics, and a single CPSS was diagnosed in each. Dogs received medical or surgical treatment without regard to signalment, clinical signs, or results of hematologic or biochemical analysis. Survival data were analyzed via a Cox regression model. RESULTS During a median follow-up period of 579 days, 18 of 126 dogs died as a result of CPSS. Dogs treated via surgical intervention survived significantly longer than did those treated medically. Hazard ratio for medical versus surgical treatment of CPSS (for the treatment-only model) was 2.9 (95% confidence interval, 1.1 to 7.2). Age at CPSS diagnosis did not affect survival. CONCLUSIONS AND CLINICAL RELEVANCE Both medical and surgical treatment can be used to achieve long-term survival of dogs with CPSS, although results of statistical analysis supported the widely held belief that surgery is preferable to medical treatment. However, the study population consisted of dogs at referral clinics, which suggested that efficacy of medical treatment may have been underestimated. Although surgical intervention was associated with a better chance of long-term survival, medical management provided an acceptable first-line option. Age at examination did not affect survival, which implied that early surgical intervention was not essential. Dogs with CPSS that do not achieve acceptable resolution with medical treatment can subsequently be treated surgically.

Magnetic resonance imaging of functional Schwann cell transplants labelled with magnetic microspheres

There is increasing interest in the use of magnetic resonance imaging (MRI) methods for tracking the fate of labelled cells in vivo post-implantation. The majority of studies have employed cell labels based on nanometer-sized ultrasmall dextran-coated iron oxide particles (USPIO), which are detected through signal hypointensity in T2-weighted images. Although sensitive to MR detection, these labels can be difficult to distinguish from other sources of signal loss in vivo and can be diluted by cell division. Recently, a micron-sized cell label has been described that is much more sensitive to MR detection and which allows detection of single labels in vivo. We show here that glial cells readily take up this label in culture and that the labelled Schwann cells can be detected in vivo by MRI following their implantation into a demyelinated lesion in the rat spinal cord. Signal loss due to the label is sufficiently great that the labelled cells can easily be distinguished from surrounding haemorrhage at the lesion site. Subsequent histological analysis of the lesion area showed that the transplanted cells were remyelinating the demyelinated axons, demonstrating that the labelled cells retained their biological function and that the majority of the label had remained within the transplanted cells.

Exogenous insulin treatment after hypofractionated radiotherapy in cats with diabetes mellitus and acromegaly.

BACKGROUND The optimal treatment for feline acromegaly has yet to be established. Surgical and medical therapies are minimally effective although radiotherapy might have greater efficacy. The purpose of this study was to review the response and outcome of cats with acromegaly and insulin-resistant diabetes mellitus (DM) to radiotherapy. HYPOTHESES That radiotherapy improves glycemic control in cats with acromegaly and that improved glycemic control is due to remission of clinical acromegaly; demonstrated by a fall in serum insulin-like growth factor-1 (IGF-1) concentrations. ANIMALS Fourteen cats with naturally occurring acromegaly. METHODS Retrospective case review; records of all cats treated for acromegaly with radiotherapy were reviewed from 1997 to 2008. Cats were selected on the basis of compatible clinical signs, laboratory features, and diagnostic imaging findings. Fourteen cats received radiotherapy, delivered in 10 fractions, 3 times a week to a total dose of 3,700 cGy. RESULTS Thirteen of 14 cats had improved diabetic control after radiotherapy. These improvements were sustained for up to 60 months. DM progressed in 2 cats and 1 did not respond. Seven cats responded before the final treatment. Ten cats were euthanized, 1 as a consequence of radiotherapy. In 8 cats in which IGF-1 was measured after treatment, changes in its concentration did not reflect the clinical improvement in glycemic control. CONCLUSIONS AND CLINICAL IMPORTANCE Radiotherapy represents an effective treatment for cats with insulin-resistant DM resulting from acromegaly. IGF-1 concentration after treatment does not provide a suitable method by which remission from either acromegaly or insulin-resistant DM may be assessed.

Long-term survival and quality of life in dogs with clinical signs associated with a congenital portosystemic shunt after surgical or medical treatment

OBJECTIVE To compare long-term survival and quality of life data in dogs with clinical signs associated with a congenital portosystemic shunt (CPSS) that underwent medical or surgical treatment. DESIGN Prospective cohort study. ANIMALS 124 client-owned dogs with CPSS. PROCEDURES Dogs received medical or surgical treatment without regard to signalment, clinical signs, or clinicopathologic results. Survival data were analyzed with a Cox regression model. Quality of life information, obtained from owner questionnaires, included frequency of CPSS-associated clinical signs (from which a clinical score was derived), whether owners considered their dog normal, and (for surgically treated dogs) any ongoing medical treatment for CPSS. A Mann-Whitney U test was used to compare mean clinical score data between surgically and medically managed dogs during predetermined follow-up intervals. RESULTS 97 dogs underwent surgical treatment; 27 were managed medically. Median follow-up time for all dogs was 1,936 days. Forty-five dogs (24 medically managed and 21 surgically managed) died or were euthanized during the follow-up period. Survival rate was significantly improved in dogs that underwent surgical treatment (hazard ratio, 8.11; 95% CI, 4.20 to 15.66) than in those treated medically for CPSS. Neither age at diagnosis nor shunt type affected survival rate. Frequency of clinical signs was lower in surgically versus medically managed dogs for all follow-up intervals, with a significant difference between groups at 4 to 7 years after study entry. CONCLUSIONS AND CLINICAL RELEVANCE Surgical treatment of CPSS in dogs resulted in significantly improved survival rate and lower frequency of ongoing clinical signs, compared with medical management. Age at diagnosis did not affect survival rate and should not influence treatment choice.

COX-2 expression and outcome in canine nasal carcinomas treated with hypofractionated radiotherapy.

The expression of cyclooxygenase isoform 2 (COX-2) in canine nasal carcinomas has been well documented. COX-2 expression has proven to be a prognostic factor in several human tumours. The aims of this study were to assess the correlation between immunohistochemical COX-2 expression and prognosis using rhinoscopic biopsies from 42 dogs with nasal carcinomas treated with hypofractionated radiotherapy, and to establish a replicable COX-2 scoring system. Ninety per cent of sections evaluated were COX-2 positive with a mean score of 6.6 (median 8.0; range 0-12). Neither COX-2 expression nor tumour type had a significant correlation with survival. There are likely to be many as yet unidentified variants which contribute to length of survival in dogs with nasal carcinomas. Immunohistochemical COX-2 expression appears unlikely to be of prognostic significance for canine nasal carcinoma.

Multiple Genetic Associations with Irish Wolfhound Dilated Cardiomyopathy

Cardiac disease is a leading cause of morbidity and mortality in dogs and humans, with dilated cardiomyopathy being a large contributor to this. The Irish Wolfhound (IWH) is one of the most commonly affected breeds and one of the few breeds with genetic loci associated with the disease. Mutations in more than 50 genes are associated with human dilated cardiomyopathy (DCM), yet very few are also associated with canine DCM. Furthermore, none of the identified canine loci explain many cases of the disease and previous work has indicated that genotypes at multiple loci may act together to influence disease development. In this study, loci previously associated with DCM in IWH were tested for associations in a new cohort both individually and in combination. We have identified loci significantly associated with the disease individually, but no genotypes individually or in pairs conferred a significantly greater risk of developing DCM than the population risk. However combining three loci together did result in the identification of a genotype which conferred a greater risk of disease than the overall population risk. This study suggests multiple rather than individual genetic factors, cooperating to influence DCM risk in IWH.

Urinary tract infections in small animals: pathophysiology and diagnosis

URINARY tract infections (UTIs) reportedly account for between 5 and 10 per cent of canine and 0-1 to 1 per cent of feline veterinary visits. In this, the first of two articles on the therapeutics of UTIs in dogs and cats, the aetiology and pathophysiology of these conditions are discussed and an approach to the diagnostic work-up is outlined. The second article, to be published in the next issue, focuses on treatment options and the long-term management of problem cases.