31st May 2013, Volume 126 Number 1375

ASL derived perfusion network shows correlation with saccadic eye movement measures in Parkinson’s disease, S.D.W. Feng1,2, M.R. MacAskill1,2, T.L. Pitcher1,2, T.R. Melzer1,2,C.F. Graham1,2, J.C. Dalrymple-Alford1,2,4 ,T.J. Anderson1,2,31New Zealand Brain Research Institute, Christchurch; 2Department of Medicine, University of Otago, Christchurch; 3Neurology Department, Christchurch Public Hospital, Christchurch; New Zealand 4Department of Psychology, University of Canterbury, Christchurch, New Zealand

Parkinson’s disease (PD) is a progressive neurodegenerative disorder affecting the motor pathway and cognitive function. ASL (arterial spin labelling) blood flow MRI is a commercially available, non-invasive method to assess perfusion changes in the brain. We have previously established that a perfusion network score correlates with cognition (assessed using the Montreal Cognitive assessment; MoCA) and motor impairment in PD (Unified Parkinson’s disease rating scale; UPDRS: see Melzer et al, Brain, 2011).

Eye movements show reduced amplitude and prolonged reaction times in PD. To further validate this perfusion pattern as a biomarker of PD status, we aim to assess the ASL network score against eye movement parameters derived from saccadic paradigms for the same subjects.

We used ASL to determine cerebral blood flow in 72 PD patients and 30 controls. A perfusion pattern was quantified using PCA and logistic regression and each subject was ascribed a single network score, indicating expression of this perfusion pattern.

Eye movement performance was assessed in the reflexive, predictive, memory guided and anti-saccade paradigms. Reaction time (latency) and initial saccade accuracy (primary gain) were measured for the tasks. The network score was assessed against these measures.

All eight measures correlated significantly with network score (positively for reaction time and negatively for accuracy), with p<0.05 and absolute r values ranging from 0.19 (network score vs predictive latency) to 0.57 (network score vs memory-guided latency).

proc1

Memory guided task:

Left: Primary gain, or initial saccade accuracy decreases with increased network score

Right: Reaction time or latency increases with increased network score

The ASL perfusion network shows a consistent pattern of correlation with eye movement measures in PD. These findings lend further objective support for the potential of an ASL derived perfusion

Reference

Melzer, T. R., Watts, R., MacAskill, M. R., Pearson, J. F., Rueger, S., Pitcher, T., Livingston, L., Graham, C., Keenan, R., Shankaranarayanan, A., Alsop, D. C., Dalrymple-Alford, J. C. & Anderson, T.J. (2011). Arterial spin labeling reveals an abnormal cerebral perfusion pattern in Parkinson’s disease. Brain, 134(3), 845-855.

The trace amine-associated receptor 1 partial agonist, RO5203648, prevents methamphetamine-induced stimulation and methamphetamine self-administration, R Cotter1, J O’Leary1, E Pei1, C Ellis1, MC Hoener2, JJ Canales1

1Department of Psychology, University of Canterbury, Christchurch, New Zealand

2F. Hoffmann-La Roche Ltd, Pharmaceuticals Division, Neuroscience Research, Basel, Switzerland

Drug addiction is a debilitating disease of the brain that poses a massive burden to society. There continues to be an enormous unmet need for treatment for drug addiction. Stimulant abuse is particularly challenging according to recent global and national epidemiological studies. The newly discovered trace amine associated receptor 1 (TAAR1) constitutes a novel receptor target for medication development in stimulant addiction. TAAR1 regulates monoamine systems in the brain, especially dopamine, and is activated directly by psychomotor stimulants, including methamphetamine (METH). The aim of the present study was to examine the effects of the newly developed TAAR1 partial agonist, RO5203648, in rodent models of METH addiction. In experiment 1, rats were administered different doses of RO5203648 (0, 1.67, 5 mg/kg i.p.) followed by METH treatment (0, 0.75, 2 mg/kg i.p.). Locomotor activity was monitored via automatised video tracking system (Viewpoint 2.5, France) in an open field. Results showed that RO5203648 dose-dependently reduced METH-induced locomotor stimulation and prevented the expression of long-term sensitization. In experiment 2, rats were trained on METH self-administration (0.5 mg/kg/infusion) and once a consistent response was obtained, pre-treatment with RO5203648 (0, 3, 10 mg/kg i.p.) was administered. Results revealed that RO5203648 effectively blocked METH self-administration. In experiment 3, different doses of RO5203648 (0.25, 0.5, 1 mg/kg/infusion) were given as a substitute for METH in the self-administration context. Data showed that RO5203648 did not exhibit METH-like properties in this paradigm. Taken together, these findings indicate that RO5203648 is able to attenuate methamphetamine-related behaviours, including locomotor stimulation, sensitization and self-administration, and underline the enormous potential of TAAR1-based medications for the treatment of stimulant abuse and addiction.

Aortic flow energetics in a porcine study of septic shock, JA Revie1, DJ Stevenson1, JG Chase1, BC Lambermont2, A Ghuysen2, P Kolh2, GM Shaw3, T Desaive21Department of Mechanical Engineering, University of Canterbury, Christchurch, 2Hemodynamic Research Laboratory, University of Liege, Liege Belgium, 3Department of Intensive Care, Christchurch Hospital, Christchurch.

Aortic pressure can be split into two components representing the storage capacitance of the arterial system, called the reservoir pressure (Pres), and flow wave phenomena in the aorta, named excess pressure (Pex). In this model, the arterial system acts as a hydraulic integrator (or reservoir), storing a portion of systolic flow, as volume, for release during diastole. The flow wave component corresponds to pressure generated from resistance aortic flow resistance. In this study, the hydraulic work of the excess (Wex) and reservoir (Wres) pressures were calculated using measurements of the aortic pressure waveform (Pao) and stroke volume (SV) from a porcine study of septic shock.

SV, Pao, and left ventricular pressures and volumes were measured in 4 pig trials. A 30 minute endotoxin infusion was used to induce septic shock. Measurements were recorded every 30 minutes for 4 hours. Further details on the experimental procedure can be found in (1).

34 subject-specific aortic models were identified from the measurements. Hydraulic work (W=∫P.Qdt) for excess and reservoir pressure were calculated and compared to left ventricular work (Wlv), end systolic elastance (Ees), afterload (Ea), and ventricular arterial coupling (Ees/Ea), calculated from ventricular pressure-volume loop analysis.

Total aortic work (Wex+Wres) compared well to Wlv (R2=0.88). However, only a weak relationship of R2 = 0.24 was found between ventricular arterial coupling (Ees/Ea) and Wex/Wres. Although a strong relationship (R2 = 0.76) was noticed between the inverse of afterload (1/Ea) and Wex. As septic shock progressed, Wres decreased, indicating the arterial system loses its ability to store blood for release during diastole, causing a flattening of the diastolic pressure. These results indicate that SV and morphological changes in Pao could be used to track important pathological changes that occur during septic shock.

Reference

Lambermont, B. et al. (2006) Artificial organs, v30, 560-564.

Identification of novel genetic variants as candidate risk factors in BRCAx hereditary breast cancer, NJ Coufal1, kConFab Investigators2, LC Walker11MacKenzie Cancer Research Group, Department of Pathology, University of Otago Christchurch, 2Peter MacCullum Cancer Centre, Melbourne, Australia.

A significant proportion of breast cancers arise in a subset of women who inherit genetic changes that increase the risk of developing the disease. However, for most familial breast cancers, there is a lack of understanding of the genetic basis for the disease. DNA copy number variants (CNVs) are a major source of genetic variation and can overlap genes sequences, triggering a range of biological changes from altered gene expression to a variety of human diseases (1, 2). The contribution of rare CNVs to the development of familial breast cancer remains unclear. This study aims to assess whether the number and/or location of germ-line CNVs is associated with disease risk in breast cancer families.

Genome-wide scans for CNVs were performed using Illumina1M SNP data from 1) 26 breast-cancer affected women from families with a history of the disease, but no mutation in the known breast cancer susceptibility genes (BRCAx), 2) 14 healthy control females, and using CNV data available from the Database of Genomic Variants. Novel case-specific CNVs that overlap candidate risk susceptibility genes have been prioritised using microarray expression and pathway analysis.

We show that the average frequency of predicted CNVs in breast cancer cases compared to the controls was increased approximately 1.2-fold. Furthermore, our study showed a significantly greater proportion of rare putative CNVs were found to contain gene coding regions in BRCAx cases compared with those in healthy controls (16% and 3%, respectively; P=0.003). We also identified rare CNVs that overlapped a total of 34 candidate genes, including ANKRD45, TBC1D5and SLC9A11, that are known to be involved in breast cancer development. CNVs identified as likely candidate moderate risk genetic variants will be validated by QPCR and assessed in family members of each proband carrier identified with the target CNV.

References

1) Schlien, A. et al. (2010) Current Opinion in Oncology v22, 55-63.
2) Stranger, B. E. et al. (2007) Science v315, 848-853.

Glycaemic control through stochastic forecasting: Implementing STAR as the standard of care in Christchurch Hospital ICU, LM Fisk1, JG Chase1, AJ Le Compte, GM Shaw21Centre for Bio-Engineering, Dept of Mechanical Engineering, University of Canterbury, Christchurch, 2Dept of Intensive Care, Christchurch Hospital, Christchurch.

Accurate glycaemic control (AGC) has been achieved in Christchurch Hospital ICU through a model-derived protocol, named SPRINT. Incidence of hypoglycaemia has been significantly reduced, with the added benefit of reduced morbidity and mortality. Stochastic TARgeted (STAR) glycaemic control forecasts changes in model-based insulin sensitivity to calculate a range of glycaemic outcomes for a treatment, creating a risk framework targeting blood glucose (BG) to 4.4-8.0 mmol/L with the aim of improving safety and performance.

Development of STAR was carried out through clinically validated virtual trials, utilizing 371 virtual patients (39,841 hours) from the SPRINT cohort. Performance was compared to clinical SPRINT and was measured as time within target glycaemic bands, and nursing workload was assessed through BG measurement frequency. Number of patients with severe (BG < 2.2mmol/L) and degree of mild (%BG < 4.0mmol/L) hypoglycaemia were used as safety metrics. On the basis of these results, a clinical pilot trial comprising 10 patients (1,486 hours) was carried out.

Incidence of severe hypoglycaemia was reduced from 14 patients in the clinical SPRINT data to 4 in the virtual trial. None were recorded during the clinical trial. Mild hypoglycaemia was reduced from 2.9% to 1.0% (virtual trial), and 1.5% (clinical trial). 91.6% BG in target range in virtual trial and 89.4% in clinical trial showed increased performance (86.0% for SPRINT results) with a reduced workload (16.0 measures/day reduced to 12.0 and 13.5 in the virtual and clinical trials, respectively). The algorithm achieving these results simply ensured the predicted risk of BG<4.4mmol/L was below 5% for a given treatment, while restricting chance of BG rising above 8.0mmol/L to 5% when. With clinical applicability tested, and the expected safety and performance benefits realised in the trial, Christchurch Hospital is implementing STAR as the standard of care in the ICU.

Neopterin synthesis in macrophage derived multinucleated giant cells, HM Burrowes1, BW Hicks2, SP Gieseg1, 1University of Canterbury, Christchurch, New Zealand, 2Deparment of Chemistry, US Air Force Academy, Colorado, USA.

Macrophages form multinucleated giant cells when confronted with foreign particles such as aggregated LDL. These giant cells, produced by the fusion of both macrophages and monocytes, can contain up to 100 nuclei. Giant cells have been shown to be present in atherosclerotic plaques, particularly around cholesterol crystals, which they appear to be endocytosing. In the laboratory, conversion of macrophages and monocytes to giant cells using IL-4 and αtocopherol causes a 200% increase in neopterin release from γ-interferon stimulated cells. Neopterin is the oxidation product of 7,8-dihydroneopterin which is synthesised in interferon-gamma stimulated human macrophage cells. Plasma neopterin levels are strongly correlated in clinical studies with the degree of cardiovascular disease (CVD) within patients. Our laboratory has previously shown that 7,8-dihydroneopterin is a potent intracellular antioxidant and down-regulator of the oxLDL binding scavenger receptor CD36.

Both macrophages and giant cells were prepared from monocytes isolated from whole human blood. Cholesterol was dissolved in 65°C ethanol and left to cool forming crystals. Neopterin and total neopterin was measured by HPLC.

When given cholesterol crystals, both giant cells and macrophages were observed to dissolve the insoluble 10-50 μm cholesterol crystals over a 24 period. The cholesterol crystals did not cause a significant loss in cell viability as measured by MTT reduction and trypan blue exclusion but did appear to cause an increase in cellular oxidant production. De novo synthesised 7,8-dihydroneopterin was observed to be rapidly oxidised directly to neopterin suggesting a different oxidation mechanism to that previous described with known intracellular oxidants such as peroxides. This ongoing study suggests that a significant amount of the neopterin detected in the plasma of CVD patients may be a result of macrophage interaction with cholesterol crystals.

Parental experiences in the neonatal intensive care unit, A Montgomery-Honger1, V E Pritchard12, C Spencer1, N Austin1, E Ballenden1, L Woodward121Canterbury Child Development Research Group, Department of Psychology, University of Canterbury, 2 Van der Veer Institute for Parkinson’s and Brain Research, New Zealand.

Many parents experience high levels of stress after the birth of a very preterm infant admitted to the neonatal intensive care unit (NICU) given the often fragile status of their infant and the numerous medical interventions necessary to stabilize the infant. However, little is known about the role of stressors external to the NICU environment and the perceptions of NICU staff. The aims of the study were: 1) to compare sources of NICU stress between mothers and fathers of VPT infants, 2) to identify daily hassles outside of the NICU, and 3) to describe staff perceptions of NICU stress.

Eleven mothers and 10 fathers of very preterm infants (<32 weeks) admitted to a level III NICU, Christchurch Women’s Hospital, and 23 NICU nurses were interviewed. The Parental Stressors Scale: NICU (PSS: NICU) determined sources of stress among parents. NICU nurses completed an adapted version of the PSS: NICU measured nursing staffs’ perceptions of parents’ experiences. Parents and staff provided information about stressors outside of the NICU that exacerbated the level of stress relating to the NICU experience, such as finances, transport and work.

Mothers reported significantly higher levels of NICU stress than fathers on the “sights and sounds”, “infant appearance”, “loss of parental role” subscales on the PSS: NICU. The most stressful item on the external stressors scale reported by parents was “fitting in everything else I have to do”. Staff perceptions differed from parents’, with NICU nurses perceiving parental experience in the NICU to be more stressful than was actually reported by parents. Findings emphasize the need for increased awareness of NICU-specific and NICU-external factors contributing to parental NICU stress. Research into the extent to which staff perceptions of parent experiences may affect the quality of staff-parent relations in the NICU is also warranted.

Identification of rare DNA copy number variants overlapping mismatch repair pathway genes in endometrial cancer patients and their potential contribution to disease risk, GL Moir-Meyer1, F Lose2, JF Pearson1, Y Tan2, The Australian National Endometrial Cancer Study Group2, Studies of Epidemiology and Risk Factors in Cancer Heredity3, DJ Thompson4, PD Pharoah3,4, AM Dunning3, DF Easton3,4, AB Spurdle2,LC Walker1,1Department of Pathology, University of Otago, Christchurch, 2Genetics and Population Health Division, Queensland Institute of Medical Research, Queensland, Australia3Department of Oncology, University of Cambridge, Strangeways Research Laboratory, Cambridge, UK, 4 Department of Public Health and Primary Care, University of Cambridge, Strangeways Research Laboratory, Cambridge, UK.

Endometrial cancer is the most common gynaecological cancer in New Zealand and the incidence is increasing as the population ages1. Genetic predictors of endometrial cancer risk that allow early detection of the disease are important for prevention and improved management strategies. Mutations in the mismatch repair genes MLH1MSH2MSH6 and PMS2 are known to confer increased risk in a proportion of endometrial cancer cases, and the mutation spectrum includes copy number variants (CNVs). There are several other genes encoding proteins that act in the mismatch repair pathway, but to date the evidence for their involvement in endometrial cancer predisposition is limited. We have utilised an existing large genetic dataset to screen for CNVs in all genes for which there is evidence for a role in the mismatch repair pathway.

Genome-wide scanning of CNVs was performed using Illumina610k single nucleotide polymorphism data from a large cohort of ~1300 endometrioid endometrial cancer cases. Fine-mapping of CNVs was carried out using four CNV calling algorithms (cnvPartition, QuantiSNP, PennCNV and GNOSIS) to increase both sensitivity and specificity of predictions. Novel CNVs were validated by quantitative PCR.

By interrogating the array data, we identified and confirmed deletions that disrupted known endometrial cancer susceptibility genes, MSH2 and MSH6. This genetic information can now be used to facilitate counselling and clinical management of these families. We also identified novel variants in several other mismatch repair pathway genes, including duplications overlappingTGFBR3 and MUTYH, anda deletion in RPA3, that are predicted to disrupt the coding sequence of TGFBR3 and RPA3. Further work is required to establish whether the CNVs are associated with risk of disease. In summary, we have identified novel aberrations in mismatch repair pathway genes that fall outside the standard testing panel for hereditary endometrial cancer.

References:
1. Ferlay, J. et al. (2010) Cancer Incidence in Five Countries, v I-IX. IARC CancerBase No.9, Lyon, France: International Agency for Research on Cancer. Available from: http://ci5.iarc.fr.
2. Win, A. K. et al. (2012) Journal of Clincial Oncology, v30, 958-964.