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1.  Role of Serine Proteases in the Regulation of Interleukin-877 during the Development of Bronchopulmonary Dysplasia in Preterm Ventilated Infants 
PLoS ONE  2014;9(12):e114524.
Rationale
The chemokine interleukin-8 is implicated in the development of bronchopulmonary dysplasia in preterm infants. The 77-amino acid isoform of interleukin-8 (interleukin-877) is a less potent chemoattractant than other shorter isoforms. Although interleukin-877 is abundant in the preterm circulation, its regulation in the preterm lung is unknown.
Objectives
To study expression and processing of pulmonary interleukin-877 in preterm infants who did and did not develop bronchopulmonary dysplasia.
Methods
Total interleukin-8 and interleukin-877 were measured in bronchoalveolar lavage fluid from preterm infants by immunoassay. Neutrophil serine proteases were used to assess processing. Neutrophil chemotaxis assays and degranulation of neutrophil matrix metalloproteinase-9 were used to assess interleukin-8 function.
Main Results
Peak total interleukin-8 and interleukin-877 concentrations were increased in infants who developed bronchopulmonary dysplasia compared to those who did not. Shorter forms of interleukin-8 predominated in the preterm lung (96.3% No-bronchopulmonary dysplasia vs 97.1% bronchopulmonary dysplasia, p>0.05). Preterm bronchoalveolar lavage fluid significantly converted exogenously added interleukin-877 to shorter isoforms (p<0.001). Conversion was greater in bronchopulmonary dysplasia infants (p<0.05). This conversion was inhibited by α-1 antitrypsin and antithrombin III (p<0.01). Purified neutrophil serine proteases efficiently converted interleukin-877 to shorter isoforms in a time- and dose-dependent fashion; shorter interleukin-8 isoforms were primarily responsible for neutrophil chemotaxis (p<0.001). Conversion by proteinase-3 resulted in significantly increased interleukin-8 activity in vitro (p<0.01).
Conclusions
Shorter, potent, isoforms interleukin-8 predominate in the preterm lung, and are increased in infants developing bronchopulmonary dysplasia, due to conversion of interleukin-877 by neutrophil serine proteases and thrombin. Processing of interleukin-8 provides an attractive therapeutic target to prevent development of bronchopulmonary dysplasia.
doi:10.1371/journal.pone.0114524
PMCID: PMC4256433  PMID: 25474412
2.  Developmental Impact of a Familial GABAA Receptor Epilepsy Mutation 
Annals of neurology  2008;64(3):284-293.
Objective
A major goal of epilepsy research is to understand the molecular and functional basis of seizure genesis. A human GABAA γ2 gene mutation (R43Q) is associated with generalized epilepsy. Introduction of this mutation into a mouse by gene targeting recapitulates the human phenotype demonstrating a strong genotype to phenotype link. GABAA receptors play a role in the moment-to-moment control of brain function and also on the long-term wiring of the brain by directing neuronal development. Our objective was to determine whether developmental expression of the mutation alters seizure susceptibility later in life.
Methods
A tetracycline-based conditional model for activation of a hypomorphic Q43 disease allele was created and validated. Seizure susceptibility was assessed using the subcutaneous pentylenetetrazole model.
Results
Seizure susceptibility was significantly reduced in mice where the Q43 allele was suppressed during development.
Interpretation
These results demonstrate that a human epilepsy-causing mutation impacts network stability during a critical developmental period. These data suggest that identification of presymptomatic children may provide a window for therapeutic intervention before overt symptoms are observed, potentially altering the course of epileptogenesis.
doi:10.1002/ana.21440
PMCID: PMC3707613  PMID: 18825662
3.  Building health systems capacity in global health graduate programs: reflections from Australian educators 
There has been increasing focus on the role of health systems in low and middle-income countries. Despite this, very little evidence exists on how best to build health systems program and research capacity in educational programs. The current experiences in building capacity in health systems in five of the most prominent global health programs at Australian universities are outlined. The strengths and weaknesses of various approaches and techniques are provided along with examples of global practice in order to provide a foundation for future discussion and thus improvements in global health systems education.
doi:10.1186/1472-698X-12-14
PMCID: PMC3489888  PMID: 22920502
4.  Temperature elevation increases GABAA-mediated cortical inhibition in a mouse model of genetic epilepsy 
Epilepsia  2010;52(1):179-184.
Summary
A missense mutation (R43Q) in the γ2 subunit of the GABAA receptor is associated with generalised (genetic) epilepsy with febrile seizures plus (GEFS+). Heterozygous GABAAγ2(R43Q) mice displayed a lower temperature threshold for thermal seizures as compared to wild type littermates. Temperature-dependent internalization of GABAAγ2(R43Q) containing receptors has been proposed as a mechanism underlying febrile seizure genesis in patients with this mutation. We tested this idea using the GABAAγ2(R43Q) knockin mouse model and analysed GABAergic miniature post synaptic inhibitory currents (mIPSCs) in acute brain slices after exposure to varying temperatures. Incubation of slices at an elevated temperature increased mIPSC amplitude in neurons from heterozygous mice with no change seen in wild type controls. [3H]Flumazenil binding measured in whole brain homogenates from mutant and control mice following elevation of body temperature showed no temperature-dependent differences in γ2-containing receptor density. Therefore, in vivo mouse data does not support earlier in vitro observations that proposed temperature-dependent internalization of γ2 R43Q containing GABAA receptors as the cellular mechanism underlying febrile seizure genesis in patients with the GABAAγ2(R43Q) mutation.
doi:10.1111/j.1528-1167.2010.02914.x
PMCID: PMC3354703  PMID: 21219304
Febrile seizures; GABAA receptor; Temperature; Receptor trafficking; Cortex; Mice
5.  Selenium and stereotypies in a mental health setting 
BMJ Case Reports  2009;2009:bcr07.2009.2053.
A 24-year-old man was referred with involuntary stereotypies of movement and thinking, of which he seemed unaware. On admission, comprehensive physical screening proved negative except for lowered selenium concentrations. Neuropsychological assessment revealed changes in performance on tests of attention, non-verbal reasoning and executive function consistent with his stereotypies. A review of his movements characterised them as possibly being consistent with non-paroxysmal kinesigenic dyskinesia. A low dose of sulpiride led to improvements in engagement and insight accompanied by modest reductions in movements.
doi:10.1136/bcr.07.2009.2053
PMCID: PMC3027968  PMID: 22132019
6.  Axon initial segment dysfunction in a mouse model of genetic epilepsy with febrile seizures plus 
The Journal of Clinical Investigation  2010;120(8):2661-2671.
Febrile seizures are a common childhood seizure disorder and a defining feature of genetic epilepsy with febrile seizures plus (GEFS+), a syndrome frequently associated with Na+ channel mutations. Here, we describe the creation of a knockin mouse heterozygous for the C121W mutation of the β1 Na+ channel accessory subunit seen in patients with GEFS+. Heterozygous mice with increased core temperature displayed behavioral arrest and were more susceptible to thermal challenge than wild-type mice. Wild-type β1 was most concentrated in the membrane of axon initial segments (AIS) of pyramidal neurons, while the β1(C121W) mutant subunit was excluded from AIS membranes. In addition, AIS function, an indicator of neuronal excitability, was substantially enhanced in hippocampal pyramidal neurons of the heterozygous mouse specifically at higher temperatures. Computational modeling predicted that this enhanced excitability was caused by hyperpolarized voltage activation of AIS Na+ channels. This heat-sensitive increased neuronal excitability presumably contributed to the heightened thermal seizure susceptibility and epileptiform discharges seen in patients and mice with β1(C121W) subunits. We therefore conclude that Na+ channel β1 subunits modulate AIS excitability and that epilepsy can arise if this modulation is impaired.
doi:10.1172/JCI42219
PMCID: PMC2912193  PMID: 20628201
7.  Relationship of proteinases and proteinase inhibitors with microbial presence in chronic lung disease of prematurity 
Thorax  2010;65(3):246-251.
Background
A proteolytic imbalance has been implicated in the development of “classical” chronic lung disease of prematurity (CLD). However, in “new” CLD this pattern has changed. This study examines the longitudinal relationship between neutrophil proteinases and their inhibitors in ventilated preterm infants and their relationship to microbial colonisation.
Methods
Serial bronchoalveolar lavage fluid was obtained from ventilated newborn preterm infants. Neutrophil elastase (NE) activity, cell counts, metalloproteinase (MMP)-9, MMP-9/TIMP-1 complex, SerpinB1 concentration and percentage of SerpinB1 and α1-antitrypsin (AAT) in complex with elastase were measured. The presence of microbial genes was examined using PCR for 16S rRNA genes.
Results
Statistically more infants who developed CLD had NE activity in at least one sample (10/20) compared with infants with resolved respiratory distress syndrome (RDS) (2/17). However, NE activity was present in a minority of samples, occurring as episodic peaks. Peak levels of MMP-9, MMP-9/TIMP-1 complex, percentage of AAT and SerpinB1 in complex and cell counts were all statistically greater in infants developing CLD than in infants with resolved RDS. Peak values frequently occurred as episodic spikes and strong temporal relationships were noted between all markers. The peak values for all variables were significantly correlated to each other. The presence of bacterial 16S rRNA genes was associated with the development of CLD and with elevated elastase and MMP-9.
Conclusion
NE activity and MMP-9 appear to be important in the development of “new” CLD with both proteinase and inhibitor concentrations increasing episodically, possibly in response to postnatal infection.
doi:10.1136/thx.2009.116061
PMCID: PMC2921268  PMID: 20335295
Chronic lung disease of prematurity; elastase; alpha-1-antitrypsin; metalloproteinases; serpinB1; proteinases; bronchopulmonary dysplasia
8.  International Proficiency Study of a Consensus L1 PCR Assay for the Detection and Typing of Human Papillomavirus DNA: Evaluation of Accuracy and Intralaboratory and Interlaboratory Agreement 
Journal of Clinical Microbiology  2003;41(3):1080-1086.
The PGMY L1 consensus primer pair combined with the line blot assay allows the detection of 27 genital human papillomavirus (HPV) genotypes. We conducted an intralaboratory and interlaboratory agreement study to assess the accuracy and reproducibility of PCR for HPV DNA detection and typing using the PGMY primers and typing amplicons with the line blot (PGMY-LB) assay. A test panel of 109 samples consisting of 29 HPV-negative (10 buffer controls and 19 genital samples) and 80 HPV-positive samples (60 genital samples and 20 controls with small or large amounts of HPV DNA plasmids) were tested blindly in triplicate by three laboratories. Intralaboratory agreement ranged from 86 to 98% for HPV DNA detection. PGMY-LB assay results for samples with a low copy number of HPV DNA were less reproducible. The rate of intralaboratory agreement excluding negative results for HPV typing ranged from 78 to 96%. Interlaboratory reliability for HPV DNA positivity and HPV typing was very good, with levels of agreement of >95% and kappa values of >0.87. Again, low-copy-number samples were more prone to generating discrepant results. The accuracy varied from 91 to 100% for HPV DNA positivity and from 90 to 100% for HPV typing. HPV testing can thus be accomplished reliably with PCR by using a standardized written protocol and quality-controlled reagents. The use of validated HPV DNA detection and typing assays demonstrating excellent interlaboratory agreement will allow investigators to better compare results between epidemiological studies.
doi:10.1128/JCM.41.3.1080-1086.2003
PMCID: PMC150263  PMID: 12624033
9.  The Campbell Collaboration  
BMJ : British Medical Journal  2001;323(7308):294-295.
PMCID: PMC1120915  PMID: 11498472
11.  CHANGES IN CELLULAR ENZYME LEVELS AND EXTRACELLULAR RELEASE OF LYSOSOMAL ACID HYDROLASES IN MACROPHAGES EXPOSED TO GROUP A STREPTOCOCCAL CELL WALL SUBSTANCE 
The Journal of Experimental Medicine  1974;139(5):1262-1282.
Mouse peritoneal macrophages exposed to type-specific polysaccharide and peptidoglycan (PPG) from group A streptococci undergo marked morphologic and biochemical changes. The cells show increases in size and an increased number of lysosomes as demonstrated by vital staining with acridine orange. There are significant elevations in the levels of both lysosomal and nonlysosomal enzymes. Higher doses of PPG cause selective release of the hydrolases into the extracellular environment with no detectable loss of cell viability. The in vitro phenomena may be relevant to understanding the role of macrophages in chronic inflammation.
PMCID: PMC2139663  PMID: 4825245

Results 1-11 (11)