Rebecca J. Webster

Regulation of epidermal growth factor receptor signaling in human cancer cells by microRNA-7

The epidermal growth factor receptor (EGFR) is frequently overexpressed in cancer and is an important therapeutic target. Aberrant expression and function of microRNAs have been associated with tumorigenesis. Bioinformatic predictions suggest that the human EGFR mRNA 3′-untranslated region contains three microRNA-7 (miR-7) target sites, which are not conserved across mammals. We found that miR-7 down-regulates EGFR mRNA and protein expression in cancer cell lines (lung, breast, and glioblastoma) via two of the three sites, inducing cell cycle arrest and cell death. Because miR-7 was shown to decrease EGFR mRNA expression, we used microarray analysis to identify additional mRNA targets of miR-7. These included Raf1 and multiple other genes involved in EGFR signaling and tumorigenesis. Furthermore, miR-7 attenuated activation of protein kinase B (Akt) and extracellular signal-regulated kinase 1/2, two critical effectors of EGFR signaling, in different cancer cell lines. These data establish an important role for miR-7 in controlling mRNA expression and indicate that miR-7 has the ability to coordinately regulate EGFR signaling in multiple human cancer cell types.

Regulation of Epidermal Growth Factor Receptor Signaling and Erlotinib Sensitivity in Head and Neck Cancer Cells by miR-7

Elevated expression and activity of the epidermal growth factor receptor (EGFR)/protein kinase B (Akt) signaling pathway is associated with development, progression and treatment resistance of head and neck cancer (HNC). Several studies have demonstrated that microRNA-7 (miR-7) regulates EGFR expression and Akt activity in a range of cancer cell types via its specific interaction with the EGFR mRNA 3′-untranslated region (3′-UTR). In the present study, we found that miR-7 regulated EGFR expression and Akt activity in HNC cell lines, and that this was associated with reduced growth in vitro and in vivo of cells (HN5) that were sensitive to the EGFR tyrosine kinase inhibitor (TKI) erlotinib (Tarceva). miR-7 acted synergistically with erlotinib to inhibit growth of erlotinib-resistant FaDu cells, an effect associated with increased inhibition of Akt activity. Microarray analysis of HN5 and FaDu cell lines transfected with miR-7 identified a common set of downregulated miR-7 target genes, providing insight into the tumor suppressor function of miR-7. Furthermore, we identified several target miR-7 mRNAs with a putative role in the sensitization of FaDu cells to erlotinib. Together, these data support the coordinate regulation of Akt signaling by miR-7 in HNC cells and suggest the therapeutic potential of miR-7 alone or in combination with EGFR TKIs in this disease.

Toward the discrimination of early melanoma from common and dysplastic nevus using fiber optic diffuse reflectance spectroscopy

We describe a study of the discrimination of early melanoma from common and dysplastic nevus using fiber optic diffuse reflectance spectroscopy. Diffuse reflectance spectra in the wavelength range 550 to 1000 nm are obtained using 400-microm core multimode fibers arranged in a six-illumination-around-one-collection geometry with a single fiber-fiber spacing of 470 microm. Spectra are collected at specific locations on 120 pigmented lesions selected by clinicians as possible melanoma, including 64 histopathologically diagnosed as melanoma. These locations are carried through to the histopathological diagnosis, permitting a spatially localized comparison with the corresponding spectrum. The variations in spectra between groups of lesions with different diagnoses are examined and reduced to features suitable for discriminant analysis. A classifier distinguishing between benign and malignant lesions performs with sensitivity/specificity of between 6469% and 7278%. Classifiers between pairs of the group common nevus, dysplastic nevus, in situ melanoma, and invasive melanoma show better or similar performance than the benign/malignant classifier, and analysis provides evidence that different spectral features are needed for each pair of groups. This indicates that multiple discriminant systems are likely to be required to distinguish between melanoma and similar lesions.

Development of aptitude at altitude

Millions of people currently live at altitudes in excess of 2500 metres, where oxygen supply is limited, but very little is known about the development of brain and behavioural function under such hypoxic conditions. We describe the physiological, cognitive and behavioural profile of a large cohort of infants (6-12 months), children (6-10 years) and adolescents (13-16 years) who were born and are living at three altitude locations in Bolivia ( approximately 500 m, approximately 2500 m and approximately 3700 m). Level of haemoglobin oxygen saturation and end-tidal carbon dioxide were significantly lower in all age groups living above 2500 metres, confirming the presence of hypoxia and hypocapnia, but without any detectable detriment to health. Infant measures of neurodevelopment and behaviour yielded comparable results across altitude groups. Neuropsychological assessment in children and adolescent groups indicated a minor reduction in psychomotor speed with increasing altitude, with no effect of age. This may result from slowing of underlying brain activity in parallel with reduced cerebral metabolism and blood flow, evidenced here by reduced cerebral blood flow velocity, particularly in the basilar artery, in children and adolescents. The proportion of European, Native American and African genetic admixture was comparable across altitude groups, suggesting that adaptation to high altitude in these children occurred in response to chronic hypoxic exposure irrespective of ethnic origin. Thus, psychomotor slowing is proposed to be an adaptive rather than a deficient trait, perhaps enabling accuracy of mental activity in hypoxic conditions.

miR-331-3p regulates expression of neuropilin-2 in glioblastoma

Aberrant expression of microRNAs (miRNAs), a class of small non-coding regulatory RNAs, has been implicated in the development and progression of high-grade gliomas. However, the precise mechanistic role of many miRNAs in this disease remains unclear. Here, we investigate the functional role of miR-331-3p in glioblastoma multiforme (GBM). We found that miR-331-3p expression in GBM cell lines is significantly lower than in normal brain, and that transient overexpression of miR-331-3p inhibits GBM cell line proliferation and clonogenic growth, suggesting a possible tumor suppressor role for miR-331-3p in this system. Bioinformatics analysis identified neuropilin-2 (NRP-2) as a putative target of miR-331-3p. Using transfection studies, we validated NRP-2 mRNA as a target of miR-331-3p in GBM cell lines, and show that NRP-2 expression is regulated by miR-331-3p. RNA interference (RNAi) to inhibit NRP-2 expression in vitro decreased the growth and clonogenic growth of GBM cell lines, providing further support for an oncogenic role for NRP-2 in high-grade gliomas. We also show that miR-331-3p inhibits GBM cell migration, an effect due in part to reduced NRP-2 expression. Finally, we identified a significant inverse correlation between miR-331-3p and NRP-2 expression in The Cancer Genome Atlas GBM cohort of 491 patients. Together, our results suggest that a loss of miR-331-3p expression contributes to GBM development and progression, at least in part via upregulating NRP-2 expression and increasing cell proliferation and clonogenic growth.

Hospitalisation with Infection, Asthma and Allergy in Kawasaki Disease Patients and Their Families: Genealogical Analysis Using Linked Population Data

Background Kawasaki disease results from an abnormal immunological response to one or more infectious triggers. We hypothesised that heritable differences in immune responses in Kawasaki disease-affected children and their families would result in different epidemiological patterns of other immune-related conditions. We investigated whether hospitalisation for infection and asthma/allergy were different in Kawasaki disease-affected children and their relatives. Methods/Major Findings We used Western Australian population-linked health data from live births (1970–2006) to compare patterns of hospital admissions in Kawasaki disease cases, age- and sex-matched controls, and their relatives. There were 295 Kawasaki disease cases and 598 age- and sex-matched controls, with 1,636 and 3,780 relatives, respectively. Compared to controls, cases were more likely to have been admitted at least once with an infection (cases, 150 admissions (50.8%) vs controls, 210 admissions (35.1%); odds ratio (OR) = 1.9, 95% confidence interval (CI) 1.4–2.6, P = 7.2×10−6), and with asthma/allergy (cases, 49 admissions (16.6%) vs controls, 42 admissions (7.0%); OR = 2.6, 95% CI 1.7–4.2, P = 1.3×10−5). Cases also had more admissions per person with infection (cases, median 2 admissions, 95% CI 1–5, vs controls, median 1 admission, 95% CI 1–4, P = 1.09×10−5). The risk of admission with infection was higher in the first degree relatives of Kawasaki disease cases compared to those of controls, but the differences were not significant. Conclusion Differences in the immune phenotype of children who develop Kawasaki disease may influence the severity of other immune-related conditions, with some similar patterns observed in relatives. These data suggest the influence of shared heritable factors in these families.

Neurophysiological evidence for cognitive and brain functional adaptation in adolescents living at high altitude

OBJECTIVE Neurophysiological methods were used to study the effects of high altitude living on brain functions in a subgroup of participants of the Bolivian Children Living at Altitude (BoCLA) project. METHODS Electroencephalogram (EEG), event-related potentials (ERP) and cerebral blood flow velocity (CBFV) were recorded in two groups of adolescents (aged 13-16 years), living either at sea-level or high altitude (~3700m). RESULTS Neuropsychological testing revealed no deficits in the high altitude group, despite significantly reduced blood oxygen saturation. In agreement, ERPs elicited by oddball target detection and choice reaction time tasks were not different between groups. In contrast, resting state EEG showed reductions in delta and beta frequency amplitudes in adolescents living at high altitude. The EEG attenuations were correlated with lower CBFV, and the EEG group differences diminished during task performance. CONCLUSIONS No indication was found for negative sequelae of chronic hypoxia in adolescents born and living at an altitude of ~3700m, rather evidence for successful neurophysiological adaptation was found under such conditions. SIGNIFICANCE Dynamic regulation of metabolic demand is one adaptive mechanism that preserves cognitive development at high altitude.

The longitudinal association of common susceptibility variants for type 2 diabetes and obesity with fasting glucose level and BMI.

BackgroundVariation in the effects of genetic variants on physiological traits over time or with age may alter the trajectories of these traits. However, few studies have investigated this possibility for variants associated with type 2 diabetes or obesity, and these show little consensus. We aimed to characterise the possible longitudinal associations of common diabetes-susceptibility variants in the KCNJ11, PPARG, TCF7L2, IGF2BP2, CDKAL1, SLC30A8 and HHEX gene loci, with fasting glucose level; and of an obesity-associated variant in the FTO gene, with body mass index (BMI).MethodsThe study analysed data from the Busselton Health Study (n = 4,554). Cross-sectional association analyses included family data and used the total association test. Longitudinal association analyses of unrelated participant data (n = 2,864) used linear mixed-effects models.ResultsIn cross-sectional analyses, we observed associations of the T allele at the IGF2BP2 single nucleotide polymorphism (SNP) rs4402960 with raised fasting glucose (p = 0.045), and the A allele at the FTO SNP rs9939609 with raised BMI (p = 0.003). Longitudinal analyses showed no significant associations between SNPs and changes in fasting glucose or BMI in the same individuals, either over mean follow-up times of 18.7 and 21.8 years respectively, or with age during adulthood.ConclusionsThere was no indication that the effects of common type 2 diabetes variants on fasting glucose varied with age during adulthood or over time.

Cognitive performance in high-altitude Andean residents compared with low-altitude populations: from childhood to older age.

OBJECTIVES To assess cognition in populations born and living at high altitude (HA; 3,700 m) and low altitude (LA; 500 m) in Bolivia, who were similar for both socioeconomic status and genetic ancestry. To determine whether HA hypoxia influences cognitive decline across the life span. METHOD In total, 191 healthy participants aged 4 to 85 years were assessed at HA (N = 94; 33; 35% male) and LA (N = 97; 46, 47% male) on a battery of cognitive tasks: fluid intelligence, attention, short- and long-term memory, and psychomotor speed. Saliva samples were obtained for evaluation of genetic ancestry. RESULTS HA participants were significantly slower on measures of processing speed and speed of attention than individuals born and living at LA. HA participants had slightly higher percentage of native Andean ancestry than LA participants, but this was not associated with cognitive performance. CONCLUSIONS This is the first study of HA residence and neurocognition across the life span. Given the physiological challenges of HA living, the impact on cognition appears to be subtle and related only to the speed of more complex cognitive operations, rather than to their accuracy. Moreover, the impact on cognition does not appear to differ with increasing age or for different degrees of genetic admixture. Further studies recruiting HA participants with a broader range of native Andean ancestry will help to address the issue of to what extent Amerindian ancestry provides neuroprotection to chronic hypoxia in those living at HA.

Engineering a portable quasi-distributed Fibre-Bragg-Grating temperature sensing system for clinical hyperthermia

We have developed a quasi-distributed temperature sensor consisting of an array of fibre Bragg gratings (FBGs), illuminated by a superluminescent diode (SLD) and interrogated by a fibre Fabry-Perot (FFP) tunable filter. This sensor has been previously tested both on agar-gel tissue phantoms and in vivo on tumours, in rabbit livers that were treated by hyperthermia. The FFP filter is controlled by a piezoelectric transducer operating in an open-loop configuration, and this introduces repeatability and long-term stability issues. Here we report the further development of this system in order to account for the FFP filter issues, and we reduce the noise levels to less than 0.035/spl deg/C rms and the long-term temperature drift below 0.1 /spl deg/C/hr.