Faculty Bibliography

Polymorphisms in the glutathione S-transferase (GST) genes, a superfamily that plays a key role in carcinogen metabolism, have been associated with an increased susceptibility to several types of cancer. We wished to evaluate whether variant allelic forms of GST isoenzymes were associated with an increased susceptibility for brain tumors and age of tumor onset. Here, we examined 394 brain tumors (221 adult and 173 pediatric cases consisting of 197 astrocytic and 197 non-astrocytic tumors) to determine the frequency of polymorphisms in the GSTM1, GSTT1, and GSTP1 genes compared to a healthy control population. Our data shows that the frequency of GST polymorphisms varies not only between adult and pediatric patients with brain tumors and healthy controls, but also between different histological subtypes of brain tumors occurring in pediatric patients. We found (i) a statistically significant increase in the frequency of the functional GSTM1 allele in high-grade pediatric astrocytomas (p < 0.002), (ii) a significant increase in the frequency of the rare GSTP1 variant Val114/Val114 in pediatric astrocytomas (p < 0.002), and (iii) a significant increase in the frequency of the rare GSTP1 Val114/Val114 genotype among pediatric tumors showing microsatellite instability (MSI) due to defects in mismatch repair (MMR) proteins (p = 0.003). Our results suggest that GSTM1 and GSTP1 polymorphisms may play a role in brain tumor susceptibility by histological subtype, particularly high-grade pediatric astrocytomas. Moreover, the presence of the genetic modifier GSTP1 Val114/Val14 genotype may begin to define a high-risk genotype for cancer susceptibility in the pediatric population

INI1/hSNF5 is a component of the ATP-dependent chromatin remodeling hSWI/SNF complex and a tumor suppressor gene of aggressive pediatric atypical teratoid and malignant rhabdoid tumors (AT/RT). To understand the molecular mechanisms underlying its tumor suppressor function, we studied the effect of reintroduction of INI1/hSNF5 into AT/RT-derived cell lines such as MON that carry biallelic deletions of the INI1/hSNF5 locus. We demonstrate that expression of INI1/hSNF5 causes G(0)-G(1) arrest and flat cell formation in these cells. In addition, INI1/hSNF5 repressed transcription of cyclin D1 gene in MON, in a histone deacetylase (HDAC)-dependent manner. Chromatin immunoprecipitation studies revealed that INI1/hSNF5 was directly recruited to the cyclin D1 promoter and that its binding correlated with recruitment of HDAC1 and deacetylation of histones at the promoter. Analysis of INI1/hSNF5 truncations indicated that cyclin D1 repression and flat cell formation are tightly correlated. Coexpression of cyclin D1 from a heterologous promoter in MON was sufficient to eliminate the INI1-mediated flat cell formation and cell cycle arrest. Furthermore, cyclin D1 was overexpressed in AT/RT tumors. Our data suggest that one of the mechanisms by which INI1/hSNF5 exerts its tumor suppressor function is by mediating the cell cycle arrest due to the direct recruitment of HDAC activity to the cyclin D1 promoter thereby causing its repression and G(0)-G(1) arrest. Repression of cyclin D1 gene expression may serve as a useful strategy to treat AT/RT

Enhanced expression of tenascin-C (TN-C) at the invasive edges of glioblastoma multiforme in close association with vascular sprouts, suggests a role for TN-C in microvascular cell migration. To test this hypothesis, we studied the migration of endothelial cells in vitro. In an aggregate migration assay, bovine retinal endothelial cells (BRECs) and human umbilical vein endothelial cells spread and migrated similarly on TN-C or fibronectin (FN). In contrast, U251 MG glioma cells migrated less on TN-C than on FN. Morphological features of U251 MG glioma cells on TN-C included poor cell spreading and short processes. In contrast, on FN, U251 MG glioma cells spread and exhibited long radial processes. Using a transmembrane migration assay, we observed that BREC adhesion was similar on TN-C or FN, whereas U251 MG glioma cells adhered better to FN than to TN-C. In addition, BRECs migrated more across the membrane toward regions coated with TN-C than FN, and conversely, U251 MG glioma cells migrated more toward FN than TN-C. Migration of endothelial and glioma cells toward TN-C or FN occurred in a dose-dependent manner and was strongly dependent on cell adhesion. In this assay, ultrastructural study revealed the migrating phenotype of the endothelial cells through the micropores of the membrane and their spread morphology on TN-C. Moreover, in situ hybridization revealed specific expression of TN-C in migrating microvascular cells in a cerebral microvascular ring assay. Finally in a phosphorylation assay, TN-C enhanced focal adhesion kinase phosphorylation of BRECs, but not of U251 MG glioma cells, and FN enhanced focal adhesion kinase phosphorylation of both BRECs and U251 MG cells. The expression of TN-C by migrating endothelial cells and the promotion of endothelial cell adhesion and migration by TN-C suggest a potential role for TN-C in pathological angiogenesis

Embryonal tumours of the central nervous system (CNS) represent a heterogeneous group of tumours about which little is known biologically, and whose diagnosis, on the basis of morphologic appearance alone, is controversial. Medulloblastomas, for example, are the most common malignant brain tumour of childhood, but their pathogenesis is unknown, their relationship to other embryonal CNS tumours is debated, and patients' response to therapy is difficult to predict. We approached these problems by developing a classification system based on DNA microarray gene expression data derived from 99 patient samples. Here we demonstrate that medulloblastomas are molecularly distinct from other brain tumours including primitive neuroectodermal tumours (PNETs), atypical teratoid/rhabdoid tumours (AT/RTs) and malignant gliomas. Previously unrecognized evidence supporting the derivation of medulloblastomas from cerebellar granule cells through activation of the Sonic Hedgehog (SHH) pathway was also revealed. We show further that the clinical outcome of children with medulloblastomas is highly predictable on the basis of the gene expression profiles of their tumours at diagnosis

Four mitochondrial DNA (mtDNA) mutations at nps 3460, 11778, 14484, and 14459 account for roughly 90% of cases of Leber hereditary optic neuropathy (LHON) and are designated as "primary" LHON mutations since they act as major predisposition factors for LHON. Although each primary mutation can arise independently on different mtDNA backgrounds during human evolution, they characteristically do not co-occur in LHON patients. We report here a family with the simultaneous occurrence of the 11778A and 14484C mutations. Neuro-ophthalmological examination of the proband, a nine-year-old Caucasian female, revealed the bilateral optic atrophy, central scotomas, and reduced visual acuity typical of LHON. Her mother had normal appearing optic discs and is today visually asymptomatic. Analysis of the proband blood mtDNA revealed that she harbored both the 11778A (heteroplasmic, 94% mutant) and the 14484C (homoplasmic mutant) mutation. This genotype was maintained in proband lymphoblasts and transmitochondrial cybrids. The mother also had both mutations, with the 14484C mutation homoplasmic in all cell types examined. However, only 31% of her blood mtDNAs carried the 11778 mutation, which segregated to essentially 100% wild-type in lymphoblast and cybrid mtDNA. Complex I-linked respiration and specific enzyme activity were consistently lowest in proband lymphoblast and cybrid mitochondria compared to those from the mother, 11778A patients, 14484C patients, or controls, thus demonstrating both a deleterious synergistic interaction between the 11778A and 14484C mutations and the magnitude of 11778A-associated complex I dysfunction. Remarkably, spontaneous vision recovery occurred in the proband, highlighting the complexities encountered when associating mtDNA genotype and complex I function with LHON expression.

The hypothesis of this study is that the sodium pump complex acts as an intracellular signal-transducing molecule in canine vascular smooth muscle cells through its interaction with other membrane and cytoskeletal proteins. We have demonstrated that 1 nm ouabain induced transactivation of the epidermal growth factor receptor (EGFR), resulting in increased proliferation and bromodeoxyuridine (BrdUrd) uptake. Immunoprecipitation and Western blotting showed that the EGFR and Src were phosphorylated within 5 min of 10(-9) m ouabain stimulation. Both ouabain-induced DNA synthesis (BrdUrd uptake) and MAPK42/44 phosphorylation were inhibited by the Src inhibitor PP2, the EGFR kinase inhibitor AG1478, the tyrosine kinase inhibitor genistein, and the MEK1 inhibitor PD98059. Ouabain concentrations higher than 1 nm had little or no stimulating effect on proliferation or BrdUrd uptake but did minimally activate ERK1/2. Thus, low concentrations of ouabain, which do not inhibit the sodium pump sufficiently to perturb the resting cellular ionic milieu, initiate a transactivational signaling cascade leading to vascular smooth muscle cell proliferation.

OBJECTIVE: To determine any differences in cerebral perfusion pressure in patients with chronic hypertension compared with those with chronic hypertension and superimposed pre-eclampsia. DESIGN: A prospective observational study. SETTING: University hospital clinic and labour and delivery suite. PARTICIPANTS: Fifteen women with chronic hypertension and 15 with superimposed pre-eclampsia. METHODS: Transcranial Doppler ultrasound was used to measure blood velocity in the middle cerebral arteries of the patients. Systemic blood pressure in the brachial artery was measured simultaneously. Middle cerebral artery. resistance index, pulsatility index, and cerebral perfusion pressure were calculated and plotted on the same axes as data from normal pregnant women. Cerebral perfusion pressure values outside of the 5th and 95th centiles were regarded as abnormal. Cerebral perfusion pressure data from the chronic hypertension and superimposed pre-eclampsia groups were also expressed in terms of the number of normative standard deviations from the mean value for normal pregnancy (Multiples of the Standard Deviation: MOS). All studies were conducted before labour, under similar conditions, and before volume expansion or treatment. Statistical analysis was by Student's t test and Fisher's exact test as appropriate with significance set at a two-tailed P<0.05. RESULTS: Patient demographics and blood pressure were not significantly different between the two groups. The resistance index and pulsatility index were not significantly different (neither absolute nor multiples of the standard deviation values). The absolute cerebral perfusion pressure was significantly higher in the patients with superimposed pre-eclampsia. The group of women with superimposed pre-eclampsia had a significantly higher mean value of cerebral perfusion pressure measured as multiples of the standard deviation from the mean value for normal pregnancy, despite there being no blood pressure difference. CONCLUSIONS: Superimposed pre-eclampsia is associated with significantly higher cerebral perfusion pressure measurements compared with women with uncomplicated chronic hypertension. This is not directly related to a higher blood pressure. The difference in cerebral perfusion pressure may be used to speculate upon the pathophysiology of the increased risk for eclampsia seen in patients with superimposed pre-eclampsia.