Gene expression analysis on small numbers of invasive cells collected by chemotaxis from primary mammary tumors of the mouse

BACKGROUND: cDNA microarrays have the potential to identify the genes involved in invasion and metastasis. However, when used with whole tumor tissue, the results average the expression patterns of different cell types. We have combined chemotaxis-based cell collection of the invasive subpopulation of cells within the primary tumor with array-based gene expression analysis to identify the genes necessary for the process of carcinoma cell invasion. RESULTS: Invasive cells were collected from live primary tumors using microneedles containing chemotactic growth factors to mimic chemotactic signals thought to be present in the primary tumor. When used with mammary tumors of rats and mice, carcinoma cells and macrophages constitute the invasive cell population. Microbeads conjugated with monoclonal anti-CD11b (Mac-1α) antibodies were used to separate macrophages from carcinoma cells. We utilized PCR-based cDNA amplification from small number of cells and compared it to the quality and complexity of conventionally generated cDNA to determine if amplified cDNA could be used with fidelity for array analysis of this cell population. These techniques showed a very high level of correlation indicating that the PCR based amplification technique yields a cDNA population that resembles, with high fidelity, the original template population present in the small number of cells used to prepare the cDNA for use with the chip. CONCLUSIONS: The specific collection of invasive cells from a primary tumor and the analysis of gene expression in these cells are is now possible. By further comparing the gene expression patterns of cells collected by invasion into microneedles with that of carcinoma cells obtained from the whole primary tumor, the blood, and whole metastatic tumors, genes that contribute to the invasive process in carcinoma cells may be identified

Genetic Background of Patients from a University Medical Center in Manhattan: Implications for Personalized Medicine

Background: The rapid progress currently being made in genomic science has created interest in potential clinical applications; however, formal translational research has been limited thus far. Studies of population genetics have demonstrated substantial variation in allele frequencies and haplotype structure at loci of medical relevance and the genetic background of patient cohorts may often be complex. Methods and Findings: To describe the heterogeneity in an unselected clinical sample we used the Affymetrix 6.0 gene array chip to genotype self-identified European Americans (N = 326), African Americans (N = 324) and Hispanics (N = 327) from the medical practice of Mount Sinai Medical Center in Manhattan, NY. Additional data from US minority groups and Brazil were used for external comparison. Substantial variation in ancestral origin was observed for both African Americans and Hispanics; data from the latter group overlapped with both Mexican Americans and Brazilians in the external data sets. A pooled analysis of the African Americans and Hispanics from NY demonstrated a broad continuum of ancestral origin making classification by race/ethnicity uninformative. Selected loci harboring variants associated with medical traits and drug response confirmed substantial within-and between-group heterogeneity. Conclusion: As a consequence of these complementary levels of heterogeneity group labels offered no guidance at the individual level. These findings demonstrate the complexity involved in clinical translation of the results from genome-wide association studies and suggest that in the genomic era conventional racial/ethnic labels are of little value.National Heart Lung and Blood Institute (NHLBI/NIH)[RO1 HL53353]Andrea and Charles Bronfman Philantropie

To what extent can headteachers be held to account in the practice of social justice leadership?

Internationally, leadership for social justice is gaining prominence as a global travelling theme. This article draws from the Scottish contribution to the International School Leadership Development Network (ISLDN) social justice strand and presents a case study of a relatively small education system similar in size to that of New Zealand, to explore one system's policy expectations and the practice realities of headteachers (principals) seeking to address issues around social justice. Scottish policy rhetoric places responsibility with headteachers to ensure socially just practices within their schools. However, those headteachers are working in schools located within unjust local, national and international contexts. The article explores briefly the emerging theoretical analyses of social justice and leadership. It then identifies the policy expectations, including those within the revised professional standards for headteachers in Scotland. The main focus is on the headteachers' perspectives of factors that help and hinder their practice of leadership for social justice. Macro systems-level data is used to contextualize equity and outcomes issues that headteachers are working to address. In the analysis of the dislocation between policy and reality, the article asks, 'to what extent can headteachers be held to account in the practice of social justice leadership?

Genome-wide association study of primary open-angle glaucoma in continental and admixed African populations.

Primary open angle glaucoma (POAG) is a complex disease with a major genetic contribution. Its prevalence varies greatly among ethnic groups, and is up to five times more frequent in black African populations compared to Europeans. So far, worldwide efforts to elucidate the genetic complexity of POAG in African populations has been limited. We conducted a genome-wide association study in 1113 POAG cases and 1826 controls from Tanzanian, South African and African American study samples. Apart from confirming evidence of association at TXNRD2 (rs16984299; OR[T] 1.20; P = 0.003), we found that a genetic risk score combining the effects of the 15 previously reported POAG loci was significantly associated with POAG in our samples (OR 1.56; 95% CI 1.26-1.93; P = 4.79 × 10-5). By genome-wide association testing we identified a novel candidate locus, rs141186647, harboring EXOC4 (OR[A] 0.48; P = 3.75 × 10-8), a gene transcribing a component of the exocyst complex involved in vesicle transport. The low frequency and high degree of genetic heterogeneity at this region hampered validation of this finding in predominantly West-African replication sets. Our results suggest that established genetic risk factors play a role in African POAG, however, they do not explain the higher disease load. The high heterogeneity within Africans remains a challenge to identify the genetic commonalities for POAG in this ethnicity, and demands studies of extremely large size

Meta-analysis of lipid-traits in Hispanics identifies novel loci, population-specific effects and tissue-specific enrichment of eQTLs

We performed genome-wide meta-analysis of lipid traits on three samples of Mexican and Mexican American ancestry comprising 4,383 individuals and followed up significant and highly suggestive associations in three additional Hispanic samples comprising 7,876 individuals. Genome-wide significant signals were observed in or near CELSR2, ZNF259/APOA5, KANK2/DOCK6 and NCAN/MAU2 for total cholesterol, LPL, ABCA1, ZNF259/APOA5, LIPC and CETP for HDL cholesterol, CELSR2, APOB and NCAN/MAU2 for LDL cholesterol and GCKR, TRIB1, ZNF259/APOA5 and NCAN/MAU2 for triglycerides. Linkage disequilibrium and conditional analyses indicate that signals observed at ABCA1 and LIPC for HDL cholesterol and NCAN/MAU2 for triglycerides are independent of previously reported lead SNP associations. Analyses of lead SNPs from the European Global Lipids Genetics Consortium (GLGC) dataset in our Hispanic samples show remarkable concordance of direction of effects as well as strong correlation in effect sizes. A meta-analysis of the European GLGC and our Hispanic datasets identified five novel regions reaching genome-wide significance: two for total cholesterol (FN1 and SAMM50), two for HDL cholesterol (LOC100996634 and COPB1) and one for LDL cholesterol (LINC00324/CTC1/PFAS). The top meta-analysis signals were found to be enriched for SNPs associated with gene expression in a tissue-specific fashion, suggesting an enrichment of tissue-specific function in lipid-associated loci

Genetic identification of a common collagen disease in Puerto Ricans via identity-by-descent mapping in a health system

Achieving confidence in the causality of a disease locus is a complex task that often requires supporting data from both statistical genetics and clinical genomics. Here we describe a combined approach to identify and characterize a genetic disorder that leverages distantly related patients in a health system and population-scale mapping. We utilize genomic data to uncover components of distant pedigrees, in the absence of recorded pedigree information, in the multi-ethnic BioMe biobank in New York City. By linking to medical records, we discover a locus associated with both elevated genetic relatedness and extreme short stature. We link the gene, COL27A1, with a little-known genetic disease, previously thought to be rare and recessive. We demonstrate that disease manifests in both heterozygotes and homozygotes, indicating a common collagen disorder impacting up to 2% of individuals of Puerto Rican ancestry, leading to a better understanding of the continuum of complex and Mendelian disease

Protein-coding variants implicate novel genes related to lipid homeostasis contributing to body-fat distribution.

Body-fat distribution is a risk factor for adverse cardiovascular health consequences. We analyzed the association of body-fat distribution, assessed by waist-to-hip ratio adjusted for body mass index, with 228,985 predicted coding and splice site variants available on exome arrays in up to 344,369 individuals from five major ancestries (discovery) and 132,177 European-ancestry individuals (validation). We identified 15 common (minor allele frequency, MAF ≥5%) and nine low-frequency or rare (MAF <5%) coding novel variants. Pathway/gene set enrichment analyses identified lipid particle, adiponectin, abnormal white adipose tissue physiology and bone development and morphology as important contributors to fat distribution, while cross-trait associations highlight cardiometabolic traits. In functional follow-up analyses, specifically in Drosophila RNAi-knockdowns, we observed a significant increase in the total body triglyceride levels for two genes (DNAH10 and PLXND1). We implicate novel genes in fat distribution, stressing the importance of interrogating low-frequency and protein-coding variants

BAMBI Regulates Angiogenesis and Endothelial Homeostasis through Modulation of Alternative TGFβ Signaling

BACKGROUND: BAMBI is a type I TGFβ receptor antagonist, whose in vivo function remains unclear, as BAMBI(-/-) mice lack an obvious phenotype. METHODOLOGY/PRINCIPAL FINDINGS: Identifying BAMBI's functions requires identification of cell-specific expression of BAMBI. By immunohistology we found BAMBI expression restricted to endothelial cells and by electron microscopy BAMBI(-/-) mice showed prominent and swollen endothelial cells in myocardial and glomerular capillaries. In endothelial cells over-expression of BAMBI reduced, whereas knock-down enhanced capillary growth and migration in response to TGFβ. In vivo angiogenesis was enhanced in matrigel implants and in glomerular hypertrophy after unilateral nephrectomy in BAMBI(-/-) compared to BAMBI(+/+) mice consistent with an endothelial phenotype for BAMBI(-/-) mice. BAMBI's mechanism of action in endothelial cells was examined by canonical and alternative TGFβ signaling in HUVEC with over-expression or knock-down of BAMBI. BAMBI knockdown enhanced basal and TGFβ stimulated SMAD1/5 and ERK1/2 phosphorylation, while over-expression prevented both. CONCLUSIONS/SIGNIFICANCE: Thus we provide a first description of a vascular phenotype for BAMBI(-/-) mice, and provide in vitro and in vivo evidence that BAMBI contributes to endothelial and vascular homeostasis. Further, we demonstrate that in endothelial cells BAMBI interferes with alternative TGFβ signaling, most likely through the ALK 1 receptor, which may explain the phenotype observed in BAMBI(-/-) mice. This newly described role for BAMBI in regulating endothelial function has potential implications for understanding and treating vascular disease and tumor neo-angiogenesis