BackgroundAbdominal aortic aneurysm (AAA) is a dilatation of the aorta affecting most frequently elderly men. Histologically AAAs are characterized by inflammation, vascular smooth muscle cell apoptosis, and extracellular matrix degradation. The mechanisms of AAA formation, progression, and rupture are currently poorly understood. A previous mRNA expression study revealed a large number of differentially expressed genes between AAA and non-aneurysmal control aortas. MicroRNAs (miRNAs), small non-coding RNAs that are post-transcriptional regulators of gene expression, could provide a mechanism for the differential expression of genes in AAA.MethodsTo determine differences in miRNA levels between AAA (n = 5) and control (n = 5) infrarenal aortic tissues, a microarray study was carried out. Results were adjusted using Benjamini-Hochberg correction (adjusted p < 0.05). Real-time quantitative RT-PCR (qRT-PCR) assays with an independent set of 36 AAA and seven control tissues were used for validation. Potential gene targets were retrieved from miRNA target prediction databases Pictar, TargetScan, and MiRTarget2. Networks from the target gene set were generated and examined using the network analysis programs, CytoScape® and Ingenuity Pathway Core Analysis®.ResultsA microarray study identified eight miRNAs with significantly different expression levels between AAA and controls (adjusted p < 0.05). Real-time qRT-PCR assays validated the findings for five of the eight miRNAs. A total of 222 predicted miRNA target genes known to be differentially expressed in AAA based on a prior mRNA microarray study were identified. Bioinformatic analyses revealed that several target genes are involved in apoptosis and activation of T cells.ConclusionsOur genome-wide approach revealed several differentially expressed miRNAs in human AAA tissue suggesting that miRNAs play a role in AAA pathogenesis.

Objectives: Abdominal aortic aneurysm (AAA), a dilatation of the infrarenal aorta, typically affects males >65 years. The pathobiological mechanisms of human AAA are poorly understood. The goal of this study was to identify novel pathways involved in the development of AAAs. Methods: A custom-designed ‘AAA-chip’ was used to assay 43 of the differentially expressed genes identified in a previously published microarray study between AAA (n = 15) and control (n = 15) infrarenal abdominal aorta. Protein analyses were performed on selected genes. Results: Altogether 38 of the 43 genes on the ‘AAA-chip’ showed significantly different expression. Novel validated genes in AAA pathobiology included ADCY7, ARL4C, BLNK, FOSB, GATM, LYZ, MFGE8, PRUNE2, PTPRC, SMTN, TMODI and TPM2. These genes represent a wide range of biological functions, such as calcium signaling, development and differentiation, as well as cell adhesion not previously implicated in AAA pathobiology. Protein analyses for GATM, CD4, CXCR4, BLNK, PLEK, LYZ, FOSB, DUSP6, ITGA5 and PTPRC confirmed the mRNA findings. Conclusion: The results provide new directions for future research into AAA pathogenesis to study the role of novel genes confirmed here. New treatments and diagnostic tools for AAA could potentially be identified by studying these novel pathways.

Highlights d E93 is required to terminate MB neurogenesis in Drosophila d E93 downregulates PI3K levels to activate MB neuroblast autophagy d E93 is a late-acting temporal factor, regulated by intrinsic Imp and Syp d Extrinsic steroid hormone signaling boosts E93 levels for neurogenesis termination

Background Bone accrual impacts lifelong skeletal health, but genetic discovery has been primarily limited to cross-sectional study designs and hampered by uncertainty about target effector genes. Here, we capture this dynamic phenotype by modeling longitudinal bone accrual across 11,000 bone scans in a cohort of healthy children and adolescents, followed by genome-wide association studies (GWAS) and variant-to-gene mapping with functional follow-up. Results We identify 40 loci, 35 not previously reported, with various degrees of supportive evidence, half residing in topological associated domains harboring known bone genes. Of several loci potentially associated with later-life fracture risk, a candidate SNP lookup provides the most compelling evidence for rs11195210 (SMC3). Variant-to-gene mapping combining ATAC-seq to assay open chromatin with high-resolution promoter-focused Capture C identifies contacts between GWAS loci and nearby gene promoters. siRNA knockdown of gene expression supports the putative effector gene at three specific loci in two osteoblast cell models. Finally, using CRISPR-Cas9 genome editing, we confirm that the immediate genomic region harboring the putative causal SNP influences PRPF38A expression, a location which is predicted to coincide with a set of binding sites for relevant transcription factors. Conclusions Using a new longitudinal approach, we expand the number of genetic loci putatively associated with pediatric bone gain. Functional follow-up in appropriate cell models finds novel candidate genes impacting bone accrual. Our data also raise the possibility that the cell fate decision between osteogenic and adipogenic lineages is important in normal bone accrual.

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To uncover novel significant association signals (p<5×10−8), genome-wide association studies (GWAS) requires increasingly larger sample sizes to overcome statistical correction for multiple testing. As an alternative, we aimed to identify associations among suggestive signals (5 × 10−8≤p<5×10−4) in increasingly powered GWAS efforts using chromatin accessibility and direct contact with gene promoters as biological constraints. We conducted retrospective analyses of three GIANT BMI GWAS efforts using ATAC-seq and promoter-focused Capture C data from human adipocytes and embryonic stem cell (ESC)-derived hypothalamic-like neurons. This approach, with its extremely low false-positive rate, identified 15 loci at p<5×10−5 in the 2010 GWAS, of which 13 achieved genome-wide significance by 2018, including at NAV1, MTIF3, and ADCY3. Eighty percent of constrained 2015 loci achieved genome-wide significance in 2018. We observed similar results in waist-to-hip ratio analyses. In conclusion, biological constraints on sub-significant GWAS signals can reveal potentially true-positive loci for further investigation in existing data sets without increasing sample size.

We investigated transcriptional control of gene expression in human abdominal aortic aneurysm (AAA). We previously identified 3274 differentially expressed genes in human AAA tissue compared to non-aneurysmal controls. Four expressed transcription factors (ELF1, ETS2, STAT5 and RUNX1) were selected for genome-wide chromatin immunoprecipitation. Transcription factor binding was enriched in 4760 distinct genes (FDR < 0.05), of which 713 were differentially expressed in AAA. Functional classification using Gene Ontology (GO), KEGG, and Network Analysis revealed enrichment in several biological processes including “leukocyte migration” (FDR = 3.09 × 10−05) and “intracellular protein kinase cascade” (FDR = 6.48 × 10−05). In the control aorta, the most significant GO categories differed from those in the AAA samples and included “cytoskeleton organization” (FDR = 1.24 × 10−06) and “small GTPase mediated signal transduction” (FDR = 1.24 × 10−06). Genes up-regulated in AAA tissue showed a highly significant enrichment for GO categories “leukocyte migration” (FDR = 1.62 × 10−11), “activation of immune response” (FDR = 8.44 × 10−11), “T cell activation” (FDR = 4.14 × 10−10) and “regulation of lymphocyte activation” (FDR = 2.45 × 10−09), whereas the down-regulated genes were enriched in GO categories “cytoskeleton organization” (FDR = 7.84 × 10−05), “muscle cell development” (FDR = 1.00 × 10−04), and “organ morphogenesis” (FDR = 3.00 × 10−04). Quantitative PCR assays confirmed a sub-set of the transcription factor binding sites including those in MTMR11, DUSP10, ITGAM, MARCH1, HDAC8, MMP14, MAGI1, THBD and SPOCK1.

Correct positioning of stem cells within their niche is essential for tissue morphogenesis and homeostasis. How stem cells acquire and maintain niche position remains largely unknown. Here, we show that a subset of brain neuroblasts (NBs) in Drosophila utilize Phosphoinositide 3-kinase (PI3-kinase) and DE-cadherin to build adhesive contact for NB niche positioning. NBs remain within their native microenvironment when levels of PI3-kinase activity and DEcadherin are elevated in NBs. This occurs through PI3-kinasedependent regulation of DE-Cadherin-mediated cell adhesion between NBs and neighboring cortex glia, and between NBs and their ganglion mother cell daughters. When levels of PI3-kinase activity and/or DE-Cadherin are reduced in NBs, NBs lose niche position and relocate to a non-native brain region that is rich in neurosecretory neurons, including those that secrete some of the Drosophila insulin-like peptides. Linking levels of PI3-kinase activity to the strength of adhesive attachment could provide cancer stem cells and hematopoietic stem cells with a means to cycle from trophicpoor to trophic-rich microenvironments.