Robert Erdman

High allelic burden of four obesity SNPs is associated with poorer weight loss outcomes following gastric bypass surgery.

Genome‐wide association and linkage studies have identified multiple susceptibility loci for obesity. We hypothesized that such loci may affect weight loss outcomes following dietary or surgical weight loss interventions. A total of 1,001 white individuals with extreme obesity (BMI >35 kg/m2) who underwent a preoperative diet/behavioral weight loss intervention and Roux‐en‐Y gastric bypass surgery were genotyped for single‐nucleotide polymorphisms (SNPs) in or near the fat mass and obesity‐associated (FTO), insulin induced gene 2 (INSIG2), melanocortin 4 receptor (MC4R), and proprotein convertase subtilisin/kexin type 1 (PCSK1) obesity genes. Association analysis was performed using recessive and additive models with pre‐ and postoperative weight loss data. An increasing number of obesity SNP alleles or homozygous SNP genotypes was associated with increased BMI (P < 0.0006) and excess body weight (P < 0.0004). No association between the amounts of weight lost from a short‐term dietary intervention and any individual obesity SNP or cumulative number of obesity SNP alleles or homozygous SNP genotypes was observed. Linear mixed regression analysis revealed significant differences in postoperative weight loss trajectories across groups with low, intermediate, and high numbers of obesity SNP alleles or numbers of homozygous SNP genotypes (P < 0.0001). Initial BMI interacted with genotype to influence weight loss with initial BMI <50 kg/m2, with evidence of a dosage effect, which was not present in individuals with initial BMI ≥50 kg/m2. Differences in metabolic rate, binge eating behavior, and other clinical parameters were not associated with genotype. These data suggest that response to a surgical weight loss intervention is influenced by genetic susceptibility and BMI.

A sequence variant associated with sortilin-1 (SORT1) on 1p13.3 is independently associated with abdominal aortic aneurysm

Abdominal aortic aneurysm (AAA) is a common human disease with a high estimated heritability (0.7); however, only a small number of associated genetic loci have been reported to date. In contrast, over 100 loci have now been reproducibly associated with either blood lipid profile and/or coronary artery disease (CAD) (both risk factors for AAA) in large-scale meta-analyses. This study employed a staged design to investigate whether the loci for these two phenotypes are also associated with AAA. Validated CAD and dyslipidaemia loci underwent screening using the Otago AAA genome-wide association data set. Putative associations underwent staged secondary validation in 10 additional cohorts. A novel association between the SORT1 (1p13.3) locus and AAA was identified. The rs599839 G allele, which has been previously associated with both dyslipidaemia and CAD, reached genome-wide significance in 11 combined independent cohorts (meta-analysis with 7048 AAA cases and 75 976 controls: G allele OR 0.81, 95% CI 0.76-0.85, P = 7.2 × 10(-14)). Modelling for confounding interactions of concurrent dyslipidaemia, heart disease and other risk factors suggested that this marker is an independent predictor of AAA susceptibility. In conclusion, a genetic marker associated with cardiovascular risk factors, and in particular concurrent vascular disease, appeared to independently contribute to susceptibility for AAA. Given the potential genetic overlap between risk factor and disease phenotypes, the use of well-characterized case-control cohorts allowing for modelling of cardiovascular disease risk confounders will be an important component in the future discovery of genetic markers for conditions such as AAA.

Association of Morbid Obesity With FTO and INSIG2 Allelic Variants

OBJECTIVE To determine whether 2 single nucleotide polymorphisms (SNPs) in the obesity genes the fat mass and obesity associated gene (FTO) and the insulin induced gene 2 (INSIG2) are associated with class III, or morbid, obesity in patients undergoing bariatric weight loss operations. DESIGN Retrospective analysis of genotype and clinical data. SETTING Large rural tertiary care health system. PATIENTS A total of 707 adult patients with a body mass index (BMI; calculated as weight in kilograms divided by height in meters squared) of at least 40 undergoing open or laparoscopic Roux-en-Y gastric bypass operations for morbid obesity or its comorbid medical problems at Geisinger Medical Center, Danville, Pennsylvania. RESULTS The mean BMI in the predominantly white female cohort was 51.2. Approximately 21% of patients were homozygous for the FTO obesity SNP variant, 13% were homozygous for the INSIG2 obesity SNP variant, and 3.4% were homozygous for both. Mean BMIs in the groups homozygous for each of these genes were not significantly different from nonhomozygotes. However, FTO/INSIG2 double homozygotes and homozygote/heterozygote pairs had significantly higher BMIs than the other groups. CONCLUSION Increased BMI in morbid obesity is associated with a combination of FTO and INSIG2 SNPs.

Matrix metalloproteinase-dependent shedding of syndecan-3, a transmembrane heparan sulfate proteoglycan, in Schwann cells

Schwann cells transiently express the transmembrane heparan sulfate proteoglycan syndecan‐3 during the late embryonic and early postnatal periods of peripheral nerve development. Neonatal rat Schwann cells released soluble syndecan‐3 into the culture medium by a process that was blocked by inhibition of endogenous matrix metalloproteinase activity. When Schwann cells were plated on a substratum that binds syndecan‐3, the released proteoglycan bound to the substratum adjacent to the cell border. Membrane‐anchored syndecan‐3 was concentrated in actin‐containing filopodia that projected from the lateral edges of the Schwann cell membrane. Membrane shedding was specific for syndecan‐3 and was not observed for the related proteoglycan syndecan‐1. Analysis of Schwann cells transfected with wild‐type and chimeric syndecan‐1 and syndecan‐3 cDNAs revealed that membrane shedding was a property of the syndecan‐3 ectodomain. Inhibition of syndecan‐3 release significantly enhanced Schwann cell adhesion and process extension on dishes coated with the non‐collagenous N‐terminal domain of α4(V) collagen, which binds syndecan‐3 and mediates heparan sulfate‐dependent Schwann cell adhesion. Matrix metalloproteinase‐dependent syndecan‐3 shedding was also observed in newborn rat peripheral nerve tissue. Syndecan‐3 shedding in peripheral nerve tissue was age specific, and was not observed during later stages of postnatal nerve development. These results demonstrate that Schwann cell syndecan‐3 is subject to matrix metalloproteinase‐dependent membrane processing, which modulates the biological function of this proteoglycan.

Regional expression of HOXA4 along the aorta and its potential role in human abdominal aortic aneurysms

BackgroundThe infrarenal abdominal aorta exhibits increased disease susceptibility relative to other aortic regions. Allograft studies exchanging thoracic and abdominal segments showed that regional susceptibility is maintained regardless of location, suggesting substantial roles for embryological origin, tissue composition and site-specific gene expression.ResultsWe analyzed gene expression with microarrays in baboon aortas, and found that members of the HOX gene family exhibited spatial expression differences. HOXA4 was chosen for further study, since it had decreased expression in the abdominal compared to the thoracic aorta. Western blot analysis from 24 human aortas demonstrated significantly higher HOXA4 protein levels in thoracic compared to abdominal tissues (P < 0.001). Immunohistochemical staining for HOXA4 showed nuclear and perinuclear staining in endothelial and smooth muscle cells in aorta. The HOXA4 transcript levels were significantly decreased in human abdominal aortic aneurysms (AAAs) compared to age-matched non-aneurysmal controls (P < 0.00004). Cultured human aortic endothelial and smooth muscle cells stimulated with INF-γ (an important inflammatory cytokine in AAA pathogenesis) showed decreased levels of HOXA4 protein (P < 0.0007).ConclusionsOur results demonstrated spatial variation in expression of HOXA4 in human aortas that persisted into adulthood and that downregulation of HOXA4 expression was associated with AAAs, an important aortic disease of the ageing population.

Novel pathways in the pathobiology of human abdominal aortic aneurysms

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.

Role of Complement Cascade in Abdominal Aortic Aneurysms

Objective—The goal of this study was to investigate the role of complement cascade genes in the pathobiology of human abdominal aortic aneurysms (AAAs). Methods and Results—Results of a genome-wide microarray expression profiling revealed 3274 differentially expressed genes between aneurysmal and control aortic tissue. Interestingly, 13 genes in the complement cascade were significantly differentially expressed between AAA and the controls. In silico analysis of the promoters of the 13 complement cascade genes showed enrichment for transcription factor binding sites for signal transducer and activator of transcription (STAT)5A. Chromatin-immunoprecipitation experiments demonstrated binding of transcription factor STAT5A to the promoters of the majority of the complement cascade genes. Immunohistochemical analysis showed strong staining for C2 in AAA tissues. Conclusion—These results provide strong evidence that the complement cascade plays a role in human AAA. Based on our microarray studies, the pathway is activated in AAA, particularly via the lectin and classical pathways. The overrepresented binding sites of transcription factor STAT5A in the complement cascade gene promoters suggest a role for STAT5A in the coordinated regulation of complement cascade gene expression.

The potential role of DNA methylation in abdominal aortic aneurysms.

Abdominal aortic aneurysm (AAA) is a complex disorder that has a significant impact on the aging population. While both genetic and environmental risk factors have been implicated in AAA formation, the precise genetic markers involved and the factors influencing their expression remain an area of ongoing investigation. DNA methylation has been previously used to study gene silencing in other inflammatory disorders and since AAA has an extensive inflammatory component, we sought to examine the genome-wide DNA methylation profiles in mononuclear blood cells of AAA cases and matched non-AAA controls. To this end, we collected blood samples and isolated mononuclear cells for DNA and RNA extraction from four all male groups: AAA smokers (n = 11), AAA non-smokers (n = 9), control smokers (n = 10) and control non-smokers (n = 11). Methylation data were obtained using the Illumina 450k Human Methylation Bead Chip and analyzed using the R language and multiple Bioconductor packages. Principal component analysis and linear analysis of CpG island subsets identified four regions with significant differences in methylation with respect to AAA: kelch-like family member 35 (KLHL35), calponin 2 (CNN2), serpin peptidase inhibitor clade B (ovalbumin) member 9 (SERPINB9), and adenylate cyclase 10 pseudogene 1 (ADCY10P1). Follow-up studies included RT-PCR and immunostaining for CNN2 and SERPINB9. These findings are novel and suggest DNA methylation may play a role in AAA pathobiology.

Immunohistochemical Analysis of the Natural Killer Cell Cytotoxicity Pathway in Human Abdominal Aortic Aneurysms

Our previous analysis using genome-wide microarray expression data revealed extreme overrepresentation of immune related genes belonging the Natural Killer (NK) Cell Mediated Cytotoxicity pathway (hsa04650) in human abdominal aortic aneurysm (AAA). We followed up the microarray studies by immunohistochemical analyses using antibodies against nine members of the NK pathway (VAV1, VAV3, PLCG1, PLCG2, HCST, TYROBP, PTK2B, TNFA, and GZMB) and aortic tissue samples from AAA repair operations (n = 6) and control aortae (n = 8) from age-, sex- and ethnicity-matched donors from autopsies. The results confirmed the microarray results. Two different members of the NK pathway, HCST and GRZB, which act at different steps in the NK-pathway, were actively transcribed and translated into proteins in the same cells in the AAA tissue demonstrated by double staining. Furthermore, double staining with antibodies against CD68 or CD8 together with HCST, TYROBP, PTK2B or PLCG2 revealed that CD68 and CD8 positive cells expressed proteins of the NK-pathway but were not the only inflammatory cells involved in the NK-pathway in the AAA tissue. The results provide strong evidence that the NK Cell Mediated Cytotoxicity Pathway is activated in human AAA and valuable insight for future studies to dissect the pathogenesis of human AAA.

Transcriptional (ChIP-Chip) Analysis of ELF1, ETS2, RUNX1 and STAT5 in Human Abdominal Aortic Aneurysm

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.