Citations

AbstractBackgroundRejection of Gal‐free (GTKO) donor pig cardiac xenografts is strongly associated with vascular non‐Gal antibody binding, endothelial cell (EC) injury, and activation and microvascular thrombosis. We adopted a pig‐to‐SCID/beige small animal transplant model to compare the pathogenicity of baboon and human anti‐pig antibody.MethodsWild‐type (GT+) or GTKO porcine coronary arteries (PCAs) were transplanted into the infrarenal aorta of SCID/beige mice. Three days after transplant, recipients were infused with anti‐pig antibody (anti‐SLA class I, an isotype control, naive or sensitized baboon serum, or naive human serum). PCAs were recovered 24 h after antibody infusion and examined using histology, immunohistochemistry, and in situ hybridization.ResultsDose‐dependent intragraft thrombosis occurred after infusion of anti‐SLA I antibody (but not isotype control) in GT+ and GTKO PCA recipients. Naive baboon serum induced thrombosis in GT+ grafts. Thrombosis was significantly reduced by pre‐treating naive baboon serum with Gal polymer and not observed when this serum was infused to GTKO PCA recipients. Naive human serum caused dose‐dependent intragraft thrombosis of GTKO PCAs. In all cases, thrombosis involved graft‐specific vascular antibody and complement deposition, macrophage adherence, EC delamination, and subendothelial thrombus formation.ConclusionsThis study provides the first direct in vivo comparison of the pathogenicity of naive human and baboon serum. The results suggest that human preformed non‐Gal antibody may have increased pathogenicity compared to baboon. This model, which showed a rejected graft histopathology similar to antibody‐mediated rejection in cardiac xenotransplantation, may be useful to assess the pathogenicity of individual protein or carbohydrate specific non‐Gal reactive antibodies.

Abstract Summary: The development of the Infinium HumanMethylation450 BeadChip enables epigenome-wide association studies at a reduced cost. One observation of the 450K data is that many CpG sites the beadchip interrogates have very large measurement errors. Including these noisy CpGs will decrease the statistical power of detecting relevant associations due to multiple testing correction. We propose to use intra-class correlation coefficient (ICC), which characterizes the relative contribution of the biological variability to the total variability, to filter CpGs when technical replicates are available. We estimate the ICC based on a linear mixed effects model by pooling all the samples instead of using the technical replicates only. An ultra-fast algorithm has been developed to address the computational complexity and CpG filtering can be completed in minutes on a desktop computer for a 450K data set of over 1000 samples. Our method is very flexible and can accommodate any replicate design. Simulations and a real data application demonstrate that our whole-sample ICC method performs better than replicate-sample ICC or variance-based method. Availability and implementation: CpGFilter is implemented in R and publicly available under CRAN via the R package ‘CpGFilter’. Contact: chen.jun2@mayo.edu or xlin@hsph.harvard.edu Supplementary information: Supplementary data are available at Bioinformatics online.

Abstract Nearly all pancreatic ductal adenocarcinomas are caused by oncogenic mutations in the KRAS gene. Pharmacological inhibition of mutant KRAS has thus far been unsuccessful in the clinical setting, precipitating a need to understand the pathways downstream of KRAS which may prove more easily targeted with small molecule inhibitors. Here we show that PI3K p110α is absolutely required for pancreatic tumorigenesis while p110β is dispensable for this process. Surprisingly, ablation of p110α does not impair the ability of KRAS to activate AKT, demonstrating that AKT activation is not sufficient for transformation. Instead we find that p110α is required for robust activation of RAC1, a small GTPase required for pancreatic metaplasia. Consistent with this, our data show that p110α is necessary for regulating epithelial expression and activation of RAC-GEFs including Vav1, Tiam1 and Ect2. Ultimately, these results define the mechanistic role of p110α in pancreatic tumorigenesis and suggest selective inhibition of this PI3K isoform as a promising therapeutic approach to treating patients with pancreas cancer. Citation Format: Kenneth K. Takeuchi, Eileen Carpenter, Claire Wu, Christopher J. Halbrook, Richard Z. Lin, Howard C. Crawford. PI3K regulation of RAC1 is required for Kras-induced pancreatic tumorigenesis. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr A04.

ABSTRACT The Wnt/β-catenin pathway is implicated in left-right (LR) axis determination; however, the underlying mechanism remains elusive. Prompted by our recent discovery that Wnt signaling regulates ciliogenesis in the zebrafish Kupffer's vesicle (KV) via Foxj1a, a ciliogenic transcription factor, we decided to elucidate functions of Foxj1a in Wnt-regulated LR pattern formation. We showed that targeted injection of wnt8a mRNA into a single cell at the 128-cell stage is sufficient to induce ectopic foxj1a expression and ectopic cilia. By interrogating the transcription circuit of foxj1a regulation, we found that both Lef1 and Tcf7 bind to a consensus element in the foxj1a promoter region. Depletion of Lef1 and Tcf7 inhibits foxj1a transcription in the dorsal forerunner cells, downregulates cilia length and number in KV, and randomizes LR asymmetry. Targeted overexpression of a constitutively active form of Lef1 also induced an ectopic protrusion that contains ectopic transcripts for sox17, foxj1a, and charon, and ectopic monocilia. Further genetic studies using this ectopic expression platform revealed two distinct functions of Foxj1a; mediating Wnt-governed monocilia length elongation as well as charon transcription. The novel Foxj1a-charon regulation is conserved in KV, and importantly, it is independent of the canonical role of Foxj1a in the biosynthesis of motile cilia. Together with the known function of motile cilia movement in generating asymmetric expression of charon, our data put forward a hypothesis that Foxj1a confers both ciliary and non-ciliary functions of Wnt signaling, which converge on charon to regulate LR pattern formation.

AbstractSuicide by shotgun is a common method of suicide with high regional variation. We sought to describe their distinct, challenging features and provide demographic and risk characteristics. We reviewed 228 gunshot wounds autopsied at Mayo Clinic from 1994 to 2014; of these, 75 (32.9%) were shotgun wounds. All were suicides and contact range. Ages ranged from 14 to 92 years old; of these, 97% were men. The majority involved the head (70.9%), were intraoral (48.2%), and had upward (73.2%) and backward (73.2%) directionality. Next most common was the chest (21.5%), with backward (94.1%) and downward (64.7%) directionality. Four involved multiple wounds. Wadding was recovered in 16 (21.3%) cases. Six (8.0%) had a survival period. Most (66.7%) took place at home. Seventy‐one percent had a known psychiatric history; 32.4% had positive toxicology. Although contact range shotgun wounds cause severe destruction, entrance wound and other characteristic are identifiable with thorough scene, autopsy, and radiographic documentation.

ABSTRACTEpithelial‐mesenchymal transition (EMT) is a process whereby epithelial cells assume mesenchymal characteristics to facilitate cancer metastasis. However, EMT also contributes to the initiation and development of primary tumors. Prior studies that explored the hypothesis that EMT gene variants contribute to epithelial ovarian carcinoma (EOC) risk have been based on small sample sizes and none have sought replication in an independent population. We screened 15,816 single‐nucleotide polymorphisms (SNPs) in 296 genes in a discovery phase using data from a genome‐wide association study of EOC among women of European ancestry (1,947 cases and 2,009 controls) and identified 793 variants in 278 EMT‐related genes that were nominally (P < 0.05) associated with invasive EOC. These SNPs were then genotyped in a larger study of 14,525 invasive‐cancer patients and 23,447 controls. A P‐value <0.05 and a false discovery rate (FDR) <0.2 were considered statistically significant. In the larger dataset, GPC6/GPC5 rs17702471 was associated with the endometrioid subtype among Caucasians (odds ratio (OR) = 1.16, 95% CI = 1.07–1.25, P = 0.0003, FDR = 0.19), whereas F8 rs7053448 (OR = 1.69, 95% CI = 1.27–2.24, P = 0.0003, FDR = 0.12), F8 rs7058826 (OR = 1.69, 95% CI = 1.27–2.24, P = 0.0003, FDR = 0.12), and CAPN13 rs1983383 (OR = 0.79, 95% CI = 0.69–0.90, P = 0.0005, FDR = 0.12) were associated with combined invasive EOC among Asians. In silico functional analyses revealed that GPC6/GPC5 rs17702471 coincided with DNA regulatory elements. These results suggest that EMT gene variants do not appear to play a significant role in the susceptibility to EOC.

Abstract Alternative NF-κB signaling is crucial for B cell activation and Ig production, and it is mainly regulated by the inhibitor of κ B kinase (IKK) regulatory complex. Dysregulation of alternative NF-κB signaling in B cells could therefore lead to hyperactive B cells and Ig overproduction. In our previous, study we found that deleted in breast cancer 1 (DBC1) is a suppressor of the alternative NF-κB pathway to attenuate B cell activation. In this study, we report that loss of DBC1 results in spontaneous overproduction of Ig in mice after 10 mo of age. Using a double mutant genetic model, we confirm that DBC1 suppresses B cell activation through RelB inhibition. At the molecular level, we show that DBC1 interacts with alternative NF-κB members RelB and p52 through its leucine zipper domain. In addition, phosphorylation of DBC1 at its C terminus by IKKα facilitates its interaction with RelB and IKKα, indicating that DBC1-mediated suppression of alternative NF-κB is regulated by IKKα. Our results define the molecular mechanism of DBC1 inhibition of alternative NF-κB activation in suppressing B cell activation.