UGT1A1 sequence variants and bilirubin levels in early postnatal life: a quantitative approach

<p>Abstract</p> <p>Background</p> <p>Fundamental to definitively identifying neonates at risk of developing significant hyperbilirubinemia is a better understanding of the genetic factors associated with early bilirubin rise. Previous genetic studies have focused on the UGT1A1 gene, associating common variation in the coding or promoter regions with qualitative assessments of bilirubin (i.e. significantly elevated or not). These studies have had conflicting results and limited success. We chose to approach the problem by focusing on the quantitative (absolute) change in bilirubin levels early in post-natal life. We apply this approach to the UGT1A1 gene - exploring the contribution of both rare and common variants to early bilirubin changes.</p> <p>Methods</p> <p>We sequenced the exons, PBREM, 5'-, and 3'- regions of the UGT1A1 gene in 80 otherwise healthy term neonates who had repeat bilirubin levels measured within the first five days of life.</p> <p>Results</p> <p>Three novel coding variants were observed, but there was no clear relationship between rare coding variants and bilirubin rise. Adjusted linear regression models fit to evaluate the relationship between changing bilirubin levels and common UGT1A1variants found that among 39 neonates whose bilirubin was resampled within 33 hours, individuals homozygous for the mutant allele of a 3'UTR SNP had significantly smaller changes in bilirubin (P = 0.003) than individuals carrying the wild-type allele.</p> <p>Conclusions</p> <p>Collectively, rare UGT1A1 coding variants do not appear to play a prominent role in determining early bilirubin levels; however common variants in the 3' UTR of UGT1A1 may modulate the early bilirubin rise. A quantitative approach to evaluating early bilirubin kinetics provides a more robust framework in which to better understand the genetics of neonatal hyperbilirubinemia.</p

Experiments with a high-density positronium gas

We have created a high-density gas of interacting positronium (Ps) atoms by irradiating a thin film of nanoporous silica with intense positron bursts and measured the Ps lifetime using a new single-shot technique. When the positrons were compressed to 3.3×1010cm-2, the apparent intensity of the orthopositronium lifetime component was found to decrease by 33%. We believe this is due to a combination of spin exchange quenching and Ps2 molecule formation associated with colliding pairs of oppositely polarized triplet positronium atoms. Our data imply an effective cross section for this process of 2.9×10-14cm-2. © 2005 The American Physical Society

Modulation of enhancer looping and differential gene targeting by Epstein-Barr virus transcription factors directs cellular reprogramming

Epstein-Barr virus (EBV) epigenetically reprogrammes B-lymphocytes to drive immortalization and facilitate viral persistence. Host-cell transcription is perturbed principally through the actions of EBV EBNA 2, 3A, 3B and 3C, with cellular genes deregulated by specific combinations of these EBNAs through unknown mechanisms. Comparing human genome binding by these viral transcription factors, we discovered that 25% of binding sites were shared by EBNA 2 and the EBNA 3s and were located predominantly in enhancers. Moreover, 80% of potential EBNA 3A, 3B or 3C target genes were also targeted by EBNA 2, implicating extensive interplay between EBNA 2 and 3 proteins in cellular reprogramming. Investigating shared enhancer sites neighbouring two new targets (WEE1 and CTBP2) we discovered that EBNA 3 proteins repress transcription by modulating enhancer-promoter loop formation to establish repressive chromatin hubs or prevent assembly of active hubs. Re-ChIP analysis revealed that EBNA 2 and 3 proteins do not bind simultaneously at shared sites but compete for binding thereby modulating enhancer-promoter interactions. At an EBNA 3-only intergenic enhancer site between ADAM28 and ADAMDEC1 EBNA 3C was also able to independently direct epigenetic repression of both genes through enhancer-promoter looping. Significantly, studying shared or unique EBNA 3 binding sites at WEE1, CTBP2, ITGAL (LFA-1 alpha chain), BCL2L11 (Bim) and the ADAMs, we also discovered that different sets of EBNA 3 proteins bind regulatory elements in a gene and cell-type specific manner. Binding profiles correlated with the effects of individual EBNA 3 proteins on the expression of these genes, providing a molecular basis for the targeting of different sets of cellular genes by the EBNA 3s. Our results therefore highlight the influence of the genomic and cellular context in determining the specificity of gene deregulation by EBV and provide a paradigm for host-cell reprogramming through modulation of enhancer-promoter interactions by viral transcription factors

Determination of Interleukin-6 and Tumor Necrosis Factor-alpha concentrations in Iranian-Khorasanian patients with preeclampsia

BACKGROUND: Our objective was to determine the role of Interleukin-6 (IL-6) and Tumor Necrosis Factor-alpha (TNF-alpha), markers of immune activation and endothelial dysfunction, in patients with preeclampsia. METHODS: Twenty four women with preeclampsia and eighteen antepartum normotensive pregnant women were recruited as controls. Serum levels of IL-6 and TNF-alpha were measured by enzyme-linked immunosorbent assay. We used independent-samples t test to assess the differences in the concentration of cytokines in preeclamptic patients and control subjects. RESULTS: IL-6 levels [mean (S.D.)] were significantly higher in preeclamptic women [5.8 (4.85) pg/ml] compared to normal pregnant women [3.01 (2.45) pg/ml] (p = 0.02). There was no significant change in concentration of TNF-alpha in preeclamptic women [53.8 (30.0) pg/ml] compared to normal pregnant women [51.9 (33.8) pg/ml] (p > 0.1). CONCLUSION: The results of this study show that IL-6 as a pro-inflammatory cytokine is present in higher concentration in women with preeclampsia. The study was undertaken in women with established preeclampsia and it is not possible to determine whether the increased concentration of IL-6 is a cause or consequence of the disease. Furthermore, these findings suggest that serum TNF-alpha level is not associated with preeclampsia

The Effect of Epstein-Barr Virus Latent Membrane Protein 2 Expression on the Kinetics of Early B Cell Infection

Infection of human B cells with wild-type Epstein-Barr virus (EBV) in vitro leads to activation and proliferation that result in efficient production of lymphoblastoid cell lines (LCLs). Latent Membrane Protein 2 (LMP2) is expressed early after infection and previous research has suggested a possible role in this process. Therefore, we generated recombinant EBV with knockouts of either or both protein isoforms, LMP2A and LMP2B (Δ2A, Δ2B, Δ2A/Δ2B) to study the effect of LMP2 in early B cell infection. Infection of B cells with Δ2A and Δ2A/Δ2B viruses led to a marked decrease in activation and proliferation relative to wild-type (wt) viruses, and resulted in higher percentages of apoptotic B cells. Δ2B virus infection showed activation levels comparable to wt, but fewer numbers of proliferating B cells. Early B cell infection with wt, Δ2A and Δ2B viruses did not result in changes in latent gene expression, with the exception of elevated LMP2B transcript in Δ2A virus infection. Infection with Δ2A and Δ2B viruses did not affect viral latency, determined by changes in LMP1/Zebra expression following BCR stimulation. However, BCR stimulation of Δ2A/Δ2B cells resulted in decreased LMP1 expression, which suggests loss of stability in viral latency. Long-term outgrowth assays revealed that LMP2A, but not LMP2B, is critical for efficient long-term growth of B cells in vitro. The lowest levels of activation, proliferation, and LCL formation were observed when both isoforms were deleted. These results suggest that LMP2A appears to be critical for efficient activation, proliferation and survival of EBV-infected B cells at early times after infection, which impacts the efficient long-term growth of B cells in culture. In contrast, LMP2B did not appear to play a significant role in these processes, and long-term growth of infected B cells was not affected by the absence of this protein. © 2013 Wasil et al

Upregulation of the cell-cycle regulator RGC-32 in Epstein-Barr virus-immortalized cells

Epstein-Barr virus (EBV) is implicated in the pathogenesis of multiple human tumours of lymphoid and epithelial origin. The virus infects and immortalizes B cells establishing a persistent latent infection characterized by varying patterns of EBV latent gene expression (latency 0, I, II and III). The CDK1 activator, Response Gene to Complement-32 (RGC-32, C13ORF15), is overexpressed in colon, breast and ovarian cancer tissues and we have detected selective high-level RGC-32 protein expression in EBV-immortalized latency III cells. Significantly, we show that overexpression of RGC-32 in B cells is sufficient to disrupt G2 cell-cycle arrest consistent with activation of CDK1, implicating RGC-32 in the EBV transformation process. Surprisingly, RGC-32 mRNA is expressed at high levels in latency I Burkitt's lymphoma (BL) cells and in some EBV-negative BL cell-lines, although RGC-32 protein expression is not detectable. We show that RGC-32 mRNA expression is elevated in latency I cells due to transcriptional activation by high levels of the differentially expressed RUNX1c transcription factor. We found that proteosomal degradation or blocked cytoplasmic export of the RGC-32 message were not responsible for the lack of RGC-32 protein expression in latency I cells. Significantly, analysis of the ribosomal association of the RGC-32 mRNA in latency I and latency III cells revealed that RGC-32 transcripts were associated with multiple ribosomes in both cell-types implicating post-initiation translational repression mechanisms in the block to RGC-32 protein production in latency I cells. In summary, our results are the first to demonstrate RGC-32 protein upregulation in cells transformed by a human tumour virus and to identify post-initiation translational mechanisms as an expression control point for this key cell-cycle regulator

Adenomyoma of the Ileum Leading to Intussusception

Adenomyoma of the ileum is a rare condition. A 68-year-old Japanese man presented with nausea and distension of the abdomen. Enhanced computed tomography of his abdomen revealed wall thickening in the ileum and dilation of the proximal small intestine. Open laparotomy was performed to find the cause of the patient's small bowel obstruction, and a tumor was found in the ileum, which had resulted in intussusception. The tumor and 20 cm of the adjacent ileum were resected. The resected specimen displayed a macroscopic appearance suggestive of a submucosal tumor. Histopathological evaluation showed duct cell proliferation and bundles of smooth muscle cells from the mucosa to the serosa, leading to a diagnosis of adenomyoma. Immunohistochemical examination found that cytokeratin 7 and carbohydrate antigen 19-9 were expressed in the duct epithelia. We report a rare case of ileal adenomyoma leading to intussusception in an adult and present the immunohistochemical evaluation of the adenomyoma

Optimizing Nervous System-Specific Gene Targeting with Cre Driver Lines: Prevalence of Germline Recombination and Influencing Factors.

The Cre-loxP system is invaluable for spatial and temporal control of gene knockout, knockin, and reporter expression in the mouse nervous system. However, we report varying probabilities of unexpected germline recombination in distinct Cre driver lines designed for nervous system-specific recombination. Selective maternal or paternal germline recombination is showcased with sample Cre lines. Collated data reveal germline recombination in over half of 64 commonly used Cre driver lines, in most cases with a parental sex bias related to Cre expression in sperm or oocytes. Slight differences among Cre driver lines utilizing common transcriptional control elements affect germline recombination rates. Specific target loci demonstrated differential recombination; thus, reporters are not reliable proxies for another locus of interest. Similar principles apply to other recombinase systems and other genetically targeted organisms. We hereby draw attention to the prevalence of germline recombination and provide guidelines to inform future research for the neuroscience and broader molecular genetics communities