Impact of the Interaction between 3′-UTR SNPs and microRNA on the Expression of Human Xenobiotic Metabolism Enzyme and Transporter Genes

Genetic variation in the expression of human XMETs leads to inter-individual variability in metabolism of therapeutic agents as well as differed susceptibility to various diseases. Recent eQTL (expression quantitative traits loci) mapping in a few human cells/tissues have identified a number of SNPs significantly associated with mRNA expression of many XMET genes. These eQTLs are therefore important candidate markers for pharmacogenetic studies. However, questions remain about whether these SNPs are causative and in what mechanism these SNPs may function. Given the important role of microRNAs in gene transcription regulation, we hypothesize that those eQTLs or their proxies in strong linkage disequilibrium (LD) altering microRNA targeting are likely causative SNPs affecting gene expression. The aim of this study is to identify eQTLs potentially regulating major XMETs via interference with microRNA targeting. To this end, we performed a genome-wide screening for eQTLs for 409 genes encoding major drug metabolism enzymes transporters and transcription factors, in publically available eQTL datasets generated from the HapMap lymphoblastoid cell lines (LCLs) and human liver and brain tissue. As a result, 308 eQTLs significantly (p<10-5) associated with mRNA expression of 101 genes were identified. We further identified 7,869 SNPs in strong LD (r2≥0.8) with these eQTLs using the 1000 Genome SNP data. Among these 8,177 SNPs, 27 are located in the 3’-UTR of 14 genes. Using two algorithms predicting microRNA-SNP interaction, we found that almost all these SNPs (26 out of 27) were predicted to create, abolish or change the target site for microRNAs in both algorithms. Many of these microRNAs were also expressed in the same tissue that the eQTL were identified. Our study provides a strong rationale for continued investigation for the functions of these eQTLs in pharmacogenetic settings

Two-Phase and Family-Based Designs for Next-Generation Sequencing Studies

The cost of next-generation sequencing is now approaching that of early GWAS panels, but is still out of reach for large epidemiologic studies and the millions of rare variants expected poses challenges for distinguishing causal from non-causal variants. We review two types of designs for sequencing studies: two-phase designs for targeted follow-up of genomewide association studies using unrelated individuals; and family-based designs exploiting co-segregation for prioritizing variants and genes.Two-phase designs subsample subjects for sequencing from a larger case-control study jointly on the basis of their disease and carrier status; the discovered variants are then tested for association in the parent study. The analysis combines the full sequence data from the substudy with the more limited SNP data from the main study. We discuss various methods for selecting this subset of variants and describe the expected yield of true positive associations in the context of an on-going study of second breast cancers following radiotherapy.While the sharing of variants within families means that family-based designs are less efficient for discovery than sequencing unrelated individuals, the ability to exploit co-segregation of variants with disease within families helps distinguish causal from non-causal ones. Furthermore, by enriching for family history, the yield of causal variants can be improved and use of identity-by-descent information improves imputation of genotypes for other family members. We compare the relative efficiency of these designs with those using unrelated individuals for discovering and prioritizing variants or genes for testing association in larger studies. While associations can be tested with single variants, power is low for rare ones. Recent generalizations of burden or kernel tests for gene-level associations to family-based data are appealing. These approaches are illustrated in the context of a family-based study of colorectal cancer

Klebsiella pneumoniae: development of carbapenem resistance due to acquisition of blaNDM-1 during antimicrobial therapy in twin infants with pneumonia

Objectives: To identify the mechanism of in vivo development of carbapenem resistance in Klebsiella pneumoniae.Methods: Seven sequential isolates of K. pneumoniae were obtained from twin infants with pneumonia. Antimicrobial susceptibility testing was performed by agar dilution. Carbapenemases including KPC and ML were initially screened using phenotypic methods, and carbapenemase-encoding genes were identified by PCR and amplicon sequencing. Plasmids of all clinical isolates and the conjugants of resistant isolates were estimated by S1 pulsed-field gel electrophoresis (PFGE). Molecular typing were conducted by PFGE of XbaI-digested genomic DNA and multilocus sequence typing (MLST). Results: For old brother, the first and third isolates were susceptible to meropenem, whereas the second and fourth isolates were resistant (MICs 16 mg/L). The first and second isolates from the young brother were susceptible to meropenem whereas the third isolate was resistant. All the resistant isolates produced NDM-1 metallo--lactamase. PFGE of XbaI-digested DNA revealed identical patterns for all the 7 isolates. All the isolates had the same sequence type named sequence type 37 (ST37). Conclusions: To our knowledge, this is the first documented case of development of carbapenem resistance in vivo mediated by NDM-1 metallo-b-lactamase in K. pneumoniae during treatment of pneumonia with meropenem

Antineuroinflammatory and neurotrophic effects of CNTF and C16 peptide in an acute experimental autoimmune encephalomyelitis rat model

Experimentalallergic encephalomyelitis (EAE) is an animal model for inflammatory demyelinating autoimmune disease, i.e., multiple sclerosis (MS). In the present study, we investigated the antineuroinflammatory/neuroprotective effects of C16, an ανβ3 integrin-binding peptide, and recombinant rat ciliary neurotrophic factor (CNTF), a cytokine that was originally identified as a survival factor for neurons, in an acute rodent EAE model. In this model, C16 peptide was injected intravenously every day for 2 weeks, and CNTF was delivered into the cerebral ventricles with Alzet miniosmotic pumps. Disease severity was assessed weekly using a scale ranging from 0 to 5. Multiple histological and molecular biological assays were employed to assess inflammation, axonal loss, neuronal apoptosis, white matter demyelination, and gliosis in the brain and spinal cord of different groups. Our results showed that the EAE induced rats revealed a significant increase in inflammatory cells infiltration, while C16 treatment could inhibit the infiltration of leukocytes and macrophages down to 2/3-1/3 of vehicle treated EAE control (P<0.05). The delayed onset of disease, reduced clinical score (P<0.01) in peak stage and more rapid recovery also were achieved in C16 treated group. Besides impairing inflammation, CNTF treatment also exerted direct neuroprotective effects, decreasing demyelination and axon loss score (P<0.05 Vs vehicle treated EAE control), and reducing the neuronal death from 40%-50% to 10%-20% (P<0.05). Both treatments suppressed the expression of cytokine tumor necrosis factor-α and interferon-when compared with the vehicle control (P<0.05). Combined treatment with C16 and CNTF produced more obvious functional recovery and neuroprotective effects than individually treatment (P<0.05). These results suggested that combination treatment with C16 and CNTF, which target different neuroprotection pathways, may be an effective therapeutic alternative to traditional therapy

Genome-wide transcriptional profiling reveals two distinct outcomes in central Nervous system infections of rabies virus

Rabies remains a major public health concern in many developing countries. The precise neuropathogenesis of rabies is unknown, though it is hypothesized to be due to neuronal death or dysfunction. Mice that received intranasal inoculation of an attenuated rabies virus (RABV) strain HEP-Flury exhibited subtle clinical signs, and eventually recovered, which is different from the fatal encephalitis caused by the virulent RABV strain CVS-11. To understand the neuropathogenesis of rabies and the mechanisms of viral clearance, we applied RNA sequencing (RNA-Seq) to compare the brain transcriptomes of normal mice versus HEP-Flury or CVS-11 intranasally inoculated mice. Our results revealed that both RABV strains altered positively and negatively the expression levels of many host genes, including genes associated with innate and adaptive immunity, inflammation and cell death. It is found that HEP-Flury infection can activate the innate immunity earlier through the RIG-I/MDA-5 signaling, and the innate immunity pre-activated by HEP-Flury or Newcastle disease virus (NDV) infection can effectively prevent the CVS-11 to invade central nervous system (CNS), but fails to clear the CVS-11 after its entry into the CNS. In addition, following CVS-11 infection, genes implicated in cell adhesion, blood vessel morphogenesis and coagulation were mainly up-regulated, while the genes involved in synaptic transmission and ion transport were significantly down-regulated. On the other hand, several genes involved in the MHC class II-mediated antigen presentation pathway were activated to a greater extent after the HEP-Flury infection as compared with the CVS-11 infection suggesting that the collaboration of CD4+ T cells and MHC class II-mediated antigen presentation is critical for the clearance of attenuated RABV from the CNS. The differentially regulated genes reported here are likely to include potential therapeutic targets for expanding the postexposure treatment window for RABV infection