Histone deacetylase inhibitor, butyrate, attenuates lipopolysaccharide-induced acute lung injury in mice

<p>Abstract</p> <p>Background</p> <p>Histone deacetylase (HDAC) inhibitors, developed as promising anti-tumor drugs, exhibit their anti-inflammatory properties due to their effects on reduction of inflammatory cytokines.</p> <p>Objective</p> <p>To investigate the protective effect of butyrate, a HDAC inhibitor, on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice.</p> <p>Methods</p> <p>ALI was induced in Balb/c mice by intratracheally instillation of LPS (1 mg/kg). Before 1 hour of LPS administration, the mice received butyrate (10 mg/kg) orally. The animals in each group were sacrificed at different time point after LPS administration. Pulmonary histological changes were evaluated by hematoxylin-eosin stain and lung wet/dry weight ratios were observed. Concentrations of interleukin (IL)-1β and tumor necrosis factor (TNF)-α in bronchoalveolar lavage fluid (BALF) and concentrations of nitric oxide (NO) and myeloperoxidase (MPO) activity in lung tissue homogenates were measured by enzyme-linked immunosorbent assay (ELISA). Expression of nuclear factor (NF)-κB p65 in cytoplasm and nucleus was determined by Western blot analysis respectively.</p> <p>Results</p> <p>Pretreatment with butyrate led to significant attenuation of LPS induced evident lung histopathological changes, alveolar hemorrhage, and neutrophils infiltration with evidence of reduced MPO activity. The lung wet/dry weight ratios, as an index of lung edema, were reduced by butyrate administration. Butyrate also repressed the production of TNF-α, IL-1β and NO. Furthermore, the expression of NF-κB p65 in nucleus was markedly suppressed by butyrate pretreatment.</p> <p>Conclusions</p> <p>Butyrate had a protective effect on LPS-induced ALI, which may be related to its effect on suppression of inflammatory cytokines production and NF-κB activation.</p

Promoter Methylation in Head and Neck Squamous Cell Carcinoma Cell Lines Is Significantly Different than Methylation in Primary Tumors and Xenografts

Studies designed to identify novel methylation events related to cancer often employ cancer cell lines in the discovery phase of the experiments and have a relatively low rate of discovery of cancer-related methylation events. An alternative algorithm for discovery of novel methylation in cancer uses primary tumor-derived xenografts instead of cell lines as the primary source of nucleic acid for evaluation. We evaluated DNA extracted from primary head and neck squamous cell carcinomas (HNSCC), xenografts grown from these primary tumors in nude mice, HNSCC-derived cell lines, normal oral mucosal samples, and minimally transformed oral keratinocyte-derived cell lines using Illumina Infinum Humanmethylation 27 genome-wide methylation microarrays. We found >2,200 statistically significant methylation differences between cancer cell lines and primary tumors and when comparing normal oral mucosa to keratinocyte cell lines. We found no statistically significant promoter methylation differences between primary tumor xenografts and primary tumors. This study demonstrates that tumor-derived xenografts are highly accurate representations of promoter methylation in primary tumors and that cancer derived cell lines have significant drawbacks for discovery of promoter methylation alterations in primary tumors. These findings also support use of primary tumor xenografts for the study of methylation in cancer, drug discovery, and the development of personalized cancer treatments

Allele-Specific, Age-Dependent and BMI-Associated DNA Methylation of Human MCHR1

Background: Melanin-concentrating hormone receptor 1 (MCHR1) plays a significant role in regulation of energy balance, food intake, physical activity and body weight in humans and rodents. Several association studies for human obesity showed contrary results concerning the SNPs rs133072 (G/A) and rs133073 (T/C), which localize to the first exon of MCHR1. The variations constitute two main haplotypes (GT, AC). Both SNPs affect CpG dinucleotides, whereby each haplotype contains a potential methylation site at one of the two SNP positions. In addition, 15 CpGs in close vicinity of these SNPs constitute a weak CpG island. Here, we studied whether DNA methylation in this sequence context may contribute to population- and age-specific effects of MCHR1 alleles in obesity. \ud Principal Findings: We analyzed DNA methylation of a 315 bp region of MCHR1 encompassing rs133072 and rs133073 and the CpG island in blood samples of 49 individuals by bisulfite sequencing. The AC haplotype shows a significantly higher methylation level than the GT haplotype. This allele-specific methylation is age-dependent. In young individuals (20â\u80\u9330 years) the difference in DNA methylation between haplotypes is significant; whereas in individuals older than 60 years it is not detectable. Interestingly, the GT allele shows a decrease in methylation status with increasing BMI, whereas the methylation of the AC allele is not associated with this phenotype. Heterozygous lymphoblastoid cell lines show the same pattern of allele-specific DNA methylation. The cell line, which exhibits the highest difference in methylation levels between both haplotypes, also shows allele-specific transcription of MCHR1, which can be abolished by treatment with the DNA\ud methylase inhibitor 5-aza-2&apos;-deoxycytidine.\ud Conclusions:We show that DNA methylation at MCHR1 is allele-specific, age-dependent, BMI-associated and affects transcription. Conceivably, this epigenetic regulation contributes to the age- and/or population specific effects reported for MCHR1 in several human obesity studies.\ud \ud doi: 10.1371/journal.pone.0017711\u

Homopolymer tract length dependent enrichments in functional regions of 27 eukaryotes and their novel dependence on the organism DNA (G+C)% composition

BACKGROUND: DNA homopolymer tracts, poly(dA).poly(dT) and poly(dG).poly(dC), are the simplest of simple sequence repeats. Homopolymer tracts have been systematically examined in the coding, intron and flanking regions of a limited number of eukaryotes. As the number of DNA sequences publicly available increases, the representation (over and under) of homopolymer tracts of different lengths in these regions of different genomes can be compared. RESULTS: We carried out a survey of the extent of homopolymer tract over-representation (enrichment) and over-proportional length distribution (above expected length) primarily in the single gene documents, but including some whole chromosomes of 27 eukaryotics across the (G+C)% composition range from 20 – 60%. A total of 5.2 × 10(7 )bases from 15,560 cleaned (redundancy removed) sequence documents were analyzed. Calculated frequencies of non-overlapping long homopolymer tracts were found over-represented in non-coding sequences of eukaryotes. Long poly(dA).poly(dT) tracts demonstrated an exponential increase with tract length compared to predicted frequencies. A novel negative slope was observed for all eukaryotes between their (G+C)% composition and the threshold length N where poly(dA).poly(dT) tracts exhibited over-representation and a corresponding positive slope was observed for poly(dG).poly(dC) tracts. Tract size thresholds where over-representation of tracts in different eukaryotes began to occur was between 4 – 11 bp depending upon the organism (G+C)% composition. The higher the GC%, the lower the threshold N value was for poly(dA).poly(dT) tracts, meaning that the over-representation happens at relatively lower tract length in more GC-rich surrounding sequence. We also observed a novel relationship between the highest over-representations, as well as lengths of homopolymer tracts in excess of their random occurrence expected maximum lengths. CONCLUSIONS: We discuss how our novel tract over-representation observations can be accounted for by a few models. A likely model for poly(dA).poly(dT) tract over-representation involves the known insertion into genomes of DNA synthesized from retroviral mRNAs containing 3' polyA tails. A proposed model that can account for a number of our observed results, concerns the origin of the isochore nature of eukaryotic genomes via a non-equilibrium GC% dependent mutation rate mechanism. Our data also suggest that tract lengthening via slip strand replication is not governed by a simple thermodynamic loop energy model

Replicating viral vector platform exploits alarmin signals for potent CD8⁺ T cell-mediated tumour immunotherapy.

Viral infections lead to alarmin release and elicit potent cytotoxic effector T lymphocyte (CTL &lt;sup&gt;eff&lt;/sup&gt; ) responses. Conversely, the induction of protective tumour-specific CTL &lt;sup&gt;eff&lt;/sup&gt; and their recruitment into the tumour remain challenging tasks. Here we show that lymphocytic choriomeningitis virus (LCMV) can be engineered to serve as a replication competent, stably-attenuated immunotherapy vector (artLCMV). artLCMV delivers tumour-associated antigens to dendritic cells for efficient CTL priming. Unlike replication-deficient vectors, artLCMV targets also lymphoid tissue stroma cells expressing the alarmin interleukin-33. By triggering interleukin-33 signals, artLCMV elicits CTL &lt;sup&gt;eff&lt;/sup&gt; responses of higher magnitude and functionality than those induced by replication-deficient vectors. Superior anti-tumour efficacy of artLCMV immunotherapy depends on interleukin-33 signalling, and a massive CTL &lt;sup&gt;eff&lt;/sup&gt; influx triggers an inflammatory conversion of the tumour microenvironment. Our observations suggest that replicating viral delivery systems can release alarmins for improved anti-tumour efficacy. These mechanistic insights may outweigh safety concerns around replicating viral vectors in cancer immunotherapy

Fractional deuteration applied to biomolecular solid-state NMR spectroscopy

Solid-state Nuclear Magnetic Resonance can provide detailed insight into structural and dynamical aspects of complex biomolecules. With increasing molecular size, advanced approaches for spectral simplification and the detection of medium to long-range contacts become of critical relevance. We have analyzed the protonation pattern of a membrane-embedded ion channel that was obtained from bacterial expression using protonated precursors and D2O medium. We find an overall reduction of 50% in protein protonation. High levels of deuteration at Hα and Hβ positions reduce spectral congestion in (1H,13C,15N) correlation experiments and generate a transfer profile in longitudinal mixing schemes that can be tuned to specific resonance frequencies. At the same time, residual protons are predominantly found at amino-acid side-chain positions enhancing the prospects for obtaining side-chain resonance assignments and for detecting medium to long-range contacts. Fractional deuteration thus provides a powerful means to aid the structural analysis of complex biomolecules by solid-state NMR

Clinical and pathological characteristics of Chinese patients with BRCA related breast cancer

Breast cancers related to BRCA mutations are associated with particular biological features. Here we report the clinical and pathological characteristics of breast cancer in Chinese women with and without BRCA mutations and of carriers of BRCA1 mutations compared to BRCA2 mutations. Two hundred and 26 high-risk Hong Kong Chinese women were tested for BRCA mutations, medical information was obtained from medical records, and risk and demographic information was obtained from personal interviews. In this cohort, 28 (12.4%) women were BRCA mutation carriers and among these carriers, 39.3% were BRCA1 and 60.7% were BRCA2 mutations. Mutation carriers were more likely to have a familial history of breast and ovarian cancer, high-grade cancers, and triple negative (TN) cancers. Prevalence of TN was 48.3% in BRCA carriers and 25.6% in non-carriers and was 67.7% in BRCA1 and 35.3% in BRCA2 carriers. Estrogen receptor (ER) negative cancer was significantly associated with BRCA1 mutations, especially in those under 40 years of age. BRCA-related breast cancer in this Chinese population is associated with family history and adverse pathological/prognostic features, with BRCA2 mutations being more prevalent but BRCA1 carriers having more aggressive and TN cancers. Compared to Caucasian populations, prevalence of BRCA2 mutations and TN cancer in BRCA2 mutation carriers in Chinese population are elevated

Painful and painless mutations of SCN9A and SCN11A voltage-gated sodium channels

Chronic pain is a global problem affecting up to 20% of the world’s population and has a significant economic, social and personal cost to society. Sensory neurons of the dorsal root ganglia (DRG) detect noxious stimuli and transmit this sensory information to regions of the central nervous system (CNS) where activity is perceived as pain. DRG neurons express multiple voltage-gated sodium channels that underlie their excitability. Research over the last 20 years has provided valuable insights into the critical roles that two channels, NaV1.7 and NaV1.9, play in pain signalling in man. Gain of function mutations in NaV1.7 cause painful conditions while loss of function mutations cause complete insensitivity to pain. Only gain of function mutations have been reported for NaV1.9. However, while most NaV1.9 mutations lead to painful conditions, a few are reported to cause insensitivity to pain. The critical roles these channels play in pain along with their low expression in the CNS and heart muscle suggest they are valid targets for novel analgesic drugs