Reading the unique DNA methylation landscape of the brain: Non-CpG methylation,hydroxymethylation, and MeCP2

DNA methylation at CpG dinucleotides is an important epigenetic regulator common to virtually all mammalian cell types, but recent evidence indicates that during early postnatal development neuronal genomes also accumulate uniquely high levels of two alternative forms of methylation, non-CpG methylation and hydroxymethylation. Here we discuss the distinct landscape of DNA methylation in neurons, how it is established, and how it might affect the binding and function of protein readers of DNA methylation. We review studies of one critical reader of DNA methylation in the brain, the Rett syndrome protein methyl CpG-binding protein 2 (MeCP2), and discuss how differential binding affinity of MeCP2 for non-CpG and hydroxymethylation may affect the function of this methyl-binding protein in the nervous system.     

Genetic analysis of the yeast NUD1 endo-exonuclease: a role in the repair of DNA double-strand breaks

The deoxyribonucleases (DNases) have been shown genetically to be important in the vital processes of DNA repair and recombination. The NUD1 gene, which codes for an endo-exonuclease of Saccharomyces cerevisiae, was analyzed for its role in the DNA double-strand break (DSB) repair processes. While the nud1 strain is only slightly sensitive to ionizing radiation, expression of the HO-endonuclease to introduce a DSB at the MAT locus in that strain results in cell death. Cell survival is inversely proportional to the duration of HO-endonuclease expression. Analysis of the surviving colonies from the nud1 strain indicated that many of the survivors are sterile and that the proportion of these sterile survivors increases with the time of HO-endonuclease expression. On the other hand, the surviving colonies from the isogenic NUD1 strain are mating-proficient. Interestingly, double mutants of nud1 rad52 are more resistant to ionizing irradiation than the rad52 strain and have a cell-survival fraction of 32% for rad52-1 nud1 and 9% for rad52::URA3 nud1 following prolonged HO-endonuclease expression, indicating that nud1 has a suppressor effect on the DSB-induced lethality in rad52. Polymerase chain reaction analysis showed that many of the nud1 survivors contained small alterations within the MAT locus, suggesting that the survivors arose through the process of non-homologous end-joining. These results suggest that the endo-exonuclease acts at a DSB to promote DNA repair via the homologous recombination pathway. Received: 20 July / 20 September 1998     

BALAD and BALAD-2 predict survival of hepatocellular carcinoma patients: a North American cohort study

BackgroundThe BALAD score and BALAD-2 class derived from bilirubin, albumin, AFP, AFP-L3, and des-gamma-carboxyprothrombin (DCP) are effective in predicting mortality in HCC, but have not been validated in North America.Methods148 HCC patients from 2000 to 2015 who had all five biomarkers tested at diagnosis were included. Hazard ratios (HR) were calculated.Results75 patients died during a median follow-up of 21.9 months. 1-and 3-year survival rates were 70.8% and 47.6%. 114 (77%) had cirrhosis. The HR (95%CI) for death were 1.24 (0.42–3.67), 1.79 (0.61–5.26), 2.83 (0.95–8.38), and 7.19 (2.26–22.91) for BALAD scores 1, 2, 3, and 4 vs. BALAD 0. The HR (95%CI) for death were 1.25 (0.65–2.40), 1.75 (0.94–3.23), and 6.20 (3.29–11.68) for BALAD-2 classes 2, 3, and 4 vs. BALAD-2 class 1. A multivariate model incorporating maximal tumor diameter, tumor number, neutrophil-lymphocyte ratio, and BALAD had HR of 1.43 (1.14–1.81) per increase of 1 BALAD score. A similar model with BALAD-2 had HR of 1.50 (1.18–1.90) per increase of 1 BALAD-2 class.ConclusionBALAD models at diagnosis can predict the survival of HCC patients in North America. AFP, AFP-L3, and DCP reflect tumor progression and metastasis of HCC and distinguish the BALAD model from other predictive models.     

The Dose-Response Effects of Consuming High Fructose Corn Syrup-Sweetened Beverages on Hepatic Lipid Content and Insulin Sensitivity in Young Adults

Increased hepatic lipid content and decreased insulin sensitivity have critical roles in the development of cardiometabolic diseases. Therefore, our objective was to investigate the dose-response effects of consuming high fructose corn syrup (HFCS)-sweetened beverages for two weeks on hepatic lipid content and insulin sensitivity in young (18–40 years) adults (BMI 18–35 kg/m²). In a parallel, double-blinded study, participants consumed three beverages/day providing 0% (aspartame: n = 23), 10% (n = 18), 17.5% (n = 16), or 25% (n = 28) daily energy requirements from HFCS. Magnetic resonance imaging for hepatic lipid content and oral glucose tolerance tests (OGTT) were conducted during 3.5-day inpatient visits at baseline and again at the end of a 15-day intervention. During the 12 intervening outpatient days participants consumed their usual diets with their assigned beverages. Significant linear dose-response effects were observed for increases of hepatic lipid content (p = 0.015) and glucose and insulin AUCs during OGTT (both p = 0.0004), and for decreases in the Matsuda (p = 0.0087) and Predicted M (p = 0.0027) indices of insulin sensitivity. These dose-response effects strengthen the mechanistic evidence implicating consumption of HFCS-sweetened beverages as a contributor to the metabolic dysregulation that increases risk for nonalcoholic fatty liver disease and type 2 diabetes.     

Spasmolytic effects, mode of action, and structure-activity relationships of stilbenoids from Nidema boothii

A CH(2)Cl(2)-MeOH (1:1) extract prepared from the whole plant of Nidema boothii inhibited spontaneous contractions (IC(50) = 6.26 +/- 2.5 microg/mL) of the guinea-pig ileum. Bioassay-guided fractionation of the active extract led to the isolation of the novel spiro compound 1, which was given the trivial name nidemone, and the new dihydrophenanthrene 3, characterized as 1,5,7-trimethoxy-9,10-dihydrophenanthrene-2,6-diol. In addition, the known stilbenoids aloifol II (2), 1,5,7-trimethoxyphenanthrene-2,6-diol (4), ephemeranthoquinone (5), gigantol (6), ephemeranthol B (7), 2,4-dimethoxyphenanthrene-3,7-diol (8), lusianthridin (9), and batatasin III (10) were obtained. The isolates were characterized structurally by spectroscopic data interpretation. Compounds 2-6, 9, and 10 induced notable concentration-dependent inhibition of the spontaneous contractions of the guinea-pig ileum with IC(50) values that ranged between 0.14 and 2.36 microM. Bibenzyl analogues 23-35 were synthesized and tested pharmacologically. The results indicated that for maximum spasmolytic activity the bibenzyls should have oxygenated substituents on both aromatic rings; on the other hand, methylation of free hydroxyl groups as well as the increment of oxygenated groups in relation to compounds 6 and 10 decreased the smooth muscle relaxant activity. It was also demonstrated that bibenzyls 6 and 10 might exert their spasmolytic action not only by a nitrergic mechanism but also by inhibiting CaM-mediated processes.     

Oral Isotretinoin as Part of the Treatment of Cutaneous Aging

BACKGROUND: A number of drugs have been used to prevent aging changes. However, studies of oral isotretinoin, the commonly used acne drug, as an antiaging drug are lacking. OBJECTIVE: To determine improvement in cutaneous aging utilizing oral isotretinoin combined with different procedures of facial rejuvenation. METHODS: Sixty patients ranging in age from 35 to 65 years, in whom additional modalities of rejuvenation were also used, were randomly assigned to receive treatment with oral isotretinoin (10-20 mg three times a week for 2 months, group A). Their results were compared with 60 patients who had undergone the same surgical procedures but with no oral isotretinoin (group B). RESULTS: All patients treated with oral isotretinoin noted improvement in wrinkles, thickness and color of the skin, size of pores, skin elasticity, tone, and reduction in pigmented lesions and mottled hyperpigmentation. A statistically significant difference was found in the improvement of group A (Wilcoxon test <0.01). Using minimal amounts of this drug, the side effects were practically negligible. CONCLUSION: Utilizing various procedures with oral isotretinoin allowed us to improve the effects of cutaneous aging. Our results using isotretinoin in these cases have been satisfactory. We believe that this is one of the first reports of the use of oral isotretinoin in intrinsic and photoaged skin.     

Identification of new meningococcal serogroup B surface antigens through a systematic analysis of neisserial genomes

The difficulty of inducing an effective immune response against the Neisseria meningitidis serogroup B capsular polysaccharide has lead to the search for vaccines for this serogroup based on outer membrane proteins. The availability of the first meningococcal genome (MC58 strain) allowed the expansion of high-throughput methods to explore the protein profile displayed by N. meningitidis. By combining a pan-genome analysis with an extensive experimental validation to identify new potential vaccine candidates, genes coding for antigens likely to be exposed on the surface of the meningococcus were selected after a multistep comparative analysis of entire Neisseria genomes. Eleven novel putative ORF annotations were reported for serogroup B strain MC58. Furthermore, a total of 20 new predicted potential pan-neisserial vaccine candidates were produced as recombinant proteins and evaluated using immunological assays. Potential vaccine candidate coding genes were PCR-amplified from a panel of representative strains and their variability analyzed using maximum likelihood approaches for detecting positive selection. Finally, five proteins all capable of inducing a functional antibody response vs N. meningitidis strain CU385 were identified as new attractive vaccine candidates: NMB0606 a potential YajC orthologue, NMB0928 the neisserial NlpB (BamC), NMB0873 a LolB orthologue, NMB1163 a protein belonging to a curli-like assembly machinery, and NMB0938 (a neisserial specific antigen) with evidence of positive selection appreciated for NMB0928. The new set of vaccine candidates and the novel proposed functions will open a new wave of research in the search for the elusive neisserial vaccine.     

Lipoprotein NMB0928 from Neisseria meningitidis serogroup B as a novel vaccine candidate

Polysaccharide-based vaccines for serogroup B Neisseria meningitidis have failed to induce protective immunity. As a result, efforts to develop vaccines for serogroup B meningococcal disease have mostly focused on outer membrane proteins (OMP). Vaccine candidates based on meningococcal OMP have emerged in the form of outer membrane vesicles (OMVs) or, more recently, purified recombinant proteins, as alternative strategies for serogroup B vaccine development. In our group, the protein composition of the Cuban OMVs-based vaccine VA-MENGOC-BC was elucidated using two-dimensional gel electrophoresis and mass spectrometry. The proteomic map of this product allowed the identification of new putative protective proteins not previously reported as components of an antimeningococcal vaccine. In the present study, we have determined the immunogenicity and protective capacity of NMB0928, one of those proteins present in the OMVs. The antigen was obtained as a recombinant protein in Escherichia coli, purified and used to immunize mice. The antiserum produced against the protein was capable to recognize the natural protein in different meningococcal strains by whole-cell ELISA and Western blotting. After immunization, recombinant NMB0928 induced bactericidal antibodies, and when the protein was administered inserted into liposomes, the elicited antibodies were protective in the infant rat model. These results suggest that NMB0928 is a novel antigen worth to be included in a broadly protective meningococcal vaccine.