Abstract Alterations of the mitochondrial DNA (mtDNA) have been described in human tumors and in other tissues in association with smoking exposure. We did quantitative PCR of cytochrome c oxidase I (Cox I) and cytochrome c oxidase II (Cox II) genes on oral rinse samples obtained from 94 patients with primary head and neck squamous cell carcinoma (HNSC) and a control group of 656 subjects. Mitochondrial DNA/nuclear DNA in saliva from HNSC patients and controls in relationship to smoking exposure, ethanol intake, and tumor stage were examined. Mean levels of Cox I and Cox II in saliva samples were significantly higher in HNSC patients: Cox I, 0.076 [95% confidence interval (95% CI), 0.06-0.09] and Cox II, 0.055 (95% CI, 0.04-0.07) in comparison with controls Cox I, 0.054 (95% CI, 0.05-0.06), P < 0.0001 and Cox II, 0.046 (95% CI, 0.04-0.05), P = 0.003 (t test). MtDNA levels were elevated in primary tumors when compared with matched, pretreatment saliva and significant correlation was noted (Cox I, r = 0.30, P = 0.005 and Cox II r = 0.33, P = 0.002, respectively, Pearson's correlation). On univariate analysis, smoking, age, HNSC diagnosis, and advanced stage of HNSC were associated with higher level of mtDNA content in saliva. Multivariate analysis showed a significant and independent association of HNSC diagnosis, age, and smoking with increasing mtDNA/nuclear DNA for Cox I and Cox II. mtDNA content alteration is associated with HNSC independently of age and smoking exposure, can be detected in saliva, and may be due to elevation in mtDNA content in primary HNSC.
Abstract Multiple mechanisms are used by the human immunodeficiency virus type 1 (HIV-1) to interfere with host-cell immune effector functions. The 27-kD Nef protein has been shown to down-modulate specific genes of the major histocompatibility complex class I (MHC-I) on the surface of infected pimary T cells, facilitating their escape from lysis by cytolytic T lymphocytes. Macrophages, as the other major immune cell type targeted by the virus, also contribute to the transmission, persistence, and pathogenesis of HIV-1. Yet, whether Nef modulates MHC-I expression on HIV-infected primary macrophages remains unclear. Currently available infectious HIV-1 molecular clones, which express a reporter gene, only infect T cells and/or do not express Nef. To overcome these limitations, we generated macrophage-tropic green fluorescent protein (GFP)-tagged HIV-1 viruses, which express the complete viral genome, and used these to assess the expression of human leukocyte antigen (HLA)-A2 on the surface of productively infected macrophages. The reporter viral genomes were replication-competent and stable, as Nef, p24 antigen, and GFP expression could be detected by immunostaining of infected, monocyte-derived macrophages (MDM) after more than 2 months postinfection. Fluorescence-activated cell sorter analyses of infected macrophages and T cells revealed that although wild-type reporter virus infection induced a statistically significant decrease in the density of surface HLA-A2, down-regulation of HLA-A2 was not seen in cells infected with reporter viruses encoding a frameshift or a single point mutation in Nef at prolines 74P and P80. The impact of Nef on HLA-A2 surface expression in MDM was also confirmed by confocal microscopy. These results suggest that the mechanisms of HLA-A2 down-modulation are similar in primary T cells and macrophages.
Candida glabrata is an opportunistic pathogen of humans, responsible for up to 30% of disseminated candidiasis. Adherence of C. glabrata to host cells is mediated by adhesin-like proteins (ALPs), about half of which are encoded in the subtelomeres. We performed a de novo assembly of two C. glabrata strains, BG2 and BG3993, using long single-molecule real-time (SMRT) reads, and constructed high-quality telomere-to-telomere assemblies of all 13 chromosomes to assess differences between C. glabrata strains. We documented variation between strains, and in agreement with earlier studies, found high (~0.5%-1%) frequencies of SNVs across the genome, including within subtelomeric regions. We documented changes in ALP gene structure and complement: there are large length differences in ALP genes in different strains, resulting from copy number variation in tandem repeats. We compared strains to characterize chromosome rearrangement events including within the poorly characterized subtelomeric regions. We show that rearrangements within the subtelomere regions all affect ALP-encoding genes, and 14/16 involve just the most terminal ALP gene. We present evidence that these rearrangements are mediated by break-induced replication. This study highlights the constrained nature of subtelomeric changes impacting ALP gene complement and subtelomere structure.
The formation of large (approximately 20-35 nm) surface hemimicelles in monolayers of semifluorinated alkanes, C(n)F(2)(n)(+1)C(m)H(2)(m)(+1) (FnHm), observed after transfer onto silicon wafers, is a general phenomenon. F6H16 and F8H14 exclusively form highly monodisperse circular hemimicelles, organized in a hexagonal array. The other FnHm investigated form both circular and elongated hemimicelles. The longer FnHm is, the larger the area fraction of elongated micelles; both the hydrocarbon block (H-block) and the fluorocarbon block (F-block) affect this area fraction. The length of the elongated micelles increases with the total length of the diblocks. The diameter of the circular micelles increases with the length of the H-block but, unexpectedly, not with that of the F-block. Model calculations account for these observations. Close examination of the circular micelles showed that they generally present a pit or a tip at their center. The width of the elongated micelles is comparable to the radius of the circular micelles, suggesting that the latter arise from a partition of elongated micelles, followed by coalescence of the edges of the resulting fragments. The elongated micelles become shorter and fewer when surface pressure increases, further suggesting a conversion of elongated into circular micelles. This conversion is reversible. The surface pressure-molecular area isotherms do not present any feature that forebears the existence of hemimicelles. The obtaining of stable surface patterns from simple, "nonpolar" molecular fluorocarbon/hydrocarbon diblocks opens a new approach for producing featured nanostructures from organic templates.
The p53 gene is frequently mutated in non-small-cell lung cancer (NSCLC); however, the effect of p53 gene mutations on patient prognosis remains unclear. Therefore, we initiated a prospective study to determine the association of p53 gene mutations with survival in patients with stage I NSCLC.Tumor samples were collected prospectively from 188 patients with operable NSCLC (stages I, II, and IIIA). p53 mutations were detected by direct dideoxynucleotide sequencing and p53 GeneChip analysis. Association of clinical and pathologic variables (e.g., alcohol consumption, sex, age, pathologic stage) with mutation of the p53 gene was determined by logistic regression. Associations between p53 mutation status, clinical and pathologic variables, and survival were assessed using a Cox proportional hazards regression model. All statistical tests were two-sided.p53 mutations were detected in 55% (104/188) of tumors. These mutations were associated with non-bronchoalveolar tumors, a history of alcohol consumption, and younger patient age. The risk of death was statistically significantly higher in patients with p53 mutations in their tumors (hazard ratio [HR] = 1.6, 95% confidence interval [CI] = 1.0 to 2.4; P =.049) than in patients with wild-type p53 in their tumors. Tumor stage, the presence of a p53 mutation, and increasing patient age were statistically significant predictors of patient death in the entire patient group; however, the statistically significant prognostic effect of p53 mutation was limited to patients with stage I NSCLC (stage I HR = 2.8, 95% CI = 1.4 to 5.6; stage II HR = 1.8, 95% CI = 0.74 to 4.4; and stage III HR = 0.70, 95% CI = 0.32 to 1.5). Among patients with stage I NSCLC, actuarial 4-year survival was statistically significantly higher in those with wild-type p53 than in those with mutant p53 (78% versus 52%, respectively; difference in 4-year survival = 26%, 95% CI = 6% to 46%; P =.009, log-rank test).Tumor p53 mutations are a statistically significant predictor of poor outcome in patients with stage I NSCLC.
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