Oxidative damage to DNA has been implicated in carcinogenesis during chronic inflammation. Epidemiological and biochemical studies suggest that one potential mechanism involves myeloperoxidase, a hemeprotein secreted by human phagocytes. In this study, we demonstrate that human neutrophils use myeloperoxidase to oxidize uracil to 5-chlorouracil in vitro. Uracil chlorination by myeloperoxidase or reagent HOCl exhibited an unusual pH dependence, being minimal at pH ϳ5, but increasing markedly under either acidic or mildly basic conditions. This bimodal curve suggests that myeloperoxidase initially produces HOCl, which subsequently chlorinates uracil by acid-or base-catalyzed reactions. Human neutrophils use myeloperoxidase and H 2 O 2 to chlorinate uracil, suggesting that nucleobase halogenation reactions may be physiologically relevant. Using a sensitive and specific mass spectrometric method, we detected two products of myeloperoxidase, 5-chlorouracil and 5-bromouracil, in neutrophil-rich human inflammatory tissue. Myeloperoxidase is the most likely source of 5-chlorouracil in vivo because halogenated uracil is a specific product of the myeloperoxidase system in vitro. In contrast, previous studies have demonstrated that 5-bromouracil could be generated by either eosinophil peroxidase or myeloperoxidase, which preferentially brominates uracil at plasma concentrations of halide and under moderately acidic conditions. These observations indicate that the myeloperoxidase system promotes nucleobase halogenation in vivo. Because 5-chlorouracil and 5-bromouracil can be incorporated into nuclear DNA, and these thymine analogs are well known mutagens, our observations raise the possibility that halogenation reactions initiated by phagocytes provide one pathway for mutagenesis and cytotoxicity at sites of inflammation.

Blood coagulation capacity increases with age in healthy individuals. Through extensive longitudinal analyses of human factor IX gene expression in transgenic mice, two essential age-regulatory elements, AE5' and AE3', have been identified. These elements are required and together are sufficient for normal age regulation of factor IX expression. AE5', a PEA-3 related element present in the 5' upstream region of the gene encoding factor IX, is responsible for age-stable expression of the gene. AE3', in the middle of the 3' untranslated region, is responsible for age-associated elevation in messenger RNA levels. In a concerted manner, AE5' and AE3' recapitulate natural patterns of the advancing age-associated increase in factor IX gene expression.

Somatic mutations induced by oxidative damage of DNA might play important roles in atherogenesis. However, the underlying mechanisms remain poorly understood. Myeloperoxidase, a heme protein expressed by select populations of artery wall macrophages, initiates one potentially mutagenic pathway by generating hypochlorous acid. This potent chlorinating agent reacts rapidly with primary amines to yield long-lived, selectively reactive N-chloramines. In the current studies, we demonstrate that myeloperoxidase produced by human macrophages differentiated in the presence of granulocyte macrophage colony-stimulating factor generates 5-chlorouracil, a mutagenic thymine analog. The primary amine taurine fails to block the reaction, suggesting that N-haloamines produced by macrophages might oxidize uracil. Model system studies demonstrated that N-chloramines convert uracil to 5-chlorouracil. Interestingly, the tertiary amine nicotine dramatically enhances uracil chlorination, suggesting that cigarette smoke might promote nucleobase oxidation by N-chloramines. To look for evidence that myeloperoxidase promotes uracil oxidation in vivo, we measured 5-chlorouracil levels in human aortic tissue, using isotope dilution gas chromatography-mass spectrometry. The level of 5-chlorouracil was 10-fold higher in atherosclerotic aortic tissue obtained during vascular surgery than in normal aortic tissue, suggesting that halogenated nucleobases produced by macrophages might contribute to atherogenesis. Because 5-chlorouracil can be incorporated into nuclear DNA, our observations raise the possibility that halogenation reactions initiated by phagocytes provide one pathway for mutagenesis, phenotypic modulation, and cytotoxicity during atherogenesis.

In reactions between styrene oxide and the ring nitrogen at the 1-position of deoxyadenosine, the epoxide is opened at both the alpha- (benzylic) and beta-carbons. The 1-substituted nucleosides formed are unstable and subsequently undergo either Dimroth rearrangement to give N6-substituted deoxyadenosines or deamination to give 1-substituted deoxyinosines. alphaN6-Substituted compounds are also formed from direct reaction at the exocyclic nitrogen. Kinetic experiments revealed that relative rates of deamination of 1-substituted deoxyadenosine-styrene oxides and 1-substituted adenosine-styrene oxides were similar. However, the rate of Dimroth rearrangement in beta1-substituted adenosine-styrene oxides was approximately 2.3-fold greater than that of beta1-substituted deoxyadenosine-styrene oxides and approximately 1.5-fold greater in alpha1-substituted adenosine-styrene oxides relative to alpha1-substituted deoxyadenosine-styrene oxides. Analysis of the products formed from reactions of styrene oxide with [3H]deoxyadenosine and [3H]deoxyadenosine incorporated into native and denatured DNA showed that the double-helical DNA structure reduced the levels of adducts formed 5-fold relative to denatured DNA but did not present a complete barrier to formation of either N6-substituted deoxyadenosine- or 1-substituted deoxyinosine-styrene oxide adducts in native DNA. Additionally, in denatured and native DNA the product distributions were altered in favor of formation of beta1-substituted deoxyinosine-styrene oxide adducts with respect to reactions of the nucleoside. The ratio of retained to inverted configuration of alphaN6-substituted products was higher in DNA than in nucleoside reactions. These experiments indicate that in addition to the N6-position, the ring nitrogen at the 1-position of deoxyadenosine is available, to some extent, for reaction in native DNA. In styrene oxide-DNA reactions, formation of 1-substituted adenines can lead to deaminated products where both Watson-Crick hydrogen-bonding sites are disrupted.

Background: We evaluated the clinical impact of cytomegalovirus (CMV) reactivation calculated in terms of the area under the curve of CMV antigenemia (CMV-AUC) on the development of invasive mold infection (IMI) in the post-engraftment phase after allogeneic hematopoietic stem cell transplantation (HSCT). Methods: Among 394 consecutive patients who underwent their first allogeneic HSCT at our center between 2007 and 2018, 335 were included after excluding patients with a past history of invasive fungal disease (IFD), the development of IFD before engraftment, engraftment failure, or early death within 30 days. CMV antigenemia (CMV-AG) was monitored weekly after engraftment and 3 or more cells/2 slides were regarded as positive. CMV-AUC was calculated by the trapezoidal method using the number of CMV-AG after logarithmic transformation and the duration in weeks and was added until negative conversion. Patients with CMV reactivation were divided into low and high CMV-AUC groups using the median value of CMV-AUC as a threshold. Results: There were 17 proven/probable IMIs including one mucormycosis and 16 probable invasive aspergillosis, and the 2-year cumulative incidence was 1.0% in the negative CMV-AUC group (n = 136), 3.3% in the low CMV-AUC group (n = 98) and 13.8% in the high CMV-AUC group (n = 101) (P = .001). In a multivariate analysis, grade II-IV acute GVHD (HR 3.74) and CMV-AUC (HR low 1.25, high 5.91) were identified as independent significant factors associated with a higher incidence of IMI. Conclusions: Cytomegalovirus kinetics evaluated in terms of CMV-AUC were significantly associated with the development of IMI in the post-engraftment phase after allogeneic HSCT. How to cite this article: Kimura S-I, Takeshita J, Kawamura M, et al. Association between the kinetics of cytomegalovirus reactivation evaluated in terms of the area under the curve of cytomegalovirus antigenemia and invasive mold infection during the post-engraftment phase after allogeneic hematopoietic stem cell transplantation. Transpl Infect Dis.

No abstract

Background: We retrospectively evaluated the association between the D-index, which reflects both the depth and duration of neutropenia, and proven/probable invasive fungal disease (IFD) early after allogeneic hematopoietic stem cell transplantation (HSCT) at our center (n = 394). Methods: The D-index was defined as the area over the neutrophil curve during neutropenia. The cumulative D-index from the start of neutropenia until the development of infection (c-D-index) was also evaluated as a real-time assessment of neutropenia. Results: There were 19 cases of early proven/probable IFD before and within 1 week after engraftment. Fifteen cases (78.9%) were seen as breakthrough infection while on empiric (n = 7), preemptive (n = 4) or prophylactic (n = 4) antifungal administration with mold-active agents. The c-D-index and lower performance status were identified as independent significant predictive factors for IFD. A receiver operating characteristic (ROC) curve analysis showed that the D-index and c-D-index were more accurate than the simple duration of neutropenia and as accurate as the duration of profound neutropenia for predicting IFD. The sensitivity, specificity, and positive and negative predictive values of the c-D-index using an appropriate cutoff (CO) value (10 644) determined by ROC curve analysis were 73.1%, 63.2%, 9.1%, and 97.9%, respectively. The advantage of the c-D-index to cumulative days of neutropenia in terms of positive and negative predictive values seemed to be small. Conclusions: The appropriate CO value for the c-D-index for predicting IFD was as high as 10 644 in allogeneic HSCT with a more frequent use of empiric antifungal therapy. The c-D-index is useful for assessing the risk of breakthrough IFD.

No abstract