Citations

Objective— We tested the hypothesis that in early hypertension (HT), increased oxidative stress leads to myocardial microvascular remodeling. Methods and Results— Pigs were studied after a 12-week observation: normal (n=8), untreated renovascular HT (n=8), or HT+chronic antioxidant supplementation (HT+A, n=6). Left ventricular muscle mass (LVMM) and myocardial blood flow (MBF) reserve were determined using electron beam computer tomography (CT), and the spatial density and tortuousity of myocardial microvessels (<500 μm) was then measured in myocardial samples with micro-CT. Myocardial microvascular morphology, oxidative stress, inflammation, and growth factor expression were determined in vitro. HT and HT+A had similarly increased arterial pressure and LVMM, but only HT showed impaired MBF response to adenosine. Compared with normal, HT had increased spatial density of myocardial microvessels, which was preserved in HT+A (111.8±7.8, 166.3±15.7, and 106.4±6.1 vessels per cm 2 , respectively). HT also showed microvascular wall thickening, increased systemic and tissue oxidative stress, inflammation, and expression of vascular endothelial growth factor and its receptor Flk-1, most of which were attenuated by antioxidants. Conclusions— Myocardial microvascular remodeling in early HT is accompanied by tissue oxidative stress, inflammation, and altered growth factor expression, and attenuated by antioxidant intervention. This study underscores a role of increased oxidative stress in modulating myocardial microvascular architecture in early HT.

Experimental studies have shown the beneficial effects of antioxidant supplementation on endothelial function in the presence of increased endogenous oxidative stress, whereas limited data are available under normal conditions. The present study tested the hypothesis that in normal pigs long-term antioxidants would have deleterious effects on the cardiovascular system. Normal domestic pigs (V, n=6) were studied 12 weeks after dietary supplementation with vitamin E (100 IU/kg per day) and vitamin C (1 g/day) and compared with normal controls (C, n=7). Myocardial perfusion and permeability index were evaluated by electron beam computed tomography after intravenous adenosine and dobutamine. Coronary endothelial function was evaluated in vitro by organ chamber and coronary tissue studied by immunoblotting and staining. Myocardial perfusion response was lower in V than in C after adenosine (10.1±4.5 versus 53.4±5.2%; P <0.01) and dobutamine (V, 78.4±8.1; C, 193.0±39.0%; P <0.05). The permeability index increased in V after adenosine (48.8±5.1%) and dobutamine (59.9±13.6%) and did not change in C. Coronary vasodilation to bradykinin and substance P was lower in V than in C. Moreover, in V, coronary nitrotyrosine and superoxide content was significantly higher than in C. The groups had similar total monomer expression of endothelial nitric oxide synthase, whereas the dimerized form, reflecting coupled enzyme, was lower in V. These findings suggest that long-term experimental antioxidant vitamin supplementation in normal pigs impairs myocardial perfusion and coronary endothelial function via an increased level of oxidative stress in the arterial wall, which may be partly related to the uncoupling of endothelial nitric oxide synthase and/or the direct prooxidant effect of vitamin radicals.

ABSTRACT Mycobacterium peregrinum consists of two taxa: types I and II. We evaluated 43 clinical type II strains from throughout the United States. They were responsible for soft-tissue and bone infections, catheter-related infections, and possible pneumonitis. By carbohydrate utilization, they were indistinguishable from type I strains, being d -mannitol and trehalose positive. However, they had a distinct susceptibility pattern that included intermediate ciprofloxacin MICs but low clarithromycin and doxycycline MICs of ≤1 μg/ml. These features were also shared by reference isolates of Mycobacterium senegalense from African bovine cases of “farcy.” By 16S rRNA gene sequencing, the type II isolates shared 100% sequence identity with M. senegalense . Partial sequencing of the type II hsp65 gene (441 bp) revealed four sequevars showing ≥98.4% identity with each other and ≥98.6% identity with the sequence of five bovine strains of M. senegalense . There was ≤97.1% identity with M. peregrinum type I isolates and other Mycobacterium fortuitum group species. Sequencing of additional gene targets including the 16S-23S rDNA internal transcribed spacer region and the rpoB gene (partial sequence) revealed a similar phylogenetic grouping. DNA-DNA hybridization showed 76 to 99% relatedness between the bovine and human strains. These studies demonstrate that type II isolates are not isolates of M. peregrinum but represent human strains of M. senegalense . This study is the first to demonstrate this species as a human pathogen. Representative human M. senegalense strains include ATCC 35755 and newly submitted strains ATCC BAA-849, ATCC BAA-850, and ATCC BAA-851.

Epigenetic inheritance of heterochromatin structure is an important cellular process whose mechanism remains elusive. In this article, we describe the identification of nine enhancers of the silencing defect of a Saccharomyces cerevisiae -PCNA mutant by screening a library of ≈4,700 viable yeast deletion mutants. Of the nine mutants identified, six ( hir1 , hir3 , sas2 , sas4 , sas5 , and sir1 ) were previously known to reduce silencing synergistically with a mutation in Cac1p, the large subunit of chromatin assembly factor 1 (CAF-1). The predicted gene products that are affected in three other mutants ( nam7 , msh2 , and rtt106 ) have not been implicated previously in silencing. Characterization of the rtt106 Δ allele revealed that it synergistically reduced heterochromatin silencing when combined with a mutation in Cac1p but not with a mutation in Asf1p (a histone H3 and H4 chaperone). Moreover, Rtt106p interacted with histones H3 and H4 both in vitro and in vivo , and it displayed a nucleosome assembly activity in vitro . Furthermore, Rtt106p interacts with CAF-1 physically through Cac1p. These biochemical and genetic data indicate that Rtt106p is a previously uncharacterized histone chaperone connecting S phase to epigenetic inheritance.

The human HtrA family of proteases consists of four members: HtrA1, HtrA2, HtrA3, and HtrA4. In humans the four HtrA homologues appear to be involved in several important functions such as cell growth, apoptosis, and inflammatory reactions, and they control cell fate via regulated protein metabolism. In previous studies it was shown that the expression of HtrA1 was ubiquitous in normal adult human tissues. Here we examined the expression of HtrA1 protein and its corresponding mRNA during mouse embryogenesis using Northern blotting hybridization, RT-PCR, and immunohistochemical staining analyses. Our results indicate that HtrA1 is expressed in a variety of tissues in mouse embryos. Furthermore, this expression is regulated in a spatial and temporal manner. Relatively low levels of HtrA1 mRNA are detected in embryos at the beginning of organogenesis (E8), and the levels of expression increase during late organogenesis (E14-E19). Our results show that HtrA1 was expressed during embryonic development in specific areas where signaling by TGFβ family proteins plays important regulatory roles. The expression of HtrA1, documented both at mRNA and protein levels by RT-PCR and immunohistochemistry in the developing nervous system, is consistent with a possible role of this protein both in dividing and postmitotic neurons, possibly via its documented inhibitory effects on TGFβ proteins. An exhaustive knowledge of the different cell- and tissue-specific patterns of expression of HtrA1 in normal mouse embryos is essential for a critical evaluation of the exact role played by this protein during development.