Postpartum fevers, a rare presentation of secondary hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a rare disease of excessive immune system activation. We report a case of HLH in a 20-year-old primigravid woman who presented with postpartum fevers. She was successfully treated with dexamethasone and anakinra, a deviation from the HLH-94 protocol, to preserve her ability to breastfeed.

Epstein-Barr virus-associated central nervous system lymphoproliferative disease in a patient with acquired immunodeficiency syndrome responsive to highly active antiretroviral therapy.

A 20-year-old man with acquired immunodeficiency syndrome (AIDS) and central nervous system (CNS) lymphoproliferative disease experienced improvement with highly active antiretroviral therapy (HAART) without radiation therapy. Our experience highlights the importance of biopsy in evaluating multifocal radiographic CNS lesions and the central role of HAART in treating AIDS-related CNS disease.

New markers of pancreatic cancer identified through differential gene expression analyses: claudin 18 and annexin A8.

BACKGROUND: New markers to distinguish benign reactive glands from infiltrating ductal adenocarcinoma of the pancreas are needed. DESIGN: The gene expression patterns of 24 surgically resected primary infiltrating ductal adenocarcinomas of the pancreas were compared with 18 non-neoplastic samples using the Affymetrix U133 Plus 2.0 Arrays and the Gene Logic GeneExpress Software System. Gene fragments from 4 genes (annexin A8, claudin 18, CXCL5, and S100 A2) were selected from the fragments found to be highly expressed in infiltrating adenocarcinomas when compared with normal tissues. The protein expression of these genes was examined using immunohistochemical labeling of tissue microarrays. RESULTS: Claudin 18 labeled infiltrating carcinomas in a membranous pattern. When compared with normal and reactive ducts, claudin 18 was overexpressed, at least focally, in 159 of 166 evaluable carcinomas (96%). Strong and diffuse claudin 18 overexpression was most often seen in well-differentiated carcinomas (P=0.02). Claudin 18 was overexpressed in 51 of 52 cases (98%) of pancreatic intraepithelial neoplasia. Annexin A8 was at least focally overexpressed in 149 of 154 evaluable infiltrating carcinomas (97%). S100 A2 was at least focally overexpressed in 118 of 154 evaluable infiltrating carcinomas (77%). Non-neoplastic glands also frequently expressed S100 A2 diminishing its potential diagnostic utility. Immunolabeling with antibodies directed against CXCL5 did not reveal any significant differences in protein expression between infiltrating adenocarcinomas and normal pancreatic ducts. CONCLUSIONS: Claudin 18 and annexin A8 are frequently highly overexpressed in infiltrating ductal adenocarcinomas when compared with normal reactive ducts, suggesting a role for these molecules in pancreatic ductal adenocarcinomas. Furthermore, these may serve as diagnostic markers, as screening tests and as therapeutic targets.

Generation and characterization of endonuclease G null mice.

Endonuclease G (endo G) is one of the most abundant nucleases in eukaryotic cells. It is encoded in the nucleus and imported to the mitochondrial intermembrane space. This nuclease is active on single- and double-stranded DNA. We genetically disrupted the endo G gene in mice without disturbing a conserved, overlapping gene of unknown function that is oriented tail to tail with the endo G gene. In these mice, the production of endo G protein is not detected, and the disruption abolishes the nuclease activity of endo G. The absence of endo G has no effect on mitochondrial DNA copy number, structure, or mutation rate over the first five generations. There is also no obvious effect on nuclear DNA degradation in standard apoptosis assays. The endo G null mice are viable and show no age-related or generational abnormalities anatomically or histologically. We infer that this highly conserved protein has no mitochondrial or apoptosis function that can discerned by the assays described here and that it may have a function yet to be determined. The early embryonic lethality of endo G null mice recently reported by others may be due to the disruption of the gene that overlaps the endo G gene.

DNA damage and aging.

The hypothesis discussed here is that a major component of aging in metazoans is oxidative damage to nuclear DNA. Such a viewpoint would be consistent with the fact that all of the thus far identified premature aging syndromes in mammals involve mutations in nuclear proteins. Several of these nuclear proteins are enzymes that are related to DNA metabolism or DNA repair. Among the single- and double-stranded DNA damage repair pathways present in eukaryotes, only one pathway often fails to restore the full information content of the genome and typically would result in a deletion of a few base pairs. This pathway is called nonhomologous DNA end joining (NHEJ) and it is a major pathway for the repair of double-strand DNA breaks. Repetitive DNA content may determine the extent to which any organism can use this pathway, and therefore, may dictate a key factor in the balance between oxidation and organismal lifespan.

Ageing, repetitive genomes and DNA damage.

The mitochondrial production of reactive oxygen species is inversely proportional to longevity in animals. A key question now is, which molecules, among those that are oxidized, affect the lifespan of the organism most significantly?

Developmental retinal apoptosis in Ku86-/- mice.

The nonhomologous DNA end-joining pathway (NHEJ), a major pathway for repairing DNA double-strand breaks (DSBs), is essential for maintaining genomic stability. Knockout animals for components in this pathway demonstrate a distinct pattern of cell death in the developing brain. Here we demonstrate that cell death is also present in the developing retina of E14.5 Ku86-deficient mouse embryos, suggesting that the increase in cell death in the retina is associated with chromosome breaks. In the adult retina, we do not find continuing apoptosis, but interestingly, we find decreased numbers of total neuronal cells. This suggests that the increased retinal apoptosis during embryogenesis causes the reduction in cell numbers observed in the adult retina. This analysis of the retina provides the first opportunity to formally test the hypothesis that embryonic apoptosis accounts for reduced total cell numbers in adult Ku86-/- mice.

Overexpression of Cu/Zn superoxide dismutase is lethal for mice lacking double-strand break repair.

The non-homologous DNA end joining (NHEJ) pathway is a major double-strand DNA break repair pathway in cells of multicellular eukaryotes. Ku is a heterodimeric protein consisting of Ku70 and Ku86, and it is thought to be the first component to bind to a broken double-strand DNA end. Mice lacking Ku86 show features of premature aging, live about 6-12 months, and show a characteristic loss of neurons in the central nervous system during development. Cells from mice lacking Ku have increased numbers of chromosome breaks, a significant fraction of which are caused by oxidative metabolism. Overexpression of the cytoplasmic Cu/Zn superoxide dismutase (SOD1) from a transgene is known to increase the number of chromosome breaks in primary cells (presumably by increasing reactive oxygen species). Here we show that SOD1 overexpression in a Ku86-/- mouse results in embryonic lethality. This striking effect is, however, subject to a strain-specific modifier. Genome-wide marker analysis is most consistent with the modifier being on mouse chromosome 13. Analysis of 10 markers on chromosome 13 suggests that the modifier is within the same region as a modifier of the murine amyotropic lateral sclerosis (ALS) phenotype when it is caused by overexpression of a mutant form of SOD1. Based on these results, we propose a model in which oxidative metabolism causes chromosome breaks, leading to neuronal death; and this neuronal death may account for that seen in NHEJ mutant animals and in mammals with SOD1-mediated ALS.

Two overlapping divergent transcription units in the human genome: the FEN1/C11orf10 locus.

Flap endonuclease 1 (FEN-1) is a nuclear enzyme involved in DNA metabolism, such as replication, repair, and recombination. Here, we report the comparative genomic organization of the chicken, mouse, and human FEN1 genes as well as the comparative organization of a small gene (C11orf10) located immediately upstream of the FEN1 gene in reverse orientation. Immunostaining revealed that the C11orf10 protein, unlike FEN-1, is located in the cytoplasm, suggesting that these two proteins do not form a physical complex. Importantly, in the human genome, the two mRNAs are overlapping (14 bp) in their 5' ends. Thus, the FEN1/C11orf10 locus is a new example of two overlapping, divergent transcription units in the human genome.

Oxygen metabolism causes chromosome breaks and is associated with the neuronal apoptosis observed in DNA double-strand break repair mutants.

Cells deficient in a major DNA double-strand break repair pathway (nonhomologous DNA end joining [NHEJ]) have increased spontaneous chromosome breaks; however, the source of these chromosome breaks has remained undefined. Here, we show that the observed spontaneous chromosome breaks are partially suppressed by reducing the cellular oxygen tension. Conversely, elevating the level of reactive oxygen species by overexpressing the antioxidant enzyme superoxide dismutase 1 (SOD1), in a transgenic mouse, increases chromosome breakage. The effect of SOD1 can also be modulated by cellular oxygen tension. The elevated chromosome breakage correlates histologically with a significant increase in the amount of neuronal cell death in Ku86(-/-) SOD1 transgenic embryos over that seen in Ku86(-/-) embryos. Therefore, oxygen metabolism is a major source of the genomic instability observed in NHEJ-deficient cells and, presumably, in all cells.

The embryonic lethality in DNA ligase IV-deficient mice is rescued by deletion of Ku: implications for unifying the heterogeneous phenotypes of NHEJ mutants.

There are two general pathways by which multicellular eukaryotes repair double-strand DNA breaks (DSB): homologous recombination (HR) and nonhomologous DNA end joining (NHEJ). All mammalian mutants in the NHEJ pathway demonstrate a lack of B and T lymphocytes and ionizing radiation sensitivity. Among these NHEJ mutants, the DNA-PK(cs) and Artemis mutants are the least severe, having no obvious phenotype other than the general defects described above. Ku mutants have an intermediate severity with accelerated senescence. The XRCC4 and DNA ligase IV mutants are the most severe, resulting in embryonic lethality. Here we show that the lethality of DNA ligase IV-deficiency in the mouse can be rescued when Ku86 is also absent. To explain the fact that simultaneous gene mutations in the NHEJ pathway can lead to viability when a single mutant is not viable, we propose a nuclease/ligase model. In this model, disrupted NHEJ is more severe if the Artemis:DNA-PK(cs) nuclease is present in the absence of a ligase, and Ku mutants are of intermediate severity, because the nuclease is less efficient. This model is also consistent with the order of severity in organismal phenotypes; consistent with chromosomal breakage observations reported here; and consistent with the NHEJ mutation identified in radiation sensitive human SCID patients.

Conversion of Allylic Alcohols into Allylic Nitromethyl Compounds via a Palladium-Catalyzed Solvolysis: An Enantioselective Synthesis of an Advanced Carbocyclic Nucleoside Precursor(1).

A two-step reaction sequence to homoallylic nitro compounds from allylic alcohols is presented. Ethoxy carbonylation of the alcohols with ethyl chloroformate provides the corresponding allylic ethyl carbonates in high yields. Exposure of these substrates to catalytic palladium(0) in CH(3)NO(2) initiates a reaction sequence, ionization-decarboxylation-nitromethylation, that culminates with the formation of nitroalkenes. The regio- and stereochemical outcomes of the nitromethyl allylation reaction can be explained by the behavior of the transient pi-allylpalladium complexes. This methodology serves as a centerpiece for the synthesis of an important carbocyclic nucleoside intermediate.