DNA topoisomerases participate in fragility of the oncogene RET

Fragile site breakage was previously shown to result in rearrangement of the RET oncogene, resembling the rearrangements found in thyroid cancer. Common fragile sites are specific regions of the genome with a high susceptibility to DNA breakage under conditions that partially inhibit DNA replication, and often coincide with genes deleted, amplified, or rearranged in cancer. While a substantial amount of work has been performed investigating DNA repair and cell cycle checkpoint proteins vital for maintaining stability at fragile sites, little is known about the initial events leading to DNA breakage at these sites. The purpose of this study was to investigate these initial events through the detection of aphidicolin (APH)-induced DNA breakage within the RET oncogene, in which 144 APHinduced DNA breakpoints were mapped on the nucleotide level in human thyroid cells within intron 11 of RET, the breakpoint cluster region found in patients. These breakpoints were located at or near DNA topoisomerase I and/or II predicted cleavage sites, as well as at DNA secondary structural features recognized and preferentially cleaved by DNA topoisomerases I and II. Co-treatment of thyroid cells with APH and the topoisomerase catalytic inhibitors, betulinic acid and merbarone, significantly decreased APH-induced fragile site breakage within RET intron 11 and within the common fragile site FRA3B. These data demonstrate that DNA topoisomerases I and II are involved in initiating APH-induced common fragile site breakage at RET, and may engage the recognition of DNA secondary structures formed during perturbed DNA replication

Hemodialysis Removes Uremic Toxins That Alter the Biological Actions of Endothelial Cells

Chronic kidney disease is linked to systemic inflammation and to an increased risk of ischemic heart disease and atherosclerosis. Endothelial dysfunction associates with hypertension and vascular disease in the presence of chronic kidney disease but the mechanisms that regulate the activation of the endothelium at the early stages of the disease, before systemic inflammation is established remain obscure. In the present study we investigated the effect of serum derived from patients with chronic kidney disease either before or after hemodialysis on the activation of human endothelial cells in vitro, as an attempt to define the overall effect of uremic toxins at the early stages of endothelial dysfunction. Our results argue that uremic toxins alter the biological actions of endothelial cells and the remodelling of the extracellular matrix before signs of systemic inflammatory responses are observed. This study further elucidates the early events of endothelial dysfunction during toxic uremia conditions allowing more complete understanding of the molecular events as well as their sequence during progressive renal failure

This publication is available at Army

authorized to DOD and DOD contractors to protect operational information from automatic dissemination under the International Exchange Program or by other means. This determination was made on 7 January 2003. Other requests will be referred to HQ TRADOC, ATTN: ATDO

Neuraminidases 3 and 4 regulate neuronal function by catabolizing brain gangliosides

Sialylated glycolipids, gangliosides play an essential role in the central nervous system regulating recognition and signaling in neurons. Metabolic blocks in processing and catabolism of gangliosides result in development of severe neurological disorders, gangliosidoses manifesting with neurodegeneration and neuroinflammation. We demonstrate that two mammalian enzymes, neuraminidases 3 and 4 play important role in catabolic processing of brain gangliosides by cleaving terminal sialic acid residues in their glycan chains. In the neuraminidase 3-4 double knockout mice, GM3 ganglioside is stored in microglia, vascular pericytes and neurons, causing micro- and astrogliosis, neuroinflammation, accumulation of lipofuscin bodies and memory loss whereas their cortical and hippocampal neurons have lower rate of neuritogenesis in vitro. Double knockout mice also have reduced levels of GM1 ganglioside and myelin in neuronal axons. Besides, neuraminidase 3 deficiency drastically increased storage of GM2 in the brain tissues of the asymptomatic mouse model of the severe human gangliosidosis, Tay-Sachs disease indicating that this enzyme is responsible for the metabolic bypass of the β-hexosaminidase A deficiency. Together, our results provide the first in vivo evidence that neuraminidases 3 and 4 have important roles in CNS function by catabolizing gangliosides and preventing their storage in lipofuscin bodies

Neuraminidases 3 and 4 regulate neuronal function by catabolizing brain gangliosides

Sialylated glycolipids, gangliosides play an essential role in the central nervous system regulating recognition and signaling in neurons. Metabolic blocks in processing and catabolism of gangliosides result in development of severe neurological disorders, gangliosidoses manifesting with neurodegeneration and neuroinflammation. We demonstrate that two mammalian enzymes, neuraminidases 3 and 4 play important role in catabolic processing of brain gangliosides by cleaving terminal sialic acid residues in their glycan chains. In the neuraminidase 3-4 double knockout mice, GM3 ganglioside is stored in microglia, vascular pericytes and neurons, causing micro- and astrogliosis, neuroinflammation, accumulation of lipofuscin bodies and memory loss whereas their cortical and hippocampal neurons have lower rate of neuritogenesis in vitro. Double knockout mice also have reduced levels of GM1 ganglioside and myelin in neuronal axons. Besides, neuraminidase 3 deficiency drastically increased storage of GM2 in the brain tissues of the asymptomatic mouse model of the severe human gangliosidosis, Tay-Sachs disease indicating that this enzyme is responsible for the metabolic bypass of the β-hexosaminidase A deficiency. Together, our results provide the first in vivo evidence that neuraminidases 3 and 4 have important roles in CNS function by catabolizing gangliosides and preventing their storage in lipofuscin bodies