Postirradiation sensitization of mammalian cells by the thiol-depleting agent dimethyl fumarate

Dimethyl fumarate (DMF) depletes intracellular glutathione (GSH) by covalent bond formation in a reaction mediated by GSH-S-transferase. Treatment of hypoxic Chinese hamster V79 cells with 5 mM DMF before irradiation radiosensitizes the cells, resulting in an enhancement ratio (ER) of about 2.7 with minimal toxicity, when the end point is clonogenic cell survival. Under the same conditions aerobic cells are sensitized, and ER of about 1.3 is found, and GSH is reduced to about 3% of control. Very similar results were obtained previously with Chinese hamster ovary (CHO) cells. In addition, new data presented here show that DMF treatment of V79 or CHO cells immediately after irradiation under hypoxic conditions sensitizes the cells, resulting in an ER of about 1.5, DMF treatment after irradiation under aerobic conditions results in an ER of 1.3, and this DMF treatment reduces protein thiols (PSH) to about 70% of control. When induction of DNA damage is measured using the neutral elution assay, treatment of V79 or CHO cells with DMF prior to irradiation under hypoxic conditions results in an ER of 1.9-2.0, but there is no enhancement of DNA damage when DMF is added after irradiation under hypoxic conditions or when cells are treated with DMF before or after irradiation under aerobic conditions. Based on these data we postulate that DMF radiosensitizes killing of hypoxic cells by two actions: depletion of GSH interferes with the chemical competition between damage fixation and repair, and depletion of PSH causes an inhibition of enzymatic repair processes. We also suggest that DMF sensitizes aerobic cells only by inhibition of enzymatic repair processes.     

Interactions of radioprotectors and oxygen in cultured mammalian cells. II. Effects of dithiothreitol on radiation-induced DNA damage and comparison with cell survival

The effects of the sulfhydryl-containing compound dithiothreitol (DTT) on radiation-induced DNA damage have been studied using two different assays: DNA unwinding hydroxyapatite chromatography and alkaline filter elution. DNA damage as measured by both assays for cells irradiated in air shows drug concentration-dependent radioprotection reaching high levels (dose reduction factor, DRF = 3) at high DTT concentrations. The pattern and degree of protection against DNA damage are the same as shown previously for cell survival. However, when cells are irradiated in hypoxia, DNA damage as measured by the unwinding technique is decreased less by low DTT concentrations than is survival, but DNA damage is decreased to a much greater extent (DRF = 3) at high concentrations of DTT (compared to DRF = 1.5 for cell survival). DNA damage as measured by the alkaline elution assay after hypoxic irradiation is decreased to a much greater extent at all concentrations of DTT with DRF = 1.6 at 1 mM and increasing to DRF = 4.5 at high levels of DTT. These results are discussed in terms of the different types of DNA damage produced in cells irradiated in air versus hypoxia and the differences in types of damage measured by the two different DNA assays and cell survival.     

Encounter of lithodid crab <Emphasis Type="Italic">Paralomis birsteini</Emphasis> on the continental slope off Antarctica,sampled by ROV

A population of stone crab (Lithodidae) was encountered on the continental slope off Antarctica in the Bellingshausen Sea between 1,123 and 1,304 m water depths using the ROV-Isis during leg 166 of the RV James Clark Ross, in January 2007. Specimens were video recorded and one specimen was retrieved by ROV for morphological and molecular identification. Based on morphology and molecular data from the mitochondrial COI gene, this specimen identified as P. birsteini, Macpherson, 1988a. The significance of the molecular data and their implications for biogeography and evolution of lithodids in the Southern Ocean are briefly discussed.     

Ullrich-Turner syndrome is not caused by haploinsufficiency of RPS4X

Ullrich-Turner syndrome (UTS) is frequently associated with monosomy X but may also occur with structural aberrations of the X and the Y chromosomes. It has been hypothesized that the ribosomal protein genes RPS4X and RPS4Y play a critical role in the prevention of UTS. Individual patients with a 46,X,i(Xq) karyotype cannot be differentiated phenotypically from 45,X UTS patients and carry three gene copies of RPS4X. Since haploinsufficiency of one or several gene(s) is thought to cause the UTS phenotype, direct assessment of RPS4X expression levels in these patients should establish whether RPS4X is involved in UTS. We have investigated fibroblasts of four 46,X,i(Xq) UTS patients with typical symptoms and a non-mosaic chromosome complement, and have found significantly increased RPS4X mRNA levels in all patients. Based on our results, we conclude that haploinsufficiency of RPS4X is not the cause of UTS.     

Accuracy of bristle placement on a leg segment in Drosophila melanogaster

Bristle positions in two rows of bristles on the basitarsus of the second leg of the fruitfly Drosophila melanogaster were analyzed in order to determine the accuracy of bristle placement within these rows. Within each row the positions of the two terminal bristles were found to be approximately equally variable, and positional variability was found to increase toward the middle of each row. Rows having fewer bristles manifested more positional variability in their midsection. These results are interpreted in terms of a possible bristle spacing mechanism involving repulsive forces between mobile bristle cells.     

Role of perforin, granzymes and the proliferative state of the target cells in apoptosis and necrosis mediated by bispecific-antibody-activated cytotoxic T cells

 Bispecific monoclonal antibodies (bi-mAb), directed against a tumor-associated antigen and the CD3 or CD28 antigen on T lymphocytes, induce activation of resting T lymphocytes and target-specific tumor cell lysis. We now show that both necrosis and apoptosis contribute to T-cell-mediated tumor cell destruction. Even though T cells up-regulate FAS/APO-1 expression upon bi-mAb stimulation, FAS/APO-1-mediated apoptosis does not contribute to bi-mAb-mediated destruction of Hodgkin’s cells. CD8⁺ lymphocytes were the most potent effectors of bi-mAb-mediated cytotoxicity and had the highest levels of mRNA coding for perforin and granzyme A and B. Ca²⁺-complexing agents, which abrogate perforin activity, led to decreased levels of necrosis, while inhibition of granzyme activity in effector or target cells had a similar effect on apoptosis. Granzyme-mediated apoptosis critically dependent on the proliferative state of the target cells, while perforin-induced necrosis was not cell-cycle-dependent. Our results underline the importance of the expression levels of perforin and granzymes in the effector T cells and of the proliferative state of the target cells in bi-mAb-mediated apoptosis and necrosis of tumor cells. Received: 5 December 1996 / Accepted: 16 January 1997     

The gene family for major urinary proteins: Expression in several secretory tissues of the mouse

The major urinary proteins (MUPs) of the mouse are encoded by a multigene family located at the Mup a locus on chromosome 4. Previous investigations have shown that the MUPs are synthesized in the liver, secreted and then excreted in the urine. We have found significant levels of MUP mRNA in several secretory tissues: the liver and the submaxillary, lachrymal and mammary glands. There are striking differences in hormonal and developmental regulation of MUP gene expression in these tissues. Furthermore, each tissue appears to express a characteristic pattern of MUP mRNAs. In particular, the lachrymal glands appear to express an entirely different set of MUP mRNAs. These results are discussed in relation to the organization of the MUP gene cluster and a possible function of the MUPs.