Eosinophilopoiesis in a murine model of allergic airway eosinophilia: involvement of bone marrow IL-5 and IL-5 receptor alpha

The airway inflammation in asthma is dominated by eosinophils. The aim of this study was to elucidate the contribution of newly produced eosinophils in airway allergic inflammation and to determine mechanisms of any enhanced eosinophilopoiesis. OVA-sensitized BALB/c mice were repeatedly exposed to allergen via airway route. Newly produced cells were identified using a thymidine analog, 5-bromo-2'-deoxyuridine, which is incorporated into DNA during mitosis. Identification of IL-5-producing cells in the bone marrow was performed using FACS. Bone marrow CD3+ cells were enriched to evaluate IL-5-protein release in vitro. Anti-IL-5-treatment (TRFK-5) was given either systemically or directly to the airways. IL-5R-bearing cells were localized by immunocytochemistry. Repeated airway allergen exposure caused prominent airway eosinophilia after three to five exposures, and increased the number of immature eosinophils in the bone marrow. Up to 78% of bronchoalveolar lavage (BAL) granulocytes were 5-bromo-2'-deoxyuridine positive. After three allergen exposures, both CD3+ and non-CD3 cells acquired from the bone marrow expressed and released IL-5-protein. Anti-IL-5 given i.p. inhibited both bone marrow and airway eosinophilia. Intranasal administration of anti-IL-5 also reduced BAL eosinophilia, partly via local effects in the airways. Bone marrow cells, but not BAL eosinophils, displayed stainable amounts of the IL-5R alpha-chain. We conclude that the bone marrow is activated by airway allergen exposure, and that newly produced eosinophils contribute to a substantial degree to the airway eosinophilia induced by allergen. Airway allergen exposure increases the number of cells expressing IL-5-protein in the bone marrow. The bone marrow, as well as the lung, are possible targets for anti-IL-5-treatment.     

Analysis of DNA status in bone marrow cells of mice internally irradiated with 90Sr

The response of bone marrow cells of CBA mice injected with 22.2, 222 and 592 kBq/animal to additional gamma-irradiation (3 Gy) for testing purposes was evaluated using SCG (Comet assay). A decrease in induction of DNA damage right after additional gamma-irradiation was determined. It correlated with bone marrow cell quantity and the tail length before additional gamma-irradiation. The results support the suggestion about the activation of DNA repair in bone marrow cells under exposure to 90Sr in vivo.     

A possible role for CXCR4 and its ligand, the CXC chemokine stromal cell-derived factor-1, in the development of bone marrow metastases in neuroblastoma

The homing of hemopoietic stem cells to the bone marrow is mediated by specific interactions occurring between CXCR4, which is expressed on hemopoietic stem cells, and its ligand, stromal cell-derived factor-1 (SDF-1), a CXC chemokine secreted by bone marrow stromal cells. In the present study we evaluated the possibility that neuroblastoma cells use a mechanism similar to that used by hemopoietic stem cells to home to the bone marrow and adhere to bone marrow stromal cells. Our study suggests that CXCR4 expression may be a general characteristic of neuroblastoma cells. SH-SY5Y neuroblastoma cells express not only CXCR4, but also its ligand, SDF-1. CXCR4 expression on SH-SY5Y neuroblastoma cells is tightly regulated by tumor cell-derived SDF-1, as demonstrated by the ability of neutralizing Abs against human SDF-1alpha to up-regulate CXCR4 expression on the tumor cells. The reduction in CXCR4 expression following short term exposure to recombinant human SDF-1alpha can be recovered as a result of de novo receptor synthesis. Recombinant human SDF-1alpha induces the migration of CXCR4-expressing SH-SY5Y neuroblastoma cells in CXCR4- and heterotrimeric G protein-dependent manners. Furthermore, SH-SY5Y cells interact at multiple levels with bone marrow components, as evidenced by the fact that bone marrow-derived constituents promote SH-SY5Y cell migration, adhesion to bone marrow stromal cells, and proliferation. These results suggest that SH-SY5Y neuroblastoma cells are equipped with adequate machinery to support their homing to the bone marrow. Therefore, the ability of neuroblastoma tumors to preferentially form metastases in the bone marrow may be influenced by a set of complex CXCR4-SDF-1 interactions.     

Regulation of eosinophilopoiesis in a murine model of asthma

Eosinophilic inflammation plays a key role in tissue damage that characterizes asthma. Eosinophils are produced in bone marrow and recent observations in both mice and humans suggest that allergen exposure results in increased output of eosinophils from hemopoietic tissue in individuals with asthma. However, specific mechanisms that alter eosinophilopoiesis in this disease are poorly understood. The current study used a well-characterized murine animal model of asthma to evaluate alterations of eosinophil and eosinophil progenitor cells (CFU-eo) in mice during initial sensitization to allergen and to determine whether observed changes in either cell population were regulated by T lymphocytes. Following the first intranasal installation of OVA, we observed sequential temporal elevation of eosinophils in bone marrow, blood, and lung. In immunocompetent BALB/c mice, elevation of bone marrow eosinophils was accompanied by transient depletion of CFU-eo in that tissue. CFU-eo rebounded to elevated numbers before returning to normal baseline values following intranasal OVA exposure. In T cell-deficient BALB/c nude (BALB/c(nu/nu)) mice, CFU-eo were markedly elevated following allergen sensitization, in the absence of bone marrow or peripheral blood eosinophilia. These data suggest that eosinophilia of asthma results from alterations in two distinct hemopoietic regulatory mechanisms. Elevation of eosinophil progenitor cells in the bone marrow is T cell independent and likely results from altered bone marrow stromal cell function. Differentiation of eosinophil progenitor cells and phenotypic eosinophilia is T cell dependent and does not occur in athymic nude mice exposed to intranasal allergen.     

Ah locus-associated differences in induction of sister-chromatid exchanges and in DNA adducts by benzo[a]pyrene in mice.

The effect of alleles of the Ah locus on the induction of sister-chromatid exchanges (SCE) was studied in C57Bl/6 and in DBA/2 mice treated twice intragastrically with benzo[a]pyrene (BP, 100 or 10 mg/kg b.w.). To measure the changes in the frequency of SCE, 2 protocols were used: in vivo in bone marrow cells after implantation of 5-bromodeoxyuridine (BrdU) tablets and in vivo/in vitro in spleen lymphocytes cultured with BrdU. On day 5 mice were killed and SCEs estimated in bone marrow cells. BP-DNA adducts in bone marrow and spleen were analyzed on day 5 after the same exposure to BP. In the spleen lymphocytes SCE frequencies were analyzed after an additional 48 h of culture. We found that at both doses of BP, the number of SCEs and BP-DNA adducts in bone marrow and in spleen cells was significantly higher in aryl hydrocarbon hydroxylase (AHH)-non-inducible (DBA/2) mice than in AHH-inducible (C57BL/6) mice. Only marginal induction of SCE was noted after the high dose of BP in C57BL/6 mice in bone marrow in vivo, whereas a highly significant increase in the frequency of SCEs was found in splenocytes in the in vivo/in vitro test. The spleen cells contained larger amounts of BP-DNA adducts and demonstrated higher absolute levels of SCEs than bone marrow cells. The sensitivity of both the in vivo/in vitro and the in vivo SCE test is high enough for assessment of Ah locus-linked differences in BP genotoxicity in mice at the prolonged time between treatment and cell preparation. The present data confirm the influence of inducibility of AHH in the intestine on the genotoxicity of BP to distal tissues after oral exposure to BP.     

Chromosomal abnormalities and sister-chromatid exchange in bone marrow cells of mice and Chinese hamsters after inhalation and intraperitoneal administration. II. Cyclophosphamide

The genotoxic effects of cyclophosphamide (CPP), a human and animal carcinogen requiring metabolic activation, were studied in bone marrow cells of mice and Chinese hamsters, analyzing chromosome abnormalities (CA) and sister-chromatid exchange (SCE) after a 2-h inhalation or a single intraperitoneal administration. In order to compare the genotoxicity after the different routes of administration in the dose range of 10-110 mg CPP/kg body weight, the systemic dose obtained by inhalation was calculated from blood concentrations and the inhalation duration after an analysis of the CPP blood kinetics. In NMRI mice the frequency of bone marrow cells with chromosome abnormalities was higher after aerosol exposure than after intraperitoneal administration of comparable CPP doses. In Chinese hamsters the CA frequency was similar with both exposure routes. Inhaled CPP was found to induce a higher frequency of CA and SCE in the bone marrow cells of mice compared to those of Chinese hamsters. The findings suggest that for genotoxins requiring metabolic activation species differences exist with respect to the influence of the route of entry and the sensitivity of bone marrow cells.     

Molecular manifestations of radiation-induced genome instability: the possibility of chemical modification

The molecular manifestations of radiation-induced genome instability-changes of the DNA structure, the excision DNA repair and the contents of the reactive oxygen forms in bone marrow cells of the repair proficient mice (CBA) and of the repair-defective (101/H) lines in the dynamics up to 185 day after ionizing radiation exposure in the dose of 1.5 Gy were studied. Is was established, that after irradiation in bone marrow cells the descendants with the decreased activity of excision DNA repair and prone to increased changes of DNA structure DHK is arised. The injection of the phenozane in concentrations causing its receptor interaction with cells, did not defend DNA of the bone marrow cells from the radiation injury after the exposure in a sublethal dose, however it exerted influence on long-term changes. Due to the phenosane of the bone marrow cells of the irradiated mice of CBA line exhibited the larger activity in a DNA repair from damages and maintenance of vitality. The bone marrow cells of male mice of repair defective 101/H line, which phenozan was entered before the irradiation, remained unfit to the remuval of DNA damages by the repair, that probably resulted the activations of the program of the maintenance of genome constancy by the apoptosis in the cells--carriers of the structural defects and the cause of animal lethality.     

Implication of Ia-positive bone marrow interstitial stem cells in the induction of unresponsiveness to canine renal allografts

The removal from stored autologous host bone marrow of a monocytoid cell population by exposure to methylprednisolone is associated with successful introduction of unresponsiveness to renal allografts in irradiated recipients reconstituted with such treated marrow. The eliminated cells are a prominent component of the canine long bone marrow interstitium and share a number of important properties with dendritic cells (DC), including size and shape; poor or nonadherence to plastic or glass surfaces; negative staining for neutral esterase, acid phosphatase, or peroxidase; nonphagocytic; Ia positive, but negative for IgG or IgM; ability to act as accessory cells in augmenting the intensity of allogeneic mixed-lymphocyte reactions. Both cell types are of bone marrow origin and are susceptible to steroids in vitro. The results suggest that the bone marrow interstitial cells identified in the course of this study may be enriched with populations of canine dendritic cell precursors and dendritic cells at various stages of differentiation. The detection of a receptor site for Helix promatia on the surface of such cells may be of usefulness in their further characterization and in the analysis of their precise role in the modulation of allogeneic unresponsiveness.     

Response kinetics of radiation-induced micronucleated reticulocytes in human bone marrow culture

The frequency of micronucleated reticulocytes (MN-RETs) in the bone marrow or peripheral blood is a sensitive indicator of cytogenetic damage. While the kinetics of MN-RET induction in rodent models following irradiation has been investigated and reported, information about MN-RET induction of human bone marrow after radiation exposure is sparse. In this report, we describe a human long-term bone marrow culture (LTBMC), established in three-dimensional (3D) bioreactors, which sustains long-term erythropoiesis. Using this system, we measured the kinetics of human bone marrow red blood cell (RBC) and reticulocyte (RET) production, as well as the kinetics of human MN-RET induction following radiation exposure up to 6Gy. Human bone marrow established in the 3D bioreactor demonstrated an average percentage of RBCs among total viable cells peaking at 21% on day 21. The average percentage of RETs among total viable cells reached a maximum of 11% on day 14, and remained above 5% by day 28, suggesting that terminal erythroid differentiation was still active. Time- and dose-dependent induction of MN-RET by gamma radiation was observed in the human 3D LTBMC, with peak values occurring at approximately 3 days following 1Gy irradiation. A trend towards delayed peak to 3-5 days post-radiation was observed with radiation doses ≥2Gy. Our data reveal valuable information on the kinetics of radiation-induced MN-RET of human bone marrow cultured in the 3D bioreactor, a synthetic bioculture system, and suggest that this model may serve as a promising tool for studying MN-RET formation in human bone marrow, thereby providing opportunities to study bone marrow genotoxicity testing, mitigating agent effects, and other conditions that are not ordinarily feasible to experimental manipulation in vivo.     

THE ROLE OF BONE MARROW OF X-IRRADIATED MICE IN THYMIC RECOVERY

The influence of the bone marrow on the repopulation of the thymus in X-irradiated mice has been investigated.
It was observed that the thymus and a certain population of bone marrow lymphocytic cells were repopulated in parallel in a cyclic fashion. This occurred either after a single exposure of mice to 400 R or after serial weekly X-ray treatments with 170 R. Lethally irradiated recipients which were grafted with bone marrow cells obtained 12-24 days after four weekly irradiations of donor mice with 170 R also exhibited a cyclic repopulation of both the thymus and the bone marrow lymphocytic population. In contrast, mice which were transplanted with bone marrow cells from unirradiated donors, containing an equal number of stem cells (CFU), exhibited a continuous rather than a cyclic recovery of both cell populations. the bone marrow stem cells of mice recovering from X-irradiation were found to have a decreased proliferative activity, since they produced significantly smaller spleen colonies in lethally irradiated recipients than marrow cells from unirradiated mice.
The results were interpreted as indicating that the bone marrow lymphocytic cells may act as thymic precursor cells and that thymic lymphopoiesis is dependent on the presence of such cells. Evidently, the production of lymphocytic cells will decrease when the stimulus for granulocyte production increases due to the limited proliferative activity of the surviving bone marrow stem cells after irradiation. This may result in a cyclic variation of the production of bone marrow lymphocytic cells and it follows that thymic lymphopoiesis will run parallel.     

Effects of cyclic pressure on bone marrow cell cultures

The present in-vitro study used bone marrow cell cultures and investigated the effects of cyclic pressure on osteoclastic bone resorption. Compared to control (cells maintained under static conditions), the number of tartrate resistant acid phosphatase (TRAP)-positive, osteoclastic cells was significantly (p<0.05) lower when, immediately upon harvesting, bone marrow cells were exposed to cyclic pressure (10-40 kPa at 1.0 Hz). In contrast, once precursors in bone marrow cells differentiated into osteoclastic cells under static culture conditions for 7 days, subsequent exposure to the cyclic pressure of interest to the present study did not affect the number of osteoclastic cells. Most important, exposure of bone marrow cells to cyclic pressure for 1 h daily for 7 consecutive days resulted in significantly (p<0.05) lower osteoclastic bone resorption and in lowered mRNA expression for interleukin-1 (IL-1) and tumor necrosisfactor-a (TNF-a), cytokines that are known activators of osteoclast function. In addition to unique contributions to osteoclast physiology, the present study provided new evidence of a correlation between mechanical loading and bone homeostasis as well as insight into the molecular mechanisms of bone adaptation to mechanical loading, namely cytokine-mediated control of osteoclast functions.     

Stem cell kinetics in spleen and bone marrow after single and fractionated irradiation of infant mice

Summary The number and type of stem cells in spleen and bone marrow of mice were determined after exposure to a single dose of 150 R on day 6, to a single dose of 500 R on day 6 or day 9 or to a fractionated dose of 150 R + 350 R on day 6 and 9. The stem cells were assayed on the basis of colony forming units (CFU) in spleen and of incorporation of iododeoxyuridine in spleen and bone marrow of lethally irradiated host mice. During the first month of life, the number of stem cells in non-irradiated mice increases markedly in bone marrow and slightly in spleen. Irradiation causes a long-lasting depression in stem cells, particularly in bone marrow and affecting preferentially erythropoietic precursor cells. Following a dose of only 150 R, the number of CFU in bone marrow is still below control levels 24 days later. An exposure to 500 R fractionated between day 6 and 9 has a markedly greater effect on stem cells in the spleen than the same dose given in a single application either at day 6 or 9.Supported by the Schutzkommission am Ministerium des Innern der BRD and contract B232-76-1BIOB of the Biology Division of the Commission of the European Community (Publikation No. 1727)     

A bicellular mechanism in the immune response to chemically defined antigens. 3. Interaction of thymus and bone marrow-derived cells

The role of thymus and bone marrow-derived cells in the in vitro response to the dinitrophenyl (DNP) determinant was studied using the millipore filter well technique for spleen organ cultures. Antibodies to DNP were assayed by the technique of inactivation of DNP-coupled T-4 bacteriophage. It was found that spleens of mice total-body irradiated at 750 R, treated with bone marrow and thymus cells after exposure and immunized against rabbit serum albumin (RSA) were able to produce antibodies to DNP when challenged in vitro with DNP-RSA. Such a response was not produced by spleen explants from x-irradiated mice treated with either thymus or bone marrow cells. Neither were antibodies to DNP produced by spleens of animals repopulated with thymus and bone marrow cells, but not immunized with the carrier. This carrier effect was manifested when the irradiated mice were treated with RSA and thymus cells 6–8 days before administration of the bone marrow cells. Yet, such an effect was not observed when the RSA and bone marrow cells were given 6–8 days before injection of the thymus cells. Thus, the thymus-derived cells appear to play the role of cells sensitive to the carrier (RSA), whereas the bone marrow seems to be involved in the production of antibodies.     

Effect over time of in-vivo administration of the polysaccharide arabinogalactan on immune and hemopoietic cell lineages in murine spleen and bone marrow.

Current evidence indicates an immunostimulating role for complex carbohydrates, i.e., polysaccharides, from several plant sources. In the present work, we determined the specific in vivo effects, with time of administration, of one such compound, a neutral arabinogalactan from larch not only on immune (lymphoid) cells, but also on natural killer (NK) lymphoid cells, as well as a variety of other hemopoietic cells in both the bone marrow and spleen of healthy, young adult mice. The latter were injected daily (i.p.) with arabinogalactan (500 microg in 0.1 ml pH 7.2 phosphate buffered saline-PBS) for 7 or 14 days. Additional, aged (1 1/2-2 yr) mice were similarly injected for 14 days only. Control mice were given the PBS vehicle in all cases, following the above injection regimen. Animals from all groups were sampled 24 h after the final injection and the immune and hemopoietic cell populations in the bone marow and spleen were assessed quantitatively. The results indicated that immediately following either 7 or 14 days of arabinogalactan administration to young, adult mice, lymphoid cells in the bone marrow were significantly decreased (p < 0.004; p < 0.001, respectively) relative to controls but remained unchanged at both time intervals in the spleen. NK cells, after 7 days of arabinogalactan exposure, were also decreased significantly in the bone marrow (p < 0.02), but unchanged in the spleen. After 14 days' exposure to the polysaccharide, NK cells in the bone marrow had returned to normal (control) levels, but were increased in the spleen (p < 0.004) to levels greater than 2-fold that of control. Among other hemopoietic cell lineages, none was influenced in the bone marrow or spleen by one-week administration of arabinogalactan; however, after two-week exposure, precursor myeloid cells and their mature (functional) progeny (granulocytes), were significantly reduced in the spleen (p < 0.043; p < 0.006, respectively), as were splenic monocytes (p < 0.001). These lineages in the bone marrow, however, remained steadfastly unaltered even after 14 days of continuous exposure to the agent. Of the vast cascade of cytokines induced in the presence of this polysaccharide, it appears that immunopoiesis- and hemopoiesis-inhibiting ones are most prevalent during at least the first two weeks of daily exposure.     

Early endotoxin tolerance is associated with alterations in bone marrow-derived macrophage precursor pools

Early endotoxin tolerance has been defined as the transient period after an initial sublethal exposure to LPS during which a normally responsive individual is rendered hyporesponsive. Little is known about the cellular mechanisms that underlie this phenomenon. In this study, an early tolerance system was established by the injection of mice with 25 micrograms of E. coli K235 LPS. Maximal hyporesponsiveness in response to a challenge injection was observed 3 to 4 days after the initial injection, and normal responsiveness returned by 8 days after the initial exposure to LPS. Further experiments described herein demonstrate that the acquisition and maintenance of the tolerant state coincides temporally with an increase in the number of macrophage progenitor cells in the bone marrow. Cell-sizing profiles of the bone marrow cells from tolerized mice indicate an enrichment for a population of cells that are significantly larger than in bone marrow preparations from control mice. By density gradient sedimentation, it was shown that the denser population of cells from tolerized mice contained the increased numbers of progenitor cells, which, by cytology, were immature monocytic cell types. The increased numbers of macrophage progenitors was sustained after a second (challenge) injection of LPS. These results indicate that early endotoxin tolerance is associated with an increase in a population of bone marrow cells that is enriched for macrophage progenitors and suggests the possibility that the lack of responsiveness observed during the hyporesponsive period is related to a failure of these immature cell types to respond to LPS.     

Combination effects of chronic cadmium exposure and gamma-irradiation on the genotoxicity and cytotoxicity of peripheral blood lymphocytes and bone marrow cells in rats

This work investigated the effects of chronic cadmium (Cd) exposure combined with γ-ray irradiation on the cytotoxicity and genotoxicity of peripheral blood cells and bone marrow cells in rats. Results showed that when the rats were exposed to low dose (LD) Cd of 0.1mg CdCl?/(kgd) for 8 and 12 weeks, the Cd concentration in blood reached to 135-140 μg/L and no toxic effects on peripheral blood lymphocytes, white blood cells (WBC) and granulocyte-monocyte (GM) progenitor cells were observed except polychromatic erythrocytes (PCE) of bone marrow. Moreover, this chronic LD Cd exposure significantly decreased irradiation-induced micronucleus (MN) formation and hypoxanthine-guanine phosphoribosyl transferase (hprt) mutation in lymphocytes and PCE, while the combination of LD Cd exposure and irradiation induced the additive metallothionein (MT) protein expression in bone marrow cells. When the rats were exposed to a high dose (HD) Cd of 0.5mg CdCl/?(kgd) for 8 and 12 weeks, the blood Cd level approached to 458-613 μg/L and an inflammatory response was induced, meanwhile, MN formation and hprt mutation were markedly increased, and the ratio of PCE/NCE (normochromatic erythrocyte) was significantly decreased. Furthermore, when the rats were exposed to HD Cd plus 2 Gy irradiation, additive toxic effects on MN formation, hprt mutation, PCE damage and GM progenitor cell proliferation were observed, while this combination treatment resulted in an obvious reduction of MT protein compared to HD Cd group. In conclusion, chronic exposure to LD Cd induced the adaptive response to irradiation in the genotoxicity of peripheral blood lymphocytes and PCE of bone marrow by the up-regulation of Cd-induced MT protein, but the combination of HD Cd exposure and irradiation generated the additive effects on the cytotoxicity and genotoxicity in peripheral blood lymphocytes and bone marrow cells.     

Early and late effects of the cytogenetic action of doxorubicin

Cytogenetic disorders in hemopoietic cells of the bone marrow were studied on mice CBA at early and late periods after exposure to MIDs of doxorubicin, an anthracycline antibiotic. It was shown that at the early period doxorubicin induced aberrations, mainly of the chromatid type, in the hemopoietic cells of the bone marrow. Instability of the genetic apparatus of the hemopoietic cells observed for 3 months of the experiment was likely to be the immediate cause of the disorders in hemopoiesis at the late periods after exposure to doxorubicin.     

Cytokine regulation of bone marrow natural suppressor cell activity in the suppression of lymphocyte function.

Natural suppressor (NS) cells, which nonspecifically suppress immune responses, are present in the spleen following exposure to radiation, chronic graft-versus-host disease, or cancer and in normal bone marrow. A model system is described which allows the study of cytokines activating and inhibiting NS cells, cytokines mediating NS activity, and NS effects on cytokine synthesis. Recombinant interleukin-3 (rIL-3) and granulocyte-macrophage colony-stimulating factor (rGM-CSF) efficiently activated NS cells present in normal bone marrow and were effective at concentrations as low as 5 U/ml. At high concentrations, GM-CSF, but not IL-3, did not activate NS cells. Recombinant interferon-gamma (rIFN-gamma) blocked the activation of bone marrow NS cells by rIL-3, but did not down-regulate NS cells once activated. The NS cells secreted one or more soluble suppressor factors, which blocked IL-2 synthesis and also inhibited IL-2-dependent T cell proliferation in the presence of excess IL-2.     

Potential role of free radicals in benzene-induced myelotoxicity and leukemia.

Occupational exposure to benzene, a major industrial chemical, has been associated with various blood dyscrasias and increased incidence of acute myelogenous leukemia in humans. It is established that benzene requires metabolism to induce its effects. Benzene exposure in humans and animals has also been shown to result in structural and numerical chromosomal aberrations in lymphocytes and bone marrow cells, indicating that benzene is genotoxic. In this review we have attempted to compile the available evidence on the role of increased free radical activity in benzene-induced myelotoxic and leukemogenic effects. Benzene administration to rodents has been associated with increased lipid peroxidation in liver, plasma, and bone marrow, as shown by an increase in the formation of thiobarbituric-acid reactive products that absorb at 535 nm. Benzene administration to rodents also results in increased prostaglandin levels indicating increased arachidonic acid peroxidation. Other evidence includes the fact that bone marrow cells and their microsomal fractions isolated from rodents following benzene-treatment have a higher capacity to form oxygen free radicals. The bone marrow contains several peroxidases, the most prevalent of which is myeloperoxidase. The peroxidatic metabolism of the benzene metabolites, phenol and hydroquinone, results in arachidonic acid peroxidation and oxygen activation to superoxide radicals, respectively. These metabolites, upon co-administration also produce a myelotoxicity similar to that observed with benzene. Recently, we have found that exposure of human promyelocytic leukemia (HL-60) cells (a cell line rich in myeloperoxidase), to the benzene metabolites, hydroquinone and 1,2,4-benzenetriol results in increased steady-state levels of 8-hydroxydeoxyguanosine a marker of oxidative DNA damage. Peroxidatic metabolism of benzene's phenolic metabolites may therefore be responsible for the increased free radical activity and toxicity produced by benzene in bone marrow. We thus hypothesize that free radicals contribute, at least in part, to the toxic and leukemogenic effects of benzene.     

Magnetic field affects thymidine kinase in vivo

Whole mice on normal or vitamin E deficient diet were immobilized by Nembutal anaesthesia and exposed to a stationary magnetic field of 1.4 tesla for up to 60 min. Thymidine kinase (TdR-K) was assayed in the high-speed supernatant of bone marrow cells which were collected into optimally adjusted nutrient medium of pH 7.3-7.4 containing 1350 mg NaHCO3 per litre and were then destroyed by sonication. In parallel, uptake of 125I-labelled 5-I-2'-deoxyuridine (125IUdR) into DNA of whole bone marrow cells, of various tissues and of the whole body was measured. The results indicate the following. The magnetic field exposure caused in bone marrow cells an increase of activity of TdR-K and of uptake of 125IUdR to about 130 per cent of control. The effect depended on immobilization of the mice in the field and on the presence of NaHCO3 in the nutrient medium used for cell collection. There was no field-induced change in body temperature. The effect on 125IUdR uptake was similar in isolated tissues and the whole body following intraperitoneal injection of the tracer. It increased to a maximum of about 135 per cent of control, during exposure times over 30 min. This effect is not explained as a result of a temporary change in the rate of cell proliferation. Vitamin E deficiency caused a depression of activity of TdR-K and of uptake of 125IUdR in bone marrow cells to about 75 per cent of control. This depression was similar to that observed after whole body gamma-irradiation with about 0.01 Gy (1 rad). The inhibitory effects of vitamin E deficiency on TdR-K were overcome by exposure to the magnetic field. Immediately after cessation of the magnetic field for 60 min, 125IUdR uptake was normal; normalization of uptake was delayed with exposure times shorter than 60 min. A 60 min exposure to the magnetic field had no long term effect on turnover of labelled cells in the mice. The data imply the non-specific control of thymidine kinase by charged molecular species and the modification of this control by the magnetic field.