FISH chromosome analysis of plutonium workers from the Sellafield nuclear facility

Chromosome analysis using a single-color FISH technique to paint three pairs of chromosomes was undertaken on a group of 46 retired plutonium workers with assessed bone marrow doses >60 mSv, 34 of whom were categorized as having robust dosimetry and 12 for whom internal doses were considered less reliable. Comparisons were made with a group of 34 workers with negligible radiation exposure and a group of 34 workers with similar recorded external gamma-ray doses but negligible internal dose. The simple translocation frequency of 17.65 +/- 1.96 x 10(-3) per genome equivalent for the 34 plutonium workers with robust dosimetry was significantly increased in comparison with that of 10.06 +/- 1.16 x 10(-3) per genome equivalent for the unirradiated control group (P = <0.001) and that of 13.55 +/- 1.43 x 10(-3) per genome equivalent for the group with similar external gamma-ray exposure (P = 0.012). Thus, although in vitro studies have indicated that the majority of alpha-particle-irradiated cells suffer complex non-transmissible chromosome damage, in vivo a significant proportion survive with simple exchanges that can be passed on to descendant cells. In contrast, the three groups demonstrated no significant differences in stable complex aberrations. No evidence of an increase in dicentrics or unstable complex aberrations associated with plutonium exposure was observed, and it can therefore be assumed that there is little, if any, ongoing irradiation of mature lymphocytes. The translocation frequency of 12.08 +/- 1.92 x 10(-3) per genome equivalent for the group of 12 plutonium workers with less reliable internal dosimetry could adequately be accounted for by age and external dose and indicates that the internal bone marrow doses are likely to have been overestimated. Cytogenetic analysis can therefore make a valuable contribution to the validation of internal doses from plutonium deposition.     

Effect of plutonium-239 on the mitotic activity of mouse bone marrow cells

Summary Mitotic index of the bone marrow cells was studied in femoral bone marrow of mice given 313 kBq²³⁹Pu kg^–1. The attention was turned to the femoral midshaft and the mitose concentration, intensified by Colcemid stathmokinetic effect, was evaluated in a sampling field from endosteal surface to the central venous canal, throughout 68 weeks. It has been found that the plutonium effect in the sampling band is rather uniform except the points in subendosteal zone early after plutonium injection, where the mitotic index was reduced in such a way that the mitotic gradient, observed in controls, was affected. The mitotic activity in femoral diaphysis of plutonium injected mice was mobilized approximately till the 30th week of contamination. Later it deteriorated progressively. The results are discussed and should not be regarded as representative for the entire bone marrow hemopoiesis.     

Radiation sensitivity of human and murine peripheral blood lymphocytes,stem and progenitor cells

Immunodeficiency is a severe side effect of radiation therapy, notably at high radiation doses. It may also impact healthy individuals exposed to environmental ionizing radiation. Although it is believed to result from cytotoxicity of bone marrow cells and of immunocompetent cells in the peripheral blood, the response of distinct bone marrow and blood cell subpopulations following exposure to ionizing radiation is not yet fully explored. In this review, we aim to compile the knowledge on radiation sensitivity of immunocompetent cells and to summarize data from bone marrow and peripheral blood cells derived from mouse and human origin. In addition, we address the radiation response of blood stem and progenitor cells. The data indicate that stem cells, T helper cells, cytotoxic T cells, monocytes, neutrophils and, at a high degree, B cells display a radiation sensitive phenotype while regulatory T cells, macrophages, dendritic cells and natural killer cells appear to be more radioresistant. No conclusive data are available for basophil and eosinophil granulocytes. Erythrocytes and thrombocytes, but not their precursors, seem to be highly radioresistant. Overall, the data indicate considerable differences in radiosensitivity of bone marrow and blood normal and malignant cell populations, which are discussed in the light of differential radiation responses resulting in hematotoxicity and related clinical implications.     

Role of cytokines in bone resorption

Osteoclastic bone resorption is modulated in humans by powerful osteotropic factors which are generated in the immediate vicinity of bone resorbing surfaces. These factors are released from marrow mononuclear cells and from some bone cells, and some are actually incorporated into the noncollagenous bone matrix from where they are released when bone is resorbed. They are likely important not only in the control of normal bone remodeling, but also in a number of disease states associated with disordered remodeling. In this review, current concepts of the effects of these factors on cells in the osteoclast lineage will be discussed.     

Bone marrow analysis of the myelodysplastic syndromes: histological and immunohistochemical features related to the evolution of overt leukemia

Bone marrow trephines from 31 patients with an initial diagnosis of myelodysplastic syndromes (MDS) were examined and analyzed histologically and immunohistochemically. In those cases terminating in overt leukemia (6/31, 19%), the number of bone marrow mast cells was significantly reduced, compared with those which did not evolve to overt leukemia. The bone marrow lymphoid cells that may participate in immunosurveillance against the proliferation of blast cells were also significantly reduced in cases terminating in overt leukemia. However, S-100 protein-positive cells, which include histiocytes and suppressor T-cells, were increased in cases terminating in overt leukemia. The results indicated that examination of the bone marrow to determine the proportions of mast cells and lymphoid cells which may be involved in host defense systems may be useful in predicting the evolution to overt leukemia in MDS. In the present series, patients with a hypocellular marrow (5/31, 16%) did not progress to overt leukemia and had a significantly lower bone marrow reticulin content, a significantly lower megakaryocyte count, a relatively higher mast cell count and a significantly higher lymphoid cell count than those with a normocellular or hypercellular marrow. These findings may reflect the initial features of MDS or, possibly, that hypocellular MDS is an independent entity with a low potential for blastic proliferation.     

Past exposure to densely ionizing radiation leaves a unique permanent signature in the genome

Speculation has long surrounded the question of whether past exposure to ionizing radiation leaves a unique permanent signature in the genome. Intrachromosomal rearrangements or deletions are produced much more efficiently by densely ionizing radiation than by chemical mutagens, x-rays, or endogenous aging processes. Until recently, such stable intrachromosomal aberrations have been very hard to detect, but a new chromosome band painting technique has made their detection practical. We report the detection and quantification of stable intrachromosomal aberrations in lymphocytes of healthy former nuclear-weapons workers who were exposed to plutonium many years ago. Even many years after occupational exposure, more than half the blood cells of the healthy plutonium workers contain large (>6 Mb) intrachromosomal rearrangements. The yield of these aberrations was highly correlated with plutonium dose to the bone marrow. The control groups contained very few such intrachromosomal aberrations. Quantification of this large-scale chromosomal damage in human populations exposed many years earlier will lead to new insights into the mechanisms and risks of cytogenetic damage.     

Inflammatory modulation of HSCs: viewing the HSC as a foundation for the immune response

Cells of the innate and adaptive immune systems are the progeny of a variety of haematopoietic precursors, the most primitive of which is the haematopoietic stem cell. Haematopoietic stem cells have been thought of generally as dormant cells that are only called upon to divide under extreme conditions, such as bone marrow ablation through radiation or chemotherapy. However, recent studies suggest that haematopoietic stem cells respond directly and immediately to infections and inflammatory signals. In this Review, we summarize the current literature regarding the effects of infection on haematopoietic stem cell function and how these effects may have a pivotal role in directing the immune response from the bone marrow.     

A comparison of the distribution of 239-Pu and calcein in the illium of the female CBA mouse

Conclusions As a model for the behaviour of plutonium in bone, calcein data must be treated with some care. It does not label the same surfaces as plutonium which results in different distribution patterns at later times. However, it might be that if in man or a long lived animal, labelling occured over a period of weeks, so that most surfaces become labelled, the resultant distribution pattern at late times would more nearly model plutonium behaviour. The single biggest difference between the behaviour of the two substances is the build up of plutonium in the bone marrow, an effect seen only slightly with calcein. The other differences noted was the redeposition of plutonium, as a consequence of recycling of the radionuclide, maintaining a concentration of plutonium on endosteal surface.     

The in vivo effects of interleukin 1. I. Bone marrow cells are induced to cycle after administration of interleukin 1

We have previously reported that interleukin 1 (IL-1) administration 20 hr before irradiation protects mice from lethal effects of radiation. The recovery of total nucleated bone marrow cells and of hematopoietic progenitor cells was enhanced in IL-1 treated, as compared to untreated, irradiated mice. This suggested that IL-1 administration may affect the cells in the bone marrow of normal mice. Intraperitoneal administration of recombinant IL-1 resulted in bone marrow cell enlargement and increased cycling of these enlarged cells. In addition, the capacity of bone marrow cells from IL-1 treated mice to proliferate in response to granulocyte macrophage-colony-stimulating factor (GM-CSF) in cell suspension cultures was enhanced. The above effects were not genetically restricted as C57BL/6, B6D2F1, C3H/HeN, and C3H/HeJ mice showed similar responses. A comparative study showed that 100 ng of IL-1 was much more effective in stimulating bone marrow cells by the above criteria than 5 micrograms GM-CSF. Since IL-1, unlike CSF, can not be demonstrated to have a direct in vitro stimulatory effect on bone marrow cells, the aforementioned in vivo effects of IL-1 are presumably mediated by other hematopoietic growth factors. We have previously shown that IL-1 induces the appearance of high titers of CSF in the serum. Consequently hematopoietic growth factors that are generated at local sites following IL-1 administration may mediate the observed cell cycling effect.     

Leukaemic peripheral blood plasma and bone marrow plasma: comparison of influence on lymphocyte proliferation

Abstract. Peripheral blood plasma from some children with untreated acute lymphoblastic leukaemia (ALL) exerted an inhibitory effect in vitro on phytohaemagglutinininduced lymphocyte transformation of normal peripheral blood lymphocytes. This occurred at concentrations beyond that required for optimal response as judged by reduction of blast cell formation and tritiated thymidine and tritiated uridine incorporation into DNA and RNA, respectively. In contrast, bone marrow plasma from these patients was non-inhibitory or contained significantly less inhibitory activity. Bone marrow plasma from the majority of healthy controls was superior to their peripheral blood plasma in enhancing phytohaemagglutinin-induced mitogenesis. The difference between an individual's bone marrow- and peripheral blood-derived plasma in enhancing proliferation of patient and healthy control cells was significantly greater amongst the patients than the healthy control group; this was attributed mainly to the increased inhibitory activity of ALL peripheral blood plasma compared with normal plasma. Medium conditioned by phytohaemagglutinin-stimulated normal peripheral blood lymphocytes was effective in neutralizing the inhibitory activity of ALL peripheral blood plasma. Taken together, these in vitro results are at least suggestive that in vivo , in healthy subjects, the rapidly proliferating cells in the bone marrow and the 'resting' blood cells in the circulation may be under the influence of a fine balance of different types and/or levels of humoral growth stimulatory and inhibitory factors and that in ALL an unstable balance of these factors exists. The decreased proliferation of circulating blast cells compared with bone marrow blasts in ALL may be attributed, at least in part, to exposure to the different levels of inhibitor(s) in the circulation and bone marrow as demonstrated in vitro by our results.     

Study on the possible factors influencing the expression of H-2 restriction specificity and Ir phenotype of antigen-specific proliferative T cells with various types of radiation chimeras

To better understand the factors described previously as influencing the manifestation of H-2 restriction specificity and Ir phenotype of T cells from radiation bone marrow chimeras, we also examined H-2 restriction specificity (Ir phenotype) of antigen (DNP-OVA, (T, G)-A-L, (H, G)-A-L)-specific proliferative T cells generated in various types of H-2 incompatible radiation chimeras prepared under our specific-pathogen-free (SPF) condition. The results indicated the following: (a) T cells generated in F1----parent bone marrow chimeras preferentially manifested host-type H-2 restriction specificity and Ir phenotype, regardless of the radiation dose (8.70 vs 11.59 Gy); (b) T cells recovered from twice-reconstituted F1----(PA----PB) chimeras manifested primary host (PB)-type Ir phenotype; (c) T cells which were recovered from (B10.Thy-1.1 X B10.BR.Thy-1.1)F1----parent (Thy-1.2) bone marrow chimeras and treated with anti-Thy-1.2 plus complement to deplete host-derived T cells still manifested preferentially the restriction specificity for host-type H-2; (d) PA-derived T cells which had differentiated in a fully allogeneic host (PB) environment of (PA + PB)----PB chimeras manifested fully allogeneic host-type Ir phenotype; (e) T cells from F1----parent chimeras that were prepared with 13-day fetal liver cells also manifested host H-2-restricted Ir phenotype; and (f) host preference for Ir phenotype of antigen-specific proliferative T cells was observed even in the case of F1----parent bone marrow chimeras reconstituted with "intact" bone marrow cells. The data suggest that thymic APCs, surviving host T cells or the source of stem cells (adult bone marrow vs 13-day fetal liver), do not necessarily play a significant role in the manifestation of H-2 restriction specificity and Ir phenotype of T cells generated in H-2 incompatible radiation chimeras.     

A model for the autosensitization autoantibody production associated with xenogeneic thymic RNA.

Normal C57BL/6 bone marrow cells cultured for 3 weeks with xenogeneic thymic RNA and syngeneic C57BL/6 antigens (immunoglobulin G or red blood cells) produced anti-immunoglobulin antibody or anti-mouse red blood cell antibody (hemolysin). Addition of both xenogeneic thymic RNA and autoantigens to bone marrow cultures was necessary to elicit autosensitization. Syngeneic thymic RNA would not substitute for xenogeneic RNA. Normal recipients inoculated with syngeneic kidney or spinal cord homogenates and xenogeneic thymic RNA developed albuminuria or motor neuropathies within 10 days. Histologic examination of tissues from these animals revealed immunoglobulin deposits on glomerular or tubular basement membranes or on myelin sheaths. These changes were not observed in tissues from control animals inoculated with only the organ homogenates. Normal mice injected with syngeneic bone marrow cells, which had been autosensitized in vitro against kidney or spinal cord homogenates, also developed albuminuria or motor neuropathies, respectively. These abnormalities were observed only if bone marrow cells had been cultured with both xenogeneic thymic RNA and autoantigens. Histologic examination of tissues from these mice also revealed immunoglobulin deposits in kidney or spinal cord tissues. These results demonstrate that xenogeneic thymic RNA can play important roles in the formation of autoantibodies.     

Long-term effects of low-level 239Pu contamination on murine bone-marrow stem cells and their progeny

The effects of long-term internal contamination with 13.3 kBq kg-1 239Pu injected intravenously were studied in 10-week-old ICR (SPF) female mice. Radiosensitivity of spleen colony-forming units (CFU-S) and 125IUdR incorporating into proliferating cells of vertebral bone marrow and spleens were determined in plutonium-treated and control animals one year after nuclide injection. The CFU-S in 239Pu-treated mice were more sensitive to X-rays (D0 = 0.52 +/- 0.01 Gy) than in controls (D0 = 0.84 +/- 0.02 Gy). 125IUdR incorporation into bone marrow and spleen cells was reduced after plutonium contamination. At one year following plutonium injection, the occurrence of chromosome aberrations was evaluated in metaphase figures of femoral bone marrow cells. The frequency of aberrations increased early after plutonium treatment, at later intervals it tended to decrease but not below the control level. While the relative numbers of vertebral marrow CFU-S decreased significantly, but only to 86 per cent of normal, cellularity of vertebral bone marrow, peripheral blood counts and survival of 239Pu-treated mice did not differ from the control data.     

Characteristics of the cohort of workers at the Mayak nuclear complex.

At Branch No. 1 of the Russian State Research Center "Biophysics Institute", a registry has been created of workers at the "Mayak" Production Association, the first nuclear complex in Russia. This registry includes 18,830 persons hired at Mayak's nuclear reactors and radiochemical and plutonium production plant between 1948 and 1972. Twenty-five percent of these workers are women. As of December 31, 1994, the vital status is known for approximately 90% of the cohort members. A total of 5,118 persons have died. The cause for 97% of total deaths has been ascertained. The cohort members were exposed to both external gamma radiation and internal radiation from incorporated plutonium. The plutonium body burden has been measured in 30% of the cohort members with potential for plutonium exposure. External gamma-ray doses were in the range from tenths of milligrays to about 10 Gy, and plutonium body burdens were up to about 260 kBq. In view of the nature of the Mayak worker cohort, it has the potential to provide reasonably precise, quantitative estimates of the long-term health effects associated with chronic low-dose-rate exposure to external gamma radiation as well as internal radiation from plutonium. However, a number of issues must be addressed before credible risk estimates can be obtained from this cohort. These issues include the development of an appropriate internal comparison group and/or external rates and separating of the effects of internal and external exposures on risk estimates.     

The mechanism for the radioprotective effects of zymosan‐A in mice

It proved that Zymosan‐A protected the haematopoietic system from radiation‐induced damage via Toll‐Like Receptor2 in our previous study. In this study, we investigated the potential mechanism for the radioprotective effects of Zymosan‐A. The mice were treated with Zymosan‐A (50 mg/kg, dissolved in NS) via peritoneal injection 24 and 2 hours before ionizing radiation. Apoptosis of bone marrow cells and the levels of IL‐6, IL‐12, G‐CSF and GM‐CSF were evaluated by flow cytometry assay. DNA damage was determined by γ‐H2AX foci assay. In addition, RNA sequencing was performed to identify differentially expressed genes (DEGs). Zymosan‐A protected bone marrow cells from radiation‐induced apoptosis, up‐regulated IL‐6, IL‐12, G‐CSF and GM‐CSF in bone marrow cells. Zymosan‐A also protected cells from radiation‐induced DNA damage. Moreover, RNA sequencing analysis revealed that Zymosan‐A induced 131 DEGs involved in the regulation of immune system process and inflammatory response. The DEGs were mainly clustered in 18 KEGG pathways which were also associated with immune system processes. Zymosan‐A protected bone marrow cells from radiation‐induced apoptosis and up‐regulated IL‐6, IL‐12, G‐CSF and GM‐CSF. Moreover, Zymosan‐A might also exhibit radioprotective effects through regulating immune system process and inflammatory response. These results provided new knowledge regarding the radioprotective effect of Zymosan‐A.     

The distribution of plutonium-214 in rodents.

Plutonium-214 citrate solution at pH 6-5 was injected intravenously or intra-peritoneally into hamsters and rats at a dose of 50 MBq kg-1 (1-35 mCi kg-1). The animals were killed 1 day or 1 week later, and tissues were removed for autoradiography and radiochemical analysis. Plutonium-241 was distributed in rats in the same way as plutonium-239, and is a suitable isotope for high-resolution tissue-section autoradiography. Plutonium deposits in cells consisted of a nuclear and a cytoplasmic component. In the hamster kidney cells, the amount associated with the nucleus was about 55 per cent of the total cellular plutonium at 24 hours after injection. Six days later, it was only about 30 per cent. Plutonium deposits were also characterized in hepatocytes, in the interstitial cells of the testes, in the cells of ovarian follicles, in chondrocytes and in bone cells, including osteoblasts and osteocytes. In bone there appeared to be both an extracellular and intracellular deposit. No evidence was found of substantial incorporation of plutonium into the mineral phase of bone.