ENDOCRINE REGULATION OF ERYTHROID LINEAGE OF HEMATOPOIESIS IN CHILDREN LIVING UNDER A LOW-DOSE RADIATION EXPOSURE AFTER THE CHORNOBYL NPP ACCIDENT. / ENDOKRYNNA REGULIaTsIIa ERYTROTsYTARNOÏ LANKY GEMOPOEZU V DITEI, IaKI ZhYVUT' NA RADIOAKTYVNO ZABRUDNENYKh TERYTORIIaKh PISLIa AVARIÏ NA ChAES.

OBJECTIVE: Elucidation of relationship between the levels of thyroid-stimulating hormone (TSH), free serum thyroxine, serum and urine cortisol and parameters of erythroid lineage of hematopoiesis to estimate the thyroid functionin children of prepubertal, pubertal, and postpubertal age permanently residing under a low-dose radiation exposureto determine the premorbid state of thyroid function. MATERIALS AND METHODS: Children aged 3 to 18 years old (n = 203) living in the most intensively radionuclide-contaminated regions of Kyiv, Zhytomyr and Chornihiv oblasts of Ukraine after the Chornobyl NPP accident wereenrolled. Complaints of ossalgia, arthralgia, fatigue, bone fractures in the history, bone dysembryogenetic stigmata,hypermobility syndrome degree, and types of somatic diseases were taken into account. Peripheral blood countparameters, biochemical indices of blood serum were studied, namely the levels of total protein, cholesterol, creatinine and alkaline phosphatase activity. Levels of the free thyroxine, pituitary TSH, serum and daily urine cortisol, anddoses of radiation exposure were determined. RESULTS: The radiation dose values in children ranged from (0.35 ± 0.09) mSv to (0.54 ± 0.12) mSv. There was nodifference between the parameters of erythroid lineage of hematopoiesis depending on radiation dose. At the levels of serum TSH up to 1.0 µIU/ml no correlation was found with cortisol levels; at TSH levels of 1.0-3.0 µIU/ml thecorrelation coefficient was r = 0.31; at TSH levels higher than 3.0 µIU/ml the correlation coefficient was r = 0.61probably indicating a compensatory role of adrenal cortex in children at risk of thyroid disease development. In children with joint hypermobility grade II there was a higher incidence of dentofacial anomalies (χ2 = 6.9), deformitiesof lower extremities (χ2 = 6.9), and dental caries (χ2 = 4.3) (p < 0.05). There was a direct correlation between theserum TSH level (over 3 µIU/ml) and micrognathia (brachygnathia) (r = 0.62) indicating the impact of thyroid disease on dentofacial development. The TSH at a level of upper limit of the reference range values may contribute toa decreased RBC count in peripheral blood, increased average volume and hemoglobin content in erythrocyte beingassociated with the initial manifestations of thyroid dysfunction. CONCLUSIONS: Abnormal endocrine regulation of hematopoiesis affects the connective tissue, stromal microenvironment of bone marrow, and accordingly the erythroid branch of hematopoiesis in children, which may be relevant inthe development and course of oncohematological diseases.

PROGNOSIS OF ACUTE LEUKEMIA DEPENDING ON THE IRON METABOLISM PARAMETERS IN CHILDREN AFTER CHORNOBYL NUCLEAR POWER PLANT ACCIDENT. / PROGNOZ PEREBIGU GOSTRYKh LEIKEMII U DITEI PISLIa AVARIÏ NA ChAES ZALEZhNO VID OBMINU ZALIZA.

OBJECTIVE: To determine the influence of iron metabolism on the prognosis of acute lymphoblastic (ALL) and (AML)myeloblastic leukemia at the different phases of chemotherapy in children after Chоrnobyl accident. MATERIALS AND METHODS: 333 children (295 - ALL, 38 - AML) were examined at the stages of chemotherapy. Thecomparison group included 93 children without leukemia. Acute leukemia variants, patients survival, relapses, thenature of disease (live child or died), iron methabolism (morphometric parameters of erythrocytes, SI, SF, STf, TS),manifestations of dyserythropoiesis, bone marrow sideroblast and patients radiation dose were taken into account. RESULTS: In 295 patients with ALL the following variants of leukemia were established: pro-B-ALL in 23, «common¼type of ALL in 224, pre-B-ALL in 29, T-ALL in 19. Thirty eight patients were diagnosed with AML (11 - M1, 19 - M2,8 - M4). Doses of radiation in patients with AL were (2.78 ± 0.10) mSv and they did not correlate with clinical andhematological parameters, disease variant. Relapse rates and shorter survival were in patients with T-ALL, pro-B-ALLand AML with SF levels > 500 ng/ml (p < 0.05). The amount of children with normochromic-normocytic anemias andmanifestations of dysplasia of erythroid lineage elements was greater in the AML than in ALL. SF content in patientswas elevated during chemotherapy and was lower than the initial one only in the remission period. Transferrin wasreliably overloaded with iron: TS (70.2 ± 2.3) % compared with the control group (32.7 ± 2.1) %. Correlationbetween TS and survival of patients was detected (rs = -0.45). Direct correlation between the number of iron granules in erythrocariocytes and SF level (rs = 0.43) was established, indicating the phenomena of ineffective erythropoiesis. CONCLUSIONS: The negative influence of iron excess in the patients body on the hemopoiesis function, manifestations of ineffective erythropoiesis and the course of acute leukemia in children have been established. Changes inferrokinetic processes in children can be the basis of leukemоgenesis development.

The influence of therapeutic radiation on the patterns of bone marrow in ovary-intact and ovariectomized mice.

BACKGROUND: The functional components of bone marrow (i.e., the hematopoietic and stromal populations) and the adjacent bone have traditionally been evaluated incompletely as distinct entities rather than the integrated system. We perturbed this system in vivo using a medically relevant radiation model in the presence or absence of ovarian function to understand integrated tissue interaction. METHODOLOGY/PRINCIPAL FINDINGS: Ovary-intact and ovariectomized mice underwent either no radiation or single fractional 16 Gy radiation to the caudal skeleton (I ± R, OVX ± R). Marrow fat, hematopoietic cellularity, and cancellous bone volume fraction (BV/TV %) were assessed. Ovariectomy alone did not significantly reduce marrow cellularity in non-irradiated mice (OVX-R vs. I-R, p = 0.8445) after 30 days; however it impaired the hematopoietic recovery of marrow following radiation exposure (OVX+R vs. I+R, p = 0.0092). The combination of radiation and OVX dramatically increases marrow fat compared to either factor alone (p = 0.0062). The synergistic effect was also apparent in the reduction of hematopoietic marrow cellularity (p = 0.0661); however it was absent in BV/TV% changes (p = 0.2520). The expected inverse relationship between marrow adiposity vs. hematopoietic cellularity and bone volume was observed. Interestingly compared with OVX mice, intact mice demonstrated double the reduction in hematopoietic cellularity and a tenfold greater degree of bone loss for a given unit of expansion in marrow fat. CONCLUSIONS/SIGNIFICANCE: Ovariectomy prior to delivery of a clinically-relevant focal radiation exposure in mice, exacerbated post-radiation adipose accumulation in the marrow space but blunted bone loss and hematopoietic suppression. In the normally coupled homeostatic relationship between the bone and marrow domains, OVX appears to alter feedback mechanisms. Confirmation of this non-linear phenomenon (presumably due to differential radiosensitivity) and demonstration of the mechanism of action is needed to provide strategies to diminish the effect of radiation on exposed tissues.

Sublethal radiation injury uncovers a functional transition during erythroid maturation.

OBJECTIVE: Clastogenic injury of the erythroid lineage results in anemia, reticulocytopenia, and transient appearance of micronucleated reticulocytes. However, the micronucleated reticulocyte dose-response in murine models is only linear to 2 Gy total body irradiation and paradoxically decreases at higher exposures, suggesting complex radiation effects on erythroid intermediates. To better understand this phenomenon, we investigated the kinetics and apoptotic response of the erythron to sublethal radiation injury. MATERIALS AND METHODS: We analyzed the response to 1 and 4 Gy total body irradiation of erythroid progenitors and precursors using colony assays and imaging flow cytometry, respectively. We also investigated cell cycling and apoptotic gene expression of the steady-state erythron. RESULTS: After 1 Gy total body irradiation, erythroid progenitors and precursors were partially depleted. In contrast, essentially all bone marrow erythroid progenitors and precursors were lost within 2 days after 4 Gy irradiation. Imaging flow cytometry analysis revealed preferential loss of phenotypic erythroid colony-forming units and proerythroblasts immediately after sublethal irradiation. Furthermore, these populations underwent radiation-induced apoptosis, without changes in steady-state cellular proliferation, at much higher frequencies than later-stage erythroid precursors. Primary erythroid precursor maturation is associated with marked Bcl-xL upregulation and Bax and Bid downregulation. CONCLUSIONS: Micronucleated reticulocyte loss after higher sublethal radiation exposures results from rapid depletion of erythroid progenitors and precursors. This injury reveals that erythroid colony-forming units and proerythroblasts constitute a particularly proapoptotic compartment within the erythron. We conclude that the functional transition of primary proerythroblasts to later-stage erythroid precursors is characterized by a shift from a proapoptotic to an antiapoptotic phenotype.

Induction of gamma-globin mRNA, erythroid differentiation and apoptosis in UVA-irradiated human erythroid cells in the presence of furocumarin derivatives.

Psoralens, also known as furocoumarins, are a class of photosensitizers largely used in the therapy of various skin diseases. In this study we have evaluated the combined effects of UVA irradiation and furocoumarins derivatives on (a) erythroid differentiation and apoptosis of human leukemia K562 cells and (b) globin gene expression in cultures of human erythroid progenitors derived from the peripheral blood. To prove the activity of a series of linear and angular furocoumarins derivatives, we employed the human leukemia K562 cell line and the two-phase liquid culture procedure for growing erythroid progenitors. Quantitative real-time reverse transcription polymerase-chain assay (Q-RT-PCR) was employed for quantification of the accumulation of globin mRNAs. The results obtained demonstrate that both linear and angular furocoumarins are strong inducers of erythroid differentiation of K562 cells. From a preliminary screening, we have selected two derivatives, 5-methoxypsoralen (5-MOP) and trimethylangelicin (TMA), for which we have investigated their mechanism of action. The cell cycle analysis showed that these derivatives induce, after irradiation, a cell cycle arrest in the G2/M phase, followed by apoptosis. Mitochondrial depolarisation and caspases activation seem to be involved in the mechanism of cell death. In erythroid precursor cells, psoralens in combination with UVA irradiation, stimulate at very low concentrations a preferential increase of gamma-globin mRNA. Altogether, these data suggest that psoralen derivatives warrant further evaluation as potential therapeutic drugs in beta-thalassemia and sickle cell anemia.

A novel polysaccharide of black soybean promotes myelopoiesis and reconstitutes bone marrow after 5-flurouracil- and irradiation-induced myelosuppression.

The aim of this study was to investigate the promotion of myelopoiesis by an active polysaccharide of black soybean (PSBS). Murine spleen cells were collected from ICR mice and conditioned media (SCM) was prepared by incubating these cells without PSBS (normal-SCM) or with PSBS in concentrations ranging from 12.5 to 100 microg/ml (PSBS-SCM). Murine bone marrow cells were treated with PSBS alone or SCM to induce the formation of colonies, including CFU-GM, CFU-GEMM, BFU-E and HPP-CFC. The concentrations of six hematopoietic growth factors contained in SCM were measured using enzyme-linked immunoassay. In the live animal experiment, PSBS was administered orally to total body-irradiated (TBI) and 5-fluorouracil (5-FU)-treated mice to assess the reconstitution of bone marrow after myelosuppression. PSBS-SCM stimulated CFU-GM, CFU-GEMM, BFU-E and HPP-CFC colony formation with 45.0, 5.0, 6.2 and 6.6-fold increases, respectively. However, neither PSBS alone nor normal-SCM had such a colony-stimulating effect. In PSBS-SCM, the levels of IL-6, IL-17, G-CSF and GM-CSF were markedly increased, but not those of IL-3 and SCF. Oral administration of PSBS in mice not only restored the leukocyte counts reduced by TBI and 5-FU treatment but also enhanced CFU-GM colony formation of bone marrow cells without a significant change in body weight. We conclude that PSBS promotes myelopoiesis activity in the bone marrow, stimulates production of various hematopoietic growth factors from spleen cells, and reconstitutes bone marrow that has been myelosuppressed by irradiation and 5-FU.