Late effects of the hematotoxic action of antineoplastic antibiotics of the anthracycline group

The morphological and functional states of hemopoiesis at late periods (up to 6 months) after using daunorubicin, carminomycin and doxorubicin, antitumor antibiotics of the anthracycline group in doses of 1/10 LD50 for 10 days were studied on 764 noninbred rats and 240 BALB/c mice. On various compensatory-functional models of hemopoiesis strain there was shown persisting inhibition of hematopoietic tissue functional activity and limited reserve reactions of granulocyto- and erythropoiesis 3 and to a lesser extent 6 months after the cytostatic action.     

Evidence for interactions between monocytes and natural killer cells in the regulation of in vitro hemopoiesis

The role of accessory cells in modulation of in vitro hemopoiesis was studied using C-mediated lysis with mAb of differing specificities. One such antibody, 25E11, which bound to all NK cells and a subset of monocytes, consistently led to enhancement of in vitro hemopoiesis when 25E11+ cells were depleted by complement or the FACS. In cultures maximally stimulated by erythropoietin and human placental conditioned medium, depletion of 25E11+ cells from beta-thalassemic PBMC resulted in up to a fourfold enhancement of multi-potential and pure erythroid colonies and a twofold enhancement of non-erythroid colonies when compared with C-only treated cells. This enhancement of in vitro hemopoiesis was also observed with cells from normal bone marrow, cord blood, and peripheral blood. The enhancement of in vitro hemopoiesis was abrogated by removal of either adherent cells or monocytes using Leu M3 antibody and C lysis. The data presented herein suggest that NK cells and a subset of monocytes may exert a negative regulatory control on both granulopoiesis and erythropoiesis. Consequently, the number of progenitor and multipotential cells in cultures of unfractionated cell populations may be greatly underestimated.     

Testosterone and estradiol modulate TSH-binding in the thyrocytes of Wistar rats: influence of age and sex.

Age and sex are important factors that influence thyroid pathophysiology. Though sex steroids are known to enhance thyrotropin (TSH) mRNA expression and incidence of thyroid tumours, there is no report on their effects on TSH action under normal physiological conditions. In the present study, the effects of testosterone (T) and estradiol (E2) on thyroidal TSH-receptor (TSH-R) concentration, and TSH-binding to thyrocytes (in vitro) were elucidated in immature and mature Wistar rats. Immature (10 days old) and adult (120 days old) rats of either sex were gonadectomized (GDX) and one group of GDX rats was treated with physiological doses of T and another with E2. Immature GDX rats were supplemented with the steroids for 10 days and adults were supplemented with the steroids for 30 days. While supplementation of steroids to immature rats was begun immediately after surgery, for adult rats it was started 10 days after gonadectomy. The rats were killed at the end of the experimental period. Gonadectomy significantly decreased serum TSH, and TSH-R concentration under in vivo condition and [125I]-TSH binding to thyrocytes under in vitro conditions. Supplementation of T to male and E2 to female GDX rats restored normality of the parameters. Thyrocytes of immature male rats challenged with linearly increasing doses of TSH or T (6.25-800 ng/ml) showed a dose-dependent increase in TSH-binding. However, thyrocytes of immature female rats challenged with T showed a gender-specific response. While there was a linear increase in TSH-binding in thyrocytes of males, a biphasic response was evident in thyrocytes of females. In the case of thyrocytes from adult rats, T induced a dose-dependent change in TSH-binding in males, which reached the peak in response to 12.5 ng T, and diminished thereafter. In contrast, E2 was inhibitory to TSH-binding to thyrocytes of adult male rats. On the other hand, E2 showed a clear gender-specific stimulation of TSH-binding in thyrocytes of females and an inhibition of the same in males. TSH and sex steroids upregulated TSH receptors in immature rats, whereas the effect was biphasic in adult rat thyrocytes. It is concluded from the present study that sex steroids modulate TSH-binding in rat thyrocytes, which may vary according to the age and sex of the animals.     

Induction of porphobilinogen oxygenase and porphobilinogen deaminase in rat blood under conditions of erythropoietic stress

Porphobilinogen is the substrate of two enzymes: porphobilinogen deaminase and porphobilinogen-oxygenase. The first one transforms it into the metabolic precursors of heme and the second diverts it from this metabolic pathway by oxidizing porphobilinogen to 5-oxopyrrolinones. Rat blood is devoid of porphobilinogen-oxygenase under normal conditions while it carries porphobilinogen-deaminase activity. When the rats were submitted to hypoxia (p_O2 = 0.42 atm) for 18 days, the activity of porphobilinogen-oxygenase appeared at the tenth day of hypoxia and reached the maximum at the 14–16th day. It decreased to a half after 2 days (half-life of the enzyme) and disappeared after 4 days of return to normal oxygen pressure. Porphobilinogen-deaminase activity increased after the first day of hypoxia, reached a maximum at the 14–16th day and did not decrease to normal values until the 15th day after return to normal oxygen pressure. The activities of both prophobilinogen-oxygenase and porphobilinogen-deaminase were induced by administration of erythropoietin. When rats were made anaemic with phenylhydrazine, porphobilinogen-oxygenase activity also appeared in the blood cells. Although the reticulocyte concentration was higher when compared to that obtained under hypoxia, the activities of the oxygenase obtained under both conditions were comparable. Porphobilinogen-deaminase activity was always closely related to the reticulocyte content. The appearance of porphobilinogen-oxygenase under the described erythropoietic conditions was due to a de novo induction of the enzyme, as shown by its inhibition with actinomycin D and cycloheximide. Porphobilinogen-oxygenase as well as porphobilinogen-deaminase were present in the rat bone marrow under normal conditions. Their activities increased in phenylhydrazine treated rats. The properties and kinetics of porphobilinogen-oxygenase from the rat blood and bone marrow were determined and found to differ in several aspects.     

Comparative studies on the characteristics of proliferation and differentiation of spleen colony-forming cells

Using a single spleen colony transplantation technique and sex chromosome typing as a natural cytogenetic marker, most spleen colony-forming cells (CFC) in adult bone marrow or fetal livers of inbred LACA or C57 mice re-established hemopoiesis in lethally irradiated mice when the spleen colonies were sampled at 13 days after transplantation. However, most of the spleen colony-forming cells in the peripheral blood of normal mice possess little potential for proliferation and are less efficient in the re-establishment of hemopoiesis in lethally irradiated mice. The CFC population is heterogeneous in the mice. From the subsequent retransplantation of colonies from colony-forming cells in the peripheral blood, the simple assessment of spleen colony-forming units (CFU-s) content, based on the number of splenic colonies, does not reliably represent the content of hemopoietic stem cells.     

Micronucleus induction after whole-body microwave irradiation of rats

Adult male Wistar rats were exposed for 2 h a day, 7 days a week for up to 30 days to continuous 2,450 MHz radiofrequency microwave (rf/MW) radiation at a power density of 5-10 mW/cm(2). Sham-exposed rats were used as controls. After ether anesthesia, experimental animals were euthanized on the final irradiation day for each treated group. Peripheral blood smears were examined for the extent of genotoxicity, as indicated by the presence of micronuclei in polychromatic erythrocytes (PCEs). The results for the time-course of PCEs indicated significant differences (P<0.05) for the 2nd, the 8th and the 15th day between control and treated subgroups of animals. Increased influx of immature erythrocytes into the peripheral circulation at the beginning of the experiment revealed that the proliferation and maturation of nucleated erythropoietic cells were affected by exposure to the 2,450 MHz radiofrequency radiation. Such findings are indicators of radiation effects on bone-marrow erythropoiesis and their subsequent effects in circulating red cells. The incidence of micronuclei/1,000 PCEs in peripheral blood was significantly increased (P<0.05) in the subgroup exposed to rf/MW radiation after eight irradiation treatments of 2 h each in comparison with the sham-exposed control group. It is likely that an adaptive mechanism, both in erythrocytopoiesis and genotoxicity appeared in the rat experimental model during the subchronic irradiation treatment.     

Embryonic and fetal hemopoiesis: an overview

Our current knowledge of embryonic and fetal hemopoiesis is critically reviewed in this article. In both murine and human systems, embryonic and fetal development is associated with multiple switching in the sites of hemopoiesis. The phenomenon is first extraembryonic, occurring in blood islands of the yolk sac. Hemopoietic stem cells (HSC) appear to derive from hemangioblasts that are of mesodermal origin. Yolk sac milieu is permissive only for erythropoiesis which proceeds synchronously and may be erythropoietin-insensitive. Yolk sac milieu is not permissive for the development of other cell lines. The final product is nucleated red cells. Yolk sac hemopoiesis is an example par excellence of primitive (as compared to definitive) form of hemopoiesis. HSC then seem to migrate via the bloodstream to the liver and spleen to seed these tissues, which then carry the burden of hemopoiesis until birth and for some time thereafter. Here also erythropoiesis predominates, but some granulopoiesis also occurs. Thus, the milieu is not totally impermissive. Hemopoiesis is in definitive form, lacking synchronicity of cell growth with the end product being anucleated cells and synthesized hemoglobin not limited to embryonic type. The site of hemopoiesis is finally transferred to the bone marrow, which is predominantly granulopoietic. Certain cellular and embryological features of these types of hemopoiesis in the context of more recent molecular understanding of stem cell homing are discussed.     

Iron metabolism in iron-deficient male quail

1. Male quails submitted 20 and 120 days to a low iron diet (7 ppm) were compared to female laying quails, exposed for 30 days to the same low iron regime, in order to compare the response of the iron metabolic control under a single (erythropoiesis) or a doubled (erythropoiesis and egg formation) iron demand. 2. Iron deposit in storage organs, the classical hematology and the intestinal iron absorption were analyzed in these animals. 3. In males, after 120 days, the iron deposits were reduced 50 and 75%, but hematological values (hematocrit and hemoglobin concentration) were normal, although in laying quails, after 30 days, an anemic condition was evident in both blood parameters and iron deposits, provoking an iron deficient erythropoiesis. 4. The enhancement of the intestinal iron uptake, confirms the anemic character of these birds.     

Red blood of cosmonauts during missions aboard the international space station (ISS)

Morphological and biochemical investigations of red blood in cosmonauts on board the International Space Station (ISS) (from the 6th to the 12th expeditions) in the space experiment program “Hematology” were carried out 30 days before the space mission (SM), at the initial (days 6–10) and final (days 160–190) stages of SM, and after the SM (immediately after SM (day 0) and on the 7th and 15th days of the adaptation period to earth conditions). A reduction of the concentration of hemoglobin after a prolonged influence of SM factors has been found, which is probably related not only to the intensity of erythropoiesis but also with the possible early removal of a part of low-quality (probably, old) erythrocytes from the bloodstream, which is confirmed by the results on the metabolism of red blood cells and the state of the cell membrane. Stimulation of erythropoiesis (increase in erythropoietin, decreased level of iron in blood, removal of low-quality and old erythrocytes) in the period of readaptation to conditions on earth is aimed at maintenance of the optimal level of red blood cells required for increased oxygen demand in tissues under the conditions of earth gravitation and enhancement of muscular load.     

Biomechanical response of arterial wall to DOCA-salt hypertension in growing and middle-aged rats

To study arterial remodeling in response to hypertension, Deoxycortico-sterone acetate (DOCA)-salt hypertension was induced in immature (aged 16 weeks) and middle-aged (48 weeks) rats, and biomechanical properties and wall dimensions of common carotid arteries were determined. Arterial segments were excised at 10 or 16 weeks postoperatively from the immature rats and at 16 weeks from the middle-aged ones. In vitro pressure-diameter tests were performed under normal (in Krebs-Ringer solution), active (norepinephrine), and passive (papaverine) conditions. Non-treated, age-matched rats (26, 32, and 64 weeks) were used to obtain control data. Wall thickness at in vivo blood pressure level was increased by hypertension at all ages; however, there were no significant changes in inner diameter. In hypertensive rats, arterial outer diameter was smaller under normal condition than under passive condition, indicating the increase of smooth muscle tone by hypertension. Diameter reduction developed by norepinephrine was increased by hypertension, which was significant above 100 mmHg; however, there were no significant differences between hypertensive and normotensive arteries, if compared at respective in vivo blood pressures. No significant differences were observed in wall stiffness at in vivo pressure. Wall hoop stress at in vivo blood pressure had a significant positive correlation with the pressure in 26-week old arteries. However, there were no differences in the stress between hypertension and normotension in 32- and 64-week old arteries. These results were essentially similar to previous ones observed in Goldblatt hypertension and in younger animals. Age-related differences in arterial wall remodeling were not clearly observed.     

THe dynamics of erythropoiesis inhibition in posttransfusion polycythaemia in the rat

The dynamics of erythropoiesis inhibition as the consequence of polycythaemia induced in rats by four to six transfusions of homologous erythrocytes was studied in detail. It was found, that, in rats with polycythaemia elicited by two transfusions of erythrocytes, erythropoiesis inhibition occurs 67h after the first transfusion and it is most pronounced in the period between 115-187 h. The hemopoietic system is not completely free from the inhibitory influence of polycythaemia up to 283 h after the first transfusion. Maintenance of a state of polycythaemia for 17 days by repeated transfusions strongly and durably inhibits the haemopoietic system. The red blood cell system remained for this whole period under the inhibitory effect.     

The mechanism of inactivation of the bird erythrocyte genome. III. Pathways of terminal differentiation of erythrocytes]

Under the conditions of acute phenylhydrazine anemia in pigeons, early erythroblasts develop in reticulocytes with basophilic cytoplasm directly by passing the stages of polychromatophilic and ortochromic erythroblasts. During the maximum activity of bone marrow erythropoiesis stimulated by anemia over 80% of reticulocytes develop in such a way. A hypothesis is put forward to the effect that there exist in birds two alternative pathways of erythropoiesis: the main pathway which ensures the slow repopulation of blood by erythrocytes under normal conditions and the reserve pathway which opens for a short time under anemia and ensures rapid increase of the erythrocyte titer in blood up to its normal value.     

Effect of erythrocyte breakdown products on mast cells and erythropoietin formation]

Experiments were conducted on CBA mice and albino rats. A study was made of the effect of erythrocyte destruction products (EDP) on the content of hemopoietic colony-forming units (CFU), differentiation of stem cells and the erythropoietin production. It was shown that 3 or 4 EDP injections to normal mice or to lethally irradiated (1000 rad) mice after the transplantation of bone marrow cells caused no changes in the CFU level of stem cells differentiation. In case of a daily (for 3 days) administration of EDP to mice before the irradiation (1000 rad) and bone marrow transplantation there was observed an increase of the colonies count in the recipients' spleen on account of the erythroid colonies. EDP injection caused no changes in the erythropoietic activity of the blood serum. A possible role of erythrocyte destruction products in the mechanism of erythropoiesis autoregulation is discussed.     

Role of the renin-angiotensin system in the adaptation to high salt intake in immature rats

The response of the renin-angiotensin system, extracellular fluid volume, plasma volume, plasma sodium and mean arterial blood pressure to an increase in salt intake (8% NaCl in the diet for 10 days) was compared in immature (20 days) and adult (80 days) rats which were either sham-operated or uninephrectomised. Salt feeding induced a significant increase in plasma sodium in immature animals, and a greater suppression of the renin-angiotensin system in immature than in adult rats, although extracellular fluid volume, plasma volume and blood pressure remained unchanged. Following uninephrectomy, however, the renin-angiotensin system was maximally suppressed in both age groups and in younger animals extracellular fluid volume, plasma volume and blood pressure were significantly increased. It is concluded that (i) the renin-angiotensin system in immature rats is more responsive to a chronically increased salt intake, (ii) this greater responsiveness partly compensates for the lower natriuretic efficiency of the kidneys of immature rats, which becomes evident after reduction of renal mass, and (iii) these events bear a relation to the higher susceptibility of prepubertal rats to the hypertensive effect of a chronically increased salt intake.     

Erythrocyte senescence and haematological changes induced by starvation in the neotropical fish traíra, Hoplias malabaricus (Characiformes, Erythrinidae)

Adult specimens of traira (Hoplias malabaricus Bloch) were subjected to long-term starvation (30 to 240 days) and re-fed for 30 days after 90 and 240 days of food deprivation. Counting of immature erythrocytes in peripheral blood showed that erythropoiesis decreased significantly during the first 30 days of food deprivation. The results suggest that a process of senescence takes place in the pre-existent red blood cells and that the cells are not replaced during starvation. After 240 days of starvation, H. malabaricus had a significantly reduced number of red blood cells, causing changes in hematocrit and blood indices (mean corpuscular volume, mean corpuscular haemoglobin and mean corpuscular haemoglobin concentration). Furthermore, during this period, the fish presented leukopenia (lymphocytopenia) and thrombocytopenia. After re-feeding, the number of leukocytes and thrombocytes recovered, but the red blood cell number remained reduced and there was a significant increase in abnormal red cell nuclei.     

Brain adaptation to chronic hypobaric hypoxia in rats.

Rats were exposed to hypobaric hypoxia (0.5 atm) for up to 3 wk. Hypoxic rats failed to gain weight but maintained normal brain water and ion content. Blood hematocrit was increased by 48% to a level of 71% after 3 wk of hypoxia compared with littermate controls. Brain blood flow was increased by an average of 38% in rats exposed to 15 min of 10% normobaric oxygen and by 23% after 3 h but was not different from normobaric normoxic rats after 3 wk of hypoxia. Sucrose space, as a measure of brain plasma volume, was not changed under any hypoxic conditions. The mean brain microvessel density was increased by 76% in the frontopolar cerebral cortex, 46% in the frontal motor cortex, 54% in the frontal sensory cortex, 65% in the parietal motor cortex, 68% in the parietal sensory cortex, 68% in the hippocampal CA1 region, 57% in the hippocampal CA3 region, 26% in the striatum, and 56% in the cerebellum. The results indicate that hypoxia elicits three main responses that affect brain oxygen availability. The acute effect of hypoxia is an increase in regional blood flow, which returns to control levels on continued hypoxic exposure. Longer-term effects of continued moderate hypoxic exposure are erythropoiesis and a decrease in intercapillary distance as a result of angiogenesis. The rise in hematocrit and the increase in microvessel density together increase oxygen availability to the brain to within normal limits, although this does not imply that tissue PO2 is restored to normal.     

Retention of hemopoiesis in tail vertebrae of newborn rats

Summary In newborn rats, the marrow cavity of tail vertebrae is hemopoietic and contains no adipose tissue. The latter develops soon after birth to replace the hemopoietic tissue within the nonexpansile volume of the marrow cavity. By transposing the tail into the warmer environment of the abdomen, hemopoiesis was retained, and the development of adipose tissue was prevented in the transposed segment, when the operation was done in preweanling but not in adult animals. Systemic stimuli of erythropoiesis (phlebotomy, induced hemolysis) acted synergistically with the temperature increment to retain hemopoiesis. The findings support the concept that adipose cells in the yellow marrow are relatively stable and once developed, they are not readily mobilized. The findings may also explain the discrepancies in the results obtained by rat tail transposition.Supported by ERDA Contract EP-78-S-03-0899, NASA Grant NSG 9061 and RCDA AM16125 awarded to Mehdi Tavassoli     

Role of the spleen in the exaggerated polycythemic response to hypoxia in chronic mountain sickness in rats.

In a rat model of chronic mountain sickness, the excessive polycythemic response to hypoxic exposure is associated with profound splenic erythropoiesis. We studied the uptake and distribution of radioactive iron and red blood cell (RBC) morphology in intact and splenectomized rats over a 30-day hypoxic exposure. Retention of (59)Fe in the plasma was correlated with (59)Fe uptake by both spleen and marrow and the appearance of (59)Fe-labeled RBCs in the blood. (59)Fe uptake in both the spleen and the marrow paralleled the production of nucleated RBCs. Splenic (59)Fe uptake was approximately 10% of the total marrow uptake under normoxic conditions but increased to 60% of the total marrow uptake during hypoxic exposure. Peak splenic (59)Fe uptake and splenomegaly occurred at the most intense phase of erythropoiesis and coincided with the rapid appearance of (59)Fe-labeled RBCs in the blood. The bone marrow remains the most important erythropoietic organ under both resting and stimulated states, but inordinate splenic erythropoiesis in this rat strain accounts in large measure for the excessive polycythemia during the development of chronic mountain sickness in chronic hypoxia.     

Micronucleus induction after whole-body microwave irradiation of rats

Adult male Wistar rats were exposed for 2 h a day, 7 days a week for up to 30 days to continuous 2450 MHz radiofrequency microwave (rf/MW) radiation at a power density of 5–10 mW/cm². Sham-exposed rats were used as controls. After ether anesthesia, experimental animals were euthanized on the final irradiation day for each treated group. Peripheral blood smears were examined for the extent of genotoxicity, as indicated by the presence of micronuclei in polychromatic erythrocytes (PCEs). The results for the time-course of PCEs indicated significant differences (P<0.05) for the 2nd, the 8th and the 15th day between control and treated subgroups of animals. Increased influx of immature erythrocytes into the peripheral circulation at the beginning of the experiment revealed that the proliferation and maturation of nucleated erythropoietic cells were affected by exposure to the 2450 MHz radiofrequency radiation. Such findings are indicators of radiation effects on bone-marrow erythropoiesis and their subsequent effects in circulating red cells. The incidence of micronuclei/1000 PCEs in peripheral blood was significantly increased (P<0.05) in the subgroup exposed to rf/MW radiation after eight irradiation treatments of 2 h each in comparison with the sham-exposed control group. It is likely that an adaptive mechanism, both in erythrocytopoiesis and genotoxicity appeared in the rat experimental model during the subchronic irradiation treatment.     

Repair processes of hemopoiesis after applying cyclophosphamide. I. Morphological changes in the bone marrow, spleen and thymus

Processes of hemopoiesis repair were monitored in rats till the day 30 after applying cyclophosphamide in doses of 100 and 200 mg X kg-1. Profound decrease of the hemopoietic activity in the bone marrow, spleen, and thymus was observed till the day 3. The reported changes were reversible, since the myelopoiesis was recovered till the day 10 in the bone marrow and the erythropoiesis in the spleen till the day 20. Significantly slower progress in repair process was observed in the lymphoid tissue; here the recovery was not completed till the day 30.