Michal Hofer

Elevation of extracellular adenosine induces radioprotective effects in mice.

The radioprotective effectiveness of the elevation of extracellular adenosine induced in mice by the combined administration of dipyridamole, a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate, a soluble adenosine pro-drug, was evaluated. Based on survival studies, endogenous hemopoietic spleen colony formation, and the postirradiation behavior of bone marrow granulocyte-macrophage colony-forming cells (GM-CFC), it was demonstrated that the combined administration of dipyridamole and AMP protects mice when given either 15 or 60 min before irradiation. It could be deduced that the radioprotective action is induced by at least two independent mechanisms: (1) protection by hypoxia as a result of the effect of the treatment on the cardiovascular system, and (2) and enhanced regeneration of the hemopoietic stem cells due to either enhanced postirradiation repair or an increased proliferation of the hemopoietic stem cells. Both of these protective mechanisms, which are able to increase the regeneration of hemopoiesis, seemed to be effective in enhancing the survival of mice given single radiation exposures, with a dose reduction factor for the LD50/30 of 1.11. The protective efficiency of the mechanisms enhancing the postirradiation recovery of hemopoiesis was also evident in experiments evaluating the survival of mice subjected to fractionated irradiation and a repeated administration of the protective agents.

Granulocyte colony-stimulating factor and drugs elevating extracellular adenosine synergize to enhance haematopoietic reconstitution in irradiated mice

Abstract: The activation of adenosine receptors has recently been demonstrated to stimulate haematopoiesis. In the present study, we investigated the ability of drugs elevating extracellular adenosine to influence curative effects of granulocyte colony‐stimulating factor (G‐CSF) in mice exposed to a sublethal dose of 4 Gy of 60Co radiation. Elevation of extracellular adenosine in mice was induced by the combined administration of dipyridamole, a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate (AMP), an adenosine prodrug. The effects of dipyridamole plus AMP, and G‐CSF, administered either alone or in combination, were evaluated. The drugs were injected to mice in a 4‐d treatment regimen starting on d 3 after irradiation and the haematopoietic response was evaluated on d 7, 10, 14, 18 and 24 after irradiation. While the effects of G–CSF on the late maturation stages of blood cells, appearing shortly after the completion of the treatment, were not influenced by dipyridamole plus AMP, positive effects of the combination therapy occurred in the post‐irradiation recovery phase which is dependent on the repopulation of haematopoietic stem cells. This was indicated by the significant elevation of counts of granulocyte‐macrophage progenitor cells (GM‐CFC) and granulocytic cells in the bone marrow (d 14), of GM‐CFC (d 14), granulocytic and erythroid cells (d 14 and 18) in the spleen, and of neutrophils (d 18), monocytes (d 14 and 18) and platelets (d 18) in the peripheral blood. These effects suggest that the repopulation potential of the combination therapy lies in a common multilineage cell population. The results of this study implicate the promising possibility to enhance the curative effects of G–CSF under conditions of myelosuppressive states induced by radiation exposure.

Drugs elevating extracellular adenosine promote regeneration of haematopoietic progenitor cells in severely myelosuppressed mice: their comparison and joint effects with the granulocyte colony-stimulating factor.

Abstract: We tested capabilities of drugs elevating extracellular adenosine and of granulocyte colony‐stimulating factor (G‐CSF) given alone or in combination to modulate regeneration from severe myelosuppression resulting from combined exposure of mice to ionizing radiation and carboplatin. Elevation of extracellular adenosine was induced by joint administration of dipyridamole (DP), a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate (AMP), serving as an adenosine prodrug. DP + AMP, G‐CSF or all these drugs in combination were administered in a 4‐d treatment regimen starting on day 3 after induction of myelosuppression. Comparable enhancements of haematopoietic regeneration due to elevation of extracellular adenosine or to action of G‐CSF were demonstrated as shown by elevated numbers of haematopoietic progenitor cells for granulocytes/macrophages (GM‐CFC) and erythrocytes (BFU‐E) in the bone marrow and spleen in early time intervals after termination of the drug treatment, i.e. on days 7 and 10 after induction of myelosuppression. Coadministration of all the drugs further potentiated the restoration of progenitor cell pools in the haematopoietic organs. The effects of the drug treatments on progenitor cells were reflected in the peripheral blood in later time intervals of days 15 and 20 after induction of myelosuppression, especially as significantly elevated numbers of granulocytes and less pronounced elevation of lymphocytes and erythrocytes. The results substantiate the potential of drugs elevating extracellular adenosine for clinical utilization in myelosuppressive states, e.g. those accompanying oncological radio‐ and chemotherapy.

Meloxicam, a Cyclooxygenase 2 Inhibitor, Supports Hematopoietic Recovery in Gamma-Irradiated Mice

Abstract Hofer, M., Pospíšil, M., Znojil, V., Holá, J., Vacek, A., Weiterová, L., Štreitová, D. and Kozub;aaik, A. Meloxicam, a Cyclooxygenase 2 Inhibitor, Supports Hematopoietic Recovery in Gamma-Irradiated Mice. Radiat. Res. 166, 556–560 (2006). Meloxicam, a selective inhibitor of cyclooxygenase 2, a nonsteroidal anti-inflammatory drug with an improved side-effects profile in terms of gastrointestinal toxicity, has been found to stimulate hematopoiesis in whole-body γ-irradiated mice. A distinct corroboration of this positive action of meloxicam is an enhancement of the recovery of hematopoietic progenitor cells committed to granulocyte-macrophage and erythroid development, which has been demonstrated in sublethally irradiated animals treated with meloxicam at a dose of 20 mg/kg administered intraperitoneally either singly 1 h before irradiation or repeatedly after radiation exposure. The results suggest that meloxicam can be added to the list of biological response modifiers that can be used in the treatment of hematopoietic damage induced by ionizing radiation.

Influence of the joint treatment with granulocyte colony-stimulating factor and drugs elevating extracellular adenosine on erythropoietic recovery following 5-fluorouracil-induced haematotoxicity in mice.

Abstract: The presented data address the problem of pleiotropic effects of granulocyte colony‐stimulating factor (G‐CSF) and suggest the ability of drugs increasing the level of extracellular adenosine to activate erythropoiesis when given jointly with G‐CSF. To demonstrate these interactions, the effects of the drugs on the recovery from erythropoietic damage induced in mice by a single dose of 5‐fluorouracil (5‐FU) were investigated. Elevation of extracellular adenosine and thus activation of adenosine receptors was induced by joint administration of dipyridamole (DP), a drug inhibiting the cellular uptake of adenosine, and adenosine monophosphate (AMP), an adenosine prodrug. The drugs were injected in a 4‐d treatment regimen starting 2 h after 5‐FU injection. Both DP+AMP and G‐CSF alone induced only weak effects. However, the combination of the three drugs produced significant elevation of erythrocytes in the peripheral blood which pertained in the post‐treatment period. Stimulation of proliferation of erythroid progenitor cells (BFU‐E) in femoral bone marrow and increased levels of reticulocytes in the peripheral blood were observed in the course of the 4‐d treatment regimen. In addition, significantly decreased mean cell haemoglobin accompanying the elevated numbers of erythrocytes in the combination‐treated mice was found. This effect could be interpreted as the result of a sublethal 5‐FU‐induced damage to erythroid progenitor and precursor cells forced to proliferate intensively by the combination therapy. The observed additivity and synergism of G‐CSF with elevated extracellular adenosine in terms of erythropoiesis is an interesting finding with potential implications in clinical practice.

Drugs elevating extracellular adenosine enhance cell cycling of hematopoietic progenitor cells as inferred from the cytotoxic effects of 5-fluorouracil

OBJECTIVE Our previous studies showed that the combined administration of drugs elevating extracellular adenosine, i.e., dipyridamole and adenosine monophosphate (AMP), enhanced hematopoiesis in normal mice and increased hematopoietic recovery in irradiated mice. In the present study, we have examined the possibility that these effects are due to the adenosine-induced cycling of the hematopoietic progenitor cells. MATERIALS AND METHODS Experiments were performed under in vivo conditions using B10CBAF1 mice. The cycling status of hematopoietic progenitor cells (CFU-S(day 10), CFC-GM, and BFU-E) was determined on the basis of their sensitivity to 5-fluorouracil (5-FU), a cycle-specific cytotoxic agent. RESULTS Pretreatment of mice with dipyridamole + AMP enhanced the cytotoxic effects of a single bolus of 5-FU at a dose of 3 mg per mouse. Sensitizing effects of drugs occurred after a delay of several hours and attained a maximum of about 40-60% reduction of the progenitor cells surviving after 5-FU alone. The period of maximum sensitization of CFU-S by the combination of dipyridamole + AMP was shifted to later time intervals as compared with the effects on CFC-GM and BFU-E. Pretreatment of mice with the drugs also aggravated the 5-FU-induced lethality. Reduction of survival was found in mice exposed to two cycles of 3 mg of 5-FU following the pretreatment with dipyridamole + AMP at a time period characterized by the highest fraction of CFU-S in the S phase. CONCLUSIONS The results suggest that adenosine receptor signaling, induced by the administration of drugs elevating extracellular adenosine, enhances cycling of the hematopoietic progenitor cells. These effects might have pharmacological implications in the therapy of blood disorders.

Radioprotective action of extracellular adenosine on bone marrow cells in mice exposed to gamma rays as assayed by the micronucleus test

Abstract Hofer, M., Mazur, L., Pospíšil, M., Weiterová, L. and Znojil, V. Radioprotective Action of Extracellular Adenosine on Bone Marrow Cells in Mice Exposed to Gamma Rays as Assayed by the Micronucleus Test. The frequency of micronucleated polychromatic erythrocytes (PCEs) in mouse bone marrow was assessed after administration of dipyridamole and/or adenosine monophosphate (AMP) to nonirradiated mice or to mice irradiated 15 min later with a sublethal dose of 6.5 Gy γ rays. In nonirradiated mice, the administration of the drugs increased the frequency of micronucleated PCEs significantly (by 108%). In contrast, in irradiated mice, the number of radiation-induced micronucleated PCEs was significantly decreased if the mice had been pretreated with dipyridamole or AMP alone (by 24% after administration of each of the compounds) and in particular after administration of the drugs in combination (by 36%).

Inhibition of cyclooxygenase 2 in mice increases production of g-csf and induces radioprotection.

Abstract Hofer, M., Pospíšil, M., Holá, J., Vacek, A., Štreitová, D. and Znojil, V. Inhibition of Cyclooxygenase 2 in Mice Increases Production of G-CSF and Induces Radioprotection. Radiat. Res. 170, 566–571 (2008). Meloxicam, a selective inhibitor of cyclooxygenase 2, was tested to determine its ability to modulate hematopoiesis and to influence survival of mid-lethally γ-irradiated mice. A single dose of meloxicam (20 mg/kg) administered to mice intraperitoneally 1 h before irradiation was shown to enhance serum levels of granulocyte colony-stimulating factor (G-CSF) during the first 24 h after irradiation, to elevate numbers of granulocytic precursor cells in bone marrow and granulocyte counts in peripheral blood on day 10 after irradiation, and to increase 30-day survival of these mice. The results provide new evidence for the protective ability of meloxicam administration to mice irradiated with mid-lethal doses and contribute to the understanding of the mechanisms of this meloxicam action by drawing attention to the possible role of increased endogenous G-CSF production.

Modulation of Animal and Human Hematopoiesis by β-Glucans: A Review

β-Glucans are cell wall constituents of bacteria, yeast, fungi, and plants. They are not expressed in mammalian cells, but they are recognized by mammalian cells as pathogen-associated molecular patterns by pattern recognition receptors and thus act as biological response modifiers. This review summarizes data on the hematopoiesis-stimulating effects of β-glucans, as well as on their ability to enhance bone marrow recovery after an injury. β-Glucans have been shown to support murine hematopoiesis suppressed by ionizing radiation or cytotoxic anti-cancer therapy. They also enhance stem cell homing and engraftment. Basically, two forms of β-glucan preparations have been investigated, namely particulate and soluble ones. β-Glucans are generally well tolerated, the particulate forms showing a higher incidence of undesirable side effects. Taken together, the hematopoiesis-stimulating properties of β-glucans predetermine these biological response modifiers to ever increasing use in human medicinal practice.

The role of adenosine receptor agonists in regulation of hematopoiesis.

The review summarizes data evaluating the role of adenosine receptor signaling in murine hematopoietic functions. The studies carried out utilized either non-selective activation of adenosine receptors induced by elevation of extracellular adenosine or by administration of synthetic adenosine analogs having various proportions of selectivity for a particular receptor. Numerous studies have described stimulatory effects of non-selective activation of adenosine receptors, manifested as enhancement of proliferation of cells at various levels of the hematopoietic hierarchy. Subsequent experimental approaches, considering the hematopoiesis-modulating action of adenosine receptor agonists with a high level of selectivity to individual adenosine receptor subtypes, have revealed differential effects of various adenosine analogs. Whereas selective activation of A1 receptors has resulted in suppression of proliferation of hematopoietic progenitor and precursor cells, that of A3 receptors has led to stimulated cell proliferation in these cell compartments. Thus, A1 and A3 receptors have been found to play a homeostatic role in suppressed and regenerating hematopoiesis. Selective activation of adenosine A3 receptors has been found to act curatively under conditions of drug- and radiation-induced myelosuppression. The findings in these and further research areas will be summarized and mechanisms of hematopoiesis-modulating action of adenosine receptor agonists will be discussed.