Fluacrypyrim Protects Hematopoietic Stem and Progenitor Cells against Irradiation via Apoptosis Prevention.

Ionizing radiation (IR)-induced hematopoietic injury has become a global concern in the past decade. The underlying cause of this condition is a compromised hematopoietic reserve, and this kind of hematopoietic injury could result in infection or bleeding, in addition to lethal mishaps. Therefore, developing an effective treatment for this condition is imperative. Fluacrypyrim (FAPM) is a recognized effective inhibitor of STAT3, which exhibits anti-inflammation and anti-tumor effects in hematopoietic disorders. In this context, the present study aimed to determine whether FAPM could serve as a curative agent in hematopoietic-acute radiation syndrome (H-ARS) after total body irradiation (TBI). The results revealed that the peritoneally injection of FAPM could effectively promote mice survival after lethal dose irradiation. In addition, promising recovery of peripheral blood, bone marrow (BM) cell counts, hematopoietic stem cell (HSC) cellularity, BM colony-forming ability, and HSC reconstituting ability upon FAPM treatment after sublethal dose irradiation was noted. Furthermore, FAPM could reduce IR-induced apoptosis in hematopoietic stem and progenitor cells (HSPCs) both in vitro and in vivo. Specifically, FAPM could downregulate the expressions of p53-PUMA pathway target genes, such as Puma, Bax, and Noxa. These results suggested that FAPM played a protective role in IR-induced hematopoietic damage and that the possible underlying mechanism was the modulation of apoptotic activities in HSCs.

rhTPO Ameliorates Radiation-Induced Long-Term Hematopoietic Stem Cell Injury in Mice.

Exposure to medium and high doses of ionizing radiation (IR) can induce long-term bone marrow (BM) suppression. We previously showed that recombinant human thrombopoietin (rhTPO) significantly promotes recovery from hematopoietic-acute radiation syndrome, but its effect on long-term BM suppression remains unknown. C57BL/6 mice were exposed to 6.5 Gy γ-rays of total body irradiation (TBI) at a dose-rate of 63.01 cGy per minute, and the mice were treated with rhTPO (100 µg; intramuscular injection) or vehicle at 2 h after TBI. All mice were killed one or two months after TBI for analysis of peripheral blood cell counts, long-term hematopoietic stem cell (HSC) frequency, and BM-derived clonogenic activity. The HSC self-renewal capacity was analyzed by BM transplantation. The levels of reactive oxygen species (ROS) production and ratios of γH2AX+ and p16, p53, and p21 mRNA in HSCs were measured by flow cytometry and real-time polymerase chain reaction, respectively. Treatment with rhTPO reduced long-term myelosuppression by improving long-term hematopoietic reconstitution (p < 0.05) after transplantation and resting state maintenance of HSCs (p < 0.05). Moreover, rhTPO treatment was associated with a sustained reduction in long-term ROS production, reduction of long-term DNA damage, diminished p53/p21 mRNA expression, and prevention of senescence after TBI. This study suggests rhTPO is an effective agent for treating IR-induced long-term BM injury because it regulates hematopoietic remodeling and HSC cycle disorder through the ROS/p53/p21/p16 pathway long term after IR.

[Effect of Recombinant Human Thrombopoietin (rhTPO) on Long-term Hematopoietic Recovery in Mice with Acute Radiation Sickness and Relative Mechanism].

OBJECTIVE: To investigate the effect and relative mechanism of Recombinant Human Thrombopoietin (rhTPO) on long-term hematopoietic recovery in mice with acute radiation sickness. METHODS: Mice were intramuscularly injected with rhTPO (100 µg/kg) 2 hours after total body irradiation with 60Co γ-rays (6.5 Gy). Moreover, six months after irradiation, peripheral blood, hematopoietic stem cells (HSC) ratio, competitive transplantation survival rate and chimerization rate, senescence rate of c-kit+ HSC, and p16 and p38 mRNA expression of c-kit+ HSC were detected. RESULTS: Six months after 6.5 Gy γ-ray irradiation, there were no differences in peripheral blood white blood cells, red blood cells, platelets, neutrophils and bone marrow nucleated cells in normal group, irradiated group and rhTPO group (P>0.05). The proportion of hematopoietic stem cells and multipotent progenitor cells in mice of irradiated group was significantly decreased after irradiation (P<0.05), but there was no significant changes in rhTPO group (P>0.05). The counts of CFU-MK and BFU-E in irradiated group were significantly lower than that in normal group, and rhTPO group was higher than that of the irradiated group(P<0.05). The 70 day survival rate of recipient mice in normal group and rhTPO group was 100%, and all mice died in irradiation group. The senescence positive rates of c-kit+ HSC in normal group, irradiation group and rhTPO group were 6.11%, 9.54% and 6.01%, respectively (P<0.01). Compared with the normal group, the p16 and p38 mRNA expression of c-kit+ HSC in the irradiated mice were significantly increased (P<0.01), and it was markedly decreased after rhTPO administration (P<0.01). CONCLUSION: The hematopoietic function of mice is still decreased 6 months after 6.5 Gy γ-ray irradiation, suggesting that there may be long-term damage. High-dose administration of rhTPO in the treatment of acute radiation sickness can reduce the senescence of HSC through p38-p16 pathway and improve the long-term damage of hematopoietic function in mice with acute radiation sickness.

[Therapeutic Effect of Single Intramuscular Administration of Recombinant Human Thrombopoietin on Rhesus Monkeys with Acute radiation Sickness].

OBJECTIVE: To confirm the therapeutic effect of recombinant human thrombopoietin (rhTPO) on rhesus monkeys irradiated with 5.0 Gy 60Co γ-ray, and provide experimental basis for clinical treatment of similar patients. METHODS: Fourteen adult rhesus monkeys were irradiated with 60Co γ-ray on both sides at the dose of 5.0 Gy (dose rate 69.2 cGy/min) to establish the acute radiation sickness model. The monkeys were divided into irradiation group (n=5), rhTPO 5 µg/kg group (n=4) and rhTPO 10 µg/kg group (n=5). Two hours after irradiation, the three groups of monkeys were injected with saline 0.1 ml/kg, rhTPO 5 µg/kg（0.1 ml/kg） and rhTPO 10 µg/kg(0.2 ml/kg), respectively. The general signs, survival, peripheral hemogram and serum biochemistry of rhesus monkeys were observed before and after irradiation, and the differences between rhTPO group and irradiation control group were compared. RESULTS: After total body irradiation with 5.0 Gy60Co γ-ray, rhesus monkeys successively showed fever, hemorrhage, sharp decrease of whole blood cell counts in peripheral blood and disorder of serum biochemical indexes. Compared with the irradiated control group, a single intramuscular injection of rhTPO 5 µg/kg or 10 µg/kg 2 hours after irradiation could improve the symptoms of fever and bleeding, increase the nadir of peripheral red blood cells and platelets counts, shorten the duration of hemocytopenia, and advance the time for blood cells to return to the pre-irradiation level. The serum biochemical results showed that rhTPO could improve the abnormality of serum biochemical indexes in rhesus monkeys induced by 5.0 Gy total body irradiation to some extent. Compared with the two administration groups, the therapeutic effect of rhTPO 10 µg/ kg was better. CONCLUSION: A single injection of rhTPO 5 µg/ kg or 10 µg/ kg 2 hours after irradiation can alleviate the injury of multilineage hematopoiesis and promote the recovery in monkeys irradiated by 5.0 Gy γ-ray. It also improves animal signs and has obvious therapeutic effect on acute radiation sickness.

[Effect of Interleukin-6 Gene Deletion on Radiation-Induced Mouse Hematopoietic Injury and Relative Mechanism].

OBJECTIVE: To study the effect of interleukin-6 (IL-6) gene deletion on radiation-induced hematopoietic injury in mice and relative mechanism. METHODS: Before and after whole body 60Co γ-ray irradiation, it was analyzed and compared that the difference of peripheral hemogram, bone marrow hematopoietic stem and progenitor cells conts in IL-6 gene knockout (IL-6-/-) and wild-type (IL-6+/+) mice and serum IL-6 and G-CSF expression levels in above- mentioned mouse were detected. Moreover, 30 days survival rate of IL-6-/- and IL-6+/+ mice after 8.0 Gy γ-ray irradiation were analyzed. RESULTS: IL-6 levels in serum of IL-6+/+ and IL-6-/- mice were respectively (98.95±3.85) pg/ml and (18.36±5.61) pg/ml, which showed a significant statistical differences (P<0.001). There were no significant differences of peripheral blood cell counts and G-CSF level in serum between IL-6+/+ and IL-6-/- mice before irradiation (P>0.05). However, the number of leukocytes, neutrophils, lymphocytes, monocytes, platelets in peripheral blood and G-CSF level in serum of IL-6-/- mice were significantly decreased at 6 h after 8.0 Gy γ-ray irradiation compared with that of IL-6+/+ mice. On days 30 after 8.0 Gy γ-ray irradiation, the survival rate of IL-6+/+ and IL-6-/- mice was 62.5% and 12.5%, and the mean survival time of dead mice was 16.0±1.0 and 10.6±5.3 days, respectively. On days 14 after 6.5 Gy γ-ray irradiation, bone marrow nucleated cells in IL-6+/+ and IL-6-/- mice were respectively (10.0±1.2)×106 and (8.3±2.2)×106 per femur. Compared with IL-6+/+ mice, the proportion of Lin-Sca-1-c-kit+ (LK) in bone marrow of IL-6-/- mice had no significant change (P>0.05), but the proportion of Lin-Sca-1+c-kit+ (LSK) was significantly decreased (P<0.05). CONCLUSION: IL-6 plays an obvious role in regulating hematopoietic radiation injury, and IL-6 deficiency can inhibit the radiation-induced increase of endogenous G-CSF level in serum, aggravates the damage of mouse hematopoietic stem cells（HSC） and the reduction of mature blood cells in peripheral blood caused by ionizing irradiation, resulting in the shortening of the survival time and significant decrease of the survival rate of mice exposed to lethal dose radiation.