| Spatholobus suberectus Dunn(SSD) is a traditional Chinese medicine for blood circulation. It has activating-hematopoiesis, antitumor, antiviral and immune regulation, antioxidant, sedation, hypnosis and other pharmacological effects. The chemical structure types of compounds in SSD are mainly flavonoids, terpenoids, sterols, anthraquinone, lactones, glycosides and other types. The previous studies show that catechin is the main active monomer of SSD’s activating-hematopoiesis effect.Radiation can cause damage to the body’s blood, immunity, digestion, reproduction and other systems, involving multiple organs. At present,it is believed that the mechanism for radiation injury is that radiation can directly lead to a variety of DNA damage, gene mutation and chromosome aberration; also, it can make the body produces a large number of oxygen free radicals (ROS) which is of high activity and low chemical stability, causing lipid peroxidation and organ injury. A large number of hematopoietic stem cells apoptosis and death caused by radiation is the main reason for the decrease of peripheral blood. Therefore, reducing the damage of oxygen free radicals to the body, promoting the improvement of the body’s hematopoietic capacity is the key to the treatment of radiation injury. So this paper is aim to study the anti-radiation mechanism of SSD and its active component catechin based on anti-oxidant and activating-hematopoiesis effect.The content of this paper is divided into four parts. Firstly, ICR mice were exposed to 6Gy irradiation, then the number of peripheral blood cells, thymus and spleen index was detected to observe the radioprotective effect of SSD and catechin. Secondly, the serum and liver SOD, GSH-Px activity and ROS, MDA level were detected to study the antioxidant effect and mechanism of SSD and catechin. The hematopoietic promoting effects were detected by observing bone marrow pathological changes and hematopoietic progenitor cell colony forming ability. The hematopoietic promoting mechanism was studied by testing the proliferation of NFS-60 cells and the phosphorylation of JAK-STAT pathway key proteins of bone marrow cells. Lastly, the effect of bone marrow cell apoptosis in mice was detected by flow cytometry. The anti-apoptotic effect and mechanism was studied by detecting the expression of apoptosis related proteins in bone marrow using western-blotting and immunohistochemistry.ICR mice were exposed to 6Gy irradiation and randomly divided into normal group, model group, positive control group (amifostine,43.6mg·kg-1,iv 30min before irradiation), SSD group (10、20、40g·kg-1) and catechin group (50、100、200mg·kg-1). The mice were administered the appropriate drugs once a day after irradiation for 28 consecutive days. Blood samples were collected from the tail end and the number of peripheral blood cells was counted before irradiation and on day 1,3,7,14,21 and 28 using a microcell counter. Changes of thymus and spleen index of mice on day7 were observed. The serum SOD, GSH-Px activity and ROS,MDA level were detected by the colorimetric method. The colony forming ability of bone marrow hematopoietic progenitor cells on day7 was detected by semi solid culture method. The proliferation of G-CSF dependent NFS-60 cells was detected by CCK8 kit and the content of G-CSF in the cells was determined by ELISA kit. The HE staining was adopted to observe the pathological changes. The phosphorylation of JAK-STAT pathway related proteins in bone marrow were detected using western-blotting.The apoptosis of bone marrow cells was detected by flow cytometry. The expression of Cleaved Caspase-3 and Bax of bone marrow cells were measured separately by western-blotting and immunohistochemistry method. SSD and catechin can both significantly revert the irradiated-induced decline in hematological parameters(RBC,WBC,PLT,Hb), improve thymus and spleen index, significantly enhance serum SOD and GSH-Px activity and decrease ROS and MDA level. The proliferation of NFS-60 cells was promoted but the intracellular G-CSF content was not increased. SSD and catechin can activate JAK2-STAT5 signaling pathway by improving the phosphorylation of JAK2, STAT5a and STAT5b. The proliferation and differentiation of hematopoietic progenitor cells in bone marrow were promoted, the apoptosis of bone marrow cells was inhibited and the expression of Cleaved Caspase-3 and Bax was significantly reduced and the expression of Bcl-2 was significantly up-regulated in SSD and catechin group. SSD and catechin have significant anti-radiation effect and its mechanism may be related to hematopoietic promoting, antioxidant and anti-apoptotic effects. |
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