The effect and underlying mechanism of melation on platelet formation and survival in a thrombocytopenic model

Abstract

Thrombocytopenia is a major treatment-related problem for cancer patients on chemo- or radiotherapy. In clinical studies, melatonin has been shown to be able to lower the frequency of thrombocytopenia in these cancer patients. However, the actual effects and underlying mechanism of melatonin on megakaryocytopoiesis, and platelet formation/survival have not been formally investigated. Our previous studies have demonstrated that serotonin can stimulate hematopoiesis and megakaryocytopoiesis (Yang et al, Stem Cells, 2007). Melatonin is an acetylated product of serotonin. To delineate the effect of melatonin on platelet production, we hypothesize that melatonin has therapeutic effects on thrombocytopenia by promoting megakaryocytopoiesis and enhancing the recovery of platelets via activation of Akt/Erk signaling. The effect of melatonin on thrombopoiesis was investigated in an irradiated mouse model (4 Gy). Eighteen mice were randomly divided into three groups (6 in each). Group 1 (normal control, N) received no irradiation or melatonin; Group 2 (model control, C) and Group 3 (melatonin, M) received 4 Gy total body irradiation. After irradiation, melatonin (10mg/kg.d) was injected intraperitoneally into Group M for consecutively 21 days. Peripheral blood platelets, white blood cells (WBC), and red blood cells (RBC) were analyzed from the three groups on day 0, 7, 14, and 21.Our results showed that melatonin enhanced the recovery of platelets and WBC counts. We also investigated the in-vitro effect of melatonin on CFU-MK formation at different doses (0-500 nM). The results showed that melatonin significantly promoted CFU-MK formation at a dose dependant manner. The maximal concentration was at 200 nM (P<0.01, n=6). The size of CFU-MK with melatonin treatment was much larger and each MK cell was more mature. Melatonin also stimulated the formation of bone marrow CFU-F. Using megakaryocytic cell CHRF-288-11, we further found that melatonin exerted protective effect on serum-free induced apoptosis of CHRF cells. The cell viability was significantly increased with the treatment of melatonin. Flow cytometric dot-plot analysis demonstrated that the population of apoptotic cells (Annexin V/PI staining) was significantly decreased with melatonin treatment (200nmol/L) for 72 hrs (n=4). It also showed trends of amelioration of caspase-3 expression and the proportion of cells containing JC-1 monomers (indicating a trend in the drop of mitochondrial membrane potential). The population of phospho-Akt and Erk1/2 were significantly increased after treatment by melatonin. Our results showed that melatonin had a promoting effect on platelet formation and survival in a thrombocytopenic model. The underlying mechanisms may be involved in directly stimulating megakaryocytopoiesis and having anti-apoptotic effect in megakaryocytes via the activation of Akt/Erk signaling.