TPO plus Tanshinone IIA has neural protective effect and this effect may be mediated via its receptor c-mpl and PI3K/Akt signaling

Thrombopoietin (TPO) is a growth factor for platelet/megakaryocytic lineage. Our previous study showed TPO has cardiac protective effect(Li et al, Circulation, 2006). TPO may also has a protective effect in neural cells. Tanshinone IIA (TIIA), isolated from Danshen (Radix Salviae Miltiorrhiza Bge), is a derivative of phenanthrenequinone, which has been widely used as anti-platelet and neural protection drug for stroke for many years in China. In this study, a hypoxic–ischemic encephalopathy model was established, followed by 2 hrs of hypoxia in 7-day-old neonatal rat pups. TPO (12.5 ug/kg) or/and TIIA (10 mg/kg) were administrated daily (IP) for two weeks. The severity of brain injury was estimated by the weight reduction of the ipsilateral cerebral hemisphere as compared to the contralateral hemisphere. There was significant less atrophy in TPO alone or TPO plus TIIA treated animals compared with that in controls at 7 and 21 days (P<0.05, n=8). The loss of neurons in the forelimb area of the cortex was also reduced after TPO or TPO plus TIIA treatment. NSE positive cells in the forelimb area of the cortex in the right hemisphere was significantly higher in the treatment group than that of the saline group (P<0.05, n=8). An improvement in sensory motor functions was also demonstrated after TPO or TPO plus TIIA treatment. Furthermore, TPO receptor (c-mpl) mRNA was identified in human cerebral hemispheres and cerebellum, and C17.2 cell line using RT-PCR. The expression of c-mpl protein was further confirmed in human cerebral hemispheres, hippocampus, cerebellum, brainstem and spinal cord using immunocytochemical staining. Its ligang TPO was also detected in human cerebrospinal fluids (n=10) by ELISA, and TPO mRNA was expressed in human cerebral hemispheres, cerebellum, and mouse neural stem cell line C17.2 by RT-PCR methods. TPO showed an anti-apoptotic effect on C17.2 cells by the AnnexinV/PI assay. Moreover, TPO activated PI3K/Akt signaling pathway which was demonstrated by Western blot. The Akt activation by TPO was inhibited by the PI3-kinase inhibitor LY294002. This study provided the evidences showing that TPO or TPO plus TIIA has neural protective effect and this effect may be mediated via its receptor c-mpl, then activated PI3K/Akt signaling.