Protective effect and its mechanism of Naotaifang on rat cortical neurons injury induced by hemoglobin.

Objective To investigate the protective effect and its mechanism of Naotaifang( Brain Blood-activating Formula, NTF)on rat cortical neurons injury induced by hemoglobin( Hb) in vitro. Methods Primary cortical neurons from newborn SD rats were cultured and induced by Hb( 10micmol/L) to simulate iron overload of neurons after ICH. The cultured neurons were randomly divided into the normal group, model group, blank serum group, 10% NTF gmedicated serum roup and deferoxamine ( DFX) group. Neuron viability was detected by CCK8 assay. The content of iron in neurons was detected by Calcein-AM staining. The protein expressions of transferring ( Tf), transferrin receptor (TfR) and divalent metal ion transporter DMT1 ) in neurons were detected by immunofluorescent staining and high content analysis (HCA). The content of lipid ROS was detected by using ELISA assay. The expressions of mRNA,Tf, TfR and DMT1 were detected with real-time PCR. Results Compared with the normal group, in the model group the neuron viability was markedly decreased(P <0.01), the iron and lipid ROS levels were significantly increased (P <0.01), and the expression of mRNA,Tf, TfR and DMT-1 were srgnificantly up-regulated(P <0.01). Compared with model group,in DFX and NTF groups, the neuron viability was srgnificantly increased (P <0.01 orP<0. 05); the levels of iron and lipid ROS were significantly decreased(P <0. 01) ; moreover, the expression levels of Tf, TfR, DMT-1 and mRNA were markedly down-regulated(P <0. 01 or P <0. 05) . No significant difference was observed between the DFX group and 10% NTF group (P >0. 05). Conclusion NTF might alleviate the iron overload and lipid ROS accumulation of Hb by regulating the iron metabolism in induced rat cortical neurons injury, so as to improve the activity of neurons. Its effect seems to be consistent with that of ferroptosis inhibitor DFX, suggesting that NTF could possibly exert neuroprotective effect by inhibiting ferroptosis of neurons after ICH. [ABSTRACT FROM AUTHOR]