Cyclovirobuxine D （CVB-D） is a compound extracted from Chinese traditional plant Buxus microphylla, which has been used for treating arrhythmia and myocardial ischemia in China. In this study, we investigated its effect on blood coagulation and thrombotic formation in mouse and rat models. The doses of CVB-D used in this study （5-20 mg/kg） prolonged clotting time （CT） in a dose-dependent manner （P〈0.01）. It also significantly prolonged thrombin time （TT）, prothrombin time （PT） and activated partial thromboplast time （aPTT） （P〈0.05 or P〈0.01） at the doses of 10-20 mg/kg. CVB-D did not affect the bleeding time （BT） compared with the control group, while warfarin significantly prolonged the bleeding time. CVB-D at the doses of 5-20 mg/kg reduced wet weight of thrombosis （P〈0.01）. This study demonstrated the anti-coagulation effect and anti-thrombosis effect of orally administered CVB-D without substantially increasing bleeding. These findings suggest that CVB-D probably can be used as an oral anti-coagulant in addition to its current applications.
Methoxyl methyl ether isoamylene quercetin （MIAQ） is one of the newly synthesized quercetin derivatives. The present study investigated the effect of MIAQ on rat aorta endothelial cells （RAECs） injured by hydrogen peroxide （H2O2）, as well as the potential mechanisms. We observed that MIAQ at 2.5-10μmol/L significantly enhanced the viability of injured RAECs, and the effect was more potent than quercetin and ct-tocopherol. However, M1AQ at the same concentration failed to show anti-oxidant activity in a cell-free system. In H2O2-injured endothelial cells treated with MIAQ （5-10μmol/L）, the level of nitric oxide （NO） and malondialdehyde was decreased, and the activities of superoxide dismutase and glutathione peroxidase was enhanced. In addition, RAECs treated with MIAQ （2.5-10 μmol/L） exhibited significant inhibiting apoptosis. In conclusion, MIAQ had protective effect on RAECs, possibly through increasing NO production and antioxidases activities, as well as inhibiting apoptosis. These findings suggest that MIAQ is possibly beneficial in the prevention of atherosclerosis and other diseases related to endothelial injury.
Recombinant ancylostoma caninum anticoagulant peptide-5 （rAcAP5） has been reported to inhibit thrombin-activatable fibrinolysis inhibitor （TAFIa） activity and have thrombolytic effect. The present study was to screen a strain expressing high-yield of rAcAP5 and to assess its thrombolytic effect on embolic middle cerebral artery occlusion （MCAO） model in rats. Codons encoding for AcAP5 were optimized. Six expression plasmids and eleven E. coli strains with different characteristics were used, a total of 66 recombinant expression strains were generated and the one with the highest yield was selected to express rAcAP5, which was purified through anion- and cation-exchange chromatography. The purity of rAcAP5 and its molecular weight were determined by HPLC and mass spectrometry, respectively. The thrombolytic effect of rAcAP5 was evaluated on embolic MCAO model in rats; regional cerebral blood flow （rCBF） was monitored with a Laser-Doppler flowmetry to test the occlusion and recanalization of MCA. The highest yield recombinant strain was C2566H/pTYB 1-rAcAP5. AcAP5 （28 mg） with 90% of purity was obtained from 1 L of cell culture. In rat embolic MCAO model, vehicle （normal saline） treatment did not change the rCBF, while treatment with rAcAP5 （50-200 μg/kg, i.v.） increased the rCBF in a dose-dependent manner. In conclusion, we prepared and characterized the rAcAP5 peptide and revealed its thrombolytic effect in embolic MCAO model and our results suggested that this peptide had the potential to be used as a thrombolytic agent.
With increasing knowledge of the molecular mechanisms of endogenous RNA interference,systemic delivery of small interfering RNA（siRNA） via targeted nanoparticles has emerged as a potential strategy for cancer gene therapy.In this study,a novel formulation[liposome-protamine-chondroitin sulfate nanoparticles（LPC-NP）]was developed for siRNA delivery by self-assembling with charge-charge interaction.The LPC-NP was further modified by DSPE-PEG_（2000） and DSPE-PEG_（2000）-T7 by the post-insertion method.T7,a transferrin-like seven-amino acid peptide,is a targeting ligand for transferrin receptor-overexpressed MCF-7 breast cancer cells.The particle size and zeta potential of LPC-NP were approximately 90 nm and ＋35 mV,respectively. It was shown that PEG modification could significantly decrease aggregation of LPC-NP in serum,and T7 peptide modified LPC-NP could significantly increase the cellular uptake and the gene-silencing effect of siRNA.In vitro cytotoxicity assay exhibited that significant cell growth inhibition was achieved in MCF-7 cells after the delivery of anti-EGFR siRNA.Our encouraging results suggested that T7-modified LPC-NP might be a promising carrier for RNAi-based tumor therapy.
