| Gliomas are a common intracranial tumor, which occupies46%of intracranial tumors, But the current methods of treatment of glioma are not satisfactory. Boron neutron capture therapy is highly selective destruction of tumor cells and low damage to the surrounding normal cells, which has become theoretically an ideal glioma treatment and has got much concern. Sodium borocaptate (Na2B12H11SH BSH) is commonly used as a boron-carrying agent in BNCT. BSH has good features in gathering the tumor cells, but can not pass through the blood-brain barrier. In recent years, the role and the effect of traditional Chinese medicine in opening blood-brain barrier, have caused much concern, such as the Angong Niuhuang Prescription. One goal of this study is the function of Angong Niuhuang Prescription openning BBB and the promiting the penetration of BSH across BBB, in order to get boron carrying agent to gather the desired concentration in the tumor and make BNCT treatment get more effective. And Mannitol is a drug used clinically for opening blood-brain barrier, the other purpose is to compare the effect of promiting the penetration of BSH across BBB by Angong Niuhuang Prescription and Mannitol.The experiments first examine the BSH distribution in the tissue concentrations of the mice. The BSH dose of176mg·kg-1was given through Tail intravenousness, blood, brain, liver and kidney were taken after lh. The ICP-AES was used to detect the concentration of B in the brains of KM mice after brain tissue was digested and filtered. The results showed that BSH were very low in the brain, it is difficult to directly through the blood-brain barrier. So Gong Niuhuang prescription was used to open up the blood-brain barrier and promit BSH through the blood-brain barrier.Evans blue staining and fluorescence microscopy were used for evaluation of the role of the Angong Niuhuang Prescription on the blood-brain barrier opening.(1) Evans blue stained Cerebral cortex:36Kuming(KM) mices were randomLy divided into0.125g·kg-1·d-1,0.25g·kg-1·d-1,0.50g·kg-1·d-1,0.75g·kg-1·d-1, which were based on the Angong Niuhuang Prescription dose. Control group was given the same volume of paraffin wax oil solution in intragastric administration. Blank groups was raised normally in the same environment. Every group was given medicine at9:00～10:00for four consecutive days. Any group except Blank was given0.05mL E.B through Tail intravenousness after30minites for the last time, And observe and compare their brain dyed blue in1hour.(2) Fluorescence microscopy:20KM mice were randomLy divided2groups. The experimental group was given medicine in0.25g·kg-1·d-1, and the control group was given the same volume of paraffin oil solution. Every group mices was given medicine at9:00～10:00for four consecutive days. Any group were given0.05mL E.B through Tail intravenousness after30minites for the last time. Then mice were killed after1h, and the brain tissues were removed. They were quickly made frozen sections, which were observed under fluorescence microscopy and the number of spot were counted.The experiment of Angong Niuhuang prescription promote BSH through BBB is divided into two parts of the dose relationship effect experiments and the time-effect relationship experiment. The trial of dose-effect relationship for the dynamic quantitative detection for promiting BSH across BBB:36KM mices were randomLy divided into0.125g·kg-1·d-1,0.25g·kg-1·d-1,0.50g·kg-1·d-1,0.75g·kg-1·d-1which were based on the Angong Niuhuang Prescription dose. Control group was given the same volume of paraffin wax oil solution in intragastric administration, Blank group was raised normally in the same environment. Every group was given medicine at9:00-10:00for four consecutive days. Any group except Blank was given BSH through Tail intravenousness after30minites for the last time, blood and brian were taken after1hour, and concentration of B in ICP-AES was detected. The time-effect relationship experiment:the experimental group was given medicine in0.25g·kg-1·d-1, the control group was given the same volume of paraffin oil solution, for four consecutive days.0.2mL BSH was given in Tail intravenous injection after30minites for the last time. brains of KM mice were taken at hour1,3,6,12h after the administration of BSH. The ICP-AES was used to detect the concentration of B in the brains of KM mice after brain tissue was digested and filtered.Compare the effect of promiting the penetration of BSH across BBB by Angong Niuhuang Prescription and Mannitol:totally72KM mice were randomLy divided into3groups. Angong Niuhuang Prescription group was given medicine in0.25g·kg-1·d-1in intragastric administration, and for four consecutive days; Mannitol group was given0.1mL in Tail intravenous injection for four consecutive days. Two groups were given BSH0.2mL in Tail intravenous injection after30minites for the last time. Control group was given the same volume of paraffin wax oil solution in intragastric administration for four days, and given BSH0.2mL in Tail intravenous injection after30minites for the last time. brains of each group were taken at hour1,3,6,12after the administration of BSH. The ICP-AES was used to detect the concentration of B in the brains of KM mice after brain tissue was digested and filtered.Found in the role of the BBB opening:(1) mice that were given a dose of0.75g·kg-1·d-1have serious toxic reactions, those with a dose of 0.50g·kg-1·d-1have slight toxic reactions.(2) The general concept of brain tissue staining:the color of mice with a dose of0.25g·kg-1·d-1and0.50g·kg-1·d-1group was deeper than that of those of0.125g·kg-1·d-1and0.75g·kg-1·d-1groups at the degree of blue staining, cerebral cortex of control group is also a touch of blue dye. It is significantly different at the cerebral cortex for four dosing groups compared with control group of blue dye from visual observation.(3) Fluorescence microscopy:A large number of oval erythema can be seen in experimental group, compared with control group having only occasionally erythema. Spot Mean of the experimental group and control group were (75.65±7.58),(28.55±3.67). There are significant differences (P<0.05).Experimental results on Angong Niuhuang prescription promoting BSH through BBB show that Dose-effect relationship experiment:10B concentrations of0.125g·kg-1·d-1,0.25g·kg-1·d-1,0.50g·kg-1·d-1,0.75g·kg-1·d-1, Blank, Controlgroups in brain were:(7.28±0.25),(9.67±0.41),(8.17±0.20),(7.42±0.32),(3.61±0.14),(3.75±0.37)μg·g-1. There are significant differences among0.25g·kg-1·d-1,0.50g·kg-1·d-1groups and control group(P<0.05). The time-effect relationship experiment: At hour1,3,6,12, the Boron concentrations of Angong Niuhuang Prescription group were (9.67±0.41),(7.96±0.28),(7.32±0.22),(6.68±0.17)μg·g-1. There are significant differences among the groups at any time(P<0.05); Angong Niuhuang Prescription group maintain stable concentrations of drug in the brain of mice.Comparison of the effect of promiting the penetration of BSH across BBB by Angong Niuhuang Prescription and Mannitol:at hour1,3,6,12the Boron concentrations of Angong Niuhuang Prescription group were (9.67±0.41),(7.96±0.28),(7.32±0.22),(6.68±0.17)μg·g-1; those of mannitol group were (7.45±0.3),(5.44±0.46),(2.55±0.33),(1.10±0.43)μg·g-1. There are significant differences between the Mannitol and control groups(p<0.05); there are significant differences between the Mannitol and Angong Niuhuang prescription groups after3h(p<0.05).Fond by the above methods of research:1. Angong Niuhuang prescription may open the blood-brain barrier, and significantly improve the BSH through the blood-brain barrier.And the dose of0.25g·kg-1·d-1is relatively safe and effective.2. Angong Niuhuang Prescription can maintain a higher concentration and a longer time in the brain, make more lasting curative effect.3. Two methods could promote the penetration of BSH across blood brain barrier. Compared with Mannitol, Angong Niuhuang Prescription can maintain high and stable concentrations of drug in the brain of mice, which is better than the effect of the Mannitol. |
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