| Objectives:Atherosclerosis is chronic inflammation on vascular wall, which is a reaction of damaging and course of repair on vascular wall. P38MAPK signal pathway is a key of controlling chronic inflammation in atherosclerosis. Interdicting P38MAPK can relieve inflammation. Our research studied DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi effect on P38MAPK activity induced by Ox-LDL in RAW264. 7 cell, in order to discuss the cell signal transduction mechanism of DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi against atherosclerosis.Methods:1. Prepared DangGui BuXue Decotion herbs and extraction of DangGui HuangQi, permuting and combining the extraction of DangGui HuangQi, prepared rabbit serum containing DangGui BuXue Decotion herbs and extraction of DangGui HuangQi after grouping.2. Prepared oxidized low-density lipoprotein.3. Cell culture; Cell activity detected by trypan blue staining method; Detected cell growth curvature and cell survival rate affected by rabbit serum through MTT method.4. Detected DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi effect on P38MAPK activity induced by Ox-LDL in RAW264. 7 cell P38MAPK and phosphor- P38MAPK protein activity change by western bolt method.Results:1. Used serous pharmacology method to prepare rabbit serum containing DangGui BuXue Decotion herbs and extraction of DangGui HuangQi.2. Recovered preservative cell and cultured cell. Cell activity detected by trypan blue staining was more than 95%. According to the cell growth curvature, chose the cell of the forth day after transferring of culture when cell grew well. Rabbit serum had nutrient action to facilitate cells' growth, but too highly concentration was bad to cells. 10% concentration had small effect on cells.3. Western blotting result:P38MAPK western blotting result : Ox-LDL group protein density area ratio is 5.3167±0.16169, compared with control group, the difference had meaning(P<0.05), which implied after Ox-LDL stimulates RAW264. 7 cell , P38MAPK protein activity increases obviously. SB203580 group compared with control group the difference had meaning, which implied SB203580 could inhibit P38MAPK protein activity. DangGui BuXue Decotion group and combination of fractions extracted from DangGui HuangQi group, compared with control group, the difference had meaning. Implied DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi could inhibit P38MAPK protein activity; Compared with Ox-LDL , the difference had meaning. Implied DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi group A and group B could could downregulate P38MAPK protein activity. Compared with SB203580, implied SB203580 was more stronger than DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi on downregulating P38MAPK protein activity. DangGui BuXue Decotion compared with combination of fractions extracted from DangGui HuangQi had no meaning, implied there was no difference between DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi in P38MAPK protein activity.Phosphor-P38MAPK western blotting result: Ox-LDL group protein density area ratio is 5. 3767±0. 36638, compared with control group, the difference had meaning(P<0.05), which implied after Ox-LDL stimulate RAW264.7 cell , Phosphor-P38MAPK protein activity increases obviously. SB203580 group(1.7233±0.11015) compared with Ox-LDL group, difference was obvious, which implied SB203580 could inhibit phosphor-P38MAPK protein activity. DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi group A and group B compared with control group, the difference had meaning. Implied DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi could inhibit phosphor-P38MAPK protein activity; For DangGui BuXue Decotion group(2.8567±0.34559) and combination of fractions extracted from DangGui HuangQi group A and group B compared with Ox-LDL , the difference had meaning. Implied DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi group A and group B could could downregulate phosphor-P38MAPK protein activity. Compared with SB203580, implied SB203580 was more stronger than DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi on downregulating phosphor-P38MAPK protein activity. DangGui BuXue Decotion compared with combination of fractions extracted from DangGui HuangQi the difference had meaning(P<0.05). Implied DangGui BuXue Decotion and extraction of DangGui HuangQi could downregulate phosphor-P38MAPK protein activity. The action of DangGui BuXue Decotion was better than extraction of DangGui HuangQi. Group A compared with Group B combination of fractions extracted from DangGui HuangQi, had no meaning, implied there was no difference between Group A compared with Group B combination of fractions extracted from DangGui HuangQi in phosphor-P38MAPK protein activity.Conclusions:100μg/ml Ox-LDL stimulated RAW264. 7cell for 15 minutes, P38MAPK signal pathway was activated obviously, P38MAPK protein and phosphor-P38MAPK protein increased greatly. SB203580 could inhibit P38MAPK protein and phosphor-P38MAPK protein activity obviously. DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi could downregulate phosphor-P38MAPK protein activity. The action of DangGui BuXue Decotion was better than extraction of DangGui HuangQi. There was no difference in downregulating phosphor-P38MAPK protein activity between group A and group B of extraction of DangGui HuangQi. Our research confirmed DangGui BuXue Decotion could downregulate phosphor-P38MAPK protein activity and relieve inflammation through blocking P38MAPK signal pathway. So DangGui BuXue Decotion and combination of fractions extracted from DangGui HuangQi could downregulate phosphor-P38MAPK protein activity to control related inflammation factor's expression and through this pathway to affect on atherosclerosis development. To block P38MAPK signal pathway may be one of mechanism of DangGui BuXue Decotion and extraction of DangGui HuangQi's action against atherosclerosis injury. |
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