Protective Effect Of Isoflavonoids From Rhizomes Iris Tectorum On Nerve Cells

Estrogen is an essential singaling molecules in nerve system and has impact ondevelopment of nerves, synthesis of neurotransmitters, survival of neurons, regenerationof myelin sheath and axons. Estrogen in plants has similar construction and function to thatin humans. In recent years, the study of estrogen in plants are becoming more and moreimportant. Especially, isoflavonoids, estrogen-like ingredients from Rhizomes Iris tectorum,proved to abolish oxygen free radical and had antioxidase activity. To observe the effect ofisoflavonoids on the injured nerve cells, cultured human neuroblastoma SH-SY5Y cells（SH-SY5Y）, being suffered from oxidative stress, anoxia or glucopenia, was employed toinvestigate the protective mechanisms in vitro.Experiment one Effects of Isoflavonoids on SH-SY5Y cells injured by oxidative stress, anoxia or glucopeniaAimTo investigate the protec effect of five ingredients from Rhizomes Iris tectorum on theSH-SY5Y cells injured by glucopenia and hypoxia-reoxygenMethods1. Cultured SH-SY5Y cells （SH-SY5Y）, suffering from glucopenia andhypoxia-reoxygen, was employed to determine time-effect curve and dose-effect curve byCCK-8method,. According to ED50of cells survival, the manner of glucopenia andhypoxia-reoxygen was set up.2. Several ingredients from rhizomes iris tectorum,such as iristectorigenin,iristectorin A, iristectorin B, tectoridin, and5,7,4′-trihydroxy-6,3′-dimethoxy isoflvonewere utilized to observe the protection on the SH-SY5Y cells injured by glucopenia andhypoxia-reoxygen and H2O2. ResultsThe survival rate of SH-SY5Y cells （SH-SY5Y） injured by H2O2at the dosage of2×10-4mol·L^-1for24hours was58.5%. The survival rate of SH-SY5Y cells （SH-SY5Y）injured by anoxia and glucopenia for4h was52.9%. There were no significant differenceamong survival rates after anoxia-reoxygen for4h. Meanwhile, the survival rate ofSH-SY5Y cells （SH-SY5Y） was56.4%. The protective effects of iristectorin A andiristectorin B are best among them, and the difference is significant when consequenceare compared with which of injured cells by H2O2（P<0.05、P<0.01）；with which of injuredcells by oxygen and glucose deprivation/regain oxygen （P<0.01、P<0.01）, Other singlebody has protective action on injured nurve cells, but the consequence is nosencestatisticsly compared with which of injured cells.Conclusioniristectorin A and iristectorin B in isoflavonoids from Rhizomes Iris tectorum haveconspicuous protective action on nervous cells.Experiment two The study of anti-oxidation mechanism of iristectorin A and iristectorin Bon SH-SY5Y cells.AimTo research the anti-oxidation mechanism of iristectorin A and iristectorin B on theSH-SY5Y cells injured by oxidative stress by hydrogen peroxide/regain oxygen and H2O2.Methods1. Blank group, drug group with puerarin, model group, group injured by oxidativestress by hydrogen peroxide and dosage group with tectoridin (20umol·L^-1,100u mol·L^-1,500μmol·L^-1) are injured by H2O2respectively.2. Blank group, drug group with puerarin, model group and dosage group withtectoridin(20umol·L^-1,100u mol·L^-1,500μmol·L^-1) are injured by oxidative stress byhydrogen peroxide/regain oxygen respectively. We use the LDH activity in extra-cellularmedia to observe the protective effect, use MDA contents and SOD, GSH-Px activities ofSH-SY5Y cells to analyse the oxidation/anti-oxidation index.Results1. In the group injured by H2O2, iristectorin A and iristectorin B can decrease LDHefflux of SH-SY5Y cell, and effectiveness is dependant on the dosage. The consequenceof iristectorin A(100μmol·L^-1,500μmol·L^-1)and iristectorin B (20μmol·L^-1,100μmol·L^-1, 500μmol·L^-1) is sence statisticsly compared with which of model group(P<0.05, P<0.01,P<0.05, P<0.05, P<0.01. Intervention group can ascend the SOD activities. Theconsequence of iristectorin A (100μmol·L^-1,500μmol·L^-1)and iristectorin B (500μmol·L^-1)is sence statisticsly compared with which of model group （P<0.05, P<0.01,P<0.05）.Intervention group can ascend the GSH-Px activities; the difference ofconsequence of iristectorin A (20μmol·L^-1,100μmol·L^-1,500μmol·L^-1)and iristectorin B(20μmol·L^-1,100μmol·L^-1,500μmol·L^-1) compared with which of model group issignificant(iristectorin A at the dosage of20μmol·L^-1, P<0.05; other intervention group,P<0.01). Intervention group can decend the MDA contents, the difference ofconsequence of iristectorin A (100μmol·L^-1,500μmol·L^-1) and iristectorin B (100μmol·L^-1,500μmol·L^-1) compared with which of model group is significant （P<0.05, P<0.01,P<0.05, P<0.01）.2. In the group injured by oxidative stress by hydrogen peroxide/regain oxygen,iristectorin A and iristectorin B can decrease LDH efflux of SH-SY5Y cell, andeffectiveness is dependant on the dosage. The consequence of iristectorin A (100μ mol·L^-1,500μmol·L^-1) and iristectorin B (100μmol·L^-1,500μmol·L^-1) is sence statisticslycompared with which of model group（P<0.05, P<0.01, P<0.01, P<0.01）. Interventiongroup can ascend the SOD activities; The consequence of iristectorin A (100μmol·L^-1,500μmol·L^-1)and iristectorin B (500μmol·L^-1) is sence statisticsly compared with whichof model group（P<0.05, P<0.05, P<0.05）. Intervention group can ascend the GSH-Pxactivities; the difference of consequence of iristectorin A (100μmol·L^-1,500μmol·L^-1) andiristectorin B (20μmol·L^-1,100μmol·L^-1,500μmol·L^-1) compared with which of modelgroup is significant(iristectorin A at the dosage of100μmol·L^-1, iristectorin B at the dosageof20μmol·L^-1,P<0.05; other intervention group, P<0.01). Intervention group can decendthe MDA contents, the difference of consequence of iristectorin A (100μmol·L^-1,500μmol·L^-1)and iristectorin B (20μmol·L^-1,100μmol·L^-1,500μmol·L^-1) compared withwhich of model group is significant （P<0.05, P<0.01, P<0.01, P<0.01, P<0.01）.Conclusioniristectorin A and iristectorin B have conspicuous protective action on cerebralischemia reperfusion nervous cells, and effectiveness is dependant on the dosage. Themechanism has to do with improving oxidation/anti-oxidation disequlibrium possibly.