Experimental Study On The Protective Effect Of Jianpitongluoyin-containing Serum On Neuronal Hypoxia Re-injury

Purpose:To observe the protective effect of Jianpi Tongluo Yin drug-containing serum on human neuroblastoma cells(SH-SY5Y)after anoxic injury,and to explore the effect of Jianpi Tongluo Yin drug-containing serum on the expression of B-cell lymphoma(Bcl-2),Bcl-2-associated X-protein(Bax)and caspase-3(Caspase-3).Analysis of Jianpi Tongluo Yin drug-containing serum can promote the expression of Bcl-2 protein,inhibit the expression of Bax and Caspase-3protein,promote the proliferation and differentiation of SH-SY5 Y cells,thus play a neuroprotective effect,and provide a theoretical basis for clinical treatment of stroke.Material and method:After one week of adaptive feeding,30 healthy male Wistar rats were fed intragastrically,and the drug-containing serum of Jianpi Tongluo Yin and Ginkgo biloba leaves were prepared.SH-SY5 Y cells were cultured in DEME medium and subcultured.When the cell density reached more than 90%.The rats were randomly divided into 6 groups:(C)in normal control group,(IR)in hypoxia-reoxygenation model group,(JL)in low concentration group of Jianpi Tongluo Yin drug-containing serum,(JM)in middle concentration group of Jianpi Tongluo Yin drug-containing serum,(JH)in high concentration group of Jianpi Tongluo Yin drug-containing serum,(GC)in positive control group of Ginkgo biloba leaves.According to the results of MTT detection,the low,medium and high concentrations of Jianpi Tongluoyin drug-containing serum were selected.After SH-SY5 Y cells were cultured for 24 hours,the number of neuronal apoptosis was detected by Hoechst staining,the apoptosis rate of nerve cells was detected by flow cytometry,the contents of Bcl-2,Bax and Caspase-3 protein were detected by Westernblot method,and the data were processed by SPSS2 6.0statistical software.Results:1.Effect of Jianpi Tongluo Yin drug-containing serum on the proliferation rate of SH-SY5 Y cells: after 24 hours of culture,the OD values of different volu-me percentages of Jianpi Tongluo Yin drug-containing serum were s ignifi-cantly different from those of the blank control group(P < 0.01).Finally,the percentage of 10%,15% and 20% drug-containing serum was taken as the conce-tration of drug-containing serum in the low,middle and high concentration group of Jianpi Tongluo Yin drug-containing serum.2.The results of Hoechst staining showed that the number of SH-SY5 Y apoptosis in the group treated with middle and high concentration of Jianpi-Tongluo Yin drug-containing serum was significantly lower than that in the model group treated with hypoxia-reoxygenation injury.3.The results of flow cytometry showed that compared with the hypoxia-reoxygenation model group,the apoptosis rate decreased in the middle and high concentration of Jianpi Tongluo Yin drug-containing ser um and the positive control group of ginkgo biloba leaves,and the difference was statistically significant(P < 0.05).4.The Westernblot results showed that under the intervention of Jianpi Tongluoyin drug-containing serum,compared with the normal contr ol group,the expression of Bax protein increased,the ratio of Bcl-2/Bax decreased signifi can-tly,and the number of Caspase-3 protein increased in the hypoxia reoxygenation model group.Compared with the hypoxia-reoxygenation model group,the expres-sion of Bcl-2 protein increased and Bax decreased significantly in the middle and high concentration group of Jianpi Tongluoyin drug-containing serum.The expres-sion of Caspase-3 protein decreased.Conclusion:1.The drug-containing serum of Jianpi Tongluo Yin has the effect of proliferation on SH-SY5 Y injured cells induced by hypoxia and reoxygenation,and when the volume fraction of drug-containing serum is between 0-20%,the cell proliferation rate increases sequentially.2.Its protective mechanism was related to i ts down-regulation of pro-apoptotic protein and increased expression of anti-apoptotic protein.