Effects Of Volatile Oil Of Frankincense On Proliferation And Apoptosis Of Rabbit Retinal Pigment Epithelial Cells And Its Molecular Mechanism

Objective Observed the effects of the volatile oil of Frankincense on the proliferation and apoptosis of retinal pigment epithelial cells in rabbits,and explored its possible molecular mechanism,so as to provide new research and development ideas for preventing and controlling PVR.Methods The effects of different concentrations of DMSO on the proliferation of rabbit RPE cells were detected by MTT assay and the concentration of DMSO in the required medium was screened.The effects of different concentrations of 5-Fu(1mg/L,10mg/L,30mg/L,50mg/L)on the proliferation of rabbit RPE cells were detected by MTT assay.Statistical analysis screened low concentration,medium concentration and high concentration of 5-Fu group were intervention of rabbit RPE cells,the use of Flow cytometry(FCM)to detect the impact of 5-Fu on its apoptosis rate,screening out the optimal concentration of the positive control drug.The water vapor reflux method was used to extract the Frankincense essential oil.Rabbit RPE cells were treated with different concentrations of frankincense volatile oil(0.15 mg/mL,0.30 mg/m L,0.45 mg/m L,0.60 mg/m L,0.75 mg/mL)for 24 h,48h and 72 h.The effect of its on OD and survival rate of rabbit RPE cells was detected by MTT assay.Statistical analysis screened low-dose,middle-dose,and high-dose Frankincense volatile oil groups.FCM method was used to detect the apoptosis rate of RPE cells in normal,vehicle and positive drug control groups and low,middle and high doses of frankincense volatile oil.Transmission electron microscopy was used to observe the effect of Frankincense volatile oil on the morphology of rabbit RPE cells.The protein expression of Bax and Bcl-2 and the ratio of Bcl-2 / Bax in RPE cells were detected by Western Blot.Results1.MTT assay of different concentrations of DMSO on rabbit RPE cell proliferation In the same time: except 0.2% DMSO intervention group,the other concentrations of DMSO group compared with the normal control group,rabbit RPE cell survival rate decreased to varying degrees,the difference was statistically significant(P <0.01).In the 0.2% DMSO intervention group,the survival rate of RPE cells in rabbits showed no significant change over time(P> 0.05).2.Effect of 5-Fu on survival rate and apoptosis rate of rabbit RPE cells by MTT assay and FCM assay Compared with the normal group,the OD value and survival rate of 5-Fu group at different concentrations decreased to some extent(P <0.01).The apoptosis rate of high-concentration 5-Fu group was significantly higher than that of other groups,the difference was statistically significant(P <0.01).3.The effect of volatile oil from frankincense on the OD and survival rate of rabbit RPE cells was detected by MTT assay Except 0.15mg/m L volatile oil from frankincense group,in other volatile oil group,the cell OD value and survival rate decreased to some extent,the difference was statistically significant(P <0.01),and with the passage of time,the cell survival rate showed a gradual decrease.4.Detection of RPE cells in rabbits by FCM and TEM At the same time,compared with the control group,apoptotic bodies and apoptotic cells were significantly increased in the positive drug control group and RPE cells in the middle and high dose groups of Frankincense volatile oil(P<0.01).The apoptotic rate increased with time(P <0.05,P <0.01).5.Western blot assay was used to detect the Bax and Bcl-2 protein expression and the ratio of Bcl-2 / Bax in rabbit RPE cells In the meantime,the relative expression of Bcl-2 / GAPDH in middle dose,high dose volatile oil from frankincense group and positive control group was significantly lower than that in normal group(P <0.01),the relative expression of Bax / GAPDH was significantly higher than that of the normal group(P <0.01),with the increase of volatile oil from frankincense dose,the relative expression of Bcl-2 protein and Bax protein decreased and increased accordingly.Compared with the control group,the positive control drug,middle dose group and high dose group volatile oil from frankincense can significantly reduce the ratio of Bcl-2 / Bax cells,that apoptosis was significantly increased(P <0.01).And the overall observation of cell apoptosis has a tendency of increasing gradually with the prolongation of the action time and the dose increase.Conclusions1.When the concentration of DMSO is less than 0.2% of the total volume,it is the concentration of the vehicle in this experiment.2.When the concentration of 5-Fu was 30 mg/L,it was the best positive drug control concentration for this experiment.3.The volatile oil of Frankincense can inhibit the proliferation of rabbit RPE cells and induce apoptosis of rabbit RPE cells by up-regulating mitochondrial signaling pathway Bax protein and down-regulating Bcl-2 protein expression.The degree of apoptosis is related to dose and time.