Rhubarb inhibits respiratory syncytial virus by regulating the miR-509-3p/Rab5c axis via NRAV
Objective To evaluate the inhibitory effect of Rhubarb Mianjian Granules on respiratory syncytial virus(RSV)in vitro,and to explore the influence of Rhubarb on the key links in the process of RSV infection and the regulation of intracellular signaling pathways.Methods A549 cells were cultured in vitro,and treated with blank control or RSV infection.At-2h,0h,2h,4h,6h and 8h of RSV infection,A549 cells were induced with high-dose Rhubarb solution,followed by detecting the relative level of RSV-N after 24h by quantitative reverse transcriptase polymerase chain reaction(qRT-PCR).The viral titer of the supernatant was determined by viral plaque assay.After 72h of RSV infection,cell viability was measured by cell counting kit-8(CCK-8)assay.In addition,A549 cells infected with RSV were treated with ribavirin,or high-dose,medium-dose and low-dose Rhubarb.After 24h of RSV infection,protein levels of RSV with Green Fluorescent Protein(RSV-GFP)and Rab5c were detected by Western blot,and relative levels of negative regulator of antiviral response(NRAV),Rab5c and RSV-N were detected by qRT-PCR.Results There was a significant difference in the survival rate of A549 cells induced with Rhubarb for 72h at varied concentrations(P＜0.01).A significant difference was detected in cell viability of A549 cells induced with 5μg/ml versus 500μg/ml(P＜0.05).There was no significant difference in the survival rate of A549 cells cultured with ribaverin for 72h at varied concentrations of(F=1.89,P=0.17).Rhubarb at 25,50 and 100μg/ml were selected as the low dose,medium dose and high dose,respectively,and the ribaverin treatment was selected at the concentration of 500μmol/L.In comparison to cell survival at-2h(2h prior to RSV infection),the survival at 2h,4h,6h and 8h was significantly elevated(all P＜0.05).In comparison to viral titer and expression level of RSV-N at-2h,those at 2h,4h,6h and 8h were significantly reduced(all P＜0.05).No significant differences were detected in the survival rate,viral titer and expression level of RSV-N between-2h and 0h(all P＞0.05).High-dose,medium-dose and low-dose Rhubarb significantly downregulated NRAV,RSV-N and Rab5c(all P＜0.05).Conclusion Rhubarb may promote the targeted degradation of Rab5c mRNA by miR-509-3p via downregulating NRAV in A549 cells,which inhibits the expression of Rab5c and slows down intracellular vesicle transport,thereby inhibiting the proliferation of RSV.
RhubarbA549 cellsrespiratory syncytial virus(RSV)Rab5cnegative regulator of antiviral response(NRAV)miR-509-3p
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