Study On The Effect Of Lianhua Qingwen Capsules Combined With Tamiflu Or Baloxavir In Inhibiting Seasonal Influenza Viruses

Influenza virus is a global pandemic,causing millions of deaths,and it has the characteristics of high mutation,existing vaccines can not be effectively protected in time,so people pay a lot of attention to the development of antiviral drugs.At present,two types of drugs have been approved for the treatment of human influenza,namely amantadine drugs and neuraminidase inhibitors.In 2014,Japan approved a capdependent endonuclease inhibitor(selective PA inhibitor),which can be used to treat new influenza viruses that do not respond to other drugs,which is named baloxavir.However,chemical drugs are prone to drug resistance mutations,so people pay attention to traditional Chinese medicine.Lianhua Qingwen has a strong antiviral effect and can inhibit a variety of viruses,and so far no drug resistance mutation has been found due to the use of Lianhua Qingwen.Therefore,strengthening the research on the antiviral effect of traditional Chinese medicine and exploring the antiviral effect of traditional Chinese medicine combined with chemical drugs has high theoretical value and clinical significance for the prevention and treatment of influenza.To explore the combination of Lianhua Qingwen capsule with Tamiflu or baroxavir on MDCK cells by determining the toxicity test of Lianhua Qingwen capsule on MDCK cells,conducting the EC50 test of Lianhua Qingwen capsule,Tamiflu and baloxavir on seasonal influenza virus,and the toxicity test of Lianhua Qingwen capsule combined with Tamiflu or baroxavir on MDCK cells.Objective:to explore the effect of Lianhua Qingwen capsule combined with Tamiflu or baloxavir on anti-seasonal influenza virus by determining the toxicity test of Lianhua Qingwen capsule on MDCK cells,conducting the EC50 test of Lianhua Qingwen capsule,Tamiflu and baloxavir on seasonal influenza virus,and the toxicity test of Lianhua Qingwen capsule combined with Tamiflu or Baloshavir on MDCK cells,in order to explore the effect of Lianhua Qingwen capsule combined with Tamiflu or Baloxavir on seasonal influenza virus.According to the results of 50%drug concentration(EC50)of Tamiflu or baloxavir against seasonal influenza virus,the synergistic effect of Tamiflu or baloxavir combined with Lianhua Qingwen capsule was analyzed and compared.1.The cytotoxicity test of Lianhua Qingwen capsule on MDCK cells was carried out.72 hours after adding Lianhua Qingwen capsule,the absorbance of the experimental group was detected by CCK-8 method.The maximum non-toxic concentration(TC0)of Lianhua Qingwen capsule was 75?g/mL by SPSS software analysis.2.The 50%drug concentration(EC50)test of Lianhua Qingwen capsule,Tamiflu and baloxavir on influenza virus infection in MDCK cells was carried out.2000TCID50/ml virus solution was mixed with drugs of different concentrations(Lianhua Qingwen capsule,Tamiflu and baloxavir).The cells were incubated at room temperature for 1 hour,and the cells were incubated in a cell incubator containing 5%CO2 at 37? for 72 hours.the blood agglutination was measured and EC50 was calculated.The EC50 values of Lianhua Qingwen capsule on H1N1,H3N2,Bv and By were 26.49 ± 11.79 ?g/mL,333.40±137.48 ?g/mL,156.69±124.14 ?g/mL and 102.78±73.54 ?g/mL,respectively.The EC50 values of Tamiflu on H1N1,H3N2,Bv and By were 1.67±2.29 ?ML,1.41±1.27 ?M/L,17.07±10.41 ?ML and 17.72±9.13 ?M/L,respectively.The EC50 values of baloxavir on H1N1,H3N2,Bv and By were 11.94±7.74 nM/L,13.28±2.23 nM/L,76.98±50.42 nM/L and 67.93±24.75 nM/L,respectively.3.The toxicity test of Lianhua Qingwen capsule combined with Tamiflu or baloxavir on MDCK cells was carried out,and the TC0 of the combination drug is Lianhua Qingwen 75 ?g/mL+Tamiflu 60 ?M/L and Lianhua Qingwen 75 ?g/mL+Baloxavir 250 nM/L.4.Through the analysis of the above experimental results,the effective inhibitory concentration of Lianhua Qingwen capsule,Tamiflu and baloxavir at the maximum non-toxic concentration was selected,and the anti-influenza virus infection test of Lianhua Qingwen capsule combined with Tamiflu or baloxavir was carried out on MDCK cells.Lianhua Qingwen capsule combined with Tamiflu showed a synergistic effect against influenza virus(H1N1,H3N2,By)infection at the cell level;Lianhua Qingwen capsule combined with baloxavir showed a synergistic effect against influenza virus(H1N1,H3N2)infection at the cell level;Lianhua Qingwen capsule combined with Tamiflu or baloxavir did not affect the anti-influenza effect of Tamiflu or baloxavir at the cell level.Significance:Through the study on the effect of Lianhua Qingwen Capsule combined with Tamiflu or Baloxavir on the inhibition of seasonal influenza virus,the change of antiviral effect produced by Chinese and Western combined drugs was revealed,and the theoretical basis was provided for the followup study on the effect of more combined drugs.