The Effect And Mechanism Of Physalin Extract Against Mycoplasma Pneumonia

Mycoplasma infection is a difficult problem in livestock and poultry production.Mycoplasma gallisepticum(MG)infection will cause a decrease in feed conversion rate,slower daily gains in broilers,and reduced performance of laying hens,which will bring huge economic losses to poultry production.At present,MG infection is mainly treated with antibiotics,but due to the irregular use of antibiotics,the emergence of drug-resistant strains makes the research and development of drugs for this disease particularly important.In recent years,natural compounds have been widely used in clinical treatment,and their advantages in the treatment of mycoplasma infections have been gradually discovered.Because many natural compounds can promote the growth of livestock and poultry,they are more advantageous in the treatment of livestock and poultry diseases.Physalis alkekengi L.var.franchetii.has been used in traditional Chinese medicine to treat respiratory diseases,and Physalin is the main active ingredient.However,there is no research report on the prevention and treatment of MG by Physalin.Therefore,this experiment established a chicken mycoplasma pneumonia model to explore the therapeutic effect of Physalin in vivo;at the same time,establish a mouse mycoplasma pneumonia infection model to explore the therapeutic effect and mechanism of Physalin in treatment of mycoplasma pneumonia,which provide practical and theoretical support for the research and development of Physalin as a new drug.Results:1.Through LPS-induced classic inflammation model of rat alveolar epithelial cells(RLE-6TN),it was found that compared with the cell control group,1 μg/m L LPS acted on RLE-6TN cells for 24 hours leads significantly reduced of the cell viability(P<0.05).The maximum non-toxic concentration(MNTC,maximum non-toxic concentration)of Physalin in RLE-6TN cells was 50 μg/m L determined by CCK8 method.LPS acting on RLE-6TN cells can significantly reduce the expression of IκB,increase the expression of NF-κB,and at the same time enhance the activation of IκB and NF-κB.LPS treatment can also increase the expression of COX2,IL-1β and related inflammatory factors(P<0.01).However,treatment with Physalin can significantly eliminate these symptoms caused by LPS(P<0.01).After the treatment of LPS,the total protein content of ERK1/2,JNK and p38 in RLE-6TN cells did not change significantly(P>0.05),but their phosphorylation levels were significantly increased(P<0.01).However,treatment with Physalin can significantly eliminate these symptoms caused by LPS(P<0.01).The anti-inflammatory effect of Physalin in vivo was verified with LPS-induced rat acute lung injury model.LPS can induce the increase in the number of white blood cells in the blood of rats,and increase the level of inflammatory factors in the rat lung tissue and the W/D ratio of the lung tissue.At the same time,it can also induce lung tissue injury and alveolar epithelial cell apoptosis.Physalin treatment can significantly eliminate these symptoms caused by LPS(P<0.01).It shows that Physalin can reduce the activation of NF-κB signaling pathway and MAPK signaling pathway,thereby reducing LPS-induced inflammatory.2.The classical oxidative damage model of RLE-6TN cells induced by H2O2 showed that compared with the cell control group,the cell viability of RLE-6TN cells was significantly reduced after 100 μmol H2O2 incubated to RLE-6TN cells for 12 h(P<0.01).10 μg/m L the new Physalin B can significantly reduce the apoptosis of rat alveolar epithelial cells induced by H2O2(P < 0.01).H2O2 treatment in vitro significantly reduced the protein expression of the Nrf2 signaling pathway in RLE-6TN cells(P<0.01),however,the new Physalin B treatment significantly eliminated these symptoms induced by H2O2(P<0.01).In the cells treated with H2O2,the expression of Bcl-2 and Bcl-x L was significantly reduced(P<0.01),the expression of Bax and p53 protein was significantly increased(P<0.01),and the new Physalin B treatment significantly eliminated these symptoms induced by H2O2(P<0.01).This indicates that the new Physalin B can reduce the oxidative damage induced by H2O2 through regulating the Nrf2/p53 signal pathway.3.Mycoplasma gallisepticum was cultured in vitro,and the effect of Physalin on the proliferation of MG in vitro was detected by RT-PCR.The results showed that 100 μg/m L of Physalin could significantly inhibit the proliferation of MG in vitro(P<0.01).The Mycoplasma gallisepticum model was induced by MG,and Physalin was administered through drinking for treatment.MG infection leads to increased nasal secretions,depression,yellowing and thickening of air sacs,congestion and swelling of the lungs,and nodular necrosis,a significant decrease in body weight,and a large number of inflammatory cell infiltration and alveolar structure in the alveolar cavity Destroy,and can induce the activation of NF-κB signaling pathway(P<0.05).However,2 g/L Physalin drinking treatment can significantly eliminate these symptoms caused by MG(P<0.05).It shows that Physalin can treat pneumonia caused by MG infection by inhibiting the proliferation of MG,repairing the structure of lung tissue and reducing inflammation through the NF-κB pathway.4.Mycoplasma pneumoniae(MP)was cultured in vitro,and the effect of Physalin on the proliferation of MP in vitro was detected by RT-PCR.The results showed that 50 μg/m L Physalin could significantly inhibit the proliferation of MP in vitro.Next,a model of MP-induced RLE-6TN cell injury was established,and the concentration of Physalin in vitro against MP-induced RLE-6TN cell injury was determined by CCK8 and RT-PCR methods.The results showed that 50 μg/m L Physalin can significantly reduce the damage of rat alveolar epithelial cells induced by MP,and inhibit the proliferation of MP in cells(P<0.01).MP was used to induce mycoplasma pneumonia model in mice,and treated by gavage Physalin.The results showed that MP infection caused a significant increase in the content of Ig G and Ig M in mice(P<0.01),a large number of inflammatory cell infiltration in the alveolar cavity and the destruction of alveolar structure.MP also activated the NF-κB signaling pathway and MAPK signaling pathway in mice,the protein expression of the Nrf2 signaling pathway was reduced,the expression of Bcl-2 and Bcl-x L was significantly reduced,and the expression of Bax and p53 protein was significantly increased(P<0.01),Physalin treatment can significantly eliminate these symptoms caused by MP.It shows that Physalin can treat MP-induced Mycoplasma Pneumonia in mice by inhibiting the proliferation of Mycoplasma pneumoniae,repairing lung tissue structure,reducing inflammation through the Nrf2/MAPK/NF-κB signaling pathway,reducing oxidative damage through the Nrf2 signaling pathway,and reducing apoptosis through the Bax signaling pathway.Conclusion:Physalin has good anti-inflammatory and antioxidant effects.Physalin can directly act on MG and MP to inhibit the proliferation of mycoplasma.Using MG and MP to stimulate target animals as a mycoplasma pneumonia model,Physalin mainly repairs lung tissue structure,reduces inflammation through the Nrf2/MAPK/NF-κB signaling pathway,reduces oxidative damage through the Nrf2 signaling pathway,and reduces apoptosis through the Bax signaling pathway to treat mycoplasma pneumonia induced by MG and MP.This result provides a new theoretical basis for the development of Physalin as an anti-mycoplasma drug and clarification of its therapeutic target.