Based On TLR4/MyD88/NF-κB Signal Pathway To Explore The Mechanism Of Yiqi Kangfei Formula On Acute Lung Injury

Objective:1.To clarify the effect of Yiqi Kangfei Formula on acute lung injury(ALI)induced by lipopolysaccharide(LPS).2.Based on TLR4/My D88/NF-κB signal pathway further explores the mechanism of Yiqi Kangfei Formula on ALI.Methods:In this experiment,60 SPF Syrian golden hamsters were selected and divided into model group,blank group,dexamethasone group,and Chinese medicine(Yiqi Kangfei Fang)low-dose,medium-dose and high-dose groups,with 10 in each group.Before modeling,blank group,model group and Chinese medicine group were given corresponding drugs for 14 days,and dexamethasone group for 4 days.After administration,LPS was injected into the trachea of the other groups except the blank group.After modeling,the general state and weight of the golden hamsters were observed.2 days later,all golden hamsters were killed for sampling.Weigh lung tissue wet weight and calculate lung index;The total protein concentration in broncho alveolar lavage fluid(BALF)was detected.HE staining was used to observe the pathological changes of lung tissue.The levels of inflammatory factors TNF-α,IL-1β and IL-6 in serum were determined by ELISA;The protein expression of TLR4,My D88 and NF-κB-p65 in lung tissue was detected by WB.Result:1.General state of the golden hamsters: the golden hamsters in the blank group had good mental state,stable breathing,smooth hair and normal activities;In the model group,the breathing rate of the ground rats was accelerated,and occasionally wheezing could be heard.The hair was haggard,the spirit was weak,and the activity was decreased.Some golden hamsters curled up into a ball,and the food intake,body weight and stool were decreased.Dexamethasone group and low-dose,medium-dose and high-dose Chinese medicine groups showed better mental state,activity and eating than model group.2.Lung index: Compared with blank group,lung index of model group was increased,the difference was statistically significant(P < 0.05).Compared with model group,lung index in dexamethasone group,low-dose,medium-dose and high-dose groups was decreased,and the difference was statistically significant(P < 0.05).Compared with dexamethasone group,there was no significant difference in low,medium and high dose groups(P > 0.05).3.Total protein concentration in BALF: Compared with blank group,total protein concentration in BALF in model group was increased,and the difference was statistically significant(P < 0.05).Compared with model group,the concentration of total protein in BALF of high dose group was decreased,and the difference was statistically significant(P< 0.05).There was no statistical difference between low and medium dose groups(P >0.05).Compared with dexamethasone group,the total protein concentration in BALF in TCM low-dose group was increased,and the difference was statistically significant(P <0.05).There was no significant difference between medium and high dose groups(P >0.05).4.General morphological changes of lung tissue: lung tissue injury is the most directly related feature of ALI.In this experiment,the gross morphology of lung tissue showed that in the blank group,the envelope of both lung lobes was intact,without color change,hemorrhagic spots,edema,etc.Compared with the blank group,the model group showed dark red pulmonary lobes,severe congestion,multiple scattered dark red hemorrhagic spots on the surface,and obvious edema.Compared with model group,morphologic changes were observed in dexamethasone group,low,medium and high dose groups.Among them,dexamethasone group and high dose group only had lung color change,no bleeding,edema and so on.5.Pathological changes of lung tissue: the blank group had normal lung tissue structure,no exudation in alveoli,no obvious swelling;Compared with the blank group,the lung tissue structure of the model group was destroyed,a large number of alveoli disappeared,and the tissue was extensively damaged.At the same time,more inflammatory cells were infiltrated and distributed,and local clusters were formed.Compared with model group,dexamethasone group and low-dose,medium-dose and high-dose Chinese medicine groups had relatively mild lung tissue lesions.There was basically no exudation in the alveoli of the dexamethasone group,but there was local enlargement.The lung tissue damage in the low-dose and medium-dose groups was moderate,and there was little or no inflammatory cell infiltration.The lung injury was very light in the high-dose group,and there was no exudation in the alveoli.6.Serum levels of inflammatory factors TNF-α,IL-1β and IL-6: Compared with blank group,serum levels of TNF-α,IL-1β and IL-6 in model group were increased,and the difference was statistically significant(P < 0.05).Compared with model group,serum TNF-α,IL-1β and IL-6 in dexamethasone group,low,medium and high dose groups were decreased,the difference was statistically significant(P < 0.05).Compared with dexamethasone group,serum levels of TNF-α and IL-1β in low-dose and medium-dose groups had no statistical difference(P > 0.05),serum levels of IL-6 in low-dose group were increased,the difference was statistically significant(P < 0.05),while serum levels of IL-6in medium-dose group were decreased,the difference was statistically significant(P <0.05).The levels of TNF-α,IL-1β and IL-6 in serum of high dose group were decreased,and the difference was statistically significant(P < 0.05).7.TLR4,My D88,NF-κB-p65 protein expression in lung tissue: Compared with blank group,the expression levels of TLR4,My D88 and NF-κB-p65 protein in lung tissue of model group were increased,and the differences were statistically significant(P < 0.05).Compared with model group,the protein expressions of TLR4,My D88 and NF-κB-p65 in lung tissues of dexamethasone group,low,medium and high dose groups were decreased,and the differences were statistically significant(P < 0.05).Compared with dexamethasone group,the protein expressions of TLR4,My D88 and NF-κB-p65 in lung tissue of low-dose Chinese medicine group were increased,and the differences were statistically significant(P< 0.05),while the protein expressions of TLR4,My D88 and NF-κB-p65 in lung tissue of medium-dose and high-dose Chinese medicine groups were decreased.The difference was statistically significant(P < 0.05).Conclusion:1.Yiqi Kangfei Formula can improve capillary membrane permeability and pulmonary edema,reduce pathological injury of lung tissue,reduce inflammatory reaction,and has a protective effect on ALI.2.Yiqi Kangfei formula may reduce the synthesis and release of inflammatory factors by inhibiting the activation of TLR4/My D88/NF-κB signal pathway,thus alleviating inflammatory response and playing a protective role against ALI.