Inhibitory Effect Of Lactoferrin On LPS-Induced Inflammation Of Primary Hepatocytes In Dairy Cows

Lactoferrin is a natural immune protein commonly found in milk and other exocrine fluids of dairy cows.Not only the entire production cycle of cows is threatened by the two major obstetric inflammation of mastitis and endometritis;at the same time,during the special transition period of dairy cows – perinatal period,the body experiences negative energy balance and metabolism occurs.Excitatory and systemic persistent subacute inflammatory stimuli have a huge impact on dairy health and performance.In this paper,we studied the inhibitory effect of lactoferrin on LPS-induced hepatic primary cell inflammatory response and its possible molecular mechanism,providing theoretical basis and technical support for the clinical anti-inflammatory application of lactoferrin.In this paper,the primary hepatocytes of dairy cows isolated by modified collagenase two-step perfusion method were used as experimental materials.The effect of LPS on the vitality of primary liver cells of dairy cows was detected by MTT assay.The expression of inflammatory factor genes in LPS-induced cells was detected by real-time PCR.The conditions of induced hepatic inflammatory injury model in dairy cows were screened.Based on the model,lactoferrin co-culture was added.The expression of key proteins and inflammatory factor genes in related inflammatory signaling pathways was detected by real-time PCR and Western Blot.The research results are as follows.1000 ng/mL LPS was applied to the primary hepatocytes of dairy cows for 24 h,but had no significant effect on cell viability,but significantly increased the expression of inflammatory factors TNF-? and IL-1?.The 1 ?g/ mL LPS was used to stimulate the primary hepatocytes of cows with 24 h,and the LPS-induced primary hepatocyte inflammatory response model was established.Lactoferrin(1,10,100 ?g/ mL)inhibits the expression of pro-inflammatory cytokines,stimulates the expression of anti-inflammatory cytokine genes,and secretes the secretion of AST and ALT from primary liver cells of dairy cows.It has an inhibitory effect.Lactoferrin(1,10,100 ?g/ mL)inhibited the expression of TLR4,CD14,MyD88,TICAM1,TICAM2 protein genes in LPS4 signaling pathway induced by LPS in dairy cows.At the same time,lactoferrin inhibited the degradation of I?B? and the phosphorylation of NF-?B p65 induced by LPS in primary liver cells of dairy cows.The molecular mechanism suggesting its anti-inflammatory effect is to inhibit the activation of the NF-?B signaling pathway by inhibiting the MyD88-dependent pathway of the TLR4 signaling pathway and the MyD88-dependent pathway(TRIF-dependent pathway).