Studies On TGF-β1,bFGF And PDGF Expression In Bovine Mammary Epithelial Cells Effected With Escherichia Coli

Escherichia coli is the most main pathogenic bacteria causing mastitis, mostly cause severe clinical mastitis, which can cause bovine mammary fibrosis. TLR2 induced cytokine and chemokine secretion in immune cells through MyD88-dependent signaling pathway. TLR4 recognize Gram-negative bacteria and E.coli lipopolysaccharide, and play a key role in mediating LPS-negative bacteria and the host response. TGF-β1, bFGF and PDGF are important cytokines in fibrosis. The aim of this study is to investigate in BMEC infected by E. coli how expressed on TGF-β1, bFGF, PDGF and their receptors, whether TLR2 and TLR4 to participate in BMEC infected by E. coli, and what roles they play in this process. In order to explore the pathogenesis of bovine mammary gland fibrosis, It can provide guidance for the prevention and treatment of bovine mammary fibrosis.In this study, BMEC were stimulated with different concentrations of heat-inactivated E. coli 105 cfu/mL,106 cfu/mL and 108 cfu/mL after infection 6h,12h,24h, and 48h through the methods of qPCR and Western blot to analyze the expression of TGF-β1, bFGF, PDGF and their receptors, By the qPCR and Western blot to analyze TLR2 and TLR4 mRNA and protein expression. By the methods of Western blot, we analyzed NF-κB and AP-1 protein expression.The results showed that the expression of TGF-β1, bFGF, PDGF and their receptors were increased in BMEC after E.coli infection. The data also exhibited that TGF-β1, TβR1, bFGF, FGFR2, PDGF-BB and PDGFRp mRNA and protein expression were increased with time-and concentration-dependent manner by time and increasing concentrations. The expression was the highest on 12h or 24h, E.coli concentrations in 106cfu/ml had highest expression. The TGF-β1 mRNA expression was the the highest on 12h, the relative expression of TβR1 protein was the highest on 24h. The highest expression of bFGF and FGFR2 were in 24h. The PDGF and PDGFRβ mRNA and protein expression were to a maximum respectively in 12 hours and 24 hours.E. coli could promote the expression of TLR2 and TLR4. The expression of TLR2 and TLR4 mRNA and protein secretion were increased in dose and time-dependent manner in BMEC stimulated with 105 cfu/mL,106 cfu/mL and 108 cfu/mL. The maximum expressions was on 24h.E. coli could promote the expression of NF-κB and AP-1 protein. The expression were increased in this process with time-and concentration-dependent manner.In conclusion, E.coli could promote the expression of TGF-β1,TβR1, bFGF FGFR2, PDGF-BB, PDGFRβ, TLR2 and TLR4. Not only TGF-β1, bFGF, PDGF and their receptors, but also TLR4 and TLR2 were involved in BMEC stimulated by E. coli, played crucial roles during this process. So we also indicated that TGF-β1, bFGF, PDGF and their receptors may play crucial roles in bovine mammary fibrosis. The expression of NF-κB and AP-1 protein were promoted in BMEC effcted by E. coli. The NF-κB and AP-1 signaling pathway were actived in the BMEC stimulated by E. coli. They may be involved in the process of bovine mammary fibrosis.