Studies On Expression Of MMPs/TIMPs And UPA System Induced By Escherichia Coli In Bovine Mammary Fibroblasts

Bvine mastitis is one of the main diseases that cause important loss of bovine milk industry,and E.coil is one of the main pathogens that induce mastitis in the bovine.The main pathological changes of mastitis in the bovine are caused by persisting injury of mammary gland,which leads to mammary fibrosis or mammary sclerosis.When the mammary tissue is damdged,bovine mammary fibroblasts(BMFB)secrete lots of extracellular matrix(ECM),and excessive accumulation of ECM leads to fibrosis.In order to explore the effect of Escherichia coli(E.coli)on the expression of MMPs/TIMPs and uPA system in BMFB,and to illuminate the role of MMPs/TIMPs and uPA system in regulating ECM.In this study,E.coli was isolated from the mammary glands of cows with mastitis to produce a heat-inactivated E.coli.BMFBs treated with heat-inactivated E.coli(105,106,and 108 cfu/mL)for 6 h.12 h,24 h,and 48 h.MMP-1/MMP-2/MMP-3/MMP-9/MMP-13/TIMP-1/TIMP-2/uPA/uPAR/PAI-1 mRNA transcriptioin levels was examined by RT-qPCR and the protein expression was examined by Western blot analysis.The result sa follow:(1)The mRNA transcriptioin levels of several MMPs(MMP-1/MMP-2/MMP-3/MMP-9/MMP-13)were basically up-regulated compared with those of the control group after the action ofthe heat-inactivated E.coil solution with differennt concentrations at each time point.The protein expression of MMP-1 was mostly down-regulated compared with the control group expect the group of 105 cfu/mL was significantly up-regulated at 6 h;the protein expression of MMP-2 showed a trend of decreasing first and the increasing compared with the control group;the protein expression of MMP-3/MMP-9/MMP-13 compared with the control group showed a trend of decreasing first and the increasing.(2)The mRNA transcriptioin levels of several TIMP-1/TIMP-2 were basically up-regulated compared with those of the control group after the action ofthe heat-inactivated E.coil solution with differennt concentrations at each time point.The protein expression of TIMP-1 was mostly down-regulated except the treatment group of 105 cfu/mL,while the other concentration treatment group was down-regulated at 6 h and 24 h,and up-regulated at 12 h and 48 h;compared with the control group,the protein expression of TIMP-2 were down-regulated except the treatment group of 108 cfu/mL was up-regulated for 24 h.(3)The mRNA transcriptional level of uPA/uPAR/PAI were basically up-regulated compared with those of the control group after the action ofthe heat-inactivated E.coil solution with differennt concentrations at each time point.The protein expression of uPA showed a trend of decreasing first and the increasing;the protein expression of uPAR showed a trend of increasing first and the decreasing;the protein expression of PAI-1 was down-regulation in the initial stage and up-regulation in 24 h compared with the control group.The study showed that heat-inactivated E.coil could promote or inhibit the mRNA transtription and the protein expression of MMPs/TIMPs and uPA system in BMFB.and promoted the involvement of MMPs/TIMPs and uPA system participate in bovine mammary fibrosis.and MMPs/TIMPs and uPA system maybe regulate ECM by E.coli-induced BMFBs,which provided the basic for MMPs/TIMPs and uPA system point to be promising therapeutic targets in bovine mammary fibrosis.