Research On The Effect Of MMP9 On Lacting Function Of Dairy Cow Mammary Epithelial Cell

The mammary gland is a delicate and complex secretory organ.Different stages of mammary gland development require fine-controlled degradation and remodeling of the extracellular matrix.Matrix metallopeptidase 9,MMP9 is considered to be a key enzyme.In order to determine the effect and possible mechanism of MMP9 gene on bovine mammary epithelial cells(BMECs),MMP9 gene overexpression vector was constructed,gene overexpression and gene silencing techniques was used to change the expression of MMP9 in BMECs.The cell proliferation of BMECs was detected by MTT method.The triglyceride content of BMECs was detected by triglyceride detection kit.The migration of BMECs was detected by cell migration assay.The expression of ?-casein and related signal molecules was detected by Western blotting or q PCR.In addition,LY-294002,U0126 or rapamycin was added to BMECs,and the expression of MMP9 and related signaling pathway molecules were detected by Western blotting.The results of MMP9 overexpression and silence experiments showed that MMP9 gene overexpression could significantly increased the expression of p-m TOR(P<0.05),m TOR(P<0.05),Bcl-2(P<0.05),Bcl-XL(P<0.05)and E-cadherin(P<0.05);MMP9 gene overexpression can also increase cell proliferation(P<0.05),expression of beta-casein(P<0.05)and triglycerides level(P<0.05).BMECs migration was promoted,but m RNA level of Bax(P<0.05)and Bak(P<0.05)was decreased.When MMP9 gene was silenced,all the indicators were opposite to those of MMP9 gene overexpression.It is speculated that MMP9 gene affects the proliferation and migration of BMECs.After adding signaling pathway inhibitors in BMECs,the results showed that LY-294002 significantly down-regulated the protein expression of p-AKT(P<0.05),AKT(P<0.05)and MMP9(P <0.05);rapamycin significantly down-regulated protein expression of p-m TOR(P<0.05),m TOR(P<0.05)and MMP9(P<0.05);U0126 significantly down-regulated protein expression of p-ERK(P<0.05),ERK(P<0.05),and MMP9(P<0.05).It is speculated that PI3K/AKT/m TOR and MAPK signaling pathways can regulate MMP9 protein expression.In summary,MMP9 gene can affect the proliferation and migration of BMECs.MMP9 can be regulated by PI3K/AKT/m TOR and MAPK signaling pathways.