HGF And G-CSF Induce The Differentiation Of Rat Bone Marrow Mesenchymal Stem Cells Into Hepatocyte-like Cells

Background and objective：There are a lot of patients with chronic liverdisease in china. Most of them can eventually lead to end-stage liverdisease. In that case, there is only poor efficacy with conventional therapyexcept orthotopic liver transplantation.Though the survival rate is85%~90%with one-year and75%~80%with three-year after operation forthem, there are a lot of issues that are limiting the develop of thetransplantion, such as:shortage of the healthy liver, heavy economic burden,immune rejection after surgery and so on. Urgently, it is particularlyimportant to find a new treatment.Research about BM-MSc has become ahot studying topic in the stem cell field.The BM-MScs can differentiateinto bone cells，osteoblasts，fat cells，hepatic cells，neurons and so on。Due to the following advantages：easily obtainable，readily cultured，lowimmunogenictity，absenting from ethical problems，the BM-MSc becamethe ideal candidate for stem cell research.Many foreign documents haverecorded that HGF，effectively regulate the BM-MSc into liver cells，hasbeen widely used in the research of BM-MSc differentiation in vitro. Hasing the ability to regulate the BM-MSc into liver cells, HGF has beenwidely used in the research of BM-MSc differentiation in vitro,as manyforeign documents recorded.G-CSF is a kind of glycoprotein withmolecular weight20-Ku.It is reported that researchers detected HGF byILISA when they cultured BM-MSc with G-CSF in vitro and they foundG-CSF can increase the BM-MSc’s activity. we research this study toinvestigate if the hepatocyte growth factor （HGF）and granlocyte-colonystimulating factor (G-CSF) could induce differentiation of rat bone marrowmesenchymal stem cells（BM-M SCs） into hepatocyte-like cells andfurther provide effective induction therapy for stem cell research.Methods:The BM-MSCs were isolated and cultured in-vitro by adherentmethod， then identified by flow cytometry.The BM-MSCs of the thirdpassage were used as four groups-c ontrol group （10%FBS）andintervationgroups（0.1μmG-CSF;20ng/mLHGF;0.1μmG-CSF+20ng/mLHGF).ThemRNA and protein were extracted on7,14,21days， then detected byRT-PCR and Western-blot.Results:The third passage BM-MSCs of intervation groups (20ng/mLHGFand0.1μmG-CSF+20ng/mLHGF) got hepatocyte like feafure afterinduction.The results tested by Flow cytometry analysis were as follows:CD34-PE0.3%,CD45-FITC0.1%,CD90-FITC99.6%,CD105-PE99.8%.The mRNA and protein of ALB and AFP were detected respetively by RT-PCR and Western-blot on day7,14,and21from inervation groups(10%FBS+20ng/mLHGF;10%FBS+0.1μmG-CSF+20ng/mLHGF),however,the amount of ALB increased as time went by,while AFPdecreased to extremely low on day21.Meaningfully,the difference betweenthe two groups is significant（P <0.05）.Conclusion: HGF can induce the differentiation of BM-MSCs intohepatocyte-like cells, which can be significantly enhanced by G-CSF.