Effects And Preliminary Mechanism Of Serum Of Severly Burned Rat On Migration Of Rat Bone Marrow Mesenchymal Stem Cells

PART ONE EFFECTS OF SERUM OF SEVERELY BURNED RAT ON PROLIFERATION AND MIGRATION OF RAT BONE MARROW MESENCHYMAL STEM CELLSObjective:Rat bone marrow mesenchymal stem cells (BMSCs) were incubated in medium containing serum of severely burned rat to simulation environment in the body after burn.To study the effects of serum of severely burned rat on proliferation and migration of BMSCs.Methods:Severely burned rat models were established and isolated the serum.Experiments were divided into normal culture group (with 10%FBS) and burn serum group(with 10% serum of severely burned rat).Proliferation of BMSCs was examined with MTT.Migration of BMSCs was observed with Transwell for different time.Results:After burn serum cultered 24h,the proliferation of BMSCs was obviously increased in burn serum group in comparison with normal cuture group(P<0.01).The migration levels of BMSCs in burn serum group for O、1、 3、6.12、24 h were respectively 0、1.0±0.7、14.6±4.6、45.0±6.4、 82.8±8.3、122.2±11.4.At 3h, the migration of BMSCs was begun,and the migration of BMSCs was higher in 6、12、24h than 3h(P<0.01).Conclusion:Serum of severely burned rat can promote the proliferation and migration of BMSCs.PART TWO PHOSHORYLATED LEVEL OF P38 AND AKT OF RAT BONE MARROW MESENCHYMAL STEM CELLS INDUCED SERUM OF SEVERELY BURNED RATObjective:To observe the phosphorylated level of p38 and Akt of rat bone marrow mesenchymal stem cells (BMSCs) induced serum of severely burned rat.Methods:Severely burned rat models were established and isolated the serum.BMSCs were cultured in BMSCs medium containing 10% serum of severely burned rat. The phosphorylated level of p38 and Akt was determined by western blotting.Results:The protein expressions of p-p38 of BMSCs in burn serum for 0、1、3、6、12、24 h were respectively 1.02±0.5、1.11±0.17、1.04±0.23、 1.02±0.20、1.16±0.22、1.0±0.19.There was no significant difference in the phosphorylated level of p38 among the observation time points in burn serum group(P>0.05). The protein expressions of p-Akt of BMSCs in burn serum for 0、1、3、12、24h were respectively 0.59±0.08、0.65±0.06、 0.83±0.22、1.10±0.21、1.22±0.19、1.62±0.30.The protein expressions of p-Akt of BMSCs increased for 6,12,24h in comparison with for 0h,and compared with that of 3h, the protein expressions of p-Akt of BMSCs was obviously increased for 12,24h (P<0.05)Conclusion:Serum of severely burned rat can increase the phosphorylated level of Akt.PART THREE MECHANISM OF SERUM OF SEVERELY BURNED RAT PROMOTE TO MIGRATION OF RAT BONE MARROW MESENCHYMAL STEM CELLSObjective:To explore the effects of phosohoinositide 3-kinase/protein kinase B(PI3K/Akt)signal pathway on migration of rat bone marrow mesenchymal stem cells(BMSCs) induced by serum of severely burned rat. Methods:Experiments were divided into 4groups according to the random number table,cells in normal culture group and burn serum group were given with the same culture condition as above;cells in burn serum+ inhibitor group were cultured with the same culture condition as in the former burn serum group correspondingly with addition of 10μmol/L LY294002; cells in burn serum+activator group were cultured with the same culture condition as in the former burn serum group correspondingly with addition of 200 ng/mL IGF-1. Migration of BMSCs was observed with Transwell. Microtubule structure was observed with Laser confocal microscope.Results:After culture 24h,compared with that of normal cuture group,the microtubule structure became depolymerized,and the number of migration of burn serum group significantly increased (P<0.01).The microtubule structure of burn serum+ activator group was more depolymerized than burn serum group,and the number of migration was higher in comparison with that burn serum group. Compared with that of burn sreum group,there was no significant difference in the microtubule structure in burn serum+ inhibitor group,but the number of migration wasobviously decreased (P <0.01)Conclusion:The IGF-1 in serum of severely burn rat may active PI3K/Akt signal pathway to induce the microtubule structure change thus enhancing the migration of BMSCs.