The Effect Of Serotonin On Proliferation Of Human Bone Marrow-derived Mesenchymal Stem Cells

Objective:To investigate the effect of serotonin (5-HT) on proliferation of human bone mesenchymal stem cells (BMSCs) in vitro.Methods:Bone marrow were inspired from ilium of donators, mononuclear cells were isolated by density gradient centrifugation and cultured in vitro. The higher purity of BMSCs were obtained by keeping the adherent cells and removal of nonadherent cells repeatedly. P3 or P4 BMSCs populations were collected, by using conditioned medium containing different concentrations of 5-HT interfered with BMSCs. Optical microscope observed cells shape variation for 24,48 and 72 hours; MTS detected and evaluated cells proliferation; Flow cytometry (FCM) detected BMSCs apoptosis rate.Results:1. Cell morphology observation:When inoculated cells, BMSCs present the circular, bright and good refraction. Common cultured cells after 24,48 and 72 hours. BMSCs began to adherent the wells, cells distribution uniformity, clear-cut, form uniform, assumed tuber shape or long spindle, cells increased proliferation speed, and mutual confluented gradually. Adding 5-HT cultured cells after 24,48 and 72 hours. Observation cells changed form gradually, cells went round, shrinks, separated with near cells; nuclear chromatin became pyknotic agglomeration, a block or chip shape change; cytoplasmic enrichment, the center transmittance reduced.2. MTS detection: At the different concentration of 5-HT (0.01,0.1,1,10,100,500,1000μM) effects, with 0% and 10% serum concentrations, compared with control, ELIASA detected the OD values reduced in all groups. Testing results showed that:At 24 and 48 hours, there were no significant different effects in BMSCs between the concentration of 5-HT (0.01,0.1,1,10,100μM) groups and control group (P>0.05), when 5-HT concentration at 500μM and 1000μM, the 5-HT groups and control group showed significant difference (P<0.05). At 72 hours, there were no significant different effects in BMSCs between the concentration of 5-HT (0.01,0.1,1,10μM) groups and control group (P>0.05), when 5-HT concentration at 100μM,500μM and 1000μM, the 5-HT groups and control group showed significant difference (P<0.05). Testing results showed that with the observation time extended,5-HT concentrations increasing, cell survival rate gradually reduced. In serum-free cases, the lowest survival rate can reach 59.5%(P<0.05).3. Flow cytometry (FCM) detection:BMSCs were treatmented by each groups of 5-HT (1,10,100,1000μM) for 48 hours, compared with control, testing results showed when 5-HT concentration at 10μM, 100μM and 1000μM, with concentrations increasing, cell apoptosis rate increased (P<0.05), when 5-HT concentration was 1000μM, apoptosis rate was 21.42%.Conclusion:5-HT had apoptosis effect in human BMSCs proliferation in vitro. No matter in a serum or serum-free cases,5-HT effected time prolonger, the lower cell survival rate; the higher 5-HT concentration, the more improve cell apoptosis rate, and in serum-free such low nutrition environment, the later 5-HT apoptosis role was more obvious.