In vitro differentiation of rabbit bone marrow mesenchymal stem cells into Schwann cells Induction effect of platelet-rich plasma

BACKGROUND: Bone marrow mesenchymal stem cells (BMSCs) differentiate into Schwann cells via specific inducers. However, the induction and culture procedures are complicated. Various growth factors have been used for induction and culture, and are likely affected by their environment. OBJECTIVE: To explore the differentiation feasibility of platelet-rich plasma-induced rabbit BMSCs, supplemented with various growth factors, into Schwann-like cells in vitro, and to examine the secretory function of Schwann-like cells. DESIGN, TIME AND SETTING: This comparison study was performed at the Experimental Animal Center, Affiliated Hospital of Qingdao University Medical College, China in October 2008. MATERIALS: Platelet-rich plasma and BMSCs were respectively obtained from the femoral vein and bone marrow of 2-month old New Zealand rabbits. The rabbit nerve growth factor ELISA kit and rabbit nerve growth factor PCR kit (Jingmei Biotech, China), as well as anti-rabbit S-100 immunofluorescence staining kit (Boster, China) were used in the present study. METHODS: BMSCs at passage three were harvested and were induced by 1 mmol/L P-mercaptoethanol and Dulbecco's-modified eagle's medium complete medium containing 35 ng/mL retinoic acid and 10% fetal bovine serum for the combination induction group, followed by incubation in complete medium supplemented with platelet-rich plasma. BMSCs in the single induction group were induced by beta-mercaptoethanol and retinoic acid, followed by incubation in L-DMEM complete medium. BMSCs in the control group were incubated in L-DMEM complete medium. MAIN OUTCOME MEASURES: Cell growth in each group was observed under an inverted microscope. Following induction, S-100 protein expression was identified by immunofluorescence staining. Nerve growth factor protein concentrations in BMSC supernatant were subsequently measured by ELISA. In addition, nerve growth factor mRNA expression in the BMSCs was determined by reverse transcription-polymerase chain reaction. RESULTS: Overall BMSC morphology was similar to Schwann cells in the combination induction group, and morphology of the majority of BMSCs was similar to Schwann cells in the single induction group 9 days after induction. In the control group, significant changes in cell morphology were not observed 9 days after induction. S-100 protein expression was detected in the combination induction and single induction groups following induction. At days 7, 9, and 11, the expression rate of S-100-positive cells was greater in the combination induction group (P < 0.05). At days 4, 7, 9, and 11, nerve growth factor levels were significantly greater in BMSC supematant in the combination induction and single induction groups compared with the control group (P < 0.05). At days 7, 9, and 11, nerve growth factor levels were significantly greater in the combination induction group compared with the single induction group (P < 0.05). At day 11, nerve growth factor mRNA expression in BMSCs was significantly greater in the combination induction group compared with the single induction group (P < 0.05). CONCLUSION: Culture conditions containing platelet-rich plasma elevated the differentiation efficiency of BMSCs into Schwann-like cells, which subsequently secreted nerve growth factor following induction.