Survival of bone marrow mesenchymal stem cells and periodontal ligament stem cells in cell sheets
10.3760/cma.j.issn.1002-0098.2014.11.010
- VernacularTitle:骨髓间充质干细胞和牙周膜干细胞膜片体内活性的动物实验
- Author:
Kangkang AN
1
;
Hongwei LIU
Author Information
1. 同济大学口腔医学院牙周科·口腔生物医学及转化医学实验室
- Keywords:
Animals,laboratory;
Mesenchymal stem cell;
Periodontal ligament;
Transplantation,heterotopic;
Cell sheet
- From:
Chinese Journal of Stomatology
2014;49(11):682-687
- CountryChina
- Language:Chinese
-
Abstract:
Objective To evaluate the survival of bone marrow mesenchymal stem cells(BMMSC) and periodontal ligament stem cells(PDLSC) in BMMSC/PDLSC cell sheets which transplanted ectopically into subcutaneous dorsum of nude mice.Methods The canine BMMSC and PDLSC from primary culture were tranfected with lentiviral vectors carrying green fluorescent protein(GFP) gene(Lentivirus-GFP) or red fluorescent protein(RFP) gene(Lentivirus-RFP) respectively.The immunophenotypes of GFP-labeled BMMSC and RFP-labeled PDLSC were identified by flow cytometry.Adipogenic and osteogenic differentiation of them were detected by alizarin red or oil red O respectively.Then,both GFP-labeled BMMSC cell sheets and RFP-labeled PDLSC cell sheets were fabricated respectively using normal culture dish(6 cm) after stimulation of extracellular matrix formation.Each was enveloped by collagen membrane (Bio-Gide) and then transplanted into the subcutaneous dorsum of nude mice.In vivo non-invasive biofluorescence imaging(BFI) was performed at 1,2,4 and 8 w post-tranplantation to trace and quantify the survival and growth of RFP-labeled PDLSC and GFP-labeled BMMSC via the BFI system of the NightOWL.The fluorescence intensity change of GFP/RFP signal was monitored and compared.The mice were sacrificed 8 weeks after cell sheets transplantation and the survival of stem cells was verified by fluorescence immunohistochemistry.Results The flow cytometry showed that GFP-labeled BMMSC positively expressed CD29,CD44,CD34,STRO-1 were 93.07％,92.84％,3.23％,67.67％,and RFP-labeled PDLSCs were 89.91％,88.47％,6.04％,74.11％,respectively.Both of them had the potency of differentiating into osteoblasts and adipocytes.The stemness of both of them was almost same.After being transplanted into nude mice,the signal strength of GFP(BMMSC) was weaker and weaker in 1,2,4 and 8 w [(83.1 ± 3.1) × 106,(65.1 ± 2.3) × 106,(51.5 ± 2.3) × 106,(33.8 ± 2.0) × 106 ph/s,respectively.].The signal strength of RFP(PDLSC) was weakenedin 1,2 and 4 w [(53.8±3.0) × 106,(42.2±2.6) × 106,(34.5±2.1) × 106 ph/s],then recovered in 8 w [(45.1 ±2.9)× 106 ph/s].The signal strength of RFP(PDLSC) was signifcantly stronger in 8 w than in 4 w (P＜ 0.01).The survival of RFP-labeled PDLSC was significant higher than that of GFP-labeled BMMSC.After 8 weeks,lots of RFP-labeled PDLSC were observed by microscope,but less GFP-labeled BMMSC were observed.Conclusions Histometric analysis revealed that the survival of stem cells in the RFP-labeled PDLSC cell sheets was significantly higher than that of in the GFP-labeled BMMSCs cell sheets.
