Effects of combination TGF-B1 transfection and platelet rich plasma (PRP) on three-dimension chondrogenic differentiation of rabbit dental pulp-derived mesenchymal stem cells

Abstract Background Platelet-rich plasma (PRP) has revealed benefits in tissue repair and regeneration, however, the effect of PRP on proliferation and chondrogenesis of mesenchymal stem cells remains controversial.This study is to evaluate the effect of different concentration PRP on proliferation and chondrogenic differentiation of rabbit bone marrow-derived mesenchymal stem cells(BMSCs). Methods PRP was obtained by centrifugation and activation, in which growth factors and cytokines were detected. BMSCs were isolated from rabbit bone marrow and characterized by flow cytometry. About 5 × 103 BMSCs were cultured in high glucose Dulbecco’s modified Eagle’s medium (HG-DMEM) with 4 different compositions: 10% fetal bovine serum, 5%PRP, 10%PRP, and 15%PRP for consecutive 7 days. Cell counting assays were performed on days 1, 3, 5, and 7 to evaluate the BMSCs proliferation. In chondrogenic differentiation, high-density cell pellets composed of 5 × 105 BMSCs were induced in 4 conditions: commercial chondrogenic medium (control), 5%PRP (HG-DMEM + 5%PRP), 10%PRP (HG-DMEM + 10%PRP), and 15%PRP (HG-DMEM + 15%PRP) for 21 days. The gene expression levels of aggrecan (ACAN), collagen type Ⅱ Alpha 1 (COL2A1), and SRY-Box Transcription Factor 9 (SOX9) in pellets were detected. Histological assessments were performed by morphologically observation and pathological stain. Independent-samples t-test and one-way analysis of variance were used in statistical analyses. Results The concentrations of growth factors and cytokines were elevated in PRP. BMSCs proliferation was enhanced in all groups, and 10% PRP revealed more obvious outcome than the others from day 5. In chondrogenic differentiation, the levels of ACAN, COL2A1, and SOX9 were lower in 3 PRP groups than control, but ACAN and SOX9 was higher in 10% PRP group than 5% and 15%. Histological examinations showed 10% PRP-treated pellets showed more regular appearance, larger size, and abundant extracellular matrix than 5% and 10% groups, but still inferior to commercial chondrogenic medium. Conclusions PRP may enhance the proliferation of rabbit BMSCs, while with limited effect on chondrogenic differentiation compared with commercial chondrogenic medium in pellets culture. Whether optimizing and modifying PRP components would lead to satisfying chondrogenesis of BMSCs, it is deserved furthermore study.

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Effect of Activated Platelet-Rich Plasma on Chondrogenic Differentiation of Rabbit Bone Marrow-Derived Mesenchymal Stem Cells

Stem Cells International ◽

10.1155/2021/9947187 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Zhen Wang ◽

Zheng Wang ◽

Bin Zhang ◽

Qinghua Zhao ◽

Yubao Liu ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Mesenchymal Stem Cells ◽

Chondrogenic Differentiation ◽

Platelet Rich Plasma ◽

Cell Counting ◽

Control Group ◽

Chondrogenic Medium ◽

Rabbit Bone ◽

Fetal Bovine

We aimed to evaluate the effect of activated platelet-rich plasma (PRP) on proliferation and chondrogenic differentiation of bone marrow-derived mesenchymal stem cells (BMSCs). Six mature male rabbits were included in this study. PRP was obtained by two-step centrifugation from whole blood, and it was activated using CaCl2 solution. BMSCs were isolated and proliferated from bone marrow of rabbits and characterized by flow cytometry. Passage 3 BMSCs were cultured in high-glucose Dulbecco’s modified Eagle’s medium (HG-DMEM) with the four different compositions for consecutive 7 days, including 10% fetal bovine serum, 5% PRP, 10% PRP, and 15% PRP. Cell counting assays were performed to evaluate the cell proliferation of BMSCs. BMSCs ( 5 × 10 5 cells/well in 6-well plates) were induced in four conditions for 21 days to chondrogenic differentiation evaluation, including commercial chondrogenic medium (control), 5% PRP (HG-DMEM+5% PRP), 10% PRP (HG-DMEM+10% PRP), and 15% PRP (HG-DMEM+15% PRP). The gene expression levels of ACAN, COL2A1, and SOX9 in pellets were detected. Morphological and pathological assessments were performed by the blind observer. After purifying, the percentages of cells with CD105(+)/CD34(−) and CD44(+)/CD45(−) were 96.5% and 92.9%, respectively. The proliferation of BMSCs was enhanced in all groups, and 10% PRP revealed more significant outcome than the others from day 5. The levels of ACAN, COL2A1, and SOX9 were lower in the three PRP groups than control group, but the levels of ACAN and SOX9 were higher in 10% PRP group than 5% and 15% PRP groups. Histological examinations showed that 10% PRP-treated pellets had more regular appearance, larger size, and abundant extracellular matrix than 5% or 10% PRP groups, but still inferior to commercial chondrogenic medium. In conclusion, our results show that PRP may enhance the proliferation of rabbit BMSCs. However, PRP have limited effect on chondrogenic differentiation in comparison with commercial chondrogenic medium in pellets culture.

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