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Published on: November 1, 2017
Regulation of human skeletal stem cells differentiation by Dlk1/Pref-1
Basem M Abdallah1, Charlotte H Jensen, Gloria Gutierrez
1Department of Endocrinology, Univerity Hospital of Odense, Odense, Denmark.
Unlabelled:
Dlk-1/Pref-1 was identified as a novel regulator of human skeletal stem cell differentiation. Dlk1/Pref-1 is expressed in bone and cultured osteoblasts, and its constitutive overexpression led to inhibition of osteoblast and adipocyte differentiation of human marrow stromal cells.
Introduction:
Molecular control of human mesenchymal stem cell (hMSC) differentiation into osteoblasts and adipocytes is not known. In this study, we examined the role of delta-like 1/preadipocyte factor-1 (Dlk1/Pref-1) in regulating the differentiation of hMSCs.
Materials And Methods:
As a model for hMSCs, we have stably transduced telomerase-immortalized hMSC (hMSC-TERT) with the full length of human Dlk1/Pref-1 cDNA and tested its effect on hMSC growth and differentiation into osteoblasts or adipocytes as assessed by cytochemical staining, FACS analysis, and real time PCR. Ex vivo calvaria organ cultures assay was used to confirm the in vitro effect of Dlk/Pref-1 on bone formation.
Results:
Dlk1/Pref-1 was found to be expressed in fetal and adult bone, hMSCs, and some osteoblastic cell lines. A retroviral vector containing the human Dlk1/Pref-1 cDNA was used to create a cell line (hMSC-dlk1) expressing high levels of Dlk1/Pref-1 protein. Overexpression of Dlk1/Pref-1 did not affect the proliferation rate of hMSC, but the ability to form mature adipocytes, mineralized matrix in vitro, and new bone formation in neonatal murine calvariae organ cultures was reduced. These effects were associated with inhibition of gene expression markers of late stages of adipocyte (adipocyte fatty acid-binding protein [aP2], peroxisome proliferator-activated receptor-gamma2 [PPARgamma2], and adiponectin [APM1]) and osteoblast differentiation (alkaline phosphatase [ALP], collagen type I [Col1], and osteocalcin [OC]). Lineage commitment markers for adipocytes (adipocyte determination and differentiation factor -1 [ADD1]) and osteoblasts (core binding factor/runt-related binding factor 2 [Cbfa1/Runx2]) were not affected.
Conclusion:
During hMSC differentiation, Dlk1/Pref-1 maintains the size of the bipotential progenitor cell pool by inhibiting the formation of mature osteoblasts and adipocytes.
Insights
Delta-like 1/preadipocyte factor-1 (Dlk1/Pref-1) regulates human skeletal stem cell differentiation. Overexpressing Dlk1/Pref-1 inhibits osteoblast and adipocyte formation, maintaining the progenitor cell pool.
Area of Science:
- Stem cell biology
- Molecular biology
- Bone biology
Background:
- The molecular mechanisms governing human mesenchymal stem cell (hMSC) differentiation into osteoblasts and adipocytes remain largely unknown.
- This study investigates the role of delta-like 1/preadipocyte factor-1 (Dlk1/Pref-1) in regulating hMSC differentiation pathways.
Purpose of the Study:
- To elucidate the function of Dlk1/Pref-1 in human skeletal stem cell differentiation.
- To determine the impact of Dlk1/Pref-1 overexpression on osteogenic and adipogenic potential of hMSCs.
Main Methods:
- Telomerase-immortalized hMSCs (hMSC-TERT) were stably transduced with human Dlk1/Pref-1 cDNA.
- Effects on hMSC proliferation and differentiation were assessed using cytochemical staining, FACS analysis, and real-time PCR.
- Ex vivo calvarial organ cultures were employed to validate in vitro findings on bone formation.
Main Results:
- Dlk1/Pref-1 expression was detected in bone, hMSCs, and osteoblastic cell lines.
- Overexpression of Dlk1/Pref-1 in hMSCs did not alter proliferation but significantly reduced adipocyte and osteoblast differentiation.
- Key gene markers for late-stage adipogenesis and osteogenesis were downregulated, while lineage commitment markers remained unaffected.
Conclusions:
- Dlk1/Pref-1 acts as a novel regulator of human skeletal stem cell differentiation.
- By inhibiting the formation of mature osteoblasts and adipocytes, Dlk1/Pref-1 preserves the bipotential progenitor cell pool.
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