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Published on: January 31, 2019
Forskolin enhances in vivo bone formation by human mesenchymal stromal cells
Joyce Doorn1, Ramakrishnaiah Siddappa, Clemens A van Blitterswijk
1Department of Tissue Regeneration, MIRA Institute for Biomedical Technology and Technical Medicine, University of Twente, Enschede, The Netherlands.
Abstract:
Activation of the protein kinase A (PKA) pathway with dibutyryl cyclic adenosine monophosphate (db-cAMP) was recently shown to enhance osteogenic differentiation of human mesenchymal stromal cells (hMSCs) in vitro and bone formation in vivo. The major drawback of this compound is its inhibitory effect on proliferation of hMSCs. Therefore, we investigated whether fine-tuning of the dose and timing of PKA activation could enhance bone formation even further, with minimum effects on proliferation. To test this, we selected two different PKA activators (8-bromo-cAMP (8-br-cAMP) and forskolin) and compared their effects on proliferation and osteogenic differentiation with those of db-cAMP. We found that all three compounds induced alkaline phosphatase levels, bone-specific target genes, and secretion of insulin-like growth factor-1, although 8-br-cAMP induced adipogenic differentiation in long-term cultures and was thus considered unsuitable for further in vivo testing. All three compounds inhibited proliferation of hMSCs in a dose-dependent manner, with forskolin inhibiting proliferation most. The effect of forskolin on in vivo bone formation was tested by pretreating hMSCs before implantation, and we observed greater amounts of bone using forskolin than db-cAMP. Our data show forskolin to be a novel agent that can be used to increase bone formation and also suggests a role for PKA in the delicate balance between adipogenic and osteogenic differentiation.
Insights
Forskolin enhances bone formation by activating the protein kinase A (PKA) pathway in human mesenchymal stromal cells (hMSCs). This approach improves bone regeneration with minimal impact on cell proliferation, offering a promising strategy for bone repair.
Area of Science:
- Biomedical Engineering
- Cell Biology
- Regenerative Medicine
Background:
- Protein kinase A (PKA) activation enhances osteogenic differentiation and bone formation.
- Dibutyryl cyclic adenosine monophosphate (db-cAMP) treatment boosts bone formation but inhibits human mesenchymal stromal cell (hMSC) proliferation.
- Optimizing PKA activation is crucial for maximizing bone formation while minimizing adverse effects on cell growth.
Purpose of the Study:
- To investigate if fine-tuning the dose and timing of PKA activation can further enhance bone formation with minimal effects on hMSC proliferation.
- To compare the effects of different PKA activators (8-bromo-cAMP (8-br-cAMP) and forskolin) with db-cAMP on hMSC proliferation and osteogenic differentiation.
Main Methods:
- Compared db-cAMP, 8-br-cAMP, and forskolin for their effects on hMSC proliferation and osteogenic differentiation markers.
- Assessed alkaline phosphatase levels, bone-specific gene expression, and insulin-like growth factor-1 secretion.
- Evaluated the in vivo bone formation capacity of forskolin-pretreated hMSCs following implantation.
Main Results:
- All tested compounds (db-cAMP, 8-br-cAMP, forskolin) induced osteogenic differentiation markers.
- 8-br-cAMP induced adipogenic differentiation in long-term cultures, making it unsuitable for in vivo studies.
- Forskolin demonstrated superior bone formation in vivo compared to db-cAMP, despite showing the most significant inhibition of hMSC proliferation in a dose-dependent manner.
Conclusions:
- Forskolin is a novel agent for increasing bone formation via PKA pathway activation.
- PKA plays a key role in balancing adipogenic and osteogenic differentiation in hMSCs.
- Optimized PKA activation strategies hold potential for enhanced bone regeneration therapies.
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