J Bone Metab.  2018 Feb;25(1):43-51. 10.11005/jbm.2018.25.1.43.

The Effect of Antidepressants on Mesenchymal Stem Cell Differentiation

• Kruk JS
• Bermeo S
• Skarratt K
• Fuller SJ
• Duque G

• KMID: 2406238
• DOI: http://doi.org/10.11005/jbm.2018.25.1.43

Abstract

BACKGROUND
Use of antidepressant medications has been linked to detrimental impacts on bone mineral density and osteoporosis; however, the cellular basis behind these observations remains poorly understood. The effect does not appear to be homogeneous across the whole class of drugs and may be linked to affinity for the serotonin transporter system. In this study, we hypothesized that antidepressants have a class- and dose-dependent effect on mesenchymal stem cell (MSC) differentiation, which may affect bone metabolism.
METHODS
Human MSCs (hMSCs) were committed to differentiate when either adipogenic or osteogenic media was added, supplemented with five increasing concentrations of amitriptyline (0.001-10 µM), venlafaxine (0.01-25 µM), or fluoxetine (0.001-10 µM). Alizarin red staining (mineralization), alkaline phosphatase (osteoblastogenesis), and oil red O (adipogenesis) assays were performed at timed intervals. In addition, cell viability was assessed using a MTT.
RESULTS
We found that fluoxetine had a significant inhibitory effect on mineralization. Furthermore, adipogenic differentiation of hMSC was affected by the addition of amitriptyline, venlafaxine, and fluoxetine to the media. Finally, none of the tested medications significantly affected cell survival.
CONCLUSIONS
This study showed a divergent effect of three antidepressants on hMSC differentiation, which appears to be independent of class and dose. As fluoxetine and amitriptyline, but not venlafaxine, affected both osteoblastogenesis and adipogenesis, this inhibitory effect could be associated to the high affinity of fluoxetine to the serotonin transporter system.

Keyword

Adipogenesis; Antidepressive agents; Mesenchymal stromal cells; Osteoblasts; Osteoporosis

MeSH Terms

Adipogenesis
Alkaline Phosphatase
Amitriptyline
Antidepressive Agents*
Bone Density
Cell Survival
Fluoxetine
Humans
Mesenchymal Stromal Cells*
Metabolism
Miners
Osteoblasts
Osteoporosis
Serotonin Plasma Membrane Transport Proteins
Venlafaxine Hydrochloride
Alkaline Phosphatase
Amitriptyline
Antidepressive Agents
Fluoxetine
Serotonin Plasma Membrane Transport Proteins
Venlafaxine Hydrochloride

Figure

• 

  Fig. 1 Effect of antidepressants on mineralization. Human mesenchymal stem cells (MSCs) were induced to differentiate into osteoblasts for 3 weeks at 37℃ in the presence of various concentrations of (A, B) amitriptyline, (C, D) venlafaxine, or (E, F) fluoxetine. Differentiating cells were also incubated with vehicle (VEH) alone, while undifferentiated cells were incubated with MSC growth media (GM) for 3 weeks as controls. Cells were then fixed and stained with Alizarin red (AR) as described in the methods. Changes in absorbance were measured relative to VEH only control. The extent of AR staining was quantified by solubilizing the stain with cetylpyridinium chloride and measuring the absorbance. Ratios were obtained by comparing absorbances with that of VEH. (A, B) Co-incubation with amitriptyline showed a modest decrease in AR staining only at the highest concentration tested, 10 µM. (C, D) Whereas co-incubation with venlafaxine showed no significant change in staining, (E, F) the addition of fluoxetine decreased staining at all concentrations tested. Undifferentiated hMSC showed no AR staining (GM bars). Data are representative of 3 to 6 independent experiments per drug. Images were taken under ×10 magnification. *P<0.05 compared to VEH-treated cells.

• 

  Fig. 2 Effect of antidepressants on alkaline phosphatase activity. Alkaline phosphatase activity was measured in human mesenchymal stem cells (MSCs) that had been induced to differentiate into osteoblasts over 3 weeks at 37℃ in the presence of various concentrations of (A) amitriptyline, (B) venlafaxine, or (C) fluoxetine. Differentiating cells were also incubated with vehicle (VEH) alone, while undifferentiated cells were incubated with MSC growth media (GM) for 3 weeks as controls. Alkaline phosphatase activity is displayed relative to VEH control. Data are representative of 3 to 6 independent experiments per drug. *P<0.05 compared to VEH-treated cells.

• 

  Fig. 3 Effect of antidepressants on adipogenesis. Human mesenchymal stem cells were induced to differentiate into adipocytes for 2 weeks at 37℃ in the presence of various concentrations of (A, B) amitriptyline, (C, D) venlafaxine, or (E, F) fluoxetine. Differentiating cells were also incubated with vehicle (VEH), while undifferentiated cells were incubated with MSC growth media (GM) for 2 weeks as controls. Cells were then fixed, stained with oil red O (ORO) and counterstained with hematoxylin as described in the methods. The extent of staining was then quantified by solubilizing the stain as described in the methods. Intensity of staining was determined by absorbance relative to VEH. Data are representative of 3 to 6 independent experiments per drug. Images were taken under ×10 magnification. *P<0.05 compared to VEH-treated cells.

• 

  Fig. 4 Assessment of cell viability. Human mesenchymal stem cells (MSC) induced to differentiate into osteoblasts (alizarin red [AR]) or adipocytes (oil red O [ORO]) in the presence of various concentrations of amitriptyline (Ami), venlafaxine (Ven), or fluoxetine (Fl). The Figure shows the effect of increasing concentrations of the drugs on cell viability. No significant effect was found at any treated conditions. Cells cultured in MSC growth media (GM) were used as control. *P<0.05 compared to differentiation media. VEH, vehicle.

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