Effect of α-pinene and thymoquinone on the differentiation of bone marrow mesenchymal stem cells into neuroprogenitor cells

Abstract

Introduction: Neurodegenerative diseases are accompanied by loss of neuronal function and integrity. Stem cell therapy is utilized to regenerate neurons to repair the damaged area. Regeneration potential of stem cells can be enhanced by using chemicals with known bioactive properties. In the current study, two bioactive compounds, α-pinene (AP) and thymoquinone (TQ) were explored for their neuronal differentiation potential of rat bone marrow mesenchymal stem cells (MSCs). Methods: MSCs were isolated, cultured and characterized immunocytochemically for the presence of specific surface markers. Optimized concentrations of both compounds (20 µM AP and 12 µM TQ) as determined by MTT assay, were used to treat MSCs in separate and combined groups. All groups were assessed for the presence of neuronal, astroglial, and germ layer markers through qPCR. Neuronal and glial protein expression were analyzed by immunocytochemistry. Results: Both compounds alone and in combination induced differentiation in MSCs with significant gene expression of neuronal markers i.e. neuron specific enolase (NSE), nestin, microtubule-associated protein 2 (MAP2), neurofilament light chain (Nefl) and Tau, and astroglial marker i.e. glial fibrillary acidic protein (GFAP). AP treated group also showed significant upregulation of endodermal and mesodermal markers indicating transition of ectoderm towards the other two germ layers. Conclusion: This study concludes that AP and TQ potentially differentiate MSCs into neuronal and astroglial lineages. However, AP treated group followed germ layer transition. Expression of neuronal as well as glial markers indicate that the differentiated neurons are at the neuroprogenitor stage and can be potential candidates for cellular therapeutics against neurodegenerative disorders.

Alpha terpineol directs bone marrow mesenchymal stem cells toward neuronal lineage through regulation of wnt signaling pathway

“…There was significant upregulation of neuron and glial specific markers that is, NSE, Tau, Nestin, NefL, MAP2 , and GFAP . Similar study has been conducted previously by our group that reported the expression of astroglial marker that is, GFAP along with the neuronal markers in differentiated MSCs 25 . Some other studies also reported the expression of GFAP in differentiated neuronal cells 26 .…”

“…However, genes of endoderm ( AFP, Sox17 , and MixL1 ) and mesoderm ( MesP1 and T Brachyury ) layers were also upregulated. At this stage, cells expressed neuronal markers representing their neuroprogenitor state, but overexpression of genes representing meso and endodermal layers simply explains the transition phenomenon from ectoderm to the other two germ layers, as reported in our previous study 25 …”

“…previously by our group that reported the expression of astroglial marker that is, GFAP along with the neuronal markers in differentiated MSCs. 25 Some other studies also reported the expression of GFAP in differentiated neuronal cells. 26 The differentiated cells were also assessed for neuronal protein expression which showed positive expression of GFAP, β-III tubulin, NeuN, and synaptophysin.…”

“…At this stage, cells expressed neuronal markers representing their neuroprogenitor state, but overexpression of genes representing meso and endodermal layers simply explains the transition phenomenon from ectoderm to the other two germ layers, as reported in our previous study. 25 We also determined the developmental state of differentiated in due course. DCX1, Stathmin and NeuroD1 are involved in various biological processes like neuronal proliferation, survival, and maturation.…”

“…Our previous study also reported that monoterpenes (alpha pinene and thymoquinone) induce differentiation of MSCs toward neuroprogenitor cells. 25 To understand the mechanism of neuronal differentiation, we analyzed signaling pathway, which plays an important role in the regulation of cell proliferation and differentiation. We observed activation of Wnt signaling after 30 min of treatment, at which point MSCs remained undifferentiated or in other words MSCs did not start differentiating toward neuronal lineage.…”