Role of RHEB in Regulating Differentiation Fate of Mesenchymal Stem Cells for Cartilage and Bone Regeneration.

Sajjad Ashraf,Soo-Hong Lee,Hansoo Park,In-Bo Han

doi:10.3390/ijms18040880

Abstract

Advances in mesenchymal stem cells (MSCs) and cell replacement therapies are promising approaches to treat cartilage and bone defects since substantial differentiation capacities of MSCs match the demands of tissue regeneration. Our understanding of the dynamic process requiring indispensable differentiation of MSCs remains limited. Herein, we describe the role of RHEB (Ras homolog enriched in brain) regulating gene signature for differentiation of human adipose derived mesenchymal stem cells (ASCs) into chondrogenic, osteogenic, and adipogenic lineages. RHEB-overexpression increases the proliferation of the ASCs. RHEB enhances the chondrogenic differentiation of ASCs in 3D culture via upregulation of SOX9 with concomitant increase in glycosaminoglycans (GAGs), and type II collagen (COL2). RHEB increases the osteogenesis via upregulation of runt related transcription factor 2 (RUNX2) with an increase in the calcium and phosphate contents. RHEB also increases the expression of osteogenic markers, osteonectin and osteopontin. RHEB knockdown ASCs were incapable of expressing sufficient SRY (Sex determining region Y)-box 9 (SOX9) and RUNX2, and therefore had decreased chondrogenic and osteogenic differentiation. RHEB-overexpression impaired ASCs differentiation into adipogenic lineage, through downregulation of CCAAT/enhancer binding protein beta (C/EBPβ). Conversely, RHEB knockdown abolished the negative regulation of adipogenesis. We demonstrate that RHEB is a novel regulator, with a critical role in ASCs lineage determination, and RHEB-modulated ASCs may be useful as a cell therapy for cartilage and bone defect treatments.

Highlights

Multipotent mesenchymal stem cells (MSCs) can potentially differentiate into multiple distinct cell lineages, including chondrogenic, osteogenic, and adipogenic [1]
We investigated whether Ras homolog enriched in brain (RHEB) plays any role in differentiation of the Adipose-derived stem cells (ASCs) into chondrogenic as well as osteogenic cells that are essential for cartilage and bone regeneration
We have demonstrated that RHEB plays a crucial role in the differentiation of ASCs, wherein it promotes chondrogenic and osteogenic differentiation but impairs adipogenesis

Summary

Introduction

Multipotent mesenchymal stem cells (MSCs) can potentially differentiate into multiple distinct cell lineages, including chondrogenic, osteogenic, and adipogenic [1]. Since their original discovery in the bone marrow, multiple additional tissue sources, including adipose depots, have been reported to contain MSCs. Adipose tissue in particular is an attractive source of MSCs because it is readily obtainable with minimal morbidity via routine liposuction procedures [2,3]. Multiple factors influence the differentiation potential of ASCs into distinct lineages, including the composition of differentiation media, the presence of growth factors, and the substrate properties of the culture plate [6,7]. The induction of ASCs differentiation into the chondrogenic lineage is usually promoted by a cocktail of Dex, Transforming growth factor (TGF-β1), ascorbic acid, and insulin-transferrin-selenium (ITS) [7,9]. The induction of ASC differentiation by numerous factors has been investigated, most studies focus only on differentiation into individual lineages rather than simultaneous chondrogenic, osteogenic, and adipogenic differentiation

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Discussion

Conclusion