Microarray Analysis of Human Adipose-Derived Stem Cells in Three-Dimensional Collagen Culture: Osteogenesis Inhibits Bone Morphogenic Protein and Wnt Signaling Pathways, and Cyclic Tensile Strain Causes Upregulation of Proinflammatory Cytokine Regulators and Angiogenic Factors

Abstract

Human adipose-derived stem cells (hASC) have shown great potential for bone tissue engineering. However, the molecular mechanisms underlying this potential are not yet known, in particular the separate and combined effects of three-dimensional (3D) culture and mechanical loading on hASC osteogenesis. Mechanical stimuli play a pivotal role in bone formation, remodeling, and fracture repair. To further understand hASC osteogenic differentiation and response to mechanical stimuli, gene expression profiles of proliferating or osteogenically induced hASC in 3D collagen I culture in the presence and absence of 10% uniaxial cyclic tensile strain were examined using microarray analysis. About 847 genes and 95 canonical pathways were affected during osteogenesis of hASC in 3D culture. Pathway analysis indicated the potential roles of Wnt/β-catenin signaling, bone morphogenic protein (BMP) signaling, platelet-derived growth factor (PDGF) signaling, and insulin-like growth factor 1 (IGF-1) signaling in hASC during osteogenic differentiation. Application of 10% uniaxial cyclic tensile strain suggested synergistic effects of strain with osteogenic differentiation media on hASC osteogenesis as indicated by significantly increased calcium accretion of hASC. There was no significant further alteration in the four major pathways (Wnt/β-catenin, BMP, PDGF, and IGF-1). However, 184 transcripts were affected by 10% cyclic tensile strain. Function and network analysis of these transcripts suggested that 10% cyclic tensile strain may play a role during hASC osteogenic differentiation by upregulating two crucial factors in bone regeneration: (1) proinflammatory cytokine regulators interleukin 1 receptor antagonist and suppressor of cytokine signaling 3; (2) known angiogenic inductors fibroblast growth factor 2, matrix metalloproteinase 2, and vascular endothelial growth factor A. This is the first study to investigate the effects of both 3D culture and mechanical load on hASC osteogenic differentiation. A complete microarray analysis investigating both the separate effect of soluble osteogenic inductive factors and the combined effects of chemical and mechanical stimulation was performed on hASC undergoing osteogenic differentiation. We have identified specific genes and pathways associated with mechanical response and osteogenic potential of hASC, thus providing significant information toward improved understanding of our use of hASC for functional bone tissue engineering applications.

Reduced serum corin levels in patients with osteoporosis

“…Recent findings of increased corin mRNA expression in bone marrow-or adipose tissuederived hMSCs that underwent osteogenic differentiation are intriguing [36,37]. We confirmed these findings in osteogenic and chondrogenic differentiation assays using human bone marrow-derived hMSCs (data not shown).…”

“…By in situ hybridization and RT-PCR, corin mRNA was detected in prehypertrophic chondrocytes and perichondrocytes in mouse developing bones and human osteosarcoma-derived cells [12]. More recently, human mesenchymal stem cells (hMSCs) from bone marrow and adipose tissues also were found to express corin mRNAs [36,37]. When the cultured hMSCs underwent osteogenic differentiation, corin mRNA expression was highly up-regulated, suggesting a potential role of corin in regulating mesenchymal stem cell differentiation in chondrogenesis and/or osteogenesis.…”

Age-Related Effects on the Potency of Human Adipose-Derived Stem Cells: Creation and Evaluation of Superlots and Implications for Musculoskeletal Tissue Engineering Applications

“…In the future it may be necessary to develop a classification system to identify predictors of differentiation potential. Although some work has been done profiling hASC using cell surface markers, 6,27 microarray surveys of gene expression, 28,29 and microRNA profiles, 12 these data have not been extensively correlated with hASC potency. It is important to note that the ages and number of donor cell lines incorporated in this study were based on the availability of donor cell lines within our laboratory bank of cells.…”

Primary cilium mechanotransduction of tensile strain in 3D culture: Finite element analyses of strain amplification caused by tensile strain applied to a primary cilium embedded in a collagen matrix

“…We recently reported their critical role in lineage specification of human adipose-derived stem cells (hASC) (Bodle et al, 2013) which can differentiate into musculoskeletal tissue similarly to MSC (Mauney et al, 2007; Ugarte et al, 2003; Weinzierl et al, 2006). We and others have previously shown that hASC exhibit mechanosensitivity with changes in proliferation and lineage specification in response to many types of mechanical stimuli (Charoenpanich et al, 2011; Hanson et al, 2009; McCullen et al, 2010; Park et al, 2011; Puetzer et al, 2013; Schatti et al, 2011; Wall et al, 2007). In particular, cyclic tensile strain, whether applied to hASC in monolayer (two-dimensional) or three-dimensional collagen culture, promotes hASC osteogenesis and cell-mediated calcium accretion (Deiderichs et al, 2010; Hanson et al, 2009; Wall et al, 2007).…”