Enhanced Osteogenic Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells by a Hybrid Hydroxylapatite/Collagen Scaffold

Abstract

Human bone marrow-derived mesenchymal stem cells (hBMSCs) and their derivative enhanced green fluorescent protein (eGFP)-hBMSCs were employed to evaluate an innovative hybrid scaffold composed of granular hydroxylapatite and collagen hemostat (Coll/HA). The cellular morphology/cytoskeleton organization and cell viability were investigated by immunohistochemistry (IHC) and AlamarBlue metabolic assay, respectively. The expression of osteopontin and osteocalcin proteins was analyzed by IHC and ELISA, whereas osteogenic genes were investigated by quantitative PCR (Q-PCR). Cell morphology of eGFP-hBMSCs was indistinguishable from that of parental hBMSCs. The cytoskeleton architecture of hBMSCs grown on the scaffold appeared to be well organized, whereas its integrity remained uninfluenced by the scaffold during the time course. Metabolic activity measured in hBMSCs grown on a biomaterial was increased during the experiments, up to day 21 (p < 0.05). The biomaterial induced the matrix mineralization in hBMSCs. The scaffold favored the expression of osteogenic proteins, such as osteocalcin and osteopontin. In hBMSC cultures, the scaffold induced up-regulation in specific genes that are involved in ossification process (BMP2/3, SPP1, SMAD3, and SP7), whereas they showed an up-regulation of MMP9 and MMP10, which play a central role during the skeletal development. hBMSCs were induced to chondrogenic differentiation through up-regulation of COL2A1 gene. Our experiments suggest that the innovative scaffold tested herein provides a good microenvironment for hBMSC adhesion, viability, and osteoinduction. hBMSCs are an excellent in vitro cellular model to assay scaffolds, which can be employed for bone repair and bone tissue engineering.

MicroRNAs Modulate Signaling Pathways in Osteogenic Differentiation of Mesenchymal Stem Cells

“…Biomaterials provide the biological structure that supports MSC osteogenic differentiation [2,[11][12][13][14][15][16][17][18]. The addition of ions has been proved to enhance the osteogenic potential of scaffolds [19].…”

Impact of Experimental Parameters on Cell–Cell Force Spectroscopy Signature

“…Delay time: different extended pauses (1,15,30,45, 60, and 120 s) were studied. The cell probe was lowered at a constant speed of 10.0 µm/s until the cell probe contacted the target cell and the preset force (setpoint force) of 1.0 nN was reached.…”

“…The use of large colloidal or flat probes in indentation experiments allows the determination of the mechanical properties of a cell, mediating the result on a large contact area [12]. At the same time, the use of a sharp tip as indenter allows detection of local changes of mechanical properties [13,14] as well as investigation of single-molecule unfolding events [15]. The stiffer cytoskeletal filaments network [16] can be characterized by AFM, as well as the softer nuclear compartment [17].…”

Long Non-coding RNAs and MicroRNAs Interplay in Osteogenic Differentiation of Mesenchymal Stem Cells

“…MSCs are promising competent biomedical candidates in the management of several bone disorders, as their use-availability (Iaquinta et al, 2019a;Mazzoni et al, 2020a). Indeed, MSCs own several important biological properties, such as the capacity to secrete molecules that can induce tissue regeneration, selfrenewal and proliferation, as well as multipotentiality, antiinflammatory and immunomodulatory effects (Pearson et al, 2015;Batta et al, 2016;Iaquinta et al, 2019a;Globig et al, 2020;Mazzoni et al, 2020b). Isolated for the first time from the bone marrow (BM) (Bianco et al, 2008), MSCs have been subsequently harvested from many different anatomical regions, such as adipose tissue (ASCs) (Mahmoudifar and Doran, 2015;Mazzoni et al, 2020a), umbilical cord (UC-MSCs) (Galderisi and Giordano, 2014), dental pulp tissues (DPSCs) (Ferro et al, 2014), and others (Fei et al, 2013;Ferro et al, 2014;Gong et al, 2014;Fierabracci et al, 2015).…”

“…cerebrospinal fluid (Cai and Cullen, 2007;Hunter et al, 2008;Gallo et al, 2012;Moldovan et al, 2013;Rotondo et al, 2013;Bononi et al, 2016;Corral-Vazquez et al, 2017;Huang et al, 2018). Furthermore, miRNAs are also being studied as biomarkers of pathophysiological conditions (Bononi et al, 2016;Hoey et al, 2019) and as targets for innovative therapeutic approaches in precision medicine (Matin et al, 2016;Mazzoni et al, 2020b). Osteogenic differentiation can be regulated by several miRNAs both positively and negatively.…”