Table 1.
Key studies on the effect of polymeric matrix stiffness on osteogenic differentiation of mesenchymal stem/progenitor cells.
| Study | Cell Source | Polymer | Modification | Modulus of Elasticity | Results |
|---|---|---|---|---|---|
| Alginate | |||||
| Zhang et al., 2020 [141] | hMSCs | Alginate–gelatin scaffold | 3D bioprinted porous scaffolds different alginate concentration (0.8%alg and 1.8%alg) and different initial cell seeding density (1.67, 5, and 15 M cells/mL) |
Soft scaffold 0.66 ± 0.08 kPa Stiff scaffold 5.4 ± 1.2 kPa |
UpregulatedALP-activity-related, 3D-bone-like-tissue-related, osteoblast-related, and early osteocyte-related gene expression |
| Freeman and Kelly, 2017 [142] | MSCs | Alginate hydrogel | 3D bioprinting matrix with varying alginate molecular weight and cross linker ratio | Osteogenic differentiation with increased ALP staining | |
| Maia et al., 2014 [143] | hMSCs | Alginate hydrogel | 3D matrix with bimodal molecular weight distribution at different polymer concentrations (1 and 2 wt.%) and RGD densities (0, 100 or 200 μM |
2 wt.% hydrogels (tan ∂ ᵙ 0.2), 1 wt.% hydrogels (tan ∂ ᵙ 0.4–0.6). |
1 wt.% alginate hydrogel matrices upregulated hMSCs osteogenic differentiation and expressed high levels of ALP and OCN |
| Collagen | |||||
| Xie et al., 2017 [146] | hMSCs | Collagen gel | Varying polymerization temperature 4, 21, and 37 °C. |
Fiber stiffness: 1.1 to 9.3 kPa Bulk stiffness: 16.4 to 151.5 Pa |
Collagen gel polymerized at 37 °C resulted in 34.1% ALP positive staining |
| Banks et al., 2014 [147] | ADSCs | Collagen–glycosaminoglycan (CG) | Chemical Crosslinking with EDAC and NHS Covalent immobilization of PDGF-BB and BMP-2 by benzophenone photolithography |
2.85 to 5 MPa | Upregulated expression of collagen 1, ALP, and OCN with increased stiffness |
| Hwang et al., 2019 [148] | hMSCs | Three bilayers of collagen/alginate nano film | 24 and 53 MPa | Increase in alkaline phosphatase activity | |
| Zhou et al., 2021 [149] | hMSCs | Nano-particulate mineralized collagen glycosaminoglycan |
Chemical crosslinking with EDAC and NHS | 3.90 −/+ 0.36 kPa | Increase in expression of ALP, collagen 1, and Runx2 |
| Tsimbouri et al., 2017 [150] | MSCs | Collagen gel | 3D collagen gel culture on the vibrational bioreactor | ~108 Pa | Increased expression of Runx2, collagen I, ALP, OPN, OCN, and BMP2. |
| Murphy et al., 2012 [151] | MSCs | Collagen/glycosaminoglycan | DHT and EDAC crosslinking | 0.5, 1, and 1.5 kPa | Osteogenic differentiation with Runx2 expression |
| Chen et al., 2015 [152] | Rat MSCs | 3D scaffold collagen and hydroxyapatite | Coated on decellularized cancellous bone | 13.00 ± 5.55 kPa, 13.87 ± 1.51 kPa, and 37.7 ± 19.6 kPa | Highest scaffold stiffness promoted higher expressions of OPN and OC |
| Chen et al., 2017 [205] | Rat MSCs | Collagen and hydroxyapatite, coated on decellularized cancellous bone | 3D oscillatory perfusion bioreactor system | 6.74 ± 1.16 kPa- 8.82 ± 2.12 kPa- 23.61 ± 8.06 kPa |
Osteogenic differentiation of MSCs |
| Gelatin | |||||
| Wan et al., 2019 [133] | PDLSCs | Gelatin | Crosslinked with variable concentrations of methacryloyl | GelMA concentrations of 10, 12, and 14 wt% stiffness 25.75 ± 1.21, 59.71 ± 8.87, and 117.82 ± 9.83 kPa, respectively | Increasing matrix stiffness increased osteogenic differentiation of PDLSCs, with upregulated expression of OCN and Runx2 |
| He et al., 2018 [134] | BMMSCs | Gelatin 3%, 6%, and 9%. | Crosslinked with transglutaminase | 9% gelatin gave rise to the highest stiffness (60.54 ± 10.45 kPa), while 3% gelatin resulted in the lowest stiffness (1.58 ± 0.42 kPa) | BMMSCs encapsulated in hydrogel with highest stiffness demonstrated the highest osteogenic differentiation |
| Van Nieuwenhove et al., 2017 [162] | ADSCs | Gelatin with variable degrees of methacrylation (GelMA 31%, GelMA 72%, and GelMA 95%) | Covalently bound to variable ratios of pentenoates modified starch (10 v% starch and 20 v% starch) | Increase in matrix stiffness promoted osteogenic differentiation of ADSCs | |
| Jiang et al., 2015 [163] | BMMSCs | GelMA encapsulating alendronate | Crosslinked by PEG diacrylate | stiffness increased from 4 to 40 kPa | Increased osteogenic differentiation of BMMSCs on stiffer hydrogel with higher alendronate concentration with upregulated ALP, collagen I, OCN, and calcium deposition |
| Sun et al., 2014 [164] | BMMSCs | Three-dimensional porous gelatin scaffolds | Crosslinked using EDC | Crosslinked scaffold demonstrated an increase in the elastic modulus from w 0.6 to ≈ 2.5 kP without any change in the scaffold internal structure | Increased stiffness increased osteogenic differentiation evidenced by increased Runx2 and OCN in vitro and increased bone formation in vivo |
| Decellularized matrix and Demineralized Bone | |||||
| Ventre et al., 2019 [165] | Murine MSCs | Decellularized MC3T3-E1-cell-derived matrix on replica from PDMS | Genipin crosslinking | Young’s modulus increased from (0.01–0.1 kPa) to (0.1–1.5 kPa). | MSCs on stiff dCDMs, revealed significant adipogenic and osteogenic differentiation potentials |
| Hu et al., 2018 [166] | BMMSC | Demineralized bone matrices | Controlling the decalcification duration (1 h, 12 h, and 5 d, respectively) | High: 66.06 ± 27.83 MPa, Medium: 26.90 ± 13.16 MPa Low: 0.67 ± 0.14 MPa |
Low stiffness scaffolds promoted osteogenesis in vitro. Subcutaneous implantation in a rat model and in a femoral condylar defect rabbit model revealed positive OCN and OPN expression |
| Hyaluronic acid (HA) | |||||
| Zhao et al., 2014 [174] | hBMMSCs | Thiol functionalized hyaluronic acid (HA) and thiol functionalized recombinant human gelatin | Crosslinked by poly (ethylene glycol) tetra-acrylate | 0.15, 1.5, and 4 kPa | Change in cell morphologies with different stiffness. Cells cultured on the 4 kPa hydrogel revealed an enhanced expression of late osteogenic genes |
| Cosgrove et al., 2016 [175] | Juvenile bovine MSCs | Methacrylated HA hydrogel | Ligation of the HAVDI adhesive peptide sequence from N-cadherin domain 1 and RGD from fibronectin | 5, 10, and 15 kPa | Lack of myosin IIA incorporated into focal adhesions hindered their maturation with increasing substrate stiffness and decreased osteogenesis |
| Dorcemus et al., 2017 [176] | hMSCs-bone-marrow-derived | Thiol-modified hyaluronan gel | Crosslinked by PEG at ratios ranging from 1:1 to 7:1 | Storage moduli from 10 to 45 Pa |
Differences between the top (cartilage-forming) and bottom (bone-forming) regions of the scaffold proved its capability for osteochondral engineering |
| Hao et al., 2018 [177] | hMSCs-bone-marrow-derived | HA carrying sulfhydryl groups and a hydrophilic polymer bearing both acrylate and tetrazine groups | Matrix metalloprotease -degradable peptidic crosslinker and adding HA conjugated with multiple copies of trans-cyclooctene (TCO) | (G’) = 180 ± 42 Pa increased to G′ = 520 ± 80 Pa | The 3D matrix tagged with a TCO- motif promoted the cells to undergo change from a rounded to spindle phenotype |
| Fibrin | |||||
| Hashemzadeh et al., 2019 [180] | hADSCs | Fibrin hydrogels embedding gold nanowires | Altering fibrinogen and thrombin concentration and incorporation of gold nanowires | With high fibrinogen and thrombin concentration, gold nanowires, promoted osteogenic differentiation | |
| Polyethylene glycol (PEG) | |||||
| Pek et al., 2010 [182] | MSCs | Thixotropic polyethylene glycol–silica (PEG–silica) nano composite gel | 3D cell culture Cell-adhesion peptide RGD (Arg–Gly–Asp) sequence immobilization |
≥75 Pa | Higher expression of the osteogenic transcription factor |
| Ye et al., 2015 [183] | Rat BMMSCs | PEG | PEG hydrogels with RGD nano-spacings of 49 and 135 nm and incubated in mixed osteogenic and adipogenic medium | Soft hydrogels (130 kPa) and stiff hydrogels (3170 kPa) | Stiff hydrogels promoted osteogenesis. Large RGD nano-spacing promoted osteogenesis |
| Steinmetz et al., 2015 [184] | hMSCs | Multilayer PEG-based hydrogel | Simple sequential photopolymerization- high RGD concentrations- dynamic mechanical stimulation | 345 kPa | Collagen I generation with mineral deposits were evident |
| Yang et al., 2020 [185] | Rat BMMSCs | PEG/silk fibroin/HA scaffold | Varying HA concentration | 80.98 to 190.51 kPa | Expression of all the osteogenesis-related markers in vitro and superior calvarial defect repair in vivo |
| Yang et al., 2016 [186] | hMSCs | PEG hydrogel | Regularly and randomly patterned photodegradable hydrogel | ∼10–12 kPa | Osteogenic differentiation of MSCs cultured on random patterns |
| Gandavarapu et al., 2014 [187] | hMSCs | PEG hydrogels | functionalized with c(RRETAWA) hydrogels through α5 integrins | ∼25 kPa | Osteogenic differentiation of hMSCs |
| Polydimethylsiloxane (PDMS) | |||||
| Xie et al., 2018 [38] | ASCs | PDMS | 1.014 ± 0.15 MPa | Osteogenic differentiation by ALP stain and upregulation of Runx2 and Osx transcriptional factors | |
| Viale-Bouroncle et al., 2014 [189] | DFCs | PDMS | Coating PDMS with fibronectin and cultured in osteogenic differentiation medium | 11 kPa | High ALP activity and accumulation of calcium on the soft substrate |
| Viale-Bouroncle et al., 2012 [190] | SHED | PDMS | Adding osteogenic differentiation medium | 93 kPa | High osteogenic differentiation |
| Wang et al., 2012 [191] | Rat MSCs | PDMS | Osteogenic medium with temperature gradient curing | 0.19 to 3.10 MPa | Calcein Blue–positive bone-nodule-like colonies |
| Vinyl polymers | |||||
| Khoramgah et al., 2020 [192] | hADSCs | Poly tetra fluoro ethylene (PTFE) and PVA with and without graphene oxide nanoparticles | 3D porous scaffolds- chemical crosslinking with small amounts of boric acids–controlled freeze-drying method | 620 and 130 kPa | Elevation in ALP activity, calcium deposition, and osteogenic-related genes expression |
| Oh et al., 2016 [193] | hBMMSCs | Cylindrical PVA/HA hydrogel | Liquid nitrogen—contacting gradual freezing–thawing method | ~20 kPa and ~200 kPa | Stiffness of ~190 kPa led to osteoblast differentiation |
| Polyesters | |||||
| Sun et al., 2019 [195] | hADSCs | Poly(ether-ester-urethane) (PEEU) containing PPDO and PCL segments | Electrospun into fiber meshes with varying PPDO to PCL weight ratios | 2.6 ± 0.8 MPa (PEEU40), 3.2 ± 0.9 MPa (PEEU50), 4.0 ± 0.9 MPa (PEEU60) 4.5 ± 0.8 MPa (PEEU70) |
Enhanced osteogenic differentiation of hADSCs with higher levels of OCN, ALP, and hydroxyapatite detected on the stiffer fiber meshes |
| Self-assembling peptides | |||||
| Hogrebe and Gooch, 2016 [203] | hMSCs | Biomimetic self-assembling peptide hydrogel containing 1 mg/mL RGD-functionalized peptide (KFE–RGD) | hMSCs were encapsulated within 3D culture and grown on top of 2D culture Adding 1:1 mixed adipogenic/osteogenic induction medium |
(G′) 10 kPa | Osteogenesis induction and alizarin red-stained calcium deposits |
| Other Polymers | |||||
| Olivares-Navarrete et al., 2017 [76] | MSCs | Methyl acrylate/methyl methacrylate polymer | Altering monomer concentration. | 0.1 MPa to 310 MPa | Chondrogenic and osteogenic differentiation when grown on substrates with less than 10 MPa stiffness |
| Wu et al., 2018 [204] | hBMMSCs | Poly(urea-urethane) (PUU)/POSS elastomeric nano-hybrid scaffolds | Thermoresponsive scaffolds indirectly 3D printed by inverse self-assembling | >10 kPa | Osteogenic differentiation |