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Controlling osteogenic stem cell differentiation via soft bioinspired hydrogels.

Jha AK, Jackson WM, Healy KE - PLoS ONE (2014)

Bottom Line: Among these factors, modulus (i.e., rigidiy) of the ECM has gained significant attention as a physical osteoinductive signal that can contribute to endochondral ossification of a cartilaginous skeletal template.To further understand the role of the matrix interactions in this process, we evaluated osteogenic differentiation of hMSCs cultured on low moduli (102, 390 or 970 Pa) poly(N-isopropylacrylamide) (p(NIPAAm)) based semi-interpenetrating networks (sIPN) modified with the integrin engaging peptide bsp-RGD(15) (0, 105 or 210 µM).These findings suggest that within a compliant and low modulus substrate, a high affinity adhesive ligand serves as a substitute for a rigid matrix to foster osteogenic differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, University of California, Berkeley, California, United States of America.

ABSTRACT
Osteogenic differentiation of human mesenchymal stem cells (hMSCs) is guided by various physical and biochemical factors. Among these factors, modulus (i.e., rigidiy) of the ECM has gained significant attention as a physical osteoinductive signal that can contribute to endochondral ossification of a cartilaginous skeletal template. However, MSCs also participate in intramembranous bone formation, which occurs de novo from within or on a more compliant tissue environment. To further understand the role of the matrix interactions in this process, we evaluated osteogenic differentiation of hMSCs cultured on low moduli (102, 390 or 970 Pa) poly(N-isopropylacrylamide) (p(NIPAAm)) based semi-interpenetrating networks (sIPN) modified with the integrin engaging peptide bsp-RGD(15) (0, 105 or 210 µM). Cell adhesion, proliferation, and osteogenic differentiation of hMSCs, as measured by alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), bone sialoprotein-2 (iBSP), and osteocalcien (OCN) protein expression, was highest on substrates with the highest modulus and peptide concentrations. However, within this range of substrate stiffness, many osteogenic cellular functions were enhanced by increasing either the modulus or the peptide density. These findings suggest that within a compliant and low modulus substrate, a high affinity adhesive ligand serves as a substitute for a rigid matrix to foster osteogenic differentiation.

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Cell adhesion and spreading on the sIPN hydrogels.(a) hMSCs were capable of adhering and spreading on the sIPNs containing the bsp-RGD(15) adhesive ligand, as assessed by f-actin stress fibers (TRITC-phalloidin; red) and nuclei (DAPI; blue). Scale bar  = 100 µm. RSM plots demonstrating the effect of sIPN stiffness (102–970 Pa) and bsp-RGD(15) adhesive ligand density (0–210 µM) on: (b) hMSC attachment after 24 hours relative to cells seeded on TCPS; (c) and area of hMSC spreading after 3 days.
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pone-0098640-g002: Cell adhesion and spreading on the sIPN hydrogels.(a) hMSCs were capable of adhering and spreading on the sIPNs containing the bsp-RGD(15) adhesive ligand, as assessed by f-actin stress fibers (TRITC-phalloidin; red) and nuclei (DAPI; blue). Scale bar  = 100 µm. RSM plots demonstrating the effect of sIPN stiffness (102–970 Pa) and bsp-RGD(15) adhesive ligand density (0–210 µM) on: (b) hMSC attachment after 24 hours relative to cells seeded on TCPS; (c) and area of hMSC spreading after 3 days.

Mentions: All of the sIPNs substrates supported hMSC viability regardless of their material parameters (Figure S1), although cell morphology (i.e., spreading) was strongly dependent on the matrix parameters (Figure 2a). Cell attachment to the substrate within 3 days of seeding was significantly lower for sIPNs with 102 Pa stiffness and 0 µM bsp-RGD(15) relative to any other matrix parameters (Figure S2a), and cell attachment appeared to correlate with both the stiffness and bsp-RGD(15) concentration in all the other sIPNs (Figure 2b). Cells exhibited primarily a rounded morphology on the substrates with either the lowest bsp-RGD(15) concentration (0 µM) or stiffness (102 Pa). Cell spreading areas on these sIPNs was negligible (Figure 2c) and significantly lower than for the other sIPNs (Figure S2b). Importantly, cell spreading on 3 of the matrices (210 µM and 970Pa; 210 µM and 390Pa; and 105 µM and 970 Pa) was higher than cell spreading on TCPS, and the greatest cell spreading was observed on matrices with the highest bsp-RGD(15) ligand density (210 µM) and stiffness (970 Pa) (Figure. S2b, S3).


Controlling osteogenic stem cell differentiation via soft bioinspired hydrogels.

Jha AK, Jackson WM, Healy KE - PLoS ONE (2014)

Cell adhesion and spreading on the sIPN hydrogels.(a) hMSCs were capable of adhering and spreading on the sIPNs containing the bsp-RGD(15) adhesive ligand, as assessed by f-actin stress fibers (TRITC-phalloidin; red) and nuclei (DAPI; blue). Scale bar  = 100 µm. RSM plots demonstrating the effect of sIPN stiffness (102–970 Pa) and bsp-RGD(15) adhesive ligand density (0–210 µM) on: (b) hMSC attachment after 24 hours relative to cells seeded on TCPS; (c) and area of hMSC spreading after 3 days.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4060996&req=5

pone-0098640-g002: Cell adhesion and spreading on the sIPN hydrogels.(a) hMSCs were capable of adhering and spreading on the sIPNs containing the bsp-RGD(15) adhesive ligand, as assessed by f-actin stress fibers (TRITC-phalloidin; red) and nuclei (DAPI; blue). Scale bar  = 100 µm. RSM plots demonstrating the effect of sIPN stiffness (102–970 Pa) and bsp-RGD(15) adhesive ligand density (0–210 µM) on: (b) hMSC attachment after 24 hours relative to cells seeded on TCPS; (c) and area of hMSC spreading after 3 days.
Mentions: All of the sIPNs substrates supported hMSC viability regardless of their material parameters (Figure S1), although cell morphology (i.e., spreading) was strongly dependent on the matrix parameters (Figure 2a). Cell attachment to the substrate within 3 days of seeding was significantly lower for sIPNs with 102 Pa stiffness and 0 µM bsp-RGD(15) relative to any other matrix parameters (Figure S2a), and cell attachment appeared to correlate with both the stiffness and bsp-RGD(15) concentration in all the other sIPNs (Figure 2b). Cells exhibited primarily a rounded morphology on the substrates with either the lowest bsp-RGD(15) concentration (0 µM) or stiffness (102 Pa). Cell spreading areas on these sIPNs was negligible (Figure 2c) and significantly lower than for the other sIPNs (Figure S2b). Importantly, cell spreading on 3 of the matrices (210 µM and 970Pa; 210 µM and 390Pa; and 105 µM and 970 Pa) was higher than cell spreading on TCPS, and the greatest cell spreading was observed on matrices with the highest bsp-RGD(15) ligand density (210 µM) and stiffness (970 Pa) (Figure. S2b, S3).

Bottom Line: Among these factors, modulus (i.e., rigidiy) of the ECM has gained significant attention as a physical osteoinductive signal that can contribute to endochondral ossification of a cartilaginous skeletal template.To further understand the role of the matrix interactions in this process, we evaluated osteogenic differentiation of hMSCs cultured on low moduli (102, 390 or 970 Pa) poly(N-isopropylacrylamide) (p(NIPAAm)) based semi-interpenetrating networks (sIPN) modified with the integrin engaging peptide bsp-RGD(15) (0, 105 or 210 µM).These findings suggest that within a compliant and low modulus substrate, a high affinity adhesive ligand serves as a substitute for a rigid matrix to foster osteogenic differentiation.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioengineering, University of California, Berkeley, California, United States of America.

ABSTRACT
Osteogenic differentiation of human mesenchymal stem cells (hMSCs) is guided by various physical and biochemical factors. Among these factors, modulus (i.e., rigidiy) of the ECM has gained significant attention as a physical osteoinductive signal that can contribute to endochondral ossification of a cartilaginous skeletal template. However, MSCs also participate in intramembranous bone formation, which occurs de novo from within or on a more compliant tissue environment. To further understand the role of the matrix interactions in this process, we evaluated osteogenic differentiation of hMSCs cultured on low moduli (102, 390 or 970 Pa) poly(N-isopropylacrylamide) (p(NIPAAm)) based semi-interpenetrating networks (sIPN) modified with the integrin engaging peptide bsp-RGD(15) (0, 105 or 210 µM). Cell adhesion, proliferation, and osteogenic differentiation of hMSCs, as measured by alkaline phosphatase (ALP), runt-related transcription factor 2 (RUNX2), bone sialoprotein-2 (iBSP), and osteocalcien (OCN) protein expression, was highest on substrates with the highest modulus and peptide concentrations. However, within this range of substrate stiffness, many osteogenic cellular functions were enhanced by increasing either the modulus or the peptide density. These findings suggest that within a compliant and low modulus substrate, a high affinity adhesive ligand serves as a substitute for a rigid matrix to foster osteogenic differentiation.

Show MeSH
Related in: MedlinePlus