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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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Relative osteogenesis on the various sIPNs.(a) Quantitative analysis of ALP activity, and (b) RUNX2, iBSP and OCN protein expression on various sIPNs. *p<0.05, ANOVA and Tukey post-hoc tests with n = 3 sIPNs.
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pone-0098640-g005: Relative osteogenesis on the various sIPNs.(a) Quantitative analysis of ALP activity, and (b) RUNX2, iBSP and OCN protein expression on various sIPNs. *p<0.05, ANOVA and Tukey post-hoc tests with n = 3 sIPNs.

Mentions: Thereafter, progression of osteogenic differentiation was assessed by quantification of ALP activity, protein expression analysis of the osteogenic transcriptional regulator RUNX2, and the secreted non-collagenous ostegenic matrix proteins iBSP and OCN [47], [48], [49]. Our results are consistent with previous studies showing maximal ALP expression between days 7 and 10 [50], [51]. sIPNs with 210 µM peptide densities and 970 Pa matrix stiffness yielded the greatest ALP expression, but sIPNs containing either of the higher matrix parameters enhanced ALP expression relative to those containing both the low (105 µM) bsp-RGD(15) peptide density and the low (390 Pa) matrix stiffness (Figure 5a). Fourteen days after the start of osteogenic differentiation, the highest expression of RUNX2 and iBSP was observed on the matrix containing the highest (210 µM) bsp-RGD(15) peptide density and the highest (970 Pa) matrix stiffness, and a similar, lower expression level was observed on the other sIPNs (Figure 5b). By contrast, the expression of OCN on all of the sIPNs was approximately the same for all sIPNs after 14 days, except for the matrix containing the lowest bsp-RGD(15) concentration (105 µM) and matrix stiffness (390 Pa). We attribute this observation to the proteins iBSP and RUNX2 being upregulated during early bone formation, whereas OCN is expressed at the later time point of osteogenesis [52].


Controlling osteogenic stem cell differentiation via soft bioinspired hydrogels.

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

Relative osteogenesis on the various sIPNs.(a) Quantitative analysis of ALP activity, and (b) RUNX2, iBSP and OCN protein expression on various sIPNs. *p<0.05, ANOVA and Tukey post-hoc tests with n = 3 sIPNs.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0098640-g005: Relative osteogenesis on the various sIPNs.(a) Quantitative analysis of ALP activity, and (b) RUNX2, iBSP and OCN protein expression on various sIPNs. *p<0.05, ANOVA and Tukey post-hoc tests with n = 3 sIPNs.
Mentions: Thereafter, progression of osteogenic differentiation was assessed by quantification of ALP activity, protein expression analysis of the osteogenic transcriptional regulator RUNX2, and the secreted non-collagenous ostegenic matrix proteins iBSP and OCN [47], [48], [49]. Our results are consistent with previous studies showing maximal ALP expression between days 7 and 10 [50], [51]. sIPNs with 210 µM peptide densities and 970 Pa matrix stiffness yielded the greatest ALP expression, but sIPNs containing either of the higher matrix parameters enhanced ALP expression relative to those containing both the low (105 µM) bsp-RGD(15) peptide density and the low (390 Pa) matrix stiffness (Figure 5a). Fourteen days after the start of osteogenic differentiation, the highest expression of RUNX2 and iBSP was observed on the matrix containing the highest (210 µM) bsp-RGD(15) peptide density and the highest (970 Pa) matrix stiffness, and a similar, lower expression level was observed on the other sIPNs (Figure 5b). By contrast, the expression of OCN on all of the sIPNs was approximately the same for all sIPNs after 14 days, except for the matrix containing the lowest bsp-RGD(15) concentration (105 µM) and matrix stiffness (390 Pa). We attribute this observation to the proteins iBSP and RUNX2 being upregulated during early bone formation, whereas OCN is expressed at the later time point of osteogenesis [52].

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