Volume Adaptation Controls Stem Cell Mechanotransduction

Luke G. Major; Andrew W. Holle; Jennifer L. Young; Matt S. Hepburn; Kwanghee Jeong; Ian L. Chin; Rowan W. Sanderson; Ji Hoon Jeong; Zachary M. Aman; Brendan F. Kennedy; Yongsung Hwang; Dong Wook Han; Hyun Woo Park; Kun Liang Guan; Joachim P. Spatz; Yu Suk Choi

doi:10.1021/acsami.9b19770

Volume Adaptation Controls Stem Cell Mechanotransduction

Luke G. Major, Andrew W. Holle, Jennifer L. Young, Matt S. Hepburn, Kwanghee Jeong, Ian L. Chin, Rowan W. Sanderson, Ji Hoon Jeong, Zachary M. Aman, Brendan F. Kennedy, Yongsung Hwang, Dong Wook Han, Hyun Woo Park, Kun Liang Guan, Joachim P. Spatz, Yu Suk Choi

Research output: Contribution to journal › Article › peer-review

43 Citations (Scopus)

Abstract

Recent studies have found discordant mechanosensitive outcomes when comparing 2D and 3D, highlighting the need for tools to study mechanotransduction in 3D across a wide spectrum of stiffness. A gelatin methacryloyl (GelMA) hydrogel with a continuous stiffness gradient ranging from 5 to 38 kPa was developed to recapitulate physiological stiffness conditions. Adipose-derived stem cells (ASCs) were encapsulated in this hydrogel, and their morphological characteristics and expression of both mechanosensitive proteins (Lamin A, YAP, and MRTFa) and differentiation markers (PPARγand RUNX2) were analyzed. Low-stiffness regions (â&circ;¼8 kPa) permitted increased cellular and nuclear volume and enhanced mechanosensitive protein localization in the nucleus. This trend was reversed in high stiffness regions (â&circ;¼30 kPa), where decreased cellular and nuclear volumes and reduced mechanosensitive protein nuclear localization were observed. Interestingly, cells in soft regions exhibited enhanced osteogenic RUNX2 expression, while those in stiff regions upregulated the adipogenic regulator PPARÎ³, suggesting that volume, not substrate stiffness, is sufficient to drive 3D stem cell differentiation. Inhibition of myosin II (Blebbistatin) and ROCK (Y-27632), both key drivers of actomyosin contractility, resulted in reduced cell volume, especially in low-stiffness regions, causing a decorrelation between volume expansion and mechanosensitive protein localization. Constitutively active and inactive forms of the canonical downstream mechanotransduction effector TAZ were stably transfected into ASCs. Activated TAZ resulted in higher cellular volume despite increasing stiffness and a consistent, stiffness-independent translocation of YAP and MRTFa into the nucleus. Thus, volume adaptation as a function of 3D matrix stiffness can control stem cell mechanotransduction and differentiation.

Original language	English
Pages (from-to)	45520-45530
Number of pages	11
Journal	ACS Applied Materials and Interfaces
Volume	11
Issue number	49
DOIs	https://doi.org/10.1021/acsami.9b19770
Publication status	Published - 2019 Dec 11

Bibliographical note

Funding Information:
L.G.M., A.W.H., and J.L.Y. contributed equally to this work. L.G.M., A.W.H., J.L.Y., and YS.C. designed and planned the study. J.L.Y., A.W.H., and J.P.S. performed SEM and pore size analysis. M.S.H., R.W.S., and B.F.K. performed OCE and stress relaxation analysis. K.J. and Z.M.A. performed Raman spectroscopy. J.H.J., Y.H., and D.-W.H. synthesized GelMA. H.W.P. and K-L. G. generated TAZ overexpressed cells. L.G.M., A.W.H., J.L.Y., and YS.C. drafted the manuscript. Y.S.C provided supervision and funding. This study work was supported by National Health and Medical Research Council Grant PG1098449 (to Y.S.C and K.-L.G), Heart Foundation Future Leader Fellowship 101173 (to Y.S.C), Department of Health, Western Australia, Merit awards–project and fellowship (to Y.S.C), Universities Australia DAAD German Research Cooperation 5744610 (to Y.S.C, I.L.C, A.W.H, J.L.Y, and J.P.S), and UWA RIGG (to Y.S.C and HW.P). The authors declare no competing financial interest.

Publisher Copyright:
© 2019 American Chemical Society.

All Science Journal Classification (ASJC) codes

Materials Science(all)

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10.1021/acsami.9b19770

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Major, L. G., Holle, A. W., Young, J. L., Hepburn, M. S., Jeong, K., Chin, I. L., Sanderson, R. W., Jeong, J. H., Aman, Z. M., Kennedy, B. F., Hwang, Y., Han, D. W., Park, H. W., Guan, K. L., Spatz, J. P., & Choi, Y. S. (2019). Volume Adaptation Controls Stem Cell Mechanotransduction. ACS Applied Materials and Interfaces, 11(49), 45520-45530. https://doi.org/10.1021/acsami.9b19770

@article{cb78dab9a5a349e989e8bac9ddb2bb77,

title = "Volume Adaptation Controls Stem Cell Mechanotransduction",

abstract = "Recent studies have found discordant mechanosensitive outcomes when comparing 2D and 3D, highlighting the need for tools to study mechanotransduction in 3D across a wide spectrum of stiffness. A gelatin methacryloyl (GelMA) hydrogel with a continuous stiffness gradient ranging from 5 to 38 kPa was developed to recapitulate physiological stiffness conditions. Adipose-derived stem cells (ASCs) were encapsulated in this hydrogel, and their morphological characteristics and expression of both mechanosensitive proteins (Lamin A, YAP, and MRTFa) and differentiation markers (PPARγand RUNX2) were analyzed. Low-stiffness regions ({\^a}&circ;¼8 kPa) permitted increased cellular and nuclear volume and enhanced mechanosensitive protein localization in the nucleus. This trend was reversed in high stiffness regions ({\^a}&circ;¼30 kPa), where decreased cellular and nuclear volumes and reduced mechanosensitive protein nuclear localization were observed. Interestingly, cells in soft regions exhibited enhanced osteogenic RUNX2 expression, while those in stiff regions upregulated the adipogenic regulator PPAR{\^I}³, suggesting that volume, not substrate stiffness, is sufficient to drive 3D stem cell differentiation. Inhibition of myosin II (Blebbistatin) and ROCK (Y-27632), both key drivers of actomyosin contractility, resulted in reduced cell volume, especially in low-stiffness regions, causing a decorrelation between volume expansion and mechanosensitive protein localization. Constitutively active and inactive forms of the canonical downstream mechanotransduction effector TAZ were stably transfected into ASCs. Activated TAZ resulted in higher cellular volume despite increasing stiffness and a consistent, stiffness-independent translocation of YAP and MRTFa into the nucleus. Thus, volume adaptation as a function of 3D matrix stiffness can control stem cell mechanotransduction and differentiation.",

author = "Major, {Luke G.} and Holle, {Andrew W.} and Young, {Jennifer L.} and Hepburn, {Matt S.} and Kwanghee Jeong and Chin, {Ian L.} and Sanderson, {Rowan W.} and Jeong, {Ji Hoon} and Aman, {Zachary M.} and Kennedy, {Brendan F.} and Yongsung Hwang and Han, {Dong Wook} and Park, {Hyun Woo} and Guan, {Kun Liang} and Spatz, {Joachim P.} and Choi, {Yu Suk}",

note = "Funding Information: L.G.M., A.W.H., and J.L.Y. contributed equally to this work. L.G.M., A.W.H., J.L.Y., and YS.C. designed and planned the study. J.L.Y., A.W.H., and J.P.S. performed SEM and pore size analysis. M.S.H., R.W.S., and B.F.K. performed OCE and stress relaxation analysis. K.J. and Z.M.A. performed Raman spectroscopy. J.H.J., Y.H., and D.-W.H. synthesized GelMA. H.W.P. and K-L. G. generated TAZ overexpressed cells. L.G.M., A.W.H., J.L.Y., and YS.C. drafted the manuscript. Y.S.C provided supervision and funding. This study work was supported by National Health and Medical Research Council Grant PG1098449 (to Y.S.C and K.-L.G), Heart Foundation Future Leader Fellowship 101173 (to Y.S.C), Department of Health, Western Australia, Merit awards–project and fellowship (to Y.S.C), Universities Australia DAAD German Research Cooperation 5744610 (to Y.S.C, I.L.C, A.W.H, J.L.Y, and J.P.S), and UWA RIGG (to Y.S.C and HW.P). The authors declare no competing financial interest. Publisher Copyright: {\textcopyright} 2019 American Chemical Society.",

year = "2019",

month = dec,

day = "11",

doi = "10.1021/acsami.9b19770",

language = "English",

volume = "11",

pages = "45520--45530",

journal = "ACS applied materials & interfaces",

issn = "1944-8244",

publisher = "American Chemical Society",

number = "49",

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T1 - Volume Adaptation Controls Stem Cell Mechanotransduction

AU - Major, Luke G.

AU - Holle, Andrew W.

AU - Young, Jennifer L.

AU - Hepburn, Matt S.

AU - Jeong, Kwanghee

AU - Chin, Ian L.

AU - Sanderson, Rowan W.

AU - Jeong, Ji Hoon

AU - Aman, Zachary M.

AU - Kennedy, Brendan F.

AU - Hwang, Yongsung

AU - Han, Dong Wook

AU - Park, Hyun Woo

AU - Guan, Kun Liang

AU - Spatz, Joachim P.

AU - Choi, Yu Suk

N1 - Funding Information: L.G.M., A.W.H., and J.L.Y. contributed equally to this work. L.G.M., A.W.H., J.L.Y., and YS.C. designed and planned the study. J.L.Y., A.W.H., and J.P.S. performed SEM and pore size analysis. M.S.H., R.W.S., and B.F.K. performed OCE and stress relaxation analysis. K.J. and Z.M.A. performed Raman spectroscopy. J.H.J., Y.H., and D.-W.H. synthesized GelMA. H.W.P. and K-L. G. generated TAZ overexpressed cells. L.G.M., A.W.H., J.L.Y., and YS.C. drafted the manuscript. Y.S.C provided supervision and funding. This study work was supported by National Health and Medical Research Council Grant PG1098449 (to Y.S.C and K.-L.G), Heart Foundation Future Leader Fellowship 101173 (to Y.S.C), Department of Health, Western Australia, Merit awards–project and fellowship (to Y.S.C), Universities Australia DAAD German Research Cooperation 5744610 (to Y.S.C, I.L.C, A.W.H, J.L.Y, and J.P.S), and UWA RIGG (to Y.S.C and HW.P). The authors declare no competing financial interest. Publisher Copyright: © 2019 American Chemical Society.

PY - 2019/12/11

Y1 - 2019/12/11

N2 - Recent studies have found discordant mechanosensitive outcomes when comparing 2D and 3D, highlighting the need for tools to study mechanotransduction in 3D across a wide spectrum of stiffness. A gelatin methacryloyl (GelMA) hydrogel with a continuous stiffness gradient ranging from 5 to 38 kPa was developed to recapitulate physiological stiffness conditions. Adipose-derived stem cells (ASCs) were encapsulated in this hydrogel, and their morphological characteristics and expression of both mechanosensitive proteins (Lamin A, YAP, and MRTFa) and differentiation markers (PPARγand RUNX2) were analyzed. Low-stiffness regions (â&circ;¼8 kPa) permitted increased cellular and nuclear volume and enhanced mechanosensitive protein localization in the nucleus. This trend was reversed in high stiffness regions (â&circ;¼30 kPa), where decreased cellular and nuclear volumes and reduced mechanosensitive protein nuclear localization were observed. Interestingly, cells in soft regions exhibited enhanced osteogenic RUNX2 expression, while those in stiff regions upregulated the adipogenic regulator PPARÎ³, suggesting that volume, not substrate stiffness, is sufficient to drive 3D stem cell differentiation. Inhibition of myosin II (Blebbistatin) and ROCK (Y-27632), both key drivers of actomyosin contractility, resulted in reduced cell volume, especially in low-stiffness regions, causing a decorrelation between volume expansion and mechanosensitive protein localization. Constitutively active and inactive forms of the canonical downstream mechanotransduction effector TAZ were stably transfected into ASCs. Activated TAZ resulted in higher cellular volume despite increasing stiffness and a consistent, stiffness-independent translocation of YAP and MRTFa into the nucleus. Thus, volume adaptation as a function of 3D matrix stiffness can control stem cell mechanotransduction and differentiation.

AB - Recent studies have found discordant mechanosensitive outcomes when comparing 2D and 3D, highlighting the need for tools to study mechanotransduction in 3D across a wide spectrum of stiffness. A gelatin methacryloyl (GelMA) hydrogel with a continuous stiffness gradient ranging from 5 to 38 kPa was developed to recapitulate physiological stiffness conditions. Adipose-derived stem cells (ASCs) were encapsulated in this hydrogel, and their morphological characteristics and expression of both mechanosensitive proteins (Lamin A, YAP, and MRTFa) and differentiation markers (PPARγand RUNX2) were analyzed. Low-stiffness regions (â&circ;¼8 kPa) permitted increased cellular and nuclear volume and enhanced mechanosensitive protein localization in the nucleus. This trend was reversed in high stiffness regions (â&circ;¼30 kPa), where decreased cellular and nuclear volumes and reduced mechanosensitive protein nuclear localization were observed. Interestingly, cells in soft regions exhibited enhanced osteogenic RUNX2 expression, while those in stiff regions upregulated the adipogenic regulator PPARÎ³, suggesting that volume, not substrate stiffness, is sufficient to drive 3D stem cell differentiation. Inhibition of myosin II (Blebbistatin) and ROCK (Y-27632), both key drivers of actomyosin contractility, resulted in reduced cell volume, especially in low-stiffness regions, causing a decorrelation between volume expansion and mechanosensitive protein localization. Constitutively active and inactive forms of the canonical downstream mechanotransduction effector TAZ were stably transfected into ASCs. Activated TAZ resulted in higher cellular volume despite increasing stiffness and a consistent, stiffness-independent translocation of YAP and MRTFa into the nucleus. Thus, volume adaptation as a function of 3D matrix stiffness can control stem cell mechanotransduction and differentiation.

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JO - ACS applied materials & interfaces

JF - ACS applied materials & interfaces

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ER -

Volume Adaptation Controls Stem Cell Mechanotransduction

Abstract

Bibliographical note

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