A preliminary comparison of two different polyacrylamide hydrogel fabrication methods demonstrate differences in stiffness measurements and adhesion abilities of osteosarcoma cells
Main Article Content
Abstract
The purpose of this research was to compare two different polyacrylamide hydrogel fabrication
methods described in the literature and assess their use in mechanotransduction studies in osteosarcoma. Both methods employ succinimide chemistry to functionalize the hydrogel surface for cell response studies, one in the form of NHS, while the other in the form of Sulfo-SANPAH.
Six hydrogels of two different stiffness were created for each method and were evaluated on their receptiveness to cell seeding and Young’s moduli with atomic force microscopy. Both hydrogel fabrication methods lack reproducibility as significant differences were observed in stiffness measurements between six hydrogels at both 0.5 kPa and 50 kPa stiffnesses. Despite the Sulfo- SANPAH method of preparation being more receptive to cell seeding and generating the expected effects on known mechanotransducers, it appeared to have larger variabilities in stiffness measurements for both 0.5 kPa and 50 kPa prepared hydrogels. Researchers may employ the Sulfo-SANPAH method to study the impact of ECM of varying stiffness on osteosarcoma cell mechanotransduction, but should remain cautious when interpreting results as a function of the expected stiffness. Studies should be accompanied by measurements of Young's modulus whenever possible.
Article Details
The Medical Research Archives grants authors the right to publish and reproduce the unrevised contribution in whole or in part at any time and in any form for any scholarly non-commercial purpose with the condition that all publications of the contribution include a full citation to the journal as published by the Medical Research Archives.
References
2. Chin L, Xia Y, Discher DE, Janmey PA. Mechanotransduction in cancer. Current Opinion in Chemical Engineering. 2016 Feb 1;11:77-84.
3. Jaalouk DE, Lammerding J. Mechanotransduction gone awry. Nature Reviews Molecular Cell Biology. 2009 Jan;10(1):63-73.
4. Varelas X, Wrana JL. Coordinating developmental signaling: novel roles for the Hippo pathway. Trends in Cell Biology. 2012 Feb 1;22(2):88-96.
5. Varelas X. The Hippo pathway effectors TAZ and YAP in development, homeostasis and disease. Development. 2014 Apr 15;141(8):1614-26.
6. Warren JS, Xiao Y, Lamar JM. YAP/TAZ activation as a target for treating metastatic cancer. Cancers. 2018 Apr;10(4):115.
7. Zucchini C, Manara MC, Cristalli C, Carrabotta M, Greco S, Pinca RS, Ferrari C, Landuzzi L, Pasello M, Lollini PL, Gambarotti M. ROCK2 deprivation leads to the inhibition of tumor growth and metastatic potential in osteosarcoma cells through the modulation of YAP activity. Journal of Experimental & Clinical Cancer Research. 2019 Dec;38(1):1-4.
8. Luu AK, Schott CR, Jones R, Poon AC, Golding B, Hamed R, Deheshi B, Mutsaers A, Wood GA, Viloria-Petit AM. An evaluation of TAZ and YAP crosstalk with TGFβ signalling in canine osteosarcoma suggests involvement of hippo signalling in disease progression. BMC Veterinary Research. 2018 Dec;14(1):1-22.
9. Luu AK, Viloria-Petit AM. Targeting Mechanotransduction in Osteosarcoma: A Comparative Oncology Perspective. International Journal of Molecular Sciences. 2020 Jan;21(20):7595.
10. Fischer RS, Myers KA, Gardel ML, Waterman CM. Stiffness-controlled three-dimensional extracellular matrices for high-resolution imaging of cell behavior. Nature Protocols. 2012 Nov;7(11):2056-66.
11. Cretu A, Castagnino P, Assoian R. Studying the effects of matrix stiffness on cellular function using acrylamide-based hydrogels. Journal of visualized experiments: JoVE. 2010(42).
12. Minaisah RM, Cox S, Warren DT. The use of polyacrylamide hydrogels to study the effects of matrix stiffness on nuclear envelope properties. In The Nuclear Envelope 2016 (pp. 233-239). Humana Press, New York, NY.
13. Kandow CE, Georges PC, Janmey PA, Beningo KA. Polyacrylamide hydrogels for cell mechanics: steps toward optimization and alternative uses. Methods in Cell Biology. 2007 Jan 1;83:29-46.
14. Dupont S, Morsut L, Aragona M, Enzo E, Giulitti S, Cordenonsi M, Zanconato F, Le Digabel J, Forcato M, Bicciato S, Elvassore N. Role of YAP/TAZ in mechanotransduction. Nature. 2011 Jun;474(7350):179-83.
15. Dupont S. Role of YAP/TAZ in cell-matrix adhesion-mediated signalling and mechanotransduction. Experimental Cell Research. 2016 Apr 10;343(1):42-53.
16. Sunyer R, Jin AJ, Nossal R, Sackett DL. Fabrication of hydrogels with steep stiffness gradients for studying cell mechanical response. PLOS One. 2012 Oct 4.