Engineering Complex Microenvironments to Manipulate and Study Cellular Mechanobiology

Links

URL: https://repository.upenn.edu/edissertations/2239/

PROQUEST: 1952167467

Abstract

Our improved understanding of cell mechano-sensing has been enabled by developments in biomaterials platforms and in vitro culture systems. These emerging systems model key aspects of the cellular niche in a controlled fashion, enabling one to ask and answer questions that could not be addressed in vivo due to the complex interactions of these multiple discrete signals. These emerging materials include key features of the native microenvironment, including complex mechanical properties, degradable domains, and native-scale feature sizes. In this thesis work, we designed a 2D methacrylated hyaluronic acid hydrogel (MeHA) system to recapitulate the multiple adhesive interactions normally present in the developing mesenchymal progenitor cell niche, including an RGD adhesive motif (from fibronectin) and the HAVDI adhesive motif (from N-Cadherin). This system allowed for the decoupled presentation of cell-ECM interactions and cell-cell interactions while simultaneously modulating other variables present in the microenvironment, such as changes in substrate stiffness. Using this system, we elucidated a new signaling pathway through which HAVDI ligation altered MSC mechanosensing of ECM stiffness, which culminated in downstream differences in cellular proliferation and differentiation. Our findings suggest that HAVDI presentation may be harnessed towards novel biomaterial design to direct cellular behavior and allow for tuning of response to substrate stiffness in regenerative medicine applications. Building from this platform, we further investigated nuclear morphology of MSCs on these 2D MeHA substrates and in native tissue, and showed that nuclear morphology and mechanotransduction were markedly different on planar substrates compared to within the native tissue niche. These findings suggested that dimensionality and the manner by which nuclear pre-stress is established in 3D regulates strain transmission to the nucleus. To better recapitulate these interactions, we cultured cells in 3D MMP-degradable HA hydrogels, and showed that these remodelable systems more accurately recapitulated the in vivo cellular response in vitro. Moving forward, work with these defined synthetic 3D niches will play a critical role in understanding of complex processes governing stem cell development and differentiation, as well as disease pathologies that arise when these processes become disrupted.

Cited Works (11)

Year	Entry
2005	Burdick JA, Chung C, Jia X, Randolph MA, Langer R. Controlled degradation and mechanical behavior of photopolymerized hyaluronic acid networks. Biomacromolecules. January–February 2005;6(1):386-391.
1997	Maniotis AJ, Chen CS, Ingber DE. Demonstration of mechanical connections between integrins, cytoskeletal filaments, and nucleoplasm that stabilize nuclear structure. Proc Natl Acad Sci USA. February 1997;94(3):849-854.
1997	Pelham RJ Jr, Wang Y-l. Cell locomotion and focal adhesions are regulated by substrate flexibility. Proc Natl Acad Sci USA. December 9, 1997;94(25):13661-13665.
2004	McBeath R, Pirone DM, Nelson CM, Bhadriraju K, Chen CS. Cell shape, cytoskeletal tension, and RhoA regulate stem cell lineage commitment. Dev Cell. April 2004;6(4):483-495.
2003	Hersel U, Dahmen C, Kessler H. RGD modified polymers: biomaterials for stimulated cell adhesion and beyond. Biomaterials. November 2003;24(24):4385-4415.
2006	Shore EM, Xu M, Feldman GJ, Fenstermacher DA, Cho T-J, Choi IH, Connor JM, Delai P, Glaser DL, LeMerrer M, Morhart R, Rogers JG, Smith R, Triffitt JT, Urtizberea JA, Zasloff M, Brown MA, Kaplan FS. A recurrent mutation in the BMP type I receptor ACVR1 causes inherited and sporadic fibrodysplasia ossificans progressiva. Nat Genet. May 2006;38(5):525-528.
2002	Kalajzic I, Kalajzic Z, Kaliterna M, Gronowicz G, Clark SH, Lichtler AC, Rowe D. Use of type I collagen green fluorescent protein transgenes to identify subpopulations of cells at different stages of the osteoblast lineage. J Bone Miner Res. January 2002;17(1):15-25.
1994	Skaggs DL, Warden WH, Mow VC. Radial tie fibers influence the tensile properties of the bovine medial meniscus. J Orthop Res. March 1994;12(2):176-185.
1997	Chen CS, Mrksich M, Huang S, Whitesides GM, Ingber DE. Geometric control of cell life and death. Science. May 30, 1997;276(5317):1425.
2006	Engler AJ, Sen S, Sweeney HL, Discher DE. Matrix elasticity directs stem cell lineage specification. Cell. August 25, 2006;126(4):P677-P689.
2003	Lutolf MP, Lauer-Fields JL, Schmoekel HG, Metters AT, Weber FE, Fields GB, Hubbell JA. Synthetic matrix metalloproteinase-sensitive hydrogels for the conduction of tissue regeneration: engineering cell-invasion characteristics. Proc Natl Acad Sci USA. April 29, 2003;100(9):5413-5418.