Carbon Nanotubes Induced Fibrogenesis on Nanostructured Substrates.

While the rapidly evolving nanotechnology has shown promise in electronics, energy, healthcare and many other fields, there is an increasing concern about the adverse health consequences of engineered nanomaterials. To accurately evaluate the toxicity of nanomaterials, in vitro models incorporated with in vivo microenvironment characteristics are desirable. This study aims to delineate the influence of nanotopography on fibrogenic response of normal human lung fibroblasts to multi-walled carbon nanotubes (MWCNTs). Nanoscale gratings and pillars of various heights were fabricated on polydimethylsiloxane substrates. Cell spreading and biomechanics were measured, and fibrogenic responses including proliferation, collagen production and reactive oxygen species generation of the fibroblasts grown on the nanostructured substrates in response to MWCNTs were assessed. It was observed that the cells could be largely stretched on shallow nanogratings, leading to stiffer cytoskeleton and nucleus, enhanced cell proliferation and collagen production, and consequently, toxic response sensitivity of the fibroblasts was undermined. In contrast, the cell spreading and stiffness could be reduced using tall, isotropic nanopillars, which significantly improved the cell toxic sensitivity to the MWCNTs. In addition to highlighting the significant influence of cell-nanotopography interactions on cell sensing CNTs, this study contributed to development of physiologically relevant in vitro models for nanotoxicology study.

Nanotopographical Modulation of Cell Function through Nuclear Deformation.

Although nanotopography has been shown to be a potent modulator of cell behavior, it is unclear how the nanotopographical cue, through focal adhesions, affects the nucleus, eventually influencing cell phenotype and function. Thus, current methods to apply nanotopography to regulate cell behavior are basically empirical. We, herein, engineered nanotopographies of various shapes (gratings and pillars) and dimensions (feature size, spacing and height), and thoroughly investigated cell spreading, focal adhesion organization and nuclear deformation of human primary fibroblasts as the model cell grown on the nanotopographies. We examined the correlation between nuclear deformation and cell functions such as cell proliferation, transfection and extracellular matrix protein type I collagen production. It was found that the nanoscale gratings and pillars could facilitate focal adhesion elongation by providing anchoring sites, and the nanogratings could orient focal adhesions and nuclei along the nanograting direction, depending on not only the feature size but also the spacing of the nanogratings. Compared with continuous nanogratings, discrete nanopillars tended to disrupt the formation and growth of focal adhesions and thus had less profound effects on nuclear deformation. Notably, nuclear volume could be effectively modulated by the height of nanotopography. Further, we demonstrated that cell proliferation, transfection, and type I collagen production were strongly associated with the nuclear volume, indicating that the nucleus serves as a critical mechanosensor for cell regulation. Our study delineated the relationships between focal adhesions, nucleus and cell function and highlighted that the nanotopography could regulate cell phenotype and function by modulating nuclear deformation. This study provides insight into the rational design of nanotopography for new biomaterials and the cell-substrate interfaces of implants and medical devices.

Three-Dimensional Microfluidic Tri-Culture Model of the Bone Marrow Microenvironment for Study of Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) initiates and progresses in the bone marrow, and as such, the marrow microenvironment is a critical regulatory component in development of this cancer. However, ALL studies were conducted mainly on flat plastic substrates, which do not recapitulate the characteristics of marrow microenvironments. To study ALL in a model of in vivo relevance, we have engineered a 3-D microfluidic cell culture platform. Biologically relevant populations of primary human bone marrow stromal cells, osteoblasts and human leukemic cells representative of an aggressive phenotype were encapsulated in 3-D collagen matrix as the minimal constituents and cultured in a microfluidic platform. The matrix stiffness and fluidic shear stress were controlled in a physiological range. The 3-D microfluidic as well as 3-D static models demonstrated coordinated cell-cell interactions between these cell types compared to the compaction of the 2-D static model. Tumor cell viability in response to an antimetabolite chemotherapeutic agent, cytarabine in tumor cells alone and tri-culture models for 2-D static, 3-D static and 3-D microfluidic models were compared. The present study showed decreased chemotherapeutic drug sensitivity of leukemic cells in 3-D tri-culture models from the 2-D models. The results indicate that the bone marrow microenvironment plays a protective role in tumor cell survival during drug treatment. The engineered 3-D microfluidic tri-culture model enables systematic investigation of effects of cell-cell and cell-matrix interactions on cancer progression and therapeutic intervention in a controllable manner, thus improving our limited comprehension of the role of microenvironmental signals in cancer biology.