The cell seeding density and spatial distribution in a 3-D scaffold are critical to the morphogenetic development of an engineered tissue. A dynamic depth-filtration seeding method was developed to improve the initial cell seeding density and spatial distribution in 3-D nonwoven fibrous matrices commonly used as tissue scaffolds. In this work, trophoblast-like ED27 cells were seeded in poly(ethylene terephthalate) (PET) matrices with various porosities (0.85-0.93). The effects of the initial concentration of cells in the suspension used to seed the PET matrix and the pore size of the matrix on the resulting seeding density and subsequent cell proliferation and tissue development were studied. Compared to the conventional static seeding method, the dynamic depth-filtration seeding method gave a significantly higher initial seeding density (2-4 x 10(7) vs 4 x 10(6) cells/cm3), more uniform cell distribution, and a higher final cell density in the tissue scaffold. The more uniform initial cell spatial distribution from the filtration seeding method also led to more cells in S phase and a prolonged proliferation period. However, both uniform spatial cell distribution and the pore size of the matrices are important to cell proliferation and morphological development in the seeded tissue scaffold. Large-pore matrices led to the formation of cell aggregates and thus might reduce cell proliferation. The dynamic depth-filtration seeding method is better in providing a higher initial seeding density and more uniform cell distribution and is easier to apply to large tissue scaffolds. A depth-filtration model was also developed and can be used to simulate the seeding process and to predict the maximum initial seeding densities in matrices with different porosities.

Myoferlin (MYOF) is a member of the evolutionarily conserved ferlin family of proteins, noted for their role in a variety of membrane processes, including endocytosis, repair, and vesicular transport. Notably, ferlins are implicated in Caenorhabditis elegans sperm motility (Fer-1), mammalian skeletal muscle development and repair (MYOF and dysferlin), and presynaptic transmission in the auditory system (otoferlin). In this paper, we demonstrate that MYOF plays a previously unrecognized role in cancer cell invasion, using a combination of mathematical modeling and in vitro experiments. Using a real-time impedance-based invasion assay (xCELLigence), we have shown that lentiviral-based knockdown of MYOF significantly reduced invasion of MDA-MB-231 breast cancer cells in Matrigel bioassays. Based on these experimental data, we developed a partial differential equation model of MYOF effects on cancer cell invasion, which we used to generate mechanistic hypotheses. The mathematical model predictions revealed that matrix metalloproteinases (MMPs) may play a key role in modulating this invasive property, which was supported by experimental data using quantitative RT-PCR screens. These results suggest that MYOF may be a promising target for biomarkers or drug target for metastatic cancer diagnosis and therapy, perhaps mediated through MMPs.cancer invasion | RNAi | partial differential equation models | metastasis A majority of cancer deaths are related not to the primary tumor itself, but rather the formation of disseminated metastases (1). Cancer spread requires that cells achieve atypical motility, which enables them to invade surrounding tissues and vessels of the blood and lymphatic systems (2-4). Thus, understanding the mechanisms and signaling processes that lead to invasive cell behavior may lead to new therapeutic approaches for controlling and treating cancer.The fundamental mechanisms of invasive cancer cell movement are largely conserved across a wide range of cell types, with some of the protease dependent and protease independent movement types demonstrated by cancer cells also seen in organisms as diverse as unicellular organisms, slime molds, and white blood cells. The ferlin family is an evolutionarily ancient family of proteins (5), which are known to affect processes crucial to migration and invasion, including membrane fusion and repair, vesicle transport, endocytosis, protein recycling and stability, and cell motility (6-13). Thus, one might expect the ferlin family to be good candidates for cancer proteins, although they have not previously been investigated in this capacity. In Caenorhabditis elegans, spermatozoa exhibit amoeboid movement, and mutations in the fer-1 gene [an orthologue of myoferlin (MYOF)] result in immobility and infertility (13). In humans, MYOF has been implicated in a variety of cellular processes, including myoblast fusion, growth factor receptor stability, endocytosis, and endothelial cell membrane repair (6,8,(10)(11)(12); however until now its role in cancer cell movement ha...

No abstract

The effects of pore size in a 3‐D polyethylene terephthalate (PET) nonwoven fibrous matrix on long‐term tissue development of human trophoblast ED27 cells were studied. Thermal compression was used to modify the porosity and pore size of the PET matrix. The pore size distributions in PET matrices were quantified using a liquid extrusion method. Cell metabolic activities, estradiol production, and cell proliferation and differentiation were studied for ED27 cells cultured in the thermally compressed PET matrices with known pore structure characteristics. In general, metabolic activities and proliferation rate were higher initially for cultures grown in the low‐porosity (LP) PET matrix (porosity of 0.849, average pore size of 30 μm in diameter) than those in the high‐porosity (HP) matrix (porosity of 0.896, average pore size of 39 μm in diameter). However, 17β‐estradiol production and cell differentiation activity in the HP matrix surpassed those in the LP matrix after 12 days. The expression levels of cyclin B1 and p27kip1 in cells revealed progressively decreasing proliferation and increasing differentiation activities for cells grown in PET matrices. Also, difference in pore size controlled the cell spatial organization in the PET matrices and contributed to the tissue development in varying degrees of proliferation and differentiation. It was also found that cells grown on the 2‐D surface behaved differently in cell cycle progression and did not show increased differentiation activities after growth had stopped and proliferation activities had lowered to a minimal level. The results from this study suggest that the 3‐D cell organization guided by the tissue scaffold is important to tissue formation in vitro. © 2000 John Wiley & Sons, Inc. Biotechnol Bioeng 70: 606–618, 2000.