Cell behavior observation and gene expression analysis of melanoma associated with stromal fibroblasts in a three‐dimensional magnetic cell culture array

Abstract

A three-dimensional (3D) multicellular tumor spheroid culture array has been fabricated using a magnetic force-based cell patterning method, analyzing the effect of stromal fibroblast on the invasive capacity of melanoma. Formation of spheroids was observed when array-like multicellular patterns of melanoma were developed using a pin-holder device made of magnetic soft iron and an external magnet, which enables the assembly of the magnetically labeled cells on the collagen gel-coated surface as array-like cell patterns. The interaction of fibroblast on the invasion of melanoma was investigated using three types of cell interaction models: (i) fibroblasts were magnetically labeled and patterned together in array with melanoma spheroids (direct-interaction model), (ii) fibroblasts coexisting in the upper collagen gel (indirect-interaction model) of melanoma spheroids, and (iii) fibroblast-sheets coexisting under melanoma spheroids (fibroblast-sheet model). The fibroblast-sheet model has largely increased the invasive capacity of melanoma, and the promotion of adhesion, migration, and invasion were also observed. In the fibroblast-sheet model, the expression of IL-8 and MMP-2 increased by 24-fold and 2-fold, respectively, in real time RT-PCR compared to the absence of fibroblasts. The results presented in this study demonstrate the importance of fibroblast interaction to invasive capacity of melanoma in the 3D in vitro bioengineered tumor microenvironment.

Effect of Vascular Formed Endothelial Cell Network on the Invasive Capacity of Melanoma Using the In Vitro 3D Co-Culture Patterning Model

“…Since there was no significant difference regardless of MCL-labeling in B16F1 cell proliferation (Fig. 2A), MCL labeling has little effect on cell-to-cell interaction [27], [28]. After 48 h of culturing, B16F1 cells in monoculture, patterned in collagen gel at seeding densities of 7.2×10 4 cells/ml (10 cells/spot, 250 µm interval pin-holder), formed spheroids at each locus (Fig.…”

“…Seeking to provide an effective, organized, and practical technique, we have developed a methodology for cell patterning in 3D using magnetic force and magnetite nanoparticles [24]–[28]. Magnetite nanoparticles embedded in cationic liposomes are used for labeling cells via electrostatic interactions between magnetic cationic liposomes (MCLs) and the target cell membrane [29].…”

Magnetic Force-Based Tissue Engineering and Regenerative Medicine

“…Single-cell patterning was achieved by adjusting the number of cells seeded, and the target cell was collected by a micromanipulator after removing the pin holder with the magnet. Okochi et al 39 have fabricated a 3D multicellular tumor spheroid culture array using a magnetic force-based cell patterning method, analyzing the effect of stromal fibroblast on the invasive capacity of melanoma.…”

In vitro skin three-dimensional models and their applications

“…Furthermore, some cells like melanocytes need the addition of a coating such as chitosan to form spheroids [23]. More recently, new methods have become commercially available that allow the aggregation of magnetised cells [24,25].…”

“…fibroblasts on the invasion of melanoma using magnetically labelled fibroblasts and melanoma to generate co-culture spheroids [25]. Another melanoma system used HaCaT spheroids as a scaffold for melanoma cells in a bioreactor system [26].…”