Spheroidal assembly and composition of human prostate cancer cells
Description
Neoplastic cells self-assemble in liquid-overlay cultures into multicellular spheroids that resemble micrometastases and avascular regions of larger tumors. A Monte Carlo simulation based on Meakin's cluster-cluster aggregation model resolved the physical mechanisms by which LNCaP human prostate cancer cells aggregate in this environment. The best-fit solution suggests that LNCaP cells aggregate with an adhesion probability of 0.5% when they migrate within a radius of influence of 180 mum. The sweeping radius of influence is indicative of cell tethering and/or chemotaxis. Aggregates form more compact structures in culture than during simulation as measured by the fractal dimension. Irregularly shaped, two-dimensional aggregates restructured through incremental cell movements into three-dimensional spheroids. Of the two cultures examined, restructuring was more pronounced for DU 145 aggregates. Motile DU 145 cells formed spheroids with a minimum cell overlay of 30% for 25-mers as estimated by simulation vs. 5% for adhesive LNCaP cells in aggregates of the same size Spheroids were then cultured in a mixed suspension within a high aspect rotating-wall vessel and static liquid-overlay plates. Mixed cultures consistently manifested phenotypic differences in morphology, viability, differentiation and expression of multidrug resistance proteins between DU 145 and LNCaP spheroids. For example, 40% +/- 12% of DU 145 cells were activity cycling 100 mum from the surface within mixed spheroids versus 0% for LNCaP spheroids; there was no significant difference in this spatial profile for static cultures. Similar differences were observed in the down regulation of prostate specific antigen (PSA) of both cancer spheroids and in increased expression of cytokeratins 8&18 of DU 145 spheroids. Poorly differentiated spheroids may be particularly responsive to mixing due to low tissue density: internuclear distance within DU 145 spheroids was approximately 5 mum larger than LNCaP spheroids. Immunostaining for P-glycoprotein is representative of this trend; average staining intensity increased 50% for DU 145 spheroid upon mixing but was unchanged for LNCaP spheroids. The effects of mixing on spheroid composition were attributed to faster interstitial diffusion. Applications of this work include the production of highly viable spheroids for drug testing and basic oncological research