Effects of simulated microgravity on DU 145 human prostate carcinoma cells

Abstract

The high aspect rotating-wall vessel (HARV) was recently designed by NASA to cultivate animal cells in an environment that simulates microgravity. This work examines the effects of HARV cultivation on DU 145 human prostate carcinoma cells. In the HARV, these prostate cells grew in suspension on Cytodex-3 microcarrier beads t o form bead aggregates with extensive three-dimensional growth between beads and on the aggregate surface. HARV and spinner-flask control cultures of DU 145 cells had similar doubling times, but the former was characterized by a higher percentage of G,-phase cells, larger bead aggregates, enhanced development of filopodia and microvilli-like structures on the aggregate surface, and stronger staining for select cytoskeletal proteins (cytokeratins 8 and 18, actin, and vimentin). When compared with static controls grown in a T-flask and Transwell insert, HARV cultures grew more slowly and differences in the cell cycle and immunostaining became more pronounced. These results suggest that HARV cultivation produced a culture that was less aggressive from the perspective of proliferation, more differentiated and less pliant than any of the three control cultures examined in this work. Possible factors effecting this change are discussed including turbulence and three-dimensional growth. 0 1996 John Wiley & Sons, Inc.

Characterization of Autocrine Growth Factors, Their Receptors and Extracellular Matrix Present in Three-Dimensional Cultures of DU 145 Human Prostate Carcinoma Cells Grown in Simulated Microgravity

“…For HARV and spinner-flask cultures, tissue aggregates were first dissociated into a single-cell suspension by exposure to DNase-free RNase (Boehringer Mannheim, Indianapolis, IN) and NONIDET P-40 (Sigma) as previously described. 11 Immunocytochemistry For the HARV and spinner flask, samples for immunocytochemistry were taken directly from the vessels, and bead aggregates were kept intact. For the Transwell, the insert itself, confluent with cells, was the sample.…”

“…11 The mode of growth was similar in the RWV and spinner flask; DU 145 cells were cultivated in both vessels on microcarrier beads in mixed medium, but there was more fluid turbulence in the spinner flask. In contrast to these mixed cultures, Transwell cultures were static and grew on a flat surface.…”

Modulating and modeling aggregation of cell-seeded microcarriers in stirred culture system for macrotissue engineering

“…As a matter of fact, microcarriers have been extensively applied in tissue engineering, especially for cell expansion purpose due to higher cell growth area in comparison with tissue culture flasks (Chung et al, 2008;Chung and Park, 2009;Eibes et al, 2010;Hong et al, 2005;Melero-Martin et al, 2006;Overstreet et al, 2003;Sart et al, 2009). While only few studies applied the cellseeded microcarriers to construct macrotissues (Palmiero et al, 2010), the self-aggregation of cell-seeded microcarriers during cultivation was documented extensively and has been implicated to influence cell growth and differentiation (Chung et al, 2008;Chung and Park, 2009;Clejan et al, 1996;Eibes et al, 2010;Muhitch et al, 2000;Qiu et al, 2001;Tang et al, 2008). Without optimal control over the aggregation, cell necrosis could happen in the center of very large aggregates (Khaoustov et al, 1999).…”

A Review of Three-Dimensional In Vitro Tissue Models for Drug Discovery and Transport Studies

“…Mechanisms of the acquired drug resistance are likely to be due to enhancement in cell-cell interactions via adhesion and soluble factors released by tumor cells, and creation of a microenvironment (e.g., low pH and hypoxia) that mimic solid tumors grown in vivo. [176][177][178] There are several methods that have been traditionally employed for culturing these spheroids, including spinner flasks, 179,180 microgravity (using rotating-wall vessels), [181][182][183][184][185][186] and spontaneous aggregation. 187,188 In spinner flasks, cells are grown in dynamic suspension in culture medium 180 yielding high numbers of spheroids without the use of a substrate.…”