A new method for the 3-D in vitro growth of human RT112bladder carcinoma cells using the alginate culture technique

Abstract

We studied the response to different in vitro culture conditions and the ability for polarization in three-dimensional (3-D) and two-dimensional (2-D) cell culture systems of the commonly used human bladder carcinoma cell line RT112. In the case of 2-D culture the cells were grown on glass or plastic coverslips, filter membranes, or bovine lens capsules. The alginate culture technique (ACT) was used to test the effects of 3-D cell culture on the polarization of the RT112 epithelial cells. Our studies show clear differences in the arrangement of the cells depending on the cultivation procedure. The RT112 cells cultured on glass and plastic supports were irregular and flattened in shape whereas the cells grown on filters or on lens capsules developed into 2-3 layers consisting of markedly polarized cells. However, ACT was superior to cell culture on either artificial supports or even on lens capsule. During the 3-D cultivation the transformed epithelial cells regenerated multicellular spheroids which maintained a tissue-like, geometrically well-ordered and highly prismatic organization. This ACT-induced morphogenesis is novel and distinct from that reported with conventional culture conditions. Microscopic investigations showed that RT112 cells grown in alginate were both much more tightly packed and more regularly organized compared to 2-D cultures. In addition, the spheroidal organized cells exhibited well developed cell-cell contacts, a distinct endoplasmatic reticulum, and a marked Golgi apparatus. In summary, ACT can be used for 3-D in vitro growth of the transformed human epithelial cell line RT112 that offers substantial advantages over conventional cell culture methods.