Morphologic Changes of Cultivated Uveal Melanoma Cells with Increasing of Passages

Abstract

 

Purpose

 

This study was conducted to investigate the morphologic changes of cultivated uveal melanoma (UM) cells with increasing number of passages.

 

Methods

 

Immediately after enucleation of an eye with mixed cell-typed UM, tumor tissue was obtained, digested with 1.25% trypsin, and cultivated in DMEM F12 with 20% FBS. The cultivated cells were passaged to reach the fifth passage. Immunocytochemistry for melan-A monoclonal antibody, as a melanoma marker, was performed to identify melanoma cells. All the experiments were done in duplicate. With increasing number of passages, morphologic changes of cultivated melanoma cells were investigated.

 

Results

 

In the first passage, there was a mixture of epithelioid melanoma cells with owl's eye nucleoli and spindle-shaped neural-like melanoma cells with dendritic processes. With increasing of passages, the predominant population of cultivated cells were epithelioid melanoma cells. However, the numbers of spindle-shaped neural-like melanoma cells were significantly reduced. Colony formation of tumor cells was observed in the third passage (Figure -1).

 

Conclusions

 

Homogeneity of cultivated melanoma cells can be obtained through increasing number of passages. It seems that differentiated neural-like melanoma cells are more vulnerable to the increasing number of passages as compared to the less differentiated epithelioid melanoma cells.

 

 

Figure 1. A- C: Cultivated UM cells in passages 1 (A), 2 (B), and 3 (C). Note the presence of epithelioid melanoma cells with owl's eye nucleoli (circle 1), neural-like melanoma cells with dendritic processes (circle 2), and colony formation of cultivated UM cells (arrow). FITC of melan-A expression in cultivated UM cells (D), DAPI of nucleus (E) and merged FITC and DAPI of UM cells (F).

 

Keywords: 589 melanoma