Formation of multinucleated cells in a Hodgkin's‐disease‐derived cell line

Abstract

The proliferative potential of multinucleated Reed-Sternberg-like cells and the process of multinuclear formation were studied on the Hodgkin's-disease-derived cell line HDLM-2. No difference in surface antigen expression was found between mono- and multinucleated cells as determined by immunolabelling with characteristic markers. After sorting and reculture of purified mononucleated cells, polykaryons emerged subsequently in these cultures, indicating that mononucleated cells give rise to multinucleated variants. The morphological observation of mitotic figures and immunostaining with the cell cycle indicators Ki-67 and BrdU provided evidence of DNA synthesis and nuclear division in multinucleated cells. The presence of mitotic figures demonstrated that multinucleated cells are able to undergo synchronous nuclear division. However, while polykaryons were clearly mitotically active and capable of DNA synthesis, the absence of telophases and the failure of active replication suggest a disturbed cytokinesis. Co-cultivation of BrdU-labelled and unlabelled populations did not lead to hybrid polykaryons with negative and positive nuclei. Therefore, multinucleated giant cell formation of HDLM-2 cells appears to involve nuclear endomitosis without cell division rather than cell fusion.

Incomplete cytokinesis and re-fusion of small mononucleated Hodgkin cells lead to giant multinucleated Reed–Sternberg cells

“…4). RS cells have been characterized as postmitotic end-stage cells (13)(14)(15). This is also true for the majority of the tracked giant HRS cells in our study, although we can show that RS cells have a residual proliferation potential after elongated lifetime and enormous size increase, whereas mononuclear giant cells have completely lost their proliferation potential (Fig.…”

“…1). These findings are in line with decades-old observations based on conventional microscopy showing that HL cell lines are heterogeneous with respect to cell size and number of nuclei per cell (6,15,26,27). Apart from a few studies investigating the proliferation potential of RS cells (13,14,28), the giant cell population has not been characterized in detail up to now.…”

“…Early experiments with HL cell lines revealed that giant RS cells have no proliferative and clonal growth potential (13)(14)(15); thus, RS cells were defined as a differentiated end-state of HL tumor cells, presumably playing a pivotal role in interaction with the tumor microenvironment in situ (16). The underlying mechanism of giant HRS cell development remained obscure, however.…”

“…Cell fusion of mononuclear Hodgkin cells has been explored as a mechanism for RS cell generation (15); however, a molecular analysis of primary HRS cells excluded the possibility that the HRS cell clone as such or the RS cells are derived from the fusion of different cells (e.g., a B cell and a non-B cell) (17). Moreover, a mixing experiment of dual fluorescent-labeled cells of the HL cell line L1236 provided evidence against cell fusion as the mechanism giving rise to RS cells (18).…”

3D Telomere FISH defines LMP1-expressing Reed–Sternberg cells as end-stage cells with telomere-poor ‘ghost' nuclei and very short telomeres

“…1,2 In about 40-50% of HL cases, Hodgkin cells (H cells) and RS cells express the Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) oncoprotein or its deletion variants, 3,4 and the risk of developing LMP1-expressing HL within a median incubation time of 4 years after symptomatic EBV infection is significantly (fourfold) increased. 5 Mononuclear H cells are the precursors of multinuclear RS cells, [6][7][8][9] and endomitotic multinucleation is associated with disturbed cytokinesis and jumping translocations 10,11 pointing to a severe telomere dysfunction. 12,13 Telomeres are the nucleoprotein complexes at the ends of chromosomes in which a number of specific proteins, either binding telomere proteins directly or a protein complex, termed 'shelterin,' is directly associated with telomeric DNA.…”