Cell fusion in the liver, revisited

Abstract

There is wide agreement that cell fusion is a physiological process in cells in mammalian bone, muscle and placenta. In other organs, such as the cerebellum, cell fusion is controversial. The liver contains a considerable number of polyploid cells: They are commonly believed to originate by genome endoreplication, although the contribution of cell fusion to polyploidization has not been excluded. Here, we address the topic of cell fusion in the liver from a historical point of view. We discuss experimental evidence clearly supporting the hypothesis that cell fusion occurs in the liver, specifically when bone marrow cells were injected into mice and shown to rescue genetic hepatic degenerative defects. Those experiments-carried out in the latter half of the last century-were initially interpreted to show "transdifferentiation", but are now believed to demonstrate fusion between donor macrophages and host hepatocytes, raising the possibility that physiologically polyploid cells, such as hepatocytes, could originate, at least partially, through homotypic cell fusion. In support of the homotypic cell fusion hypothesis, we present new data generated using a chimera-based model, a much simpler model than those previously used. Cell fusion as a road to polyploidization in the liver has not been extensively investigated, and its contribution to a variety of conditions, such as viral infections, carcinogenesis and aging, remains unclear.

Keywords: Cell fusion; Chimeras; Extracellular vesicles; Hepatocytes; Lineage tracing; TdTomato.

Figure 1

Schematic representation of the Cre: TdTomato approach here described. A: Depiction of the transgene carried by the tdTomato mouse (JAX 007905). The transgene is inserted in the Rosa26 locus and contains a Tomato gene (tdTomato) under the control of a constitutive promoter. Between the promoter and the gene there is a stop cassette flanked by two loxP sequences in the same orientation that prevents tdTomato transcription in this configuration. If the stop cassette is removed by exposure to the Cre recombinase, the tdTomato gene becomes expressed and its expression is maintained through the cell life; B: The generation of mouse chimeras by aggregation of one Cre+ morula (Cre, black-circled cells) derived from CMV-Cre mice (JAX 006054) with a tdTomato one (iTom, red-circled cells) is shown. The aggregated morulae are transferred to females and the progeny analyzed at three months of age. tdTomato expression should occur only if a Cre+ cell fuses with a tdTomato one (full red hepatocyte), whereas if no fusion occurs only tdTomato-negative cells are seen.

Figure 2

Immunohistochemical analysis of tdTomato expression in livers of chimeras and controls. For tdTomato detection, 3 μm slices were stained with anti-RFP antibody (ab124754: Abcam, Cambridge, United Kingdom) diluted 1:100 in PBS containing 0.05% Tween 20. Detection was performed using Mach 1 HRP-polymer (Biocare Medical, Concord, CA, United States) incubation followed by the revelation with Betazoid DAB (Biocare Medical). A: C57BL/6J wild-type mouse; B: Inactive tdTomato mouse; C: Cre+/tdTomato+ double transgenic mouse; D-G: Cre:tdTomato chimera 1 and 2, respectively. Wild type and Inactive Tomato mice are completely negative; Cre+/tdTomato+ are completely positive due to activation of the tdTomato gene in all cells. In the chimeras, many cells are negative, but a fraction shows clear expression of the tdTomato gene, which has been activated by the co-expression of Cre in fused cells. E and G: higher magnification views of the boxed area in D and F respectively.