Radiation damage increases Purkinje neuron heterokaryons in neonatal cerebellum.

Recent reports have demonstrated that bone marrow-derived cells (BMDCs) can fuse with cells in brain, liver, muscle, heart, intestine and kidney.1–9 The factors that trigger in vivo cell fusion and the functional consequences of these events remain largely unknown. Cell fusion transfers genetic and other cellular components from blood cells into cells of other tissues, a process that could contribute to rescuing cells that have suffered genetic damage. In lethal genetic liver and kidney deficiencies, transplantation of disease-free BMDCs that fused with mutant hepatocytes or renal tubular proximal cells rescued both cell types, resulting in increased survival.3,9 Studies in brain have demonstrated that transplantation of bone marrow carrying green fluorescent protein (GFP) into radiated recipients is followed by the appearance of GFP-labeled Purkinje neurons. While initial studies suggested that these were new neurons formed by transdifferentiation,10 more recent work indicates that the labeled Purkinje neurons had two nuclei and were formed by fusion with BMDCs. 1,2, 11–13 It has been suggested that these fusion events could be induced by cerebellar damage caused by whole body radiation, a procedure that depletes the host bone marrow in preparation for transplantation. Recent works suggest that while radiation is not required for the formation of binucleated Purkinje neurons,11,13 general inflammatory responses can increase the number of fusion events.11–13 Radiation-induced damage in the cerebellum is minimal in adults compared to neonates. Low doses of radiation have multiple effects on the development of the cerebellum: the number of granule and basket cells is significantly reduced,14–20 Purkinje neurons are misaligned and have abnormal dendrites,14–16,21,22 and the overall size of the cerebellum is decreased along with foliar and lobular malformations.14,15,21,22 Functional deficiencies following early postnatal radiation of the cerebellum include tremor, ataxia, hypoactivity, and cognitive deficits.16,21–25 It is not known if cell fusion is affected by radiation in neonatal animals. In this study, we asked whether radiation increases fusion events between BMDCs and Purkinje neurons during early postnatal (P4) development. We grafted BMDCs carrying GFP and Cre recombinase into animals carrying a floxed LacZ gene. Upon cell fusion, the Cre in the BMDCs recombines the loxP sequences upstream of the reporter LacZ, allowing its activation and, therefore, the detection of fusion events.2 Cell fusion was confirmed by the presence of two nuclei in a Purkinje neuron. We compared the number of fused Purkinje neurons after engraftment of donor bone marrow in radiated or non-radiated neonatal mice. We also determined the frequency of cell fusion events in head-radiated transgenic mice that carry Cre recombinase under a hematopoietic-specific promoter and floxed LacZ reporter gene in all cells. The results indicate that radiation damage dramatically increases the incidence of fusion events between BMDCs and Purkinje neurons in the neonatal cerebellum. Cell fusion may be a consequence of BMDCs infiltration after radiation-induced damage or/and to direct or indirect radiation damage in Purkinje neurons. These observations are relevant to understand cerebellar damage following therapeutic radiation in young individuals.