Presumptive germ cells derived from mouse pluripotent somatic cell hybrids.
The fusion of embryonic stem (ES) cells with differentiated somatic cells is an approach that reverses a somatic cell nucleus to a state of pluripotency. The resulting ES-somatic cell hybrids (ES-SCH) retain most of the properties of ES cells: differentiate into multiple cell types and have the ability to produce embryoid bodies (EB) and chimeras. However, it is still unknown whether ES-SCH will be able to complete the differentiation into germ cells (GC) in vitro similar to ES cells. Here, we show that near diploid ES-SCH, obtained by the fusion of mouse ES and spleen cells, were able to differentiate in vitro into presumptive GC. Differentiation of ES-SCH was induced through EB formation and by the addition of retinoic acid. Presumptive GC obtained reacted positively with anti-EMA, Vasa, Fragilis and Dazl antibodies and expressed GC-specific genes, such as Vasa, Stella, Dazl, Piwil 2, Tex14, Bmp8b, Tdrd1 and Rnf17. Fluorescent in situ hybridization analysis indicates chromosome reduction in the GC-like cells. Expression of meiotic and postmeiotic GC-specific genes such as Haprin, Acrosin, Scyp1, Scyp3 and Stra-8 were also detected. Transmission electron microscopy confirmed ES-SCH differentiation into presumptive GC. The presence of several autosomes and the X chromosome originated from the "somatic" partner did not prevent ES-SCH differentiation towards presumptive GC. Overall our study suggests an interesting in vitro model, which allows the study GC differentiation in reprogrammed somatic cells.
Lavagnolli
Thais M C
TM
Department of Genetics and Morphology, Federal University of São Paulo, São Paulo-SP, Brazil.
Fonseca
Simone A S
SA
Serafim
Rui C
RC
Pereira
Virgínia S
VS
Santos
Enrico J C
EJ
Abdelmassih
Soraya
S
Kerkis
Alexandre
A
Kerkis
Irina
I
D000818:Animals; D002454:Cell Differentiation / Q000502:physiology*; D049109:Cell Proliferation; D002478:Cells, Cultured; D005260:Female; D005854:Germ Cells / Q000166:cytology*; D006822:Hybrid Cells / Q000166:cytology*; D007124:Immunoenzyme Techniques; D017404:In Situ Hybridization, Fluorescence; D051379:Mice; D046529:Microscopy, Electron, Transmission; D039904:Pluripotent Stem Cells / Q000166:cytology*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013154:Spleen / Q000166:cytology / Q000502:physiology
D012333:RNA, Messenger