Differential expression of notch signaling-related transcripts accompanies pro-thymocyte proliferation and phenotype transition induced by epidermal growth factor plus insulin in fetal thymus organ cultures.
Thymus regression upon stressing stimuli, such as infectious diseases, is followed by organ reconstitution, paralleling its development in ontogeny. A narrow window of thymus development was here studied, encompassing the pro-T lymphoid precursor expansion during specification stages, by the use of epidermal growth factor plus insulin (INS) in murine fetal thymus organ cultures. Aiming to disclose signaling pathways related to these stages, cultured thymus lobes had their RNA extracted, for the search of transcripts differentially expressed using RNAse protection assays and reverse transcriptase-polymerase chain reactions. We found no difference that could explain INS-driven thymocyte growth, in the pattern of transcripts for death/proliferation mediators, or for a series of growth factor receptors and transcriptional regulators known as essential for thymus development. Thymocyte suspensions from cultured lobes, stained for phenotype analysis by fluorescence activated cell sorting, showed a decreased staining for Notch1 protein at cell surfaces upon INS addition. We analyzed the expression of Notch-related elements, and observed the recruitment of a specific set of transcripts simultaneous and compatible with INS-driven thymocyte growth, namely, transcripts for Notch3, for its ligand Jagged2, and for Deltex1, a mediator of a poorly characterized alternative pathway downstream of the Notch receptor.
Laboratório de Biologia Molecular Maury Miranda, Instituto de Biofísica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, RJ, 21949-900, Brazil. csondermannf@starmedia.com
Claudia Sondermann
CS
Freitas
Sergio Ranto
SR
Dalmau
Eliana
E
Abdelhay
D000818:Animals; D002454:Cell Differentiation; D004815:Epidermal Growth Factor / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D051880:Receptors, Notch; D020133:Reverse Transcriptase Polymerase Chain Reaction; D015398:Signal Transduction / Q000235:genetics*; D013601:T-Lymphocytes / Q000378:metabolism / Q000502:physiology*; D013950:Thymus Gland / Q000166:cytology* / Q000254:growth & development / Q000378:metabolism
D007328:Insulin; D008565:Membrane Proteins; D051880:Receptors, Notch; D004815:Epidermal Growth Factor