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Dissertation
Signal transduction and regulation of gene expression during replicative senescence of human fibroblasts
Doctor of Philosophy (Ph.D.), Medical College of Pennsylvania and Hahnemann University
Jan 1999
DOI:
https://doi.org/10.17918/00009634
Abstract
The focus of the work presented here was to gain insight on the mechanism underlying the inability of replicatively senescent fibroblasts to complete events of the G0/G1 phase of the cell cycle and to, ultimately, proliferate. In chemically-defined media activation of the IGF-I receptor (IGF-IR) is an absolute requirement for growth of early passage WI-38 fibroblasts. Recent evidence suggests that, in addition to the previously described signaling systems, IGF-IR controls cell growth through a novel signaling pathway activated at its C-terminus. We identified a novel gene (DBI-1) that is suppressed in response to activation of the IGF-IR C-terminus and used it as a marker to assess whether this pathway is functional in senescent cells. DBI-1 was expressed at similar levels by both young and senescent cultures of WI-38 cells, suggesting that the signaling pathway that leads to suppression of DBI-1 is intact in senescent cells. Another pathway crucial for transmission of mitogenic signals in response to activation of the IGF-IR as well as of other receptors-tyrosine kinases, is the Raf/MEK/ERK signaling cascade. We observed that stimuli that activate the ERK pathway, such as serum and the antioxidant NGA, fail to induce c-fos expression in senescent WI-38 cells. We also found that activation of ERKs is impaired in senescent cells and that in late passage cells there is an accumulation of unphosphorylated (inactive) ERK2 molecules. The senescenoe-associated decline in ERK activation is presumably caused by the decreased MEK activity since MEK phosphorylation is also lower in late passage cells. Additionally, we observed that both the phosphorylation and the DNA binding activity of the transcription factor p62TCF/Elk-1 is impaired in senescent cells. These results suggest that in senescent cells there are impairments in the activation of signaling molecules that are normally activated during the early G1 phase of the cell cycle. We also observed that senescent cells attain a G0 state distinct from that of young cells since late passage WI-38 cells lose the ability to express the quiescence-specific gene EPC-1. Downregulation of EPC-1 is also a characteristic of replicatively senescent cultures of dermal fibroblasts established from donors of various ages, even though there is no correlation between EPC-1 mRNA abundance and donor age. In summary, our data suggest that the G0/G1 phases are distinct between young and senescent cells. Furthermore, our data are in agreement with the hypothesis that failure of senescent cells to attain a normal G0 state and dysregulation of mitogenic pathways that should be activated in early G1, contributes to their inability to proliferate.
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Details
- Title
- Signal transduction and regulation of gene expression during replicative senescence of human fibroblasts
- Creators
- Maria Tresini
- Contributors
- Vincent J. Cristofalo (Advisor) - Drexel University, Medical College of Pennsylvania and Hahnemann University (1993-1996, 1998-2002)
- Awarding Institution
- Medical College of Pennsylvania and Hahnemann University
- Degree Awarded
- Doctor of Philosophy (Ph.D.)
- Publisher
- Medical College of Pennsylvania and Hahnemann University; Philadelphia, Pennsylvania
- Number of pages
- xiii, 195 pages
- Resource Type
- Dissertation
- Language
- English
- Academic Unit
- School of Medicine (1993-1996, 1998-2002); Medical College of Pennsylvania and Hahnemann University (1993-1996, 1998-2002)
- Other Identifier
- 991021888741704721