MUTER, JOANNE,RUTH (2011) The Application of a Statistical Model Investigating Reactive Oxygen Species in Premature Ageing Syndromes. Masters thesis, Durham University.
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The premature ageing syndromes Hutchison-Gilford Progeria Syndrome (HGPS) and Restrictive Dermopathy (RD) are rare genetic disorders that result in greatly accelerated ageing. RD is neonatal fatal, whereas children suffering from HGPS age approximately 8 times faster than normal individuals and die during their second decade of life -usually from either stroke or heart disease.
The underlying genetic causes of these diseases affect the post-translational processing of the protein lamin A. Lamin A is a Class V intermediate filament protein which resides in the cell nucleus, where it forms a cage-like structure against mechanical strain and is involved in transcriptional regulation, protecting the cell’s genome from damage.
HGPS is caused by a mutation in LMNA, the gene encoding lamin A, which results in a truncated mutant protein termed progerin. RD, on the other hand, has genetic roots in the mutation of the metalloprotease Zmpste24, which has a vital role in the processing of prelamin A. The mutation
results in the accumulation of immature prelamin A at the nuclear membrane.
As a result of increased understanding of the defects in HGPS arising from LMNA mutations a number of therapeutic routes are currently being trialled, but with little success. Work in my supervisor’s laboratory has suggested that defective forms of lamin A may lead to the production
of reactive oxygen species (ROS) in cells. ROS can cause a range of damage in cells including DNA damage. This type of damage eventually leads to cellular senescence which is thought to be a precursor to normal ageing.
The results put forth here suggest that primary dermal fibroblasts from these premature ageing syndromes show defects in both DNA damage repair and ROS buffering ability. This is shown from my statistical modelling of immunofluorescence data showing DNA double strand break
repair over time. RD and HGPS fibroblasts showed persistent γ-H2AX foci after DNA damage induction with oxidative stress, yet showed improved repair after induction with a direct DNA break inducing agent, etoposide.
Proliferative markers indicated a reduced proliferative capacity in premature ageing fibroblasts, which was heightened after treatment with oxidative stress. Taken together these results suggest that RD and HGPS fibroblasts more readily enter a state of cellular senescence when
exposed to oxidising conditions than control fibroblasts from both and young and old donors.
Phenotypical aberrant cellular morphology associated with these diseases was shown to be independent of the defect in ROS buffering ability, supporting evidence showing improvements in abnormal nuclear shape by treatment with farnesyltransferase inhibitors (FTIs) have little
impact on the DNA damage repair defects associated with these diseases.
From the results here, N-acetyl cysteine (NAC), a powerful antioxidant, has shown to be effective in reducing the proportion of irreparable DNA damage in HGPS and RD fibroblasts. It has also been shown to increase repair rates by actively reducing ROS levels as detected by flow
cytometry. NAC treatment therefore shows beneficial properties in treating certain phenotypes associated with progeria and may require consideration as a cooperative form of treatment.
The implications of these results are that HGPS is a useful model for normal ageing as fibroblasts from an elderly donor showed signs not dissimilar to those of HGPS, albeit in milder form. These included an increase in the number of persistent γ-H2AX foci and increased ROS levels. This
suggests that activation of the HGPS cryptic splice site in normal ageing may account for increased age-associated genomic instability Thus it is tempting to speculate that lamin A is a modulator of both premature and normal ageing through its regulation of cellular oxidative
|Item Type:||Thesis (Masters)|
|Award:||Master of Science|
|Keywords:||laminopathy, DNA damage, reactive oxygen species, progeria|
|Faculty and Department:||Faculty of Science > Biological and Biomedical Sciences, School of|
|Copyright:||Copyright of this thesis is held by the author|
|Deposited On:||01 Jun 2011 11:24|