View
224
Download
0
Category
Preview:
Citation preview
Rare, premature aging disease
Progressive diseaseImbalance
connective tissueCharacteristics
Short stature Scleroderma-like skin Progressive joint
contracture Atherosclerosis
HUTCHINSON-GILFORD PROGERIA SYNDROME (HGPS)
Premature death by heart attack, stroke, or atherosclerotic disease Average age of 13
Progerin – altered version of lamin A protein Point mutation Chromosome 11
Depression of the enzyme MMP-3 contributes to HGPS
HUTCHINSON-GILFORD PROGERIA SYNDROME (HGPS)
Also known as lamin A/CProtein encoded by LMNA geneFunctions
Aides in chromatic organization, DNA replication, transcription, and repair
Provides structure for the nuclear envelopeLMNA gene encodes for the protein prelamin A
Prelamin A has a farnesyl group attached to it’s end
LMNA PROTEIN
Normal process Progeria process
Farnesyl group is removed
Farnesyl group stays attached
Prelamin A is coded Progerin is coded
Nothing attached to nucleus
Progerin attached to nucleus
Normal nucleus Abnormal shaped nucleus
Family of enzymes that degrade the extracellular matrix (ECM)Maintain proper balance between ECM synthesis and degradation
Family includes MMP-1 through MMP-28This paper focuses on MMP-2, -3, and -9
MMP-3 has the broadest substrate specificityDegrade most of the basement membraneHelps rebuild connective tissue
MATRIX METALLOPROTEINASES (MMP)
Is MMP-3 mRNA and MMP-3 protein regulation defective in HGPS patients?
Does the production of MMP-3 in HGPS cells change over time?
If changes are present, are they MMP-3 specific or general to the MMP family?
GOALS
Cell Lines Obtained skin fibroblasts from the Progeria Research
Foundation Cell and Tissue Bank and the Coriell Cell Repository
HGPS and non-HGPS lines Ages 2, 3, 9, 10, and 13
Western Blot To detect lamin A/C, prelamin A, and progerin
Real-Time Reverse Transcription PCR To amplify genes MMP-2, -3, and -9 and β-actin
MMP protein levels MMP-2 and -9 used Gel Zymography MMP-3 used Enzyme-Linked Immunosorbent Assay (ELISA)
METHODS
Statistical MethodsLinear Mixed Model
Used to show MMP mRNA levels relative to β-actin
Used to show MMP-3 protein levelsPearson’s Correlation
Used to see relationship between number and MMP expression
Standard t-testUsed to show any other results
METHODS
Western BlotHGPS lines – produced prelamin A, lamin A,
lamin C, and progerinDonor age-matched counterparts – produced
prelamin A, lamin A, and lamin C
RESULTS
Reverse Transcription PCR
MMP-3 mRNA – 47-fold lower (p=.0107)
MMP-2 mRNA – 4.8-fold lower (p=.0275)
MMP-9 mRNA – not significantly diff erent
Significant donor age-dependent decline in HGPS fibroblasts MMP-3 p<.001 MMP-2 p<.003
RESULTS
ELISA MMP-3 protein levels reduced 10-fold in HGPS fibroblasts More significant with increasing donor age
Gelatin Zymography MMP-2 and -9 protein levels not significantly different
RESULTS
Is MMP-3 mRNA and MMP-3 protein regulation defective in HGPS patients? Yes – reduced amounts in primary dermal fibroblasts
Does the production of MMP-3 in HGPS cells change over time? Yes – significant decline in both mRNA and protein levels
Suggests a correlation with disease severity Suggests altered balance in connective tissue remodeling
If changes are present, are they MMP-3 specific or general to the MMP family? MMP-3 is specifically downregulated in HGPS
DISCUSSION
MMP-3 could be a potential biomarker to aide in the process of finding treatments and improving existing ones
What are other potential biomarkers for HGPS?
Are these helpful in other diseases affected by the MMP-3 enzyme
RESEARCH PROPOSAL
al., M. A. (2008). Phenotype and Course of Hutchinson-Gildord Progeria Syndrome. The New England Journal of Medicine , 592-604.
Halaschek-Wiener, J., & Brooks-Wilson, A. (2007). Progeria of Stem Cells: Stem Cell Exhaustion in Hutchinson-Gildfor Progeria Syndrome. Journal of Gerontology , 3-8.
Harten, I. A., Zahr, R. S., Lemire, J. M., Machan, J. T., Moses, M. A., Doiron, R. J., et al. (2011). Age-Dependent Loss of MMP-3 in Hutchinson-Gilford Progeria Syndrome. Journal of Gerontology: Biological Sciences , 1201-1207.
Rastogi, R., & Mohan, S. C. (2010). Progeria Syndrome: A Case Report. Indian Journal of Orthopaedics , 1-9.
Tortora, G. J., Funke, B. R., & Case, C. L. (2010). Microbiology: An Introduction. San Francisco: Pearson Benjamin Cummings.
Wamer, H. R. (2008). Research on Hutchinson-Gilford Progeria Syndrome. Journal of Gerontology: Biological Sciences , 775-776.
BIBLIOGRAPHY
Recommended