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Age Dependent Loss of MMP-3 in Hutchinson-Gilford Progeria Syndrome

Age Dependent Loss of MMP-3 in Hutchinson-Gilford Progeria Syndrome. Presented by: Mari Springer. Hutchinson-Gilford progeria syndrome (HGPS). Rare, premature aging disease Progressive disease Imbalance connective tissue Characteristics Short stature Scleroderma-like skin

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Age Dependent Loss of MMP-3 in Hutchinson-Gilford Progeria Syndrome

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  1. Age Dependent Loss of MMP-3 in Hutchinson-Gilford Progeria Syndrome Presented by: Mari Springer

  2. Hutchinson-Gilford progeria syndrome (HGPS) • Rare, premature aging disease • Progressive disease • Imbalance connective tissue • Characteristics • Short stature • Scleroderma-like skin • Progressive joint contracture • Atherosclerosis

  3. 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

  4. LMNA protein • Also known as lamin A/C • Protein encoded by LMNA gene • Functions • Aides in chromatic organization, DNA replication, transcription, and repair • Provides structure for the nuclear envelope • LMNA gene encodes for the protein prelamin A • Prelamin A has a farnesyl group attached to it’s end

  5. Matrix Metalloproteinases (MMP) • Family of enzymes that degrade the extracellular matrix (ECM) • Maintain proper balance between ECM synthesis and degradation • Family includes MMP-1 through MMP-28 • This paper focuses on MMP-2, -3, and -9 • MMP-3 has the broadest substrate specificity • Degrade most of the basement membrane • Helps rebuild connective tissue

  6. Goals • 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?

  7. Methods • 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)

  8. methods • Statistical Methods • Linear Mixed Model • Used to show MMP mRNA levels relative to β-actin • Used to show MMP-3 protein levels • Pearson’s Correlation • Used to see relationship between number and MMP expression • Standard t-test • Used to show any other results

  9. Results • Western Blot • HGPS lines – produced prelamin A, lamin A, lamin C, and progerin • Donor age-matched counterparts – produced prelamin A, lamin A, and lamin C

  10. 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 different • Significant donor age-dependent decline in HGPS fibroblasts • MMP-3 p<.001 • MMP-2 p<.003

  11. 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

  12. Discussion • 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

  13. Research Proposal • 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

  14. bibliography • 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.

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