Presented by: Mari Springer

AGE DEPENDENT LOSS OF MMP-3 IN HUTCHINSON-

GILFORD PROGERIA SYNDROME

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. 

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