Analysis of protease levels in brain samples from Alzheimer’s patients

Abstract Proteolytic processing of membrane proteins is a tightly regulated activity needed for cell signaling and receptor recycling. At the same time it is a fragile process aberrantly deregulated in neurodegenerative diseases [1]. In Alzheimer’s disease (AD) ADAMs metalloproteases, - and -secretases, cathepsins and meprins are the main proteases involved in the complex processing of the amyloid  precursor protein (APP), which is strictly linked to the genesis of the disease [2,3]. In this study we analyze mRNA and protein levels, using qPCR and western blotting (WB), of pivotal metalloproteases, in frontal cortices of AD patients in comparison to non-AD control subjects, to verify whether in pathological conditions there is a imbalance in the expression of ADAM10 and ADAM17, ADAMTS1, CTSL, MEP1A, MEP1B and MMP9. The analysis show that, between the two α-secretases, ADAM10 mRNA expression is significantly higher (p<0.05) in AD samples in comparison to the control group; while ADAM17 mRNA levels do not significantly differ between AD and controls. The mRNAs expression analysis on other metalloproteases shows a statistically significant different expression (p<0.05) only for ADAMTS1 and MEP1B: 2-fold and 1,62-fold higher in AD samples than in the control group. The mRNA expression of CTSL, MEP1A and MMP9 is not different between the groups. Looking at protein levels, WB experiments shows that ADAMTS1 and MEP1B protein expression is significantly higher in AD samples than in control subjects: 4-fold (p<0,0001) and 1.9-fold higher (p<0,01), respectively. These data indicate that in AD there is a significant overexpression of metalloproteases involved in APP processing, and this increment is controlled at trascriptional level. Whether these variations are cause or consequence of the disease is still to be determined.