Inhibition of cathepsin B reduces β-amyloid production in regulated secretory vesicles of neuronal chromaffin cells: evidence for cathepsin B as a candidate β-secretase of Alzheimer's disease

Abstract

The regulated secretory pathway of neurons is the major source of extracellular A beta that accumulates in Alzheimer's disease (AD). Extracellular A beta secreted from that pathway is generated by beta-secretase processing of amyloid precursor protein (APP). Previously, cysteine protease activity was demonstrated as the major beta-secretase activity in regulated secretory vesicles of neuronal chromaffin cells. In this study, the representative cysteine protease activity in these secretory vesicles was purified and identified as cathepsin B by peptide sequencing. Immunoelectron microscopy demonstrated colocalization of cathepsin B with A beta in these vesicles. The selective cathepsin B inhibitor, CA074, blocked the conversion of endogenous APP to A beta in isolated regulated secretory vesicles. In chromaffin cells, CA074Me (a cell permeable form of CA074) reduced by about 50% the extracellular A beta released by the regulated secretory pathway, but CA074Me had no effect on A beta released by the constitutive pathway. Furthermore, CA074Me inhibited processing of APP into the COOH-terminal beta-secretase-like cleavage product. These results provide evidence for cathepsin B as a candidate beta-secretase in regulated secretory vesicles of neuronal chromaffin cells. These findings implicate cathepsin B as beta-secretase in the regulated secretory pathway of brain neurons, suggesting that inhibitors of cathepsin B may be considered as therapeutic agents to reduce A beta in AD.

Protease Pathways in Peptide Neurotransmission and Neurodegenerative Diseases

“…Results indicating cathepsin B as a β-secretase in regulated secretory vesicles (Hook et al, 2005), combined with evidence for BACE 1 as β-secretase for basal and constitutive secretion of Aβ, are consistent with the unifying hypothesis for distinct proteolytic pathways for β-secretase processing of APP (Fig. 7).…”

“…The regulated secretion of neuronal Aβ is induced by receptor-mediated activation of neurons (Nitsch et al, 1993;Farber et al, 1995;Wolf et al, 1995;Efthimiopoulos et al, 1996;Jolly-Tornetta et al, 1998;Hook et al, 2002). Compared to the major regulated secretory pathway for Aβ production and secretion, a minor portion of Aβ undergoes basal secretion from the constitutive secretory pathway (Hook et al, 2002(Hook et al, , 2005.…”

“…Notably, cathepsin B was recently identified as the primary β-secretase in chromaffin secretory vesicles for production of Aβ (Hook et al, 2005). Identification was achieved by purification with a β-secretase specific cleavage site fluorescent assay, affinity labeling, and peptide microsequencing.…”

“…In contrast, the aspartyl protease BACE 1 represents the β-secretase in the minor constitutive secretory pathway for basal secretion of Aβ (Hussain et al, 1999;Sinha et al, 1999;Vassar et al, 1999;Yan et al, 1999;Lin et al, 2000). Since the regulated secretory pathway is the primary source of secreted, extracellular Aβ from active neurons (Hook et al, 2002(Hook et al, , 2005, cathepsin B in regulated secretory vesicles is likely to be a key protease for production of neurotoxic Aβ in brain. The hypothesis that cathepsin B participates as β-secretase in the regulated secretory pathway of brain neurons indicates that inhibitors of cathepsin B should be explored as novel therapeutic agents to reduce Aβ in AD.…”

6-Shogaol has anti-amyloidogenic activity and ameliorates Alzheimer’s disease via CysLT1R-mediated inhibition of cathepsin B

“…In primary cultures of chromaffin cells, the cell permeable cysteine protease inhibitor E64d was also shown to inhibit the production of the APPderived COOH-terminal b-secretase cleavage product. Previous reports have also concluded that cathepsin B and CysLT1R are involved in AD development and progression [11]. However, the relationship between cathepsin B and CysLT1R activity is not clear.…”