Selective Neutral pH Inhibitor of Cathepsin B Designed Based on Cleavage Preferences at Cytosolic and Lysosomal pH Conditions.

Cathepsin B is a cysteine protease that normally functions within acidic lysosomes for protein degradation, but in numerous human diseases, cathepsin B translocates to the cytosol having neutral pH where the enzyme activates inflammation and cell death. Cathepsin B is active at both the neutral pH 7.2 of the cytosol and the acidic pH 4.6 within lysosomes. We evaluated the hypothesis that cathepsin B may possess pH-dependent cleavage preferences that can be utilized for design of a selective neutral pH inhibitor by (1) analysis of differential cathepsin B cleavage profiles at neutral pH compared to acidic pH using multiplex substrate profiling by mass spectrometry (MSP-MS), (2) design of pH-selective peptide-7-amino-4-methylcoumarin (AMC) substrates, and (3) design and validation of Z-Arg-Lys-acyloxymethyl ketone (AOMK) as a selective neutral pH inhibitor. Cathepsin B displayed preferences for cleaving peptides with Arg in the P2 position at pH 7.2 and Glu in the P2 position at pH 4.6, represented by its primary dipeptidyl carboxypeptidase and modest endopeptidase activity. These properties led to design of the substrate Z-Arg-Lys-AMC having neutral pH selectivity, and its modification with the AOMK warhead to result in the inhibitor Z-Arg-Lys-AOMK. This irreversible inhibitor displays nanomolar potency with 100-fold selectivity for inhibition of cathepsin B at pH 7.2 compared to pH 4.6, shows specificity for cathepsin B over other cysteine cathepsins, and is cell permeable and inhibits intracellular cathepsin B. These findings demonstrate that cathepsin B possesses pH-dependent cleavage properties that can lead to development of a potent, neutral pH inhibitor of this enzyme.

Cathepsin B in neurodegeneration of Alzheimer's disease, traumatic brain injury, and related brain disorders.

Investigations of Alzheimer's disease (AD), traumatic brain injury (TBI), and related brain disorders have provided extensive evidence for involvement of cathepsin B, a lysosomal cysteine protease, in mediating the behavioral deficits and neuropathology of these neurodegenerative diseases. This review integrates findings of cathepsin B regulation in clinical biomarker studies, animal model genetic and inhibitor evaluations, structural studies, and lysosomal cell biological mechanisms in AD, TBI, and related brain disorders. The results together indicate the role of cathepsin B in the behavioral deficits and neuropathology of these disorders. Lysosomal leakage occurs in AD and TBI, and related neurodegeneration, which leads to the hypothesis that cathepsin B is redistributed from the lysosome to the cytosol where it initiates cell death and inflammation processes associated with neurodegeneration. These results together implicate cathepsin B as a major contributor to these neuropathological changes and behavioral deficits. These findings support the investigation of cathepsin B as a potential drug target for therapeutic discovery and treatment of AD, TBI, and TBI-related brain disorders.

Radiative-transfer modeling of spectra of planetary regoliths using cluster-based dense packing modifications.

In remote sensing of planetary bodies, the development of analysis techniques that lead to quantitative interpretations of datasets has relatively been deficient compared to the wealth of acquired data, especially in the case of regoliths with particle sizes on the order of the probing wavelength. Radiative transfer theory has often been applied to the study of densely packed particulate media like planetary regoliths, but with difficulty; here we continue to improve theoretical modeling of spectra of densely packed particulate media. We use the superposition T-matrix method to compute the scattering properties of an elementary volume entering the radiative transfer equation by modeling it as a cluster of particles and thereby capture the near-field effects important for dense packing. Then, these scattering parameters are modified with the static structure factor correction to suppress the irrelevant far-field diffraction peak rendered by the T-matrix procedure. Using the corrected single- scattering parameters, reflectance (and emissivity) is computed via the invariant-imbedding solution to the scalar radiative transfer equation. We modeled the emissivity spectrum of the 3.3 µm particle size fraction of enstatite, representing a common regolith component, in the mid-infrared (~5 - 50 µm). The use of the static structure factor correction coupled with the superposition T-matrix method produced better agreement with the corresponding laboratory spectrum than the sole use of the T-matrix method, particularly for volume scattering wavelengths (transparency features). This work demonstrates the importance of proper treatment of the packing effects when modeling semi-infinite densely packed particulate media using finite, cluster-based light scattering models.

[Did a 16th century Christian missionary observe that the Japanese were collectivists?].

Volpi (2004) pointed out that Alessandro Valignano, a 16th century Christian missionary, had considered the Japanese extreme collectivists. According to Volpi, his remark was based on Valignano's reports (1583, 1592) edited by Alvares-Taladriz (1954). However, it is highly questionable whether Volpi examined these texts directly because the information about them provided by Volpi involved many serious errors. A thorough inspection of Valignano's translated reports found no mention of Japanese collectivism. On the contrary, he had actually reported exceedingly individualistic behaviors of Japanese warriors. Such behaviors are consistent with what is widely known about the 16th century Civil Wars in Japan. It has thus turned out that no reliable evidence is present for the alleged observation by Valignano.

Proteoglycan from salmon nasal cartridge [corrected] promotes in vitro wound healing of fibroblast monolayers via the CD44 receptor.

Proteoglycans (PGs) are involved in various cellular functions including cell growth, adhesion, and differentiation; however, their physiological roles are not fully understood. In this study, we examined the effect of PG purified from salmon nasal cartilage (SNC-PG) on wound closure using tissue-cultured cell monolayers, an in vitro wound-healing assay. The results indicated that SNC-PG significantly promoted wound closure in NIH/3T3 cell monolayers by stimulating both cell proliferation and cell migration. SNC-PG was effective in concentrations from 0.1 to 10µg/ml, but showed much less effect at higher concentrations (100-1000µg/ml). The effect of SNC-PG was abolished by chondroitinase ABC, indicating that chondroitin sulfates (CSs), a major component of glycosaminoglycans (GAGs) in SNC-PG, are crucial for the SNC-PG effect. Furthermore, chondroitin 6-sulfate (C-6-S), a major CS of SNC-PG GAGs, could partially reproduce the SNC-PG effect and partially inhibit the binding of SNC-PG to cells, suggesting that SNC-PG exerts its effect through an interaction between the GAGs in SNC-PG and the cell surface. Neutralization by anti-CD44 antibodies or CD44 knockdown abolished SNC-PG binding to the cells and the SNC-PG effect on wound closure. These results suggest that interactions between CS-rich GAG-chains of SNC-PG and CD44 on the cell surface are responsible for the SNC-PG effect on wound closure.

[Beneficial effect of high-dose etidronate on periarticular calcinosis associated with arterial and periarticular basic calcium phosphate (BCP) crystal deposition disease].

Basic calcium phosphate (BCP) deposition disease is a crystal-induced inflammation syndrome. We report a beneficial effect of high-dose etidronate, one of the bisphosphonates, on periarticular calcinosis in a case of arterial and periarticular BCP crystal deposition disease. A 55-year-old woman was referred to St. Marianna University Hospital because of a 30-year history of recurrent acute periarthritis. The X-ray photographs revealed periarticular calcinosis of the hand and foot, wrist, and knee joints without bony erosions and the calcification of carotid and popliteal arteries. The biopsied specimen from periarticular tissue showed Alizalin-red and von Kossa staining-positive crystals. With the diagnosis of BCP crystal-induced periarthritis, she has been treated with colchicine and probenecid. This regimen was partially effective in terms of a decrease in the frequency of periarthritic attack. Despite the treatment, periarticular calcinosis increased in size and number. She was treated with 800 mg of etidronate 3 months after the etidronate therapy; periarticular calcinosis was diminished although arterial calcification was unchanged. This case suggests that high-dose etidronate might be useful for heterotopic calcification associated with rheumatic diseases.