No abstract

Amyloid is a term for extracellular protein fibril deposits that have characteristic tinctorial and structural properties. Heparan sulphate, or the heparan sulphate proteoglycan perlecan, has been identified in all amyloids and implicated in the earliest stages of inflammation-associated (AA) amyloid induction. Heparan sulphate interacts with the AA amyloid precursor and the beta-peptide of Alzheimer's amyloid, imparting characteristic secondary and tertiary amyloid structural features. These observations suggest that molecules that interfere with this interaction may prevent or arrest amyloidogenesis. We synthesized low-molecular-weight (135-1,000) anionic sulphonate or sulphate compounds. When administered orally, these compounds substantially reduced murine splenic AA amyloid progression. They also interfered with heparan sulphate-stimulated beta-peptide fibril aggregation in vitro.

Aims/hypothesis Islet amyloid in type 2 diabetes contributes to loss of beta cell mass and function. Since islets are susceptible to oxidative stress-induced toxicity, we sought to determine whether islet amyloid formation is associated with induction of oxidative stress. Methods Human islet amyloid polypeptide transgenic and non-transgenic mouse islets were cultured for 48 or 144 h with or without the antioxidant N-acetyl-L-cysteine (NAC) or the amyloid inhibitor Congo Red. Amyloid deposition, reactive oxygen species (ROS) production, beta cell apoptosis, and insulin secretion, content and mRNA were measured.Results After 48 h, amyloid deposition was associated with increased ROS levels and increased beta cell apoptosis, but no change in insulin secretion, content or mRNA levels. Antioxidant treatment prevented the rise in ROS, but did not prevent amyloid formation or beta cell apoptosis. In contrast, inhibition of amyloid formation prevented the induction of oxidative stress and beta cell apoptosis. After 144 h, amyloid deposition was further increased and was associated with increased ROS levels, increased beta cell apoptosis and decreased insulin content. At this time-point, antioxidant treatment and inhibition of amyloid formation were effective in reducing ROS levels, amyloid formation and beta cell apoptosis. Inhibition of amyloid formation also increased insulin content. Conclusions/interpretation Islet amyloid formation induces oxidative stress, which in the short term does not mediate beta cell apoptosis, but in the longer term may feed back to further exacerbate amyloid formation and contribute to beta cell apoptosis.

Development of inhibitors specific for heme oxygenases (HOs) should aid our understanding of the HO system and facilitate future therapeutic applications. The crystal structure of human HO-1 complexed with 1-(adamantan-1-yl)-2-(1H-imidazol-1-yl)ethanone (3) was determined. This inhibitor binds to the HO-1 distal pocket such that the imidazolyl moiety coordinates with heme iron while the adamantyl group is stabilized by a hydrophobic binding pocket. Distal helix flexibility, coupled with shifts in proximal residues and heme, acts to expand the distal pocket, thus accommodating the bulky inhibitor without displacing heme. Inhibitor binding effectively displaces the catalytically critical distal water ligand. Comparison with the binding of 2-[2-(4-chlorophenyl)ethyl]-2-[1H-imidazol-1-yl)methyl]-1,3-dioxolane (2) revealed a common binding mode, despite differing chemical structures beyond the imidazolyl moiety. The inhibitor binding pocket is flexible, yet contains well-defined subpockets to accommodate appropriate functional groups. On the basis of these structural insights, we rationalize binding features to optimize inhibitor design.

Fibroblast apoptosis plays a crucial role in normal and pathological scar formation and therefore we studied whether the putative apoptosis-inducing factor curcumin affects fibroblast apoptosis and may function as a novel therapeutic. We show that 25-μM curcumin causes fibroblast apoptosis and that this could be inhibited by co-administration of antioxidants N-acetyl-l-cysteine (NAC), biliverdin or bilirubin, suggesting that reactive oxygen species (ROS) are involved. This is supported by our observation that 25-μM curcumin caused the generation of ROS, which could be completely blocked by addition of NAC or bilirubin. Since biliverdin and bilirubin are downstream products of heme degradation by heme oxygenase (HO), it has been suggested that HO-activity protects against curcumin-induced apoptosis. Interestingly, exposure to curcumin maximally induced HO-1 protein and HO-activity at 10–15 μM, whereas, at a concentration of >20-μM curcumin HO-1-expression and HO-activity was negligible. NAC-mediated inhibition of 25-μM curcumin-induced apoptosis was demonstrated to act in part via restored HO-1-induction, since the rescuing effect of NAC could be reduced by inhibiting HO-activity. Moreover pre-induction of HO-1 using 5-μM curcumin protected fibroblasts against 25-μM curcumin-induced apoptosis. On a functional level, fibroblast-mediated collagen gel contraction, an in vitro wound contraction model, was completely prevented by 25-μM curcumin, while this could be reversed by co-incubation with NAC, an effect that was also partially HO-mediated. In conclusion, curcumin treatment in high doses (>25 μM) may provide a novel way to modulate pathological scar formation through the induction of fibroblast apoptosis, while antioxidants, HO-activity and its effector molecules act as a possible fine-tuning regulator.

No abstract

Heme oxygenase-1 (HO-1), a member of the heat shock protein family, plays a key role as a sensor and regulator of oxidative stress. Herein, we identify HO-1 as a biomarker and potential therapeutic target for advanced prostate cancer (PCA). Immunohistochemical analysis of prostate tissue using a progression tissue microarray from patients with localized PCA and across several stages of disease progression revealed a significant elevation of HO-1 expression in cancer epithelial cells, but not in surrounding stromal cells, from hormonerefractory PCA (HRPCA) compared with hormone-responsive PCA and benign tissue. Silencing the ho-1 gene in HRPCA cells decreased the HO-1 activity, oxidative stress, and activation of the mitogen-activated protein kinase-extracellular signalregulated kinase/p38 kinase. This coincided with reduced cell proliferation, cell survival, and cell invasion in vitro, as well as inhibition of prostate tumor growth and lymph node and lung metastases in vivo. The effect of ho-1 silencing on these oncogenic features was mimicked by exposure of cells to a novel selective small-molecule HO-1 inhibitor referred to as OB-24. OB-24 selectively inhibited HO-1 activity in PCA cells, which correlated with a reduction of protein carbonylation and reactive oxygen species formation. Moreover, OB-24 significantly inhibited cell proliferation in vitro and tumor growth and lymph node/lung metastases in vivo. A potent synergistic activity was observed when OB-24 was combined with Taxol. Together, these results establish HO-1 as a potential therapeutic target for advanced PCA. [Cancer Res 2009;69(20):8017-24]

. Can. J. Chem. 69, 1273Chem. 69, (1991.AK1401 is a mutant of Pseudomonas aeruginosa strain PA01 (serotype 05) that does not express 0-antigen, but does express "A-band" lipopolysaccharide (LPS). The polysaccharide portion of the A-band LPS (A-PS) from AK1401 was found to consist mainly of D-rhamnose, with smaller amounts of 3-0-methylrhamnose, ribose, mannose, glucose, and a 3-0-methylhexose. 'H nuclear magnetic resonance spectra of the intact A-PS indicated that the main structural feature was a repeating trisaccharide of a-D-rhamnose having the following structure:The 'H NMR spectrum of the repeating unit was completely assigned through the use of 2D shift-correlated and relayed coherence transfer NMR spectroscopy. ' The linkage positions and sequence of residues were found by nuclear Overhauser enhancement difference spectroscopy. Le AK1401 est une espkce mutante du Pselcdomonas aeruginosa de la lignte PA01 (strotype 0 5 ) qui n'exprime pas de 0-antigkne, mais qui exprime un lipopolysaccharide (LPS) de abande-A),. On a trouvC que la portion polysaccharide du LPS de bande-A du AK1401 est principalement formCe de D-rhamnose, avec des quantitCs plus faibles de 3-0-mtthylrhamnose, de ribose, de mannose, de glucose et d'un 3-0-mtthylhexose. Les spectres de rtsonance magnCtique nucltaire du 'H du PS-A indiquent que la caracttristique structurale principale est un motif trisaccharide de a-D-rhamnose, de structure : Faisant appel2 la RMN en 2D avec corrtlation des dtplacernents et transfert de cohCrence par relais, on a fait une attribution complkte du spectre RMN du 'H du motif. On a dttermint les positions de liaison et la stquence des rtsidus par l'effet Overhauser nuclCaire differentiel.