Nonoxidative protein glycation is implicated in familial amyotrophic lateral sclerosis with superoxide dismutase-1 mutation

To assess a role for oxidative stress in the pathogenesis of amyotrophic lateral sclerosis (ALS), we analyzed the immunohistochemical localization of 8-hydroxy-2′-deoxyguanosine (OHdG) as a nucleic acid oxidation product, acrolein-protein adduct and 4-hydroxy-2-nonenal (HNE)-protein adduct as lipid peroxidation products, N ^ɛ-carboxymethyl-lysine (CML) as a lipid peroxidation or protein glycoxidation product, pentosidine as a protein glycoxidation product, and imidazolone and pyrraline as nonoxidative protein glycation products in the spinal cord of three familial ALS patients with superoxide dismutase-1 (SOD1) A4V mutation, six sporadic ALS patients, and six age-matched control individuals. The spinal cord sections of the control cases did not show any distinct immunoreactivities for these examined products. In the familial ALS cases, intense immunoreactivities for pyrraline and CML were confined to the characteristic Lewy body-like hyaline inclusions, and imidazolone immunoreactivity was located in the cytoplasm of the residual motor neurons. No significant immunoreactivities for other examined products were detected in the familial ALS spinal cords. In the sporadic ALS cases, intense immunoreactivities for pentosidine, CML and HNE-protein adduct were seen in the cytoplasm of the degenerated motor neurons, and OHdG immunoreactivity was located in the cell nuclei of the residual neurons and glial cells. The present results indicate that oxidative reactions are involved in the disease processes of sporadic ALS, while there is no evidence for increased oxidative damage except for CML deposition in the familial ALS spinal cords. Furthermore, it is likely that the accumulation of pyrraline and imidazolone supports a nonoxidative mechanism in SOD1-related motor neuron degeneration. Received: 18 August 1999 / Revised, accepted: 17 November 1999     

Reconstructed skin modified by glycation of the dermal equivalent as a model for skin aging and its potential use to evaluate anti-glycation molecules

Glycation is a slow chemical reaction which takes place between amino residues in protein and a reducing sugar. In skin this reaction creates new residues or induces the formation of cross-links (advanced glycation end products or AGEs) in the extracellular matrix of the dermis. Formation of such cross-links between macromolecules may be responsible for loss of elasticity or modification of other properties of the dermis observed during aging. We had previously developed a reconstructed skin model which enabled us to study the consequences of matrix alteration by preglycation of the collagen and have reported several modifications of interest induced by glycation in the dermal and epidermal compartments of reconstructed skin as well as at the level of the dermal-epidermal junction. For example we showed that collagen IV and laminin were increased in the basement membrane zone and that alpha6 and beta1 integrins in epidermis were expanded to suprabasal layers. The aim of this new study was to look at the biological effects of glycation inhibitors like aminoguanidine in the skin model. Aminoguanidine was mixed with collagen in the presence of ribose as reducing sugar, and immunostaining was used to visualize its effects on AGE Products and biological markers. After aminoguanidine treatment, we found a low amount of AGE products and a possible return to the normal pattern of distribution of markers in skin constructs as compared to those treated with ribose only. Interestingly similar results were also obtained, although to a lesser extent, with a blueberry extract. In conclusion the glycation inhibitory effect has been functionally demonstrated in the reconstructed skin model and it is shown that this model can be used to assess anti-glycation agents.