title: Lipid peroxidation-derived 4-hydroxynonenal-modified proteins accumulate in human facial skin fibroblasts during ageing in vitro.
abstract: The reactive aldehyde, 4-hydroxynonenal (HNE), is recognized as a product of lipid peroxidation, which binds to macromolecules, in particular proteins. HNE-modified proteins (HNE-MP) have been shown to accumulate during ageing, generally by using polyclonal antibodies, which increase the possibility of detecting false positives. Therefore, we have used a genuine monoclonal antibody specific for HNE-His adducts of proteins/peptides, which were revealed by immunoblotting method for whole-cell HNE-MP measurements in serially passaged human facial skin fibroblasts undergoing ageing in vitro. There was a significant increase in the levels of HNE-MP in serially passaged cells approaching a near senescent state at high passage level (P-61), as compared with low passage level (P-11) young and middle-aged (P-27) cells. However, if the cells were analyzed soon after re-initiation from the frozen samples with little further passaging, the amount of HNE-MP was low even in relatively high passage level (P-37) cells, which is an indication of selective elimination of cells with high molecular damage during the process of thawing and re-initiation in culture. This pilot study on normal human facial skin fibroblasts shows that HNE-MP detection by monoclonal antibody-based dot blot method can be used as a marker for age-related accumulation of lipid peroxidative molecular damage, and could be useful for testing and monitoring the effects of potential skin care products on ageing parameters.
authors.affiliation: Laboratory of Cellular Ageing, Department of Molecular Biology and Genetics, Aarhus University, DK8000, Aarhus, Denmark, molekylemanden@hotmail.com.
authors.forename: Peter
authors.initials: P
authors.lastname: Jørgensen
authors.forename: Lidija
authors.initials: L
authors.lastname: Milkovic
authors.forename: Neven
authors.initials: N
authors.lastname: Zarkovic
authors.forename: Georg
authors.initials: G
authors.lastname: Waeg
authors.forename: Suresh I S
authors.initials: SI
authors.lastname: Rattan
mesh_terms: D000447:Aldehydes / Q000378:metabolism*; D015415:Biomarkers / Q000378:metabolism; D001706:Biopsy; D016922:Cellular Senescence / Q000502:physiology*; D005260:Female; D005347:Fibroblasts / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D015151:Immunoblotting; D066298:In Vitro Techniques; D015227:Lipid Peroxidation / Q000502:physiology*; D008858:Microscopy, Phase-Contrast; D008875:Middle Aged; D010865:Pilot Projects; D012867:Skin / Q000378:metabolism* / Q000473:pathology; D015595:Skin Aging / Q000473:pathology / Q000502:physiology
chemical_list: D000447:Aldehydes; D015415:Biomarkers; C027576:4-hydroxy-2-nonenal