ABSTRACT
Lipid peroxidation generates reactive aldehydes, most notably hydroxynonenal (HNE), which covalently binds amino acid residue side chains leading to protein inactivation and insolubility. Specific adducts of lipid peroxidation have been demonstrated to be intimately associated with pathological lesions of Alzheimer's disease (AD), suggesting that oxidative stress is a major component in the disease. Here, we examined the HNE-cross-linking modifications by using an antibody specific for a lysine-lysine cross-link. Since in a prior study we noted no immunolabeling of neuritic plaques or neurofibrillary tangles but instead found strong labeling of axons, we focused this study on axons. Axonal labeling was examined in mouse sciatic nerve, and immunoblotting showed the cross-link was restricted to neurofilament heavy and medium subunits, which while altering migration, did not indicate larger NF aggregates, indicative of intermolecular cross-links. Examination of mice at various ages showed the extent of modification remaining relatively constant through the life span. These findings demonstrate lipid-cross-linking peroxidation primarily involves lysine-rich neurofilaments and is restricted to intramolecular cross-links.
Subject(s)
Aldehydes/chemistry , Neurofilament Proteins/chemistry , Neurofilament Proteins/metabolism , Sciatic Nerve/metabolism , Animals , Antibodies/immunology , Fluorescence , Lysine/chemistry , Lysine/immunology , Mice , Mice, Inbred Strains , Sciatic Nerve/chemistry , Sciatic Nerve/cytologySubject(s)
Graft Rejection/immunology , Interleukin-2/physiology , Killer Cells, Natural/immunology , Lung Transplantation/immunology , Lymphocyte Activation , Animals , Bronchoalveolar Lavage Fluid/cytology , Bronchoalveolar Lavage Fluid/immunology , Cyclosporine/therapeutic use , Cytotoxicity, Immunologic , Dogs , Humans , Interleukin-2/pharmacology , Recombinant Proteins/pharmacology , Transplantation, HomologousABSTRACT
cDNA coding for an antifreeze protein (LS-12) in the longhorn sculpin, Myoxocephalus octodecimspinosis, was prepared from liver mRNA using reverse transcriptase-polymerase chain reaction coupled with 3' and 5' RACE procedures. This cDNA contains 609 base pairs, including a 384-bp open reading frame which codes for a 128-residue LS-12 precursor protein. The predicted amino acid sequence of the mature LS-12 corresponds exactly to the amino acid sequence obtained from Edman degradation [G. Deng, D.W. Andrews, R.A. Laursen, FEBS Lett., 402, 1997, pp. 17-20]. The 20 residues preceding mature LS-12 are predicted to be a signal sequence, which is presumably cleaved off before the mature, 108-residue protein is secreted into the circulatory system. This is the first report of a cDNA sequence from M. octodecimspinosis.
Subject(s)
Antifreeze Proteins, Type IV , Fish Proteins , Proteins/chemistry , Amino Acid Sequence , Animals , Base Sequence , Cloning, Molecular , Liver/chemistry , Molecular Sequence Data , Protein Precursors/chemistry , Protein Sorting Signals/chemistry , Sequence Analysis, DNAABSTRACT
Sperm function was assessed in 19 men 3-10 years after cessation of gossypol treatment and 2-9 years after recovery of normal sperm density. Nineteen normal fertile men of similar age served as the controls. The zona-free hamster egg-sperm penetration assay (SPA) revealed a highly significant difference (P less than 0.01) in the penetration rates between the treated and the control groups. Hormone assays indicated that there were no significant differences in circulating levels of luteinizing hormone (LH) and testosterone (T) between the two groups, but the follicle stimulating hormone (FSH) level of the treated group was significantly higher than that of the controls (P less than 0.05). The results showed that sperm function in the treated group was lower than that in the control, which may be a result of persistent gossypol-mediated damage to the testes.
