Particulate phase cigarette smoke increases MnSOD, NQO1, and CINC-1 in rat lungs

Loss of antioxidant/oxidant homeostasis perpetuates inflammation in the lungs and may contribute to the development of COPD and lung cancer. Cigarette smoke (CS) is a primary source of airway oxidative stress and recruits inflammatory cells into smokers' lungs. However, whether these consequences are attributable to a specific or the collective fraction of CS is unknown. We investigated whether the particulate or the gas phase of CS would alter expression of the antioxidant enzymes MnSOD and NQO1 or CINC-1. Sprague Dawley rats were exposed to sham (n = 10) or the particulate phase (PP; n = 10) or gas phase (n = 10) of a Kentucky reference cigarette (1R4F) for 2 h/d for 28 d, after which animals were sacrificed and the lower left lobe of the lung was removed. Immunoblots for SOD and NQO1 revealed that lungs exposed to PP had higher MnSOD/actin and NQO1/actin ratios than either sham-or gas phase-treated animals. In contrast, CuZnSOD remained unchanged. In PP-exposed animals, CINC-1 was 3-fold higher than in sham-exposed animals. The increases in MnSOD and NQO1 protein were associated with increases in total SOD, NQO1, and MPO activities. These data provide evidence that the PP of CS alters oxidant/antioxidant homeostasis in the lungs and participates in the pathogenesis of CS-induced lung diseases such as COPD and cancer.     

Not all perlecans are created equal: interactions with fibroblast growth factor (FGF) 2 and FGF receptors

Human basement membrane heparan sulfate proteoglycan (HSPG) perlecan binds and activates fibroblast growth factor (FGF)-2 through its heparan sulfate (HS) chains. Here we show that perlecans immunopurified from three cellular sources possess different HS structures and subsequently different FGF-2 binding and activating capabilities. Perlecan isolated from human umbilical arterial endothelial cells (HUAEC) and a continuous endothelial cell line (C11 STH) bound similar amounts of FGF-2 either alone or complexed with FGFRalpha1-IIIc or FGFR3alpha-IIIc. Both perlecans stimulated the growth of BaF3 cell lines expressing FGFR1b/c; however, only HUAEC perlecan stimulated those cells expressing FGFR3c, suggesting that the source of perlecan confers FGF and FGFR binding specificity. Despite these differences in FGF-2 activation, the level of 2-O- and 6-O-sulfation was similar for both perlecans. Interestingly, perlecan isolated from a colon carcinoma cell line that was capable of binding FGF-2 was incapable of activating any BaF3 cell line unless the HS was removed from the protein core. The HS chains also exhibited greater bioactivity after digestion with heparinase III. Collectively, these data clearly demonstrate that the bioactivity of HS decorating a single PG is dependent on its cell source and that subtle changes in structure including secondary interactions have a profound effect on biological activity.     

Overexpression of a kinase-inactive ATR protein causes sensitivity to DNA-damaging agents and defects in cell cycle checkpoints.

ATR, a phosphatidylinositol kinase-related protein homologous to ataxia telangiectasia mutated (ATM), is important for the survival of human cells following many forms of DNA damage. Expression of a kinase-inactive allele of ATR (ATRkd) in human fibroblasts causes increased sensitivity to ionizing radiation (IR), cis-platinum and methyl methanesulfonate, but only slight UV radiation sensitivity. ATRkd overexpression abrogates the G2/M arrest after exposure to IR, and overexpression of wild-type ATR complements the radioresistant DNA synthesis phenotype of cells lacking ATM, suggesting a potential functional overlap between these proteins. ATRkd overexpression also causes increased sensitivity to hydroxyurea that is associated with microtubule-mediated nuclear abnormalities. These observations are consistent with uncoupling of certain mitotic events from the completion of S-phase. Thus, ATR is an important component of multiple DNA damage response pathways and may be involved in the DNA replication (S/M) checkpoint.     

Neanderthal skeleton from Tabun: U-series data by gamma-ray spectrometry

The Neanderthal hominid Tabun C1, found in Israel by Garrod & Bate, was attributed to either layer B or C of their stratigraphic sequence. We have used gamma-ray spectrometry to determine the²³⁰Th/²³⁴U and²³¹Pa/²³⁵U ratios of two bones from this skeleton, the mandible and a femur. The ages calculated from these ratios depend on the uranium uptake history of the bones. Assuming a model of early U (EU) uptake the age of the Tabun C1 mandible is 34±5 ka. The EU age of the femur is 19±2 ka. The femur may have experienced continuous (linear) U uptake which would give an age of 33±4 ka, in agreement with the mandible's EU age, but implies marked inhomogeneity in U uptake history at the site. These new age estimates for the skeleton suggest that it was younger than deposits of layer C. This apparent age is less than those of other Neanderthals found in Israel, and distinctly younger than the ages of the Skhul and Qafzeh burials. This suggests that Neanderthals did not necessarily coexist with the earliest modern humans in the region. All of the more complete Neanderthal fossils from Israel are now dated to the cool period of the last glacial cycle, suggesting that Neanderthals may have arrived in this region as a result of the southward expansion of their habitable range. The young age determined for the Tabun skeleton would suggest that Neanderthals survived as late in the Levant as they did in Europe.     

A 55 kDa antigen of Pneumocystis carinii: analysis of the cellular immune response and characterization of the gene

Rat-derived Pneumocystis carinii contains a major antigen complex of 45–55 kDa. The fusion protein of a cDNA encoding the 3′ portion of the 55 kDa antigen, which had previously been shown to be recognized by serum antibodies of exposed subjects, was investigated for its ability to stimulate a cellular immune response. Rats exposed to P. carinii via the environment exhibited a vigorous proliferative response to the antigen whereas unexposed rats did not. The full-length cDNA for a 55 kDa antigen was cloned and found to contain a 1245 bp open reading frame capable of encoding a 414-amino-acid peptide. The gene encoding this protein contained a single 39 bp intron and transcribed a 1.45 kb RNA message. The cloning and characterization of the 55 kDa antigen gene will allow production of the specific immunological reagents necessary to characterize this molecule and study its role in the biology and pathogenesis of P. carinii.     

Purified eukaryotic initiation factor 2 from calf liver consists of two polypeptide chains of 48,000 and 38,000 daltons.

Eukaryotic initiation factor 2 (eIF-2), which specifically binds Met-tRNAMetf and forms stable ternary complexes with GTP, has been purified from ribosomal salt wash proteins from calf liver. The purified factor exhibits only two polypeptide bands of Mr = 48,000 and 38,000 following electrophoresis in 15% polyacrylamide gels in the presence of sodium dodecyl sulfate. Densitometric tracings show the two polypeptides are present in a molar ratio of 1:1. This suggests a Mr = 86,000 for the native enzyme, a value which agrees with the apparent molecular weight determined by physical methods. Less pure preparations of eIF-2 show additional polypeptide bands, including 50,000- and 46,000-dalton bands, all of which can be removed by further purification without affecting the activity of eIF-2.