Hiroshi Takei

Alterations of protein kinase C, 8-hydroxydeoxyguanosine, and K-ras oncogene in rat lungs exposed to passive smoking

To investigate the effects of exposure to sidestream cigarette smoke (CS) on the initiation and promotion of lung cancer, two groups of 8 or 10 rats were exposed to CS for a 1 h period twice a day for 8, 12, or 20 weeks. The protein kinase C (PKC) activity of the lung exhibited significant changes of 120, 86 and 81% in the CS groups, compared with the respective control group values in the three exposure periods. The in vitro activation of PKC by the active oxygens was efficiently eliminated by hydroxyl radical scavengers, indicating that hydroxyl radicals are responsible for the PKC activation. For the alterations in the lung nucleus caused by passive smoking, the 12- and 20-week exposure CS groups showed significant increases in the accumulation of 8-hydroxydeoxyguanosine. One rat with K-ras activation by G:C transversion (GGT-->GCT) at codon 12 was found among 26 rats of the CS groups in the three exposure periods. These results show that active oxygens introduced by passive smoking may contribute to K-ras activation as an initiator of a tumor model, possibly through the oxygen-induced DNA damage, and may also contribute to an initial activation and the subsequent down-regulation of PKC as a promoter.

AMINO ACID SEQUENCES OF HEMOGLOBIN BETA CHAINS OF FIVE SPECIES OF PINNIPEDS : NEOPHOCA CINEREA, OTARIA BYRONIA, EUMETOPIAS JUBATUS, PUSA HISPIDA, AND PAGOPHILUS GROENLANDICA

Pinnipeds (Otariidae, Odobenidae, and Phocidae) in the order Carnivora have one or two types (Hb I and Hb II) of hemoglobin components. These hemoglobins consist of identicalβ chains and differentα chains. We determined the complete amino acid sequences of the hemoglobinβ chain of three species of Otariidae (Australian sea lion, South American sea lion, and northern sea lion) and two species of Phocidae (ringed seal and harp seal) from intactβ chain and chemical cleavage fragments. The sequences are similar toβ chains of the already known sequences of pinnipeds. These sequences were compared with those of other carnivores (Mustelidae, Ursidae, Canidae, and Felidae) and adult human hemoglobinβ chain. Using Artiodactyla (pig) as an outgroup, we find that the tree constructed by means of phylogenetic analysis shows that Odobenidae is closest to Otariidae, and that Otariidae and Odobenidae are closer to Mustelidae than to Phocidae.

Expression of PDGF and C‐myc in Atherosclerotic Lesions in Cholesterol‐Fed Chicken

Platelet-derived growth factor (PDGF) has been reported to be a potent mitogen for mesenchymal cells such as smooth muscle cells and glial ce1ls.l Several reports2,’ have indicated that protooncogenes such as c-myc and c-fos may be involved in the mechanisms of atherosclerosis. The aim of this study is to investigate the role of PDGF and c-myc on the development of atherosclerotic lesions in chickens, which serve as a useful experimental model for the study of athero~clerosis.~ Experimental animals consisted of two groups. Six 4-month-old roosters were fed an atherogenic diet that contained 2% cholesterol and 10% corn oil for 3 months. Five age-matched normal roosters were fed basal diet. The thoracic and abdominal aortas were collected and histologically examined. Immunostaining was performed by the avidin-biotin-peroxidase complex (ABC) (Vector Labs., Burlingame, Calif.) method5 using primary antibodies of PDGF-A, PDGF-B (genzyme), PDGF receptor, Pan-myc (Cambridge Research Biochemicals Ltd.). In situ hybridization was carried out according to the method of Coxet ~ 1 . ~ Probes for detecting the PDGF-A, PDGF-B, PDGF-A receptor, and PDGF-B receptor mRNAs were purchased from Oncogene Science, Inc., and were biotinylated with the terminal labeling kit (Enzo Biochem, Inc.). The probe for c-myc mRNA was prepared by inserting the Sma I cut fragment (exon 11) of pMyc6.514 (offered by Japanese Cancer Research Resources Bank) into plasmid pSP65. According to the modified method of Melton et al.,’ biotinylated RNA probes were synthesized in vitro using sP6 RNA polymerase, the linearized plasmid DNA, and Biotin 1 I-UTP. Biotinylated probes on slides were visualized with Blu GENEm (BRL). A lipid-rich thickened intimal lesion was more frequently seen in the thoracic aortas than in the abdominal aortas of cholesterol-fed roosters. Immunohistochemicat examination disclosed no significant expression of PDGF-A, -B, PDGF receptors, and c-myc in the entire aorta of normal roosters. In cholesterol-fed roostcrs, the intense reaction of PDGF-B, PDGF receptor (PDGF-R), and c-myc was seen in lipid-rich thickened intimal lesions of the entire aorta while no significant reaction of PDGF-A was observed in the same lesions. In situ hybridization study demonstrated

Promotive effect of elastase on regression of aortic atherosclerosis in cholesterol-fed quails.

In order to test the anti-atherosclerotic function of elastase, 44 Japanese quails, 40 d of age, were used in this study. An atherogenic diet contained 15% corn oil and 2% cholesterol. Elaszym was orally administered at a dose of 6,000 EL units per kg body weight 3 times a week for 3 months. After 3 months feeding the atherogenic diet was discontinued. Moderate hypercholesterolemia and marked lipid-rich aortic lesions were noted in the group which was fed the atherogenic diet for 3 months. The thickened intima was composed of fibroblasts and alpha-1-anti-trypsin, S-100 protein, calmodulin and elastase were strongly demonstrated. Withdrawal of the atherogenic diet resulted in marked improvement of the serum cholesterol level, and slight reduction of the degree of the intimal thickening of the thoracic aorta. Elastase treatment after the withdrawal of atherogenic diet induced significant regression of the aortic lesions of the thoracic aorta. These results suggest that Elaszym possesses the promotive effect on regression of atherosclerotic lesions.