The metalloproteinase ADAM‐12 regulates bronchial epithelial cell proliferation and apoptosis

Abstract. Objectives: The ADAMs (a disintegrin and metalloproteinase) enzymes compose a family of membrane‐bound proteins characterized by their multi‐domain structure and ADAM‐12 expression is elevated in human non‐small cell lung cancers. The aim of this study was to investigate the roles played by ADAM‐12 in critical steps of bronchial cell transformation during carcinogenesis. Materials and methods: To assess the role of ADAM‐12 in tumorigenicity, BEAS‐2B cells were transfected with a plasmid encoding human full‐length ADAM‐12 cDNA, and then the effects of ADAM‐12 overexpression on cell behaviour were explored. Treatment of clones with heparin‐binding epidermal growth factor (EGF)‐like growth factor (HB‐EGF) neutralizing antibodies as well as an EGFR inhibitor allowed the dissection of mechanisms regulating cell proliferation and apoptosis. Results: Overexpression of ADAM‐12 in BEAS‐2B cells promoted cell proliferation. ADAM‐12 overexpressing clones produced higher quantities of HB‐EGF in their culture medium which may rely on membrane‐bound HB‐EGF shedding by ADAM‐12. Targeting HB‐EGF activity with a neutralizing antibody abrogated enhanced cell proliferation in the ADAM‐12 overexpressing clones. In sharp contrast, targeting of amphiregulin, EGF or transforming growth factor‐α failed to influence cell proliferation; moreover, ADAM‐12 transfectants were resistant to etoposide‐induced apoptosis and the use of a neutralizing antibody against HB‐EGF activity restored rates of apoptosis to be similar to controls.Conclusions: ADAM‐12 contributes to enhancing HB‐EGF shedding from plasma membranes leading to increased cell proliferation and reduced apoptosis in this bronchial epithelial cell line.     

Hep G2 cells as a resource for metabolic studies: lipoprotein, cholesterol, and bile acids

Hep G2, a liver cell line derived from a human hepatoblastoma that is free of known hepatotropic viral agents, has been found to express a wide variety of liver-specific metabolic functions. Among these functions are those related to cholesterol and triglyceride metabolism. Confluent Hep G2 monolayers express normal low-density lipoprotein (LDL) receptors and continue to internalize and metabolize chylomicrons, very low-density lipoproteins (VLDL), LDL, and high-density lipoproteins. In lipoprotein-free medium, apolipoproteins A-I, A-II, B, C, and E accumulate in the medium together with cholesterol, cholesteryl ester, triglyceride, and all the primary bile acids. The regulation of their synthesis and secretion is not fully known and their interrelationships have not been established. Because Hep G2 cells express these and other components of cholesterol and triglyceride metabolism, they are a microcosm for studying the central role of the liver.     

The formation of a test system for assessing the safety of different types of pertussis preparations: their cytotoxic action on mouse thymocytes in vitro

The test for the evaluation of the toxicity of different types of pertussis preparations as manifested by their in vitro influence on mouse thymic cells (T test) has been finally worked out. The use of the T test has made it possible to reveal the nonstandard character of the production lots of adsorbed diphtheria-pertussis-tetanus vaccines, both whole-cell vaccine and Japanese acellular vaccine. The degree of the in vitro damaging action of pertussis preparations on mouse thymic cells greatly depends on the residual content of Bordetella pertussis nontoxoidized toxin which, in contrast to B. pertussis lipopolysaccharide and filamentous hemagglutinin, produces pronounced cytotoxic action on mouse thymic cells.     

Hydrogen production by rumen holotrich protozoa: effects of oxygen and implications for metabolic control by in situ conditions

Experiments with washed suspensions of holotrich protozoa (Isotricha spp. and Dasytricha ruminantium) showed that both organisms have an efficient O2-scavenging capability (apparent Km values 2.3 and 0.3 microM, respectively). Reversible inhibition of H2 production increased almost linearly with increasing O2 up to 1.5 microM; higher levels of O2 gave irreversible inhibition. In situ determinations of H2, CH4, O2 and CO2 in ovine rumen liquor, using a membrane inlet mass spectrometer probe, indicated that O2 was present before feeding at 1-1.5 microM and decreased to undetectable levels (less than 0.25 microM) within 25 min after feeding. A transient increase in O2 concentration after feeding occurred only in defaunated animals and resulted in suppression of CH4 and CO2 production. The presence of washed holotrich protozoa decreases the O2 sensitivity of CH4 production by suspensions of a cultured methanogenic bacterium Methanosarcina barkeri. It is concluded that holotrich protozoa play a role in ruminal O2 utilization as well as in the production of fermentation end products (especially short-chain volatile fatty acids) utilized by the ruminant and H2 utilized by methanogenic bacteria. These hydrogenosome-containing protozoa thus both control patterns of fermentation by influencing O2 levels, and are themselves regulated by the low ambient O2 concentrations they experience in the rumen.     

Effect of psychotropic substances on the development of alcoholic motivation in noninbred white rats

The experiments have shown the capacity of antidepressant amitriptylin (0.5 mg/kg, i. p.) and tranquilizer phenazepam (0.1 mg/kg i. p.) to normalize the adaptive behaviour and almost completely prevent the development of alcohol motivation in animals with insufficient adaptive behaviour. It was established that in animals initially rejecting alcohol, chronic treatment with these drugs as well as d-amphetamine promoted alcohol motivation. The results obtained have proved our earlier hypothesis that preclinical search for drugs for the prevention and treatment of early stages of alcoholism is possible only in animals pre-selected according to their inclination to experimental alcoholism.     

Demonstration of the presence of G-proteins in hepatic microsomal fraction

The presence of G-proteins in isolated hepatic microsomal vesicles is demonstrated. The G-proteins were identified by their capacity to be ADP-ribosylated by cholera and pertussis toxins. Cholera toxin identified 42 and 45 kDa proteins, corresponding to alpha s-1 and alpha s-2, respectively. Pertussis toxin identified a 40 kDa protein corresponding to alpha i. The microsomal G-proteins are identical to the corresponding G proteins of the plasma membrane, but are present in different proportions; the microsomes have considerably less alpha s proteins than the plasma membrane.