Chondromodulin I is a bone remodeling factor

Chondromodulin I (ChM-I) was supposed from its limited expression in cartilage and its functions in cultured chondrocytes as a major regulator in cartilage development. Here, we generated mice deficient in ChM-I by targeted disruption of the ChM-I gene. No overt abnormality was detected in endochondral bone formation during embryogenesis and cartilage development during growth stages of ChM-I(-/-) mice. However, a significant increase in bone mineral density with lowered bone resorption with respect to formation was unexpectedly found in adult ChM-I(-/-) mice. Thus, the present study established that ChM-I is a bone remodeling factor.     

Isolation and characterization of thermophilic bacilli degrading cinnamic, 4-coumaric,and ferulic acids

Thirty-four thermophilic Bacillus sp. strains were isolated from decayed wood bark and a hot spring water sample based on their ability to degrade vanillic acid under thermophilic conditions. It was found that these bacteria were able to degrade a wide range of aromatic acids such as cinnamic, 4-coumaric, 3-phenylpropionic, 3-(p-hydroxyphenyl)propionic, ferulic, benzoic, and 4-hydroxybenzoic acids. The metabolic pathways for the degradation of these aromatic acids at 60 degrees C were examined by using one of the isolates, strain B1. Benzoic and 4-hydroxybenzoic acids were detected as breakdown products from cinnamic and 4-coumaric acids, respectively. The beta-oxidative mechanism was proposed to be responsible for these conversions. The degradation of benzoic and 4-hydroxybenzoic acids was determined to proceed through catechol and gentisic acid, respectively, for their ring fission. It is likely that a non-beta-oxidative mechanism is the case in the ferulic acid catabolism, which involved 4-hydroxy-3-methoxyphenyl-beta-hydroxypropionic acid, vanillin, and vanillic acid as the intermediates. Other strains examined, which are V0, D1, E1, G2, ZI3, and H4, were found to have the same pathways as those of strain B1, except that strains V0, D1, and H4 had the ability to transform 3-hydroxybenzoic acid to gentisic acid, which strain B1 could not do.     

Carbon autonomy of reproductive shoots of Siberian alder (<Emphasis Type="Italic">Alnus hirsuta</Emphasis> var.<Emphasis Type="Italic"> sibirica</Emphasis>)

Carbon autonomy of current-year shoots in flowering, and of current-year shoots plus 1-year-old shoots (1-year-old shoot system) in fruiting of Siberian alder (Alnus hirsuta var. sibirica) was investigated using a stable isotope of carbon, ¹³C. The current-year shoot and 1-year-old shoot systems were fed ¹³CO₂ and the atom% excess of ¹³C in flowers and fruits was determined. The majority of photosynthate allocated to flower buds was originally assimilated in the leaves of the flowering current-year shoots. Of all the current-year shoots on fruiting 1-year-old shoots, only those nearest to the fruits allocated the assimilated photosynthate to fruit maturation. These results indicate that the current-year shoots and 1-year-old shoot systems are carbon-autonomous units for producing flowers and maturing fruits, respectively.     

Stable Transfection of GM1 Synthase Gene into GM1-Deficient NG108-15 Cells,CR-72 Cells,Rescues the Responsiveness of TRK-Neurotrophin Receptor to Its Ligand,NGF

Previous studies from this laboratory and others have suggested the evidences that acidic glycosphingolipid, ganglioside GM1 (GM1), is an endogenous regulator of high affinity nerve growth factor receptor, Trk, which is an essential factor for the normal development and differentiation of neuronal cells by forming a complex with Trk. The present study was aimed to examine whether Trk expressed in cells that are deficient in endogenous GM1 due to the mutation of GM1 synthase gene (NG-CR72 cells) is responsive to its ligand nerve growth factor and how genetic restoration of GM1 synthase gene by a stable transfection of the gene affects the function of the Trk protein. The data clearly showed that (1) confocal lazor microscopic studies disclosed NG-CR72 cells are really deficient in GM1, (2) stable transfection of GM1 synthase cDNA into these cells (NG-CR72G cells) restores the expression of GM1 in the cells, and (3) Trk protein is expressed in NG-CR72 cells but its location seemed not to be on the plasma membrane, whereas we clearly observed that the Trk protein is expressed on the plasma membrane in NG-CR72G cells. (4) NGF did not elicit the autophosphorylation of the Trk protein in GM1 deficient NG-CR72 cells but did elicit the activation of the Trk protein in NG-CR72G cells with an activation of mitogen activated protein kinase. These studies strongly suggested that GM1 is necessary for the normal expression of the Trk protein function and for normal targeting of the Trk protein to the plasma membrane.     

Predominant growth of Alcanivorax strains in oil-contaminated and nutrient-supplemented sea water

We found that bacteria closely related to Alcanivorax became a dominant bacterial population in petroleum-contaminated sea water when nitrogen and phosphorus nutrients were supplied in adequate quantity. The predominance of Alcanivorax bacteria was demonstrated under three experimental conditions: (i) in batch cultures of sea water containing heavy oil; (ii) in columns packed with oil-coated gravel undergoing a continuous sea water flow; and (iii) in a large-scale tidal flux reactor that mimics a beach undergoing tidal cycles with fresh sea water. These results suggest that bacteria related to Alcanivorax are major players in the bioremediation of oil-contaminated marine environments.     

Interferon-α and interleukin-12 gene therapy of cancer: interferon-α induces tumor-specific immune responses while interleukin-12 stimulates non-specific killing

Cytokine gene therapy is applied in clinical studies of tumors, and IFN-alpha and IL-12 are widely used for cancer immunotherapy. Using a poorly immunogenic murine colorectal cancer cell line, MC38, we compared antitumor effects of IFN-alpha and IL-12. Transduced MC38 cell lines expressing IFN-alpha or IL-12 (MC38-IFNalpha or MC38-IL12, respectively) were established using retroviral vectors. Transduction of IFN-alpha or IL-12 gene to MC38 cells significantly reduced tumorigenicity in immunocompetent mice. When tumor-free mice initially injected with MC38-IFNalpha or MC38-IL12 cells were reinjected contralaterally with wild-type MC38 cells (MC38-WT) after 35 days, 7 of 12 or 2 of 12 mice rejected MC38-WT cells, respectively. In therapy-model mice with established tumor derived from MC38-WT cells, inoculation of gene-transduced cells significantly suppressed growth of the tumor in MC38-IFNalpha-inoculated groups, but not in the IL-12-inoculated group. Immunohistologic and flow cytometric analyses showed marked infiltration of CD8(+) cells in wild-type tumors of mice inoculated with IFN-alpha-expressing cells. Leukocyte-depletion experiments implicated CD8(+) T cells in tumor rejection induced by IFN-alpha-transduction; both CD8(+) T cells and natural killer cells were implicated in the more modest antitumor effect from IL-12 expression. To investigate induction of tumor-specific immune responses, we stimulated splenocytes from tumor-free mice twice in vitro with genetically modified MC38 cells. In vitro stimulations with MC38-IFNalpha cells induced definite MC38-specific lysis, but not stimulations with MC38-IL-12 cells. Injecting combination of MC38-IFNalpha and MC38-IL-12 cells caused an additive antitumor effect in the therapy model. These data suggested that IFN-alpha induces cytotoxic T lymphocytes and elicits long-lasting tumor-specific immunity, whereas IL-12 seems to stimulate non-specific killing. With additional refinements, combined IFN-alpha and IL-12 gene therapy might warrant clinical trials.