A novel anti-tumor protein extracted from Meretrix meretrix Linnaeus induces cell death by increasing cell permeability and inhibiting tubulin polymerization

Discovery and development of new pharmaceuticals from marine organisms are attracting increasing interest. Several agents derived from marine organisms are under preclinical and clinical evaluation as potential anticancer drugs. We extracted and purified a novel anti-tumor protein from the coelomic fluid of Meretrix meretrix Linnaeus by ammonium sulphate fractionation, ion exchange and hydrophobic interaction chromatography. The molecular weight of the highly purified protein, designated MML, was 40 kDa as determined by SDS-PAGE analysis. MML exhibited significant cytotoxicity to several cancer cell types, including human hepatoma BEL-7402, human breast cancer MCF-7 and human colon cancer HCT116 cells. However, no inhibitory effect was found when treating murine normal fibroblasts NIH3T3 and benign human breast MCF-10A cells with MML. The cell death induced by MML was characterized by cell morphological changes. The induction of apoptosis of BEL-7402 cells by MML was weak by DNA ladder assay. The possible mechanisms of its anti-tumor effect might be the changes in cell membrane permeability and inhibition of tubulin polymerization. MML may be developed as a novel, highly selective and effective anti-cancer drug.

Suspension culture of gametophytes of transgenic kelp in a photobioreactor

Transgenic Laminaria japonica gametophytes producing a recombinant tissue-type plasminogen activator (rtPA) protein, which is an effective third-generation thrombolytic agent for acute myocardial infarction (AMI), were cultured in an illuminated bubble column bioreactor. A maximum final dry cell weight of 1120 mg l(-1) was obtained in batch culture with an initial dry cell weight of 126 mg l(-1) and with aeration rate of 1.2 l air min(-1) l(-1) culture, nitrate at 1.5 mM and phosphate at 0.17 mM. The yield of rtPA was 56 mu g g(-1) dry cell wt. This is the first report regarding cultivation of a transgenic macroalga in a bioreactor.

Haematopoietic lineage cell-specific protein 1 (HS1) promotes actin-related protein (Arp) 2/3 complex-mediated actin polymerization

HS1 (haematopoietic lineage cell-specific gene protein 1), a prominent substrate of intracellular protein tyrosine kinases in haematopoietic cells, is implicated in the immune response to extracellular stimuli and in cell differentiation induced by cytokines. Although HS1 contains a 37-amino acid tandem repeat motif and a C-terminal Src homology 3 domain and is closely related to the cortical-actin-associated protein cortactin, it lacks the fourth repeat that has been shown to be essential for cortactin binding to filamentous actin (F-actin). In this study, we examined the possible role of HS1 in the regulation of the actin cytoskeleton. Immunofluorescent staining demonstrated that HS1 co-localizes in the cytoplasm of cells with actin-related protein (Arp) 2/3 complex, the primary component of the cellular machinery responsible for de novo actin assembly. Furthermore, recombinant HS1 binds directly to Arp2/3 complex with an equilibrium dissociation constant (K-d) of 880 nM. Although HS1 is a modest F-actin-binding protein with a Kd of 400 nM, it increases the rate of the actin assembly mediated by Arp2/3 complex, and promotes the formation of branched actin filaments induced by Arp2/3 complex and a constitutively activated peptide of N-WASP (neural Wiskott-Aldrich syndrome protein). Our data suggest that HS1, like cortactin, plays an important role in the modulation of actin assembly.

Density-dependent effects on seston dynamics and rates of filtering and biodeposition of the suspension-cultured scallop Chlamys farreri in a eutrophic bay (northern China): An experimental study in semi-in situ flow-through systems

Effects of stocking density on seston dynamics and filtering and biodeposition by the suspension-cultured Zhikong scallop Chlamys farreri Jones et Preston in a eutrophic bay (Sishili Bay, northern China), were determined in a 3-month semi-field experiment with continuous flow-through seawater from the bay. Results showed that the presence of the scallops could strongly decrease seston and chlorophyll a concentrations in the water column. Moreover, in a limited water column, increasing scallop density could cause seston depletion due to scallop's filtering and biodeposition process, and impair scallop growth. Both filtration rate and biodeposition rate of C. farreri showed significant negative correlation with their density and positive relationship with seston concentration. Calculation predicts that the daily removal of suspended matter from water column by the scallops in Sishili Bay ecosystem can be as high as 45% of the total suspended matter; and the daily production of biodeposits by the scallops in early summer in farming zone may amount to 7.78 g m(-2), with daily C, N and P biodeposition rates of 3.06 x 10(-1), 3.86 x 10(-2) and 9.80 x 10(-3) g m(-2), respectively. The filtering and biodeposition by suspension-cultured scallops could substantially enhance the deposition of total suspended particulate material, suppress accumulation of particulate organic matter in water column, and increase the flux of C, N and P to benthos, strongly enhancing pelagic-benthic coupling. It was suggested that the filtering-biodeposition process by intensively suspension-cultured bivalve filter-feeders could exert strong top-down control on phytoplankton biomass and other suspended particulate material in coastal ecosystems. This study also indicated that commercially suspension-cultured bivalves may simultaneously and potentially aid in mitigating eutrophication pressures on coastal ecosystems subject to anthropogenic N and P loadings, serving as a eutrophic-environment bioremediator. The ecological services (e.g. filtering capacity, top-down control, and benthic-pelagic coupling) functioned by extractive bivalve aquaculture should be emphasized in coastal ecosystems. (c) 2005 Elsevier B.V. All rights reserved.

A two-stage process with temperature-shift for enhanced anthocyanin production in strawberry cell suspension cultures

A two-stage process with temperature-shift has been developed to enhance the anthocyanin yield in suspension cultures of strawberry cells. The effect of the temperature-shift interval and the shift-time point was investigated for the optimization of this strategy. In this process, strawberry cells were grown at 30 degrees C (the optimum temperature for cell growth) for a certain period as the first stage, with the temperature shifted to a lower temperature for the second stage. In response to the temperature shift-down, anthocyanin synthesis was stimulated and a higher content could be achieved than that at both boundary temperatures but cell growth was suppressed. When the lower boundary temperature was decreased, cell growth was lowered and a delayed, sustained maximum anthocyanin content was achieved. Anthocyanin synthesis was strongly influenced by the shift-time point but cell growth was not. Consequently, the maximum anthocyanin content of 2.7 mg.g-fresh cell(-1) was obtained on day 9 by a temperature-shift from 30 degrees C, after 3-d culture, to 15 degrees C. The highest anthocyanin yield of 318 mg.L-1 on day 12 was achieved when the temperature was shifted from 30 degrees C, after 5-d culture, to 20 degrees C. For a global optimization of both the yield and productivity, the optimum anthocyanin yield and productivity of 272 mg.L-1 and 30.2 mg.L-1.d(-1) on day 9 were achieved by a two-stage culture with a temperature-shift from 30 degrees C after 3 d to 20 degrees C.

Isospecific polymerization of styrene with modified ziegler-type catalysts

Effects of various kinds of additives as well as aging of the catalyst on the polymerization of styrene catalyzed by TiCl4/MgCl2-AlEt3 system have been studied. Experiments show that in toluene the isotacticity of polystyrene can be up to 83% for aged catalyst, whereas when the catalyst is not aged. non-stereospecific polymer is the main product. When PCl3 is used as an additive, the catalyst system gives high activity and isotacticity. The use of a mixture of AlEt3/H2O (1: 1 mole ratio) as a cocatalyst is also efficient. The catalyst [TiCl4-PCl3/MgCl2-AlEt3/H2O] displays high activity and product isotacticity (94%) with an average molecular weight up to 2 X 10(-6). When Co(acac)(3) is added to to [TiCl4/MgCl2-AlEt3] catalyst after it was aged, the isotacticity can be up to 97%. (C) 2001 Elsevier Science Ltd. All rights reserved.

Instability of anthocyanin accumulation in Vitis vinifera L. var. Gamay Freaux suspension cultures

The inherent instability of metabolite production in plant cell culture-based bioprocessing is a major problem hindering its commercialization. To understand the extent and causes of this instability, this study was aimed at understanding the variability of anthocyanin accumulation during long-term subcultures, as well as within subculture batches, in Vitis vinifera cell cultures. Therefore, four cell line suspensions of Vitis vinitera L. var. Gamay Freaux, A, B, C and D, originated from the same callus by cell-aggregate cloning, were established with starting anthocyanin contents of 2.73 +/- 0.15, 1.45 +/- 0.04, 0.77 +/- 0.024 and 0.27 +/- 0.04 CV (Color Value)/g-FCW (fresh cell weight), respectively. During weekly subculturing of 33 batches over 8 months, the anthocyanin biosynthetic capacity was gradually lost at various rates, for all four cell lines, regardless of the significant difference in the starting anthocyanin content. Contrary to this general trend, a significant fluctuation in the anthocyanin content was observed, but with an irregular cyclic pattern. The variabilities in the anthocyanin content between the subcultures for the 33 batches, as represented by the variation coefficient (VC), were 58, 57, 54, and 84% for V vinifera cell lines A, B, C and D, respectively. Within one subculture, the VCs from 12 replicate flasks for each of 12 independent subcultures were averaged, and found to be 9.7%, ranging from 4 to 17%. High- and low-producing cell lines, VV05 and VV06, with 1.8-fold differences in their basal anthocyanin contents, exhibited different inducibilities to L-phenylalanine feeding, methyl jasmonate and light irradiation. The low-producing cell line, showed greater potential in enhanced the anthocyanin production.