Anaerobic digestion of alfalfa silage with recirculation of process liquid.

Process liquid recirculation initially stimulated one-phase anaerobic digestion of alfalfa silage in two semi-continuously fed and stirred tank reactors. Thus, with increased pH, alkalinity and stability it was possible to increase the organic loading rate to 3 g VS L(-1) d(-1), as compared to 2.25 g VS L(-1) d(-1) in a control reactor without recirculation. However, the recirculation of liquid eventually caused an accumulation of organic and inorganic substances, leading to an inhibition of hydrolysis and methanogenesis. This inhibition of microbial activity was prevented in one of the processes by replacing 50% of the recirculated process liquid with water during the second half of the operation period. A multiple linear regression model of principal components using seven input variables explained the variance in output variables nearly as well as the original model using all 23 measured input variables. The results show that it is necessary to adjust the degree of liquid recirculation to reach an optimal process.

Fibroblast heterogeneity in collagenolytic response to colchicine.

Matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are important in various physiological and pathological conditions, including those that involve homeostasis of collagen. Drug induced regulation of MMP-1, other MMPs and TIMPs is critical in treatment of various diseases, e.g. the use of the plant alkaloid, colchicine. One possible factor that might explain the failure in colchicine-treatment of some patients is interindividual variability on the cellular level. To investigate the possible individual heterogeneity in response to colchicine, we studied the effect of colchicine-induced synthesis of collagenase from 32 different human skin fibroblast strains derived from both healthy individuals as well as individuals with different skin diseases. We showed that colchicine induced an increased synthesis of collagenase in 22 of 32 cases. This heterogeneity occurred in fibroblasts from healthy as well as diseased individuals. To determine if colchicine also affected the fibroblast synthesis of gelatinase, stromelysin and tissue inhibitors of MMPs, we investigated several individuals from a single family. The results showed that both colchicine responsive and non-responsive fibroblasts with respect to collagenase synthesis responded to colchicine by an increased stromelysin synthesis, while the synthesis of gelatinase and TIMP-1 were unaffected. As a whole, our results indicate that individual heterogeneity in collagenase response to colchicine treatment may partly explain some of the controversial results obtained with colchicine as a drug.

Colchicine induces membrane-associated activation of matrix metalloproteinase-2 in osteosarcoma cells in an S100A4-independent manner.

Like the metastasis-associated protein S100A4, matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) are important in physiological and pathological conditions. Previously, we showed that S100A4 is involved in the regulation of MMPs and TIMPs, and in the present work we have investigated whether the anti-inflammatory and microtubule-disrupting drug colchicine has an effect on the expression of these proteins in osteosarcoma cell lines (OHS) with high and low levels of S100A4. Colchicine treatment of the various OHS cells resulted in an increased expression of MT1-MMP and TIMP-2 mRNA, and a corresponding increase of these two proteins in isolated cell membranes. Colchicine-treated cells produced more of the activated form of MMP-2 than control cells. However, the drug did not affect the amount of MMP-2 and TIMP-1 mRNA or protein, and it reduced the S100A4 mRNA expression. Isolated cell membranes from the colchicine-treated cells were more effective in activating exogenous proMMP-2 than membranes from control cells, and inhibitory studies indicated that it was the colchicine-induced increase in MT1-MMP that caused the increased activation of endogenous MMP-2. A peptide inhibitor of nuclear factor kappaB nuclear translocation, SN50, blocked the colchicine-induced activation of proMMP-2 and reduced the synthesis of MMP-2 in colchicine-treated cells, but not in control cells. It can be concluded that colchicine modulates the expression of MT1-MMP and TIMP-2 and hence the activation of proMMP-2 independently of the S100A4 level in osteosarcoma cells.

Effects of glucose overloading on microbial community structure and biogas production in a laboratory-scale anaerobic digester.

This study characterizes the response of the microbial communities of a laboratory-scale mesophilic biogas process, fed with a synthetic substrate based on cellulose and egg albumin, to single pulses of glucose overloading (15 or 25 times the daily feed based on VS). The microbial biomass and community structure were determined from analyses of membrane phospholipids. The ratio between phospholipid fatty acids (PLFAs; eubacteria and eucaryotes) and di-ethers (PLEL; archaea) suggested that methanogens constituted 4-8% of the microbial biomass. The glucose addition resulted in transient increases in the total biomass of eubacteria while there were only small changes in community structure. The total gas production rate increased, while the relative methane content of the biogas and the alkalinity decreased. However, the biomass of methanogens was not affected by the glucose addition. The results show that the microbial communities of biogas processes can respond quickly to changes in the feeding rate. The glucose overload resulted in a transient general stimulation of degradation rates and almost a doubling of eubacterial biomass, although the biomass increase corresponded to only 7% of the glucose C added.

S100A4 regulates membrane induced activation of matrix metalloproteinase-2 in osteosarcoma cells.

To study the role of the metastasis associated protein S100A4, an osteosarcoma cell line (OHS) with a high level of this protein was transfected with a vector containing a ribozyme that degrades S100A4 mRNA and, as controls, OHS cells were transfected with the vector alone. We have followed up our previous investigation (Bjørnland et al. 1999) by a detailed investigation of these cell lines' synthesis of MMP and TIMP proteins at different cell densities. It is shown that the cell lines with a low S100A4 level produced a reduced amount of immunoreactive MMP-2 at cellular subconfluence, while at confluence there was no difference compared to the control cells. The cell lines with a reduced S100A4 level produced less of the activated form of MMP-2 (62-kDa) and less TIMP-1 than the corresponding control cells, independent of cell density. Isolated cell membranes from cell lines with a reduced S100A4 level contained less MT1-MMP, MMP-2 and TIMP-2 compared to the control cells. Activation of exogenously added proMMP-2 was less effective with the former membrane preparations. It appeared that the mechanism behind the S100A4 dependent activation of proMMP-2 varied with cell density, as SN50, a peptide inhibitor of NF-kappaB nuclear translocation reduced the activation of MMP-2 at low cell density, but had no effect at high cell density. Thus, one of the mechanisms by which S100A4 may exert its effect on metastasis of some tumors is by regulating the MMP-2 activity.