Gelsolin Induces Colorectal Tumor Cell Invasion via Modulation of the Urokinase-Type Plasminogen Activator Cascade

Gelsolin is a cytoskeletal protein which participates in actin filament dynamics and promotes cell motility and plasticity. Although initially regarded as a tumor suppressor, gelsolin expression in certain tumors correlates with poor prognosis and therapy-resistance. In vitro, gelsolin has anti-apoptotic and pro-migratory functions and is critical for invasion of some types of tumor cells. We found that gelsolin was highly expressed at tumor borders infiltrating into adjacent liver tissues, as examined by immunohistochemistry. Although gelsolin contributes to lamellipodia formation in migrating cells, the mechanisms by which it induces tumor invasion are unclear. Gelsolin's influence on the invasive activity of colorectal cancer cells was investigated using overexpression and small interfering RNA knockdown. We show that gelsolin is required for invasion of colorectal cancer cells through matrigel. Microarray analysis and quantitative PCR indicate that gelsolin overexpression induces the upregulation of invasion-promoting genes in colorectal cancer cells, including the matrix-degrading urokinase-type plasminogen activator (uPA). Conversely, gelsolin knockdown reduces uPA levels, as well as uPA secretion. The enhanced invasiveness of gelsolin-overexpressing cells was attenuated by treatment with function-blocking antibodies to either uPA or its receptor uPAR, indicating that uPA/uPAR activity is crucial for gelsolin-dependent invasion. In summary, our data reveals novel functions of gelsolin in colorectal tumor cell invasion through its modulation of the uPA/uPAR cascade, with potentially important roles in colorectal tumor dissemination to metastatic sites.     

Substrate specificity of the deazaflavin-dependent nitroreductase from Mycobacterium tuberculosis responsible for the bioreductive activation of bicyclic nitroimidazoles

The bicyclic 4-nitroimidazoles PA-824 and OPC-67683 represent a promising novel class of therapeutics for tuberculosis and are currently in phase II clinical development. Both compounds are pro-drugs that are reductively activated by a deazaflavin (F(420)) dependent nitroreductase (Ddn). Herein we describe the biochemical properties of Ddn including the optimal enzymatic turnover conditions and substrate specificity. The preference of the enzyme for the (S) isomer of PA-824 over the (R) isomer is directed by the presence of a long hydrophobic tail. Nitroimidazo-oxazoles bearing only short alkyl substituents at the C-7 position of the oxazole were reduced by Ddn without any stereochemical preference. However, with bulkier substitutions on the tail of the oxazole, Ddn displayed stereospecificity. Ddn mediated metabolism of PA-824 results in the release of reactive nitrogen species. We have employed a direct chemiluminescence based nitric oxide (NO) detection assay to measure the kinetics of NO production by Ddn. Binding affinity of PA-824 to Ddn was monitored through intrinsic fluorescence quenching of the protein facilitating a turnover-independent assessment of affinity. Our results indicate that (R)-PA-824, despite not being turned over by Ddn, binds to the enzyme with the same affinity as the active (S) isomer. This result, in combination with docking studies in the active site, suggests that the (R) isomer probably has a different binding mode than the (S) with the C-3 of the imidazole ring orienting in a non-productive position with respect to the incoming hydride from F(420). The results presented provide insight into the biochemical mechanism of reduction and elucidate structural features important for understanding substrate binding.     

Application of motion analysis system in pre-impact fall detection

The purpose of this study is to investigate unique features of body segments in fall and activities of daily living (ADL) to make automatic detection of fall in its descending phase before the impact. Thus, fall-related injuries can be prevented or reduced by deploying feedback systems before the impact. In this study, the authors propose the following hypothesis: (1) thigh segment normally does not go beyond certain threshold angle to forward and sideways directions in ADL and (2) even if it does, the angular characteristics measured at torso and thigh differ from one another in ADL whereas in the case of fall, they become congruent. These two factors can be used to distinguish fall from ADL in its inception. Vicon 3-D motion analysis system was used in this study. High level of correlation between thigh and torso segments (corr > 0.99) was found for fall activities and low correlation coefficients (mean corr for lateral movements is 0.2338 and for sagittal movements is -0.665) were observed in ADL. By applying the hypothesis, all simulated falls could be detected with no false alarms and around 700ms lead-time before the impact was achieved in pre-impact fall detection. It is the longest lead-time obtained so far in pre-impact fall detection.     

Biodegradation of dimethyl phthalate by Sphingomonas sp. isolated from phthalic-acid-degrading aerobic granules

This study isolated nine strains of aerobic phenol-degrading granules. These isolates (I1–I9) were characterized using 16S rRNA gene sequencing, with γ-Proteobacteria as the dominant strains in the aerobic granules. While most strains demonstrated either high phenol-degrading capabilities or auto-aggregation capabilities, three isolates, I2, I6, and I8 showed both features. These findings contradict the previous view that auto-aggregation and phenol degradation are mutually exclusive in aerobic granules. Strains I2 and I8 independently formed single-culture aerobic granules except for I3. Anti-microbial activity test results indicated that strains I2 and I8 inhibited growth of strain I3. However, co-culturing I3 with I2 or I8 helped to form granules.     

Influence of starvation time on formation and stability of aerobic granules in sequencing batch reactors

Three sequencing batch reactors, R1, R2 and R3, with a 1.5-h, 4-h and 8-h cycle time, respectively, were used to cultivate aerobic granules with the same synthetic wastewater containing 1000 mg l(-1) COD. As the initial COD concentrations in the cycles were the same, three different cycle times led to three different starvation times in repeated cycles of the three reactors. It was found that 63 cycles were needed to form granules with the longest starvation time in R3 while it took 256 cycles in R1 with the shortest starvation time. However, as far as the formation time was concerned, granules were formed on day 16 with 1.5-h cycle time while on day 21 with 8-h cycle time, which indicated that a shorter cycle time with a shorter starvation time speeded up the granulation. This was mainly due to the stronger hydraulic selection pressure at shorter cycle time. However, it was found that granules formed with cycle time of 1.5h were unstable. Fluffy granules with poor settling ability were observed in R1 in the 4th month, which led to the collapse of R1 after 160-day of operation. Granules in R2 and R3 showed good stability during the long-term operation. Therefore, a reasonable starvation time was necessary to maintain the long-term stability of aerobic granules.     

Effect of Iron Hydroxide on Phosphate Removal during Anaerobic Digestion of Activated Sludge

The addition of iron (III) hydroxide during methanogenic digestion of activated sludge by anaerobic sludge displaying an iron-reducing activity resulted in a microbial reduction of iron (III) with the formation of iron (II), capable of precipitating phosphates. The feasibility of eliminating 66.6 to 99.6% of the dissolved phosphate at initial concentrations of 1000 to 3500 mg PO^3- ₄/l by adding 6420 mg/l iron (III) hydroxide into a reactor for anaerobic fermentation of activated sludge was analyzed. The optimal ratio of iron (III) added to dissolved phosphate removed (mg) providing a 95% removal amounted to 2 : 1. These results may be used in new technology for anaerobic wastewater treatment with phosphate removal.     

Bacillus naphthovorans sp. nov. from oil-contaminated tropical marine sediments and its role in naphthalene biodegradation

. A Bacillus sp., designated as strain MN-003, was isolated as the dominant cultivatable naphthalene-degrading organism from oil-contaminated tropical marine sediments. Strain MN-003 is strictly aerobic, rod-shaped, Gram-positive, catalase positive, oxidase negative, and forms endospores. Strain MN-003 grew at salinities ranging from 0.28 to 7.00% and temperatures ranging from 15 to 41°C. Phylogenetic analyses reveal that strain MN-003 is most similar to Bacillus sp. VAN14, with a 16S rRNA sequence identity of 97.9%. Based on taxonomic and 16S rRNA data, strain MN-003 was named Bacillus naphthovorans sp. nov. When grown with naphthalene as sole carbon source, strain MN-003 had a maximal specific growth rate (µmax) of 0.32ǂ.03 h^–1, and a half-saturation constant (Ks) of 22.3dž.2 µM. A batch study of the tropical marine sediments enriched with naphthalene showed that cells of the Bacillus genus grew to become dominant members of the microbial community. The bacilli comprised 39.5Lj.5% of the microbial fraction after 20 days of enrichment.     

Partial ischemia and proteinuria during isolated kidney perfusion is accompanied by the release of vascular [35S]heparan sulfate

The isolated kidney perfused with modified Krebs-Henseleit buffer with amino acids yields heavy proteinuria associated with reduced ATP levels characteristic of partial ischemia. These conditions are associated with a similar perfusion time dependent release of degraded vascular [35S]heparan sulfate proteoglycan into the perfusate solution which included a 60% loss of [35S]macromolecular material from the glomerulus after 2h of perfusion. Small amounts of [35S]macromolecular material were found in the urine and lymph. These results demonstrate that partial ischemia promotes a specific response in the overall renal vasculature, probably involving oxygen reactive metabolites, that results in the preferential release of heparan sulphate from the basement membrane and endothelial cells on the luminal side of the capillary wall.     

Isomeric, anti-rhamnose antibodies having specificity for rhamnose-containing, streptococcal heteroglycans

L-Rhamnose (6-deoxy-L-mannose) is a constituent carbohydrate unit of microbial, immunogenic heteroglycans and lipopolysaccharides, and often functions as the immunodeterminant group of such immunogens. Two types of anti-rhamnose antibody have now been isolated by affinity chromatography of immune sera obtained from rabbits immunized with vaccines of Streptococcus mutans, strain KI-R, and Streptococcus pneumoniae, type 32. The antibodies of one type were directed at a glycan of L-rhamnose, D-glucose, and D-galactose in the cell wall of S. mutans, and those of the other type, against a capsular glycan of L-rhamnose and D-glucose from S. pneumoniae. The two types of anti-rhamnose antibody were immunologically distinct, and showed no reciprocal cross-reactivity. Additional properties of the two types of antibody were determined; thus, both types of antibody were of the IgG class of immunoglobulins, both possessed molecular weights of 1.45 X 10(5), and both consisted of multiple or isomeric forms.