Alterations in the chondroitin sulfate chain in human osteoarthritic cartilage of the knee.

OBJECTIVE: To determine whether the structure of chondroitin sulfate (CS) in cartilage is reflected by the degree of cartilage degeneration in patients with osteoarthritis (OA) of the knee and to determine how CS biosynthesis affects cartilage degeneration. DESIGN: Two osteoarthritic cartilage samples were obtained from medial femoral condyle (MFC) and lateral femoral condyle (LFC) of 24 knees with end-stage OA. The samples were assigned to two groups as follows: lesion and remote cartilage were adjacent to and remote from the osteoarthritic cartilage, respectively. Histological grade was determined according to the Mankin score. The CS concentration and chain length were determined using high-performance liquid chromatography (HPLC) and gel filtration chromatography, respectively. Expression of the gene encoding CS glycosyltransferase was evaluated using a real-time quantitative polymerase chain reaction (qPCR) assay. These results were compared between lesion and remote cartilage. RESULTS: The Mankin score indicated that lesion cartilage was more degraded compared with remote cartilage. Although the CS levels varied among individuals, the mean CS concentration and chain length were significantly lower and shorter in lesion cartilage than in remote cartilage, respectively (concentration: 12.04 vs 14.84 µg/mg wet weight, P = 0.021; chain length: 5.36 vs 6.19 kDa, P = 0.026). Three genes encoding CS glycosyltransferases (CHPF, CSGALNACT1, CSGALNACT2) were expressed at lower levels in lesion cartilage. CONCLUSIONS: In the osteoarthritic knee, the CS concentration and chain length were reduced closer to the more degraded cartilage with decreasing CS glycosyltransferase gene expression. Inhibition of CS glycosyltransferase gene expression may reduce CS chain length, which may contribute to OA progression.

Enhanced removal of chemical oxygen demand, nitrogen and phosphorus using the ameliorative anoxic/anaerobic/oxic process and micro-electrolysis.

Synthetic wastewater was treated using a novel system integrating the reversed anoxic/anaerobic/oxic (RAAO) process, a micro-electrolysis (ME) bed and complex biological media. The system showed superior chemical oxygen demand (COD), total nitrogen (TN) and total phosphorus (TP) removal rates. Performance of the system was optimised by considering the influences of three major controlling factors, namely, hydraulic retention time (HRT), organic loading rate (OLR) and mixed liquor recirculation (MLR). TP removal efficiencies were 69, 87, 87 and 83% under the HRTs of 4, 8, 12 and 16 h. In contrast, HRT had negligible effects on the COD and TN removal efficiencies. COD, TN and TP removal efficiencies from synthetic wastewater were 95, 63 and 87%, respectively, at an OLR of 1.9 g/(L·d). The concentrations of COD, TN and TP in the effluent were less than 50, 15 and 1 mg/L, respectively, at the controlled MLR range of 75-100%. In this system, organics, TN and TP were primarily removed from anoxic tank regardless of the operational conditions.

Quantification of hepatitis C virus in patients treated with peginterferon-alfa 2a plus ribavirin treatment by COBAS TaqMan HCV test.

Extremely low levels of serum hepatitis C virus (HCV) RNA can be detected by COBAS TaqMan HCV test. To investigate whether the COBAS TaqMan HCV test is useful for measuring rapid virological response (RVR) and early virological response (EVR) to predict sustained virological response (SVR), we compared the virological response to PEG-IFN-alfa 2a plus RBV in 76 patients infected with HCV genotype 1 when undetectable HCV RNA by the COBAS TaqMan HCV test was used, with those when below 1.7 log IU/mL HCV RNA by COBAS TaqMan HCV test was used, which corresponded to the use of traditional methods. Among the 76 patients, 28 (36.8%) had SVR, 13 (17.1%) relapsed, 19 (25.0%) did not respond, and 16 (21.0%) discontinued the treatment due to side effects. The positive predictive values for SVR based on undetectable HCV RNA by COBAS TaqMan HCV test at 24 weeks after the end of treatment [10/10 (100%) at week 4, 21/23 (91.3%) at week 8 and 26/33 (78.7%) at week 12] were superior to those based on <1.7 log IU/mL HCV RNA [17/19 (89.4%) at week 4, 27/38 (71.0%) at week 8, and 27/43 (62.7%) at week 12]. The negative predictive values for SVR based on <1.7 log IU/mL HCV RNA by COBAS TaqMan HCV test [46/57 (80.7%) at week 4, 37/38 (97.3%) at week 8, and 32/33 (96.9%) at week 12] were superior to those based on undetectable HCV RNA [48/66 (72.7%) at week 4, 46/53 (86.7%) at week 8, and 41/43 (95.3%) at week 12]. The utilization of both undetectable RNA and <1.7 log IU/mL HCV RNA by COBAS TaqMan HCV test is useful and could predict SVR and non-SVR patients with greater accuracy.

Effects of variations of extracellular polymeric substances and soluble microbial products on activated sludge properties during anaerobic storage.

Anaerobic storage always takes place in wastewater treatment plants or sludge handling processes. Although extracellular polymeric substances (EPS) and soluble microbial products (SMP) are well known to be the two important chemical parameters to influence sludge properties, few studies have demonstrated the coexisting effects of the two biopolymers on sludge characteristics during anaerobic storage of activated sludge. The experiments, conducted with acetate as the substrate at 35 degrees C and 25 degrees C and with glucose at 35 degrees C, indicated that EPS content decreased significantly in the first two or three days and later slightly increased in all runs, while SMP concentration varied gradually from 66-105mg l(-1) to 330-420mg 1(-1), increasing almost linearly with anaerobic storage time. All the components in EPS decreased after anaerobic storage, especially protein, which was the dominant organic compound in both EPS and SMP with more than 60% of the total amounts of the two biopolymers. Protein and DNA in SMP increased to a large extent and mainly contributed to the three-to five- fold increase in the total amount of SMP, while carbohydrates in SMP remained at a relatively low level. From statistical analysis, some major correlations could be obtained: a weak correlation existed between SMP and EPS; SMP were much more significantly correlated than EPS with sludge properties (biomass, specific resistance to filtration, sludge volume index and relative hydrophobicity of sludge surface). The results indicated that SMP, not EPS, significantly influence the sludge dewatering and settling properties, being deteriorated by more SMP production from cell lysis during anaerobic storage.

Effects of nutrient addition on phenol biodegradation rate in biofilm reactors for hypersaline wastewater treatment.

Hypersaline wastewater, generated by many industrial activities, is difficult to treat through conventional biological processes. In this kind of hypersaline environment, complex nutrients are needed for the normal growth of many microorganisms. For this paper, the organisms were taken from a municipal wastewater treatment plant and acclimated to 15% salinity in a biofilm treatment process (Sequencing Batch Biofilm Reactor) during the treatment of phenol-containing synthetic wastewater. They are used to evaluate the effects of nutrient addition on the phenol biodegradation rate. Adding yeast extract, glucose, KCl and four mineral nutrients into the shaking flasks containing wastewater and cultivating organisms, revealed phosphate as the crucial nutrient stimulating phenol biodegradation at 15% salinity. The operation results of the sequencing batch biofilm reactor indicates that phosphate content increased up to five times the original level can increase the phenol removal rate by 150%. A 99% phenol removal efficiency could be achieved by shortening the reaction time in the biofilm reactor from 40 h to 16 h, compared with basic nutrients added. In this article we applied an applicable and effective shaking-culture method to determine nutrient requirements in biotreatment processes without stopping the running reactors.

Biodegradation of microcystis and microcystins by indigenous nanoflagellates on biofilm in a practical treatment facility.

The potential for degradation of Microcystis and microcystins by organisms in biofilm in a practical biological treatment facility combined with conventional treatment processes was examined. Viable cells of Microcystis viridis, which produced microcystin LR, RR and YR, were degraded in 6 to 10 days by the addition of biofilm collected in early summer, mid-summer and autumn. Monas sp. grew remarkably well compared with other organisms in the samples of all seasons. Therefore, we conclude that Monas sp. was indigenous animals in the biological treatment facility and was able to degrade Microcystis cells. Some previous studies reported the predation of Microcystis cells by Monas guttula in pure culture conditions (monoxenic culture). We clarified that Monas sp. can grow and prey on Microcystis cells even in mixed culture including many kinds of other organisms in biofilm in a practical treatment facility. Also, intracellular microcystin LR, RR and YR in Microcystis cells could be degraded simultaneously with high reduction of Microcystis cells by organisms in biofilm.

A novel control process of cyanobacterial bloom using cyanobacteriolytic bacteria immobilized in floating biodegradable plastic carriers.

A process using a floating carrier for immobilization of cyanobacteriolytic bacteria, B.cereus N-14, was proposed to realize an effective in situ control of natural floating cyanobacterial blooms. The critical concentrations of the cyanobacteriolytic substance and B.cereus N-14 cells required to exhibit cyanobacteriolytic activity were investigated. The results indicated the necessity of cell growth to produce sufficiently high amounts of the cyanobacteriolytic substance to exhibit its activity and also for conditions enabling good contact between high concentrations of the cyanobacteriolytic substance and cyanobacteria. Floating biodegradable plastics made of starch were applied as a carrier material to maintain close contact between the immobilized cyanobacteriolytic bacteria and floating cyanobacteria. The floating starch-carriers could eliminate 99% of floating cyanobacteria in 4 d. Since B.cereus N-14 could produce the cyanobacteriolytic substance under the presence of starch and some amino acids, the cyanobacteriolytic activity could be attributed to carbon source fed from starch carrier and amino acids eluted from lysed cyanobacteria. Therefore, the effect of using a floating starch-carrier was confirmed from both view points as a carrier for immobilization and a nutrient source to stimulate cyanobacteriolytic activity. The new concept to apply a floating carrier immobilizing useful microorganisms for intensive treatment of a nuisance floating target was demonstrated.

New in-vivo calibration phantoms and their performance.

New in-vivo calibration phantoms (anthropometric phantoms) were developed to meet the needs for Japanese standard phantoms. Two important characteristics of these phantoms were that (1) they were designed using Japanese body size survey data, and (2) they were designed so that they can be adapted to various positions or geometries. The performance of these phantoms was tested with respect to body size, activity distribution along the axis, and counting efficiency. The actual dimensions of the anthropometric phantoms were compared with the survey data. Most items (31 of 47) indicated good agreement between the actual values and the survey data for the adult anthropometric phantom. The activity distribution for the anthropometric phantoms was compared with that for block phantoms that simulate a uniform activity distribution. The anthropometric phantoms have some gaps in their joints. The measurement results, however, indicated that these gaps did not significantly affect the overall accuracy of the measurements. Differences in counting efficiency between the block phantoms and the anthropometric phantoms for the same age were no more than 6%.

Degradation of microcystin by biofilm in practical treatment facility.

Potential for degradation of microcystin by biofilm was examined by some batch experiments using biofilm scraped from practical biological treatment facility combined with conventional treatment processes. The viable cells of Microcystis viridis, which produced microcystin LR, RR and YR were degraded at 6 to 10 days by the addition of biofilm. Biofilm collected in summer season had especially higher potential for degradation of Microcystis with complete degradation at 6 days. In all seasons, Monas spp. grew remarkably, accompanied with the higher decrease of the viable cells of Microcystis and the micro-animals were considered as a main predator for Microcystis cells. Intracellular microcystin LR, RR, YR were degraded simultaneously with high reduction of Microcystis cells. Dissolved microcystin LR of 1,000 microg l(-1) was effectively degraded by indigenous aquatic bacteria on biofilm during 5 days, the degradability became higher with the increase in the concentration of microcystin LR. From the results of our research, it was clarified that the aggregated microorganisms consisting of biofilm had high potential for degradation of intracellular and dissolved microcystin.

Characterization of an algae-lytic substance secreted by Bacillus cereus, an indigenous bacterial isolate from Lake Kasumigaura.

In order to develop a practical biological strategy for water bloom-control in Lake Kasumigaura, indigenous algae-lytic bacteria targeting Microcystis spp. (the dominant cyanobacteria), was first carried out. Some basic lytic characteristics of the isolates concerning the biological and physico-chemical factors in actual water environments were then investigated. Fifteen algae-lytic bacteria were successfully isolated using the double layer method. The isolate showing the highest lytic activity was identified as Bacillus cereus based on the 16SrRNA sequence. Extracellular products of B. cereus were found to be responsible for algae-lytic activity. Algae-lytic assay tests using bacterial supernatants pre-treated under several conditions indicated that the majority of the effective algae-lytic substances were either hydrophilis or hydrophobics with a carbon number less than 18, and with molecular weight less than 2000 Da. The algae-lytic activity of the bacterial supernatant was found to be enhanced under alkaline conditions, but it was lost under acidic conditions. This pH specific characteristic is advantageous for application in water bloom environments where the pH is usually in the alkaline region.

Measurements of summer radon and its progeny concentrations along with environmental gamma dose rates in Taiwan.

The concentrations of 222Rn (radon) and its progeny with surrounding environmental gamma-dose rates were measured simultaneously inside and outside of buildings at 10 locations around Taipei and Hualien in Taiwan. For summer radon in Taiwan, indoor concentrations were estimated to be about 20 Bq m(-3) with about 90 nSv h- of environmental gamma, and outdoors, about 10 Bq m(-3) with about 70 nSv h(-1). The equilibrium factors were calculated to be 0.2-0.3 indoors and 0.3-0.4 outdoors. Indoor radon concentration had a weak positive correlation with gamma-dose rate. Since there is a possibility that high radon concentrations exist indoors during the cool season in Taiwan because of extremely low ventilation rates in the dwellings, a winter survey in January through February will be needed for future estimation of the annual effective dose.

Development status of Terumo implantable left ventricular assist system.

We have been developing an implantable left ventricular assist system (T-ILVAS) featuring a magnetically suspended centrifugal pump (MSCP) since 1995. In vitro and in vivo studies using a prototype MSCP composed of a polycarbonate housing and impeller (196 ml) have demonstrated long-term durability and excellent blood compatibility for up to 864 days, and excellent stability of the magnetic bearing of the MSCP. These preliminary results strongly suggested that the magnetic bearing of the MSCP is reliable and is a most feasible mechanism for a long-term circulatory assist device. We have recently devised a clinical version pump made of titanium (180 ml) with a new position sensor mechanism and a wearable controller with batteries. Cadaver fit study confirmed that the Type IV pump could be implanted in a small patient with a body surface area as small as 1.3. The in vitro performance tests of the Type IV pump demonstrated excellent hydrodynamic performances with an acceptable hemolysis rate. New position sensors for the titanium housing showed more uniform sensor outputs of a magnetic bearing than in the prototype polycarbonate pump. The Type IV pump then was evaluated in vivo in 6 sheep at the Oxford Heart Centre. Four sheep were electively sacrificed at 3 months and were allowed to survive for more than 6 months for long-term evaluation. In this particular series of experiments, no anticoagulant/antiplatelet regimen was utilized except for a bolus dose of heparin during surgery. There was a left ventricular mural thrombi around the inflow cannula in 1 sheep. Otherwise, there was no mechanical failure nor sign of thromboembolism throughout the study.

Use of artificial neural network in the prediction of algal blooms.

A model to quantify the interactions between abiotic factors and algal genera in Lake Kasumigaura, Japan was developed using artificial neural network technology. Results showed that the timing and magnitude of algal blooms of Microcystis, Phormidium and Synedra in Lake Kasumigaura could be successfully predicted. As for the newly occurring dominant Oscillatoria, results were not satisfactory. The evaluation of the importance of factors showed that Microcystis, Phormidium, Oscillatoria and Synedra were alkalophilic. The algal proliferation for Microcystis, Oscillatoria and Synedra decrease due to the increase in total nitrogen, while the growth of Phormidium is enhanced with more nitrogen. In addition, the algal density is affected by zooplankton grazing but with the exception of Phormidium due to it being poor food source. Algal responses to the orthogonal combinations of the external environmental factors, chemical oxygen demand, pH, total nitrogen and total phosphorus at three levels were modeled. Various combinations of environmental factors enhance the proliferation of some algae while other combinations inhibit bloom formation.

Seasonal variation in frequency of dividing cells of freshwater phytoplankters.

The frequency of dividing cells (FDC) of a natural phytoplankton community was studied seasonally in a freshwater pond to which water directly from Lake Kasumigaura was supplied. Phytoplankton samples were collected every 3 h at three depths (5, 30 and 75 cm) over 24 h periods in each of the four seasons and examined for FDC, cell size and abundance. Dominant phytoplankton species were Synedra ulna, Cyclotella kutzingiana, Chlamydomonas cingulata and Oscillatoria tenuis with Melosira italica, Cryptomonas ovata, and Synechococcus sp. being seasonally abundant. All species showed a similar FDC pattern related to the light-dark period that is a moderately phased division with the maximum FDC between 21:00-03:00.

Discovery of a non-peptide small molecule that selectively mimics the biological actions of calcitonin.

Calcitonin (CT), a 32-amino acid peptide hormone secreted mainly from the thyroid gland, plays an important role in maintaining bone homeostasis. To discover non-peptide small molecules with biological actions similar to those of CT, a cell-based screening of an in-house chemical library was performed and a pyridone derivative (SUN B8155) was identified. Like CT, it elevated cyclic AMP (cAMP) levels in T47D and UMR106-06 cells which endogenously express human and rat CT receptor, respectively. SUN B8155 also stimulated cAMP formation in cells expressing recombinant human CT receptor, but not in those expressing human parathyroid hormone/parathyroid hormone-related peptide receptor. Accumulation of cAMP in T47D cells was blocked by a selective antagonist of CT receptor, salmon CT(8-32), whereas SUN B8155 did not displace the specific binding of [(125)I]CT to the receptor. Our results suggested that the compound selectively interacts with the CT receptor by a mechanism similar to but probably different from that of CT itself. In rats, intraperitoneal administration of SUN B8155 significantly lowered serum calcium levels, like CT. Our results demonstrate, for the first time, that the biological activities of the newly identified small molecule can mimic that of CT, acting via the CT receptor.

N-methyl-D-aspartate receptors regulate a group of transiently expressed genes in the developing brain.

Mammalian brain development requires the transmission of electrical signals between neurons via the N-methyl-d-aspartate (NMDA) class of glutamate receptors. However, little is known about how NMDA receptors carry out this role. Here we report the first genes shown to be regulated by physiological levels of NMDA receptor function in developing neurons in vivo: NMDA receptor-regulated gene 1 (NARG1), NARG2, and NARG3. These genes share several striking regulatory features. All three are expressed at high levels in the neonatal brain in regions of neuronal proliferation and migration, are dramatically down-regulated during early postnatal development, and are down-regulated by NMDA receptor function. NARG2 and NARG3 appear to be novel, while NARG1 is the mammalian homologue of a yeast N-terminal acetyltransferase that regulates entry into the G(o) phase of the cell cycle. The results suggest that highly specific NMDA receptor-dependent regulation of gene expression plays an important role in the transition from proliferation of neuronal precursors to differentiation of neurons.

Long-term evaluation of extracorporeal shock-wave lithotripsy for cholesterol gallstones.

BACKGROUND: Extracorporeal shock-wave lithotripsy (ESWL) is a treatment that preserves the gallbladder. Problems after ESWL treatment include stone recurrence and the development of biliary symptoms. METHODS: Two hundred and sixty-two patients with cholesterol-type gallstones, the best indication for ESWL treatment, and 42 control patients with cholesterol-type gallstones who received no treatment entered this study. We evaluated the factors associated with recurrence of gallstones after stone clearance and the development of biliary symptoms after ESWL treatment. RESULTS: The 3-, 5- and 7-year cumulative probabilities of gallstone recurrence were 20.6, 27.1 and 33.1%, respectively, with the recurrence probability significantly lower in patients with good gallbladder contractility. In patients with recurrence, ursodeoxycholic acid (UDCA) treatment was effective. In 69 patients with residual gallstones, the 3-, 5- and 7-year cumulative risks of biliary symptoms were 17.3, 24.9 and 30.5%, respectively. With residual gallstones, the risk of biliary symptoms developing was significantly lower in patients with a < or = 3 mm fragment size at the end of ESWL treatment and in those treated consistently with UDCA for 6 months or more after treatment with ESWL. The risk of biliary symptoms was significantly lower in ESWL-treated patients with residual stones who had a < or = 3 mm fragment size after treatment compared to those of control patients. CONCLUSIONS: Ursodeoxycholic acid was effective in clearing stones in patients with gallstone recurrence. In patients with residual stones, the fragmentation of stones to < or = 3 mm and UDCA administration effectively reduced the risk of subsequent biliary symptoms.

Crystallization and preliminary X-ray analysis of chondroitin sulfate ABC lyases I and II from Proteus vulgaris.

Chondroitin sulfate ABC lyases (E.C. 4.2.2.4) are broad-specificity glycosaminoglycan-degrading enzymes. Their preferred substrates are chondroitin sulfate and dermatan sulfate, which are broken down to short oligosaccharides. Proteus vulgaris produces two such lyases, ABC lyase I and II, with molecular weights of 112-113 kDa. Diffraction-quality crystals of both enzymes have been obtained by the hanging-drop vapour-diffusion method. ABC lyase I crystallizes in space group P2(1)2(1)2(1), with unit-cell parameters a = 49.3, b = 95.1, c = 230.0 A, Z = 4, and diffracts to 1.9 A resolution. Crystals of ABC lyase II belong to space group P1, with unit-cell parameters a = 64.2, b = 64.3, c = 142.1 A, alpha = 95.7, beta = 98. 1, gamma = 95.5 degrees, Z = 2; diffraction extends to at least 2.1 A.