Effect of the addition of rice straw on microbial community in a sewage sludge digester.

Rice straw was added to a sewage sludge digester and its effects on methane production, dewatering characteristics, and microbial communities in the digested sludge were examined by a continuous digestion experiment under mesophilic conditions (35 °C). Stable gas generation was monitored in all digestion experiments. Methane yield from raw sludge, chopped rice straw and softened rice straw were estimated to be 0.27, 0.18 and 0.26 NL/g total solids load, respectively. The capillary suction time of digested sludge was decreased by the addition of rice straw. Archaeal and bacterial communities in the sludge were elucidated by PCR-DGGE (polymerase chain reaction--denaturing gradient gel electrophoresis) targeting 16S rRNA genes. The Shannon index of DGGE profiles indicated that bacterial diversity increased with the addition of softened rice straw. DNA sequences of significant bands of the digested sludge were most closely related to Methanosaeta concilii (97.4% identity) and Methanoculleus bourgensis (100% identity). Meanwhile, those in the co-digested sludge with rice straw were most closely related to Methanosarcina barkeri (98.4% identity) and Methanoculleus bourgensis (99.3% identity). Although both Methanosaeta spp. and Methanosarcina spp. metabolize acetate to methane, Methanosarcina spp. have a competitive advantage at acetate concentrations of >70 mg/L. Results suggested that the quantity of acetate produced during rice straw degradation may change the archaeal community.

Insulin resistance is a risk factor for the progression of chronic kidney disease.

OBJECTIVE: Insulin resistance may contribute to the pathogenesis of hypertension and progressive chronic kidney disease (CKD), however, few clinical studies have explored the role of insulin resistance in predicting the deterioration of renal function in CKD patients. MATERIALS AND METHODS: Enrolled in the study were non-diabetic hypertensive patients with CKD Stage 3. Insulin resistance was assessed by a homeostasis model assessment of insulin resistance (HOMA-R) measured at the entry to the study. Patients were followed for 3 years and comparisons of renal and metabolic parameters were made in conjunction with HOMA-R between entry and the end of the study period. The insulin-resistant (IR) group was defined as patients with HOMA-R 2.0 and more, and the insulin-sensitive (IS) group as those with HOMA-R < 2.0. RESULTS: Blood pressure in both groups was equally controlled below 130/80 mmHg throughout the observation period. The degree of insulin resistance HOMA-R and immunoreactive insulin (IRI) remained unchanged in the IS group, however, both were ameliorated in the IR group (HOMA-R, from 3.4 +/- 1.5 - 3.0 +/- 1.1, p = 0.022 and IRI, from 14.4 +/- 6.1 microU/ml - 12.6 +/- 6.8 microU/ml, p = 0.012). Creatinine clearance (CCr) and estimated glomerular filtration rate (e-GFR) decreased and serum creatinine (Cr) concentration increased in all patients. The decline in CCr calculated as the slope of the reciprocal of serum Cr concentration (1/Cr) was greater in the IR group (0.007 +/- 0.004 (1/Cr/dl/mg/month) than in the IS group (0.003 +/- 0.002 (1/Cr/dl/mg/month), p < 0.001). Linear regression analysis showed that the slope of 1/Cr was negatively correlated with HOMA-R, IRI, BMI, respectively. Furthermore, stepwise regression analysis showed that the independent variables to explain the decline in renal function were HOMA-R and IRI. CONCLUSION: Insulin resistance is a significant risk factor for the deterioration of renal function in hypertensive non-diabetic patients with CKD.

T-type calcium channel blockade as a therapeutic strategy against renal injury in rats with subtotal nephrectomy.

T-type calcium channel blockers have been previously shown to protect glomeruli from hypertension by regulating renal arteriolar tone. To examine whether blockade of these channels has a role in protection against tubulointerstitial damage, we used a stereo-selective T-type calcium channel blocker R(-)-efonidipine and studied its effect on the progression of this type of renal injury in spontaneously hypertensive rats that had undergone subtotal nephrectomy. Treatment with racemic efonidipine for 7 weeks significantly reduced systolic blood pressure and proteinuria. The R(-)-enantiomer, however, had no effect on blood pressure but significantly reduced proteinuria compared to vehicle-treated rats. Both agents blunted the increase in tubulointerstitial fibrosis, renal expression of alpha-smooth muscle actin and vimentin along with transforming growth factor-beta (TGF-beta)-induced renal Rho-kinase activity seen in the control group. Subtotal nephrectomy enhanced renal T-type calcium channel alpha1G subunit expression mimicked in angiotensin II-stimulated mesangial cells or TGF-beta-stimulated proximal tubular cells. Our study shows that T-type calcium channel blockade has renal protective actions that depend not only on hemodynamic effects but also pertain to Rho-kinase activity, tubulointerstitial fibrosis, and epithelial-mesenchymal transitions.

Transcriptional analysis of two Rhodobacter capsulatus ferredoxins by translational fusion to Escherichia coli lacZ.

Plasmids which contained the translational fusion of Escherichia coli lacZ to Rhodobacter capsulatus ferredoxin genes, fdxN and fdxA, were constructed. Effects of growth conditions on the expression of each ferredoxin were analyzed by measuring the beta-galactosidase activity in R. capsulatus which harbored a corresponding plasmid. Transcription of fdxN::lacZ, the ferredoxin I fusion gene, was regulated at least 100-fold by either NH4+ or O2 but not by illumination, confirming that fdxN belongs to the nif-gene family. Transcription of fdxA::lacZ, the ferredoxin II fusion gene, however, was constant under all the conditions surveyed, suggesting that the protein has some constitutive function(s).

Biomimetic configurational arrays of hydroxyapatite nanocrystals on bio-organics.

Biomimetic configurational arrays of hydroxyapatite (HAp) nanocrystals on several bio-organics, collagen (Col), chondroitin sulfate (ChS), and their mixture, were comparatively studied. The nanocomposites of HAp/Col, HAp/ChS, and HAp/Col/ChS were synthesized through a precipitation method with calcium hydroxide suspension and phosphoric acid solution containing Col, ChS, and their mixture, respectively. The (0 0 2) diffraction of the synthesized HAp crystals on a Col fiber showed an around 60 arching angle, while that on a ChS fiber showed just around 10 degrees. The same configurational arrays of HAp crystals could also be obtained on the mixture of Col and ChS fibers. The different electron diffraction patterns of the HAp crystals on the Col and ChS fibers were explained by the different macromolecular configurations of the Col and ChS fibrils which compose their fibers. The results may be applicable to develop a bone substitute which mimics the peculiar configurational arrays of HAp crystals found in bone and their detailed organic composition.

Two distinct ferredoxins from Rhodobacter capsulatus: complete amino acid sequences and molecular evolution.

Two distinct ferredoxins were purified from Rhodobacter capsulatus SB1003. Their complete amino acid sequences were determined by a combination of protease digestion, BrCN cleavage and Edman degradation. Ferredoxins I and II were composed of 64 and 111 amino acids, respectively, with molecular weights of 6,728 and 12,549 excluding iron and sulfur atoms. Both contained two Cys clusters in their amino acid sequences. The first cluster of ferredoxin I and the second cluster of ferredoxin II had a sequence, CxxCxxCxxxCP, in common with the ferredoxins found in Clostridia. The second cluster of ferredoxin I had a sequence, CxxCxxxxxxxxCxxxCM, with extra amino acids between the second and third Cys, which has been reported for other photosynthetic bacterial ferredoxins and putative ferredoxins (nif-gene products) from nitrogen-fixing bacteria, and with a unique occurrence of Met. The first cluster of ferredoxin II had a CxxCxxxxCxxxCP sequence, with two additional amino acids between the second and third Cys, a characteristics feature of Azotobacter-[3Fe-4S] [4Fe-4S]-ferredoxin. Ferredoxin II was also similar to Azotobacter-type ferredoxins with an extended carboxyl (C-) terminal sequence compared to the common Clostridium-type. The evolutionary relationship of the two together with a putative one recently found to be encoded in nifENXQ region in this bacterium [Moreno-Vivian et al. (1989) J. Bacteriol. 171, 2591-2598] is discussed.

[Aeromonas hydrophila septicemia with necrotizing myofascitis in a patient with hypoplastic leukemia].

A 54-year-old male was admitted to Kawasaki Medical School Hospital with the complaint of fever. His diagnosis of hypoplastic leukemia had been made one year ago. After the admission, cecal mass with pain and high fever were noted. Four days later, he suddenly lost consciousness and died. Aeromonas hydrophila was isolated from blood cultures and also from the myofascitis specimen. Autopsy specimen of the iliopsoas muscle showed necrotizing myofascitis. The specimen obtained from the cecum showed submucosal hemorrhage with edema and these findings were compatible to ischemic colitis. This pathogen is widely distributed in nature, especially in water fields. Therefore, it would be advised to consider the Aeromonas hydrophila as one of the pathological organisms pathognomonic for the septicemia, when one may see febrile and gastrointestinal symptoms in a patient with hematological malignancies.

Examination of gas desorption by B4C resin for use in neutron scattering experiment.

Gas desorption rates for several types of B(4)C resins were investigated using a throughput method. The investigation was particularly focused on determining the out gas composition, effects of dry air, grain size (density) effects on the gas desorption rates. It is found that water is the main component of out gas and that dry air can effectively reduce gas desorption.

Crystal growth of carbonate apatite using a CaCO3 flux.

Single crystals of carbonate apatite were grown using a CaCO3 flux under an Ar gas pressure of 55 MPa. The crystals obtained were observed by scanning electron microscopy, optical microscopy and X-ray diffraction. Electron probe microanalyses and thermal analyses were performed. CO3 ions in planar triangle form replaced both OH sites and PO4 tetrahedral sites in the apatite structure: in particular, the OH sites were perfectly substituted by CO3 ions using this method.

Preparation and mechanical properties of calcium phosphate/copoly-L-lactide composites.

New artificial bone materials were prepared using calcium phosphates, hydroxyapatite and beta-tricalicum phosphate, and copoly-L-lactide, CPLA. Calcium phosphate powder and CPLA were mixed at 453 K for 10 min with various mixing ratios. Scanning electron microscope observations indicated that the composites of beta-tricalicum phosphate and CPLA were homogeneously dispersed and highly adhesive. Young's modulus of the composites was the same as bone, and bending strength was over half that of bone. The improvement of Young's modulus compared to the original two materials was due to a composite effect. The composites are expected to be usable as artificial bone materials.

The Surface Structure of Hydroxyapatite Single Crystal and the Accumulation of Arachidic Acid.

Single crystals of hydroxyapatite (HAp) were grown by a flux method using beta-tricalcium phosphate and Ca(OH)(2) under hot isostatic pressure. After chemical etching by 0.05 N HCl aqueous solution, the HAp crystal surfaces were observed by an atomic force microscope (AFM) and shown dominantly to consist of steps with heights corresponding to a lattice distance d(100) or its 2/3. Arachidic acid films were accumulated on the etched HAp crystal by a Langmuir-Blodgett method. From section analyses by AFM, a distance between the carboxyl groups of arachidic acid and the HAp surface was estimated to be approximately 0.04 nm, sufficiently adjacent for a chemical interaction to take place. Copyright 2000 Academic Press.

Composition and structure of apatite formed on organic polymer in simulated body fluid with a high content of carbonate ion.

Apatite layer was formed on polyethyleneterephthalate (PET) substrate by the following biomimetic process. The PET substrate was placed on granular particles of a CaO, SiO2-based glass in simulated body fluid (SBF) with ion concentrations nearly equal to those of human blood plasma to form apatite nuclei on their surfaces. The apatite nuclei was then grown into a continuous layer by subsequently soaking the substrate in SBF under air or CO2 atmosphere in which CO2 partial pressure in the ambient was adjusted to 14.8 kPa to increase the content of carbonate ion to a level nearly equal to that of blood plasma. The increase in the content of carbonate ions in SBF changed the Ca/P atomic ratio of the apatite from 1.51 to 1.63, content of CO(3)2- ions from 2.64 to 4.56 wt %, and lattice constants a from 94.32 to 94.23 nm and c from 68.70 to 68.83 nm, respectively. The Ca/P ratio and lattice constants of the apatite formed in SBF under CO2 atmosphere were approximately identical to those of bone apatite, i.e. Ca/P atomic ratio 1.65, content of CO(3)2- ion 5.80 wt % and lattice constants a 94.20 and c 68.80 nm. This indicates that an apatite with composition and structure nearly identical to those of bone apatite can be produced in SBF by adjusting its ion concentrations including the content of carbonate ions to be equal to those of blood plasma.

Genetic analysis of functional differences among distinct ferredoxins in Rhodobacter capsulatus.

Rhodobacter capsulatus has been known to possess two ferredoxins (I and II) with distinct physicochemical and structural properties: ferredoxin I is a 2[4Fe-4S] type and the other is a [3Fe-4S] [4Fe-4S] type. To analyze their possible functional differences, their genes (fdxN and fdxA) were cloned, sequenced, and subjected to interposon mutagenesis experiments. The former gene was adjacent to a gene encoding a chloroplast-type [2Fe-2S] ferredoxin (fdxC). Mutants with inactivated fdxN and/or fdxC were obtained, and they showed virtually no growth under nitrogen-fixing conditions. Complementation experiments confirmed that both fdxN and fdxC were required for nitrogen fixation. On the other hand, we have not been able to disrupt fdxA under the screening conditions surveyed, including conditions that do not require nitrogenase activity for growth, suggesting that ferredoxin II could have an unknown essential role(s). These indicate functional differences among multiple ferredoxins in one bacterium other than in cyanobacterial heterocysts and indispensability of certain ferredoxins in nitrogen fixation other than Rhizobium meliloti FdxN.

Composition and structure of the apatite formed on PET substrates in SBF modified with various ionic activity products.

An apatite layer was formed on polyethyleneterephthalate (PET) substrates by the following biomimetic process. PET substrates were placed on granular particles of a CaO-SiO2-based glass in simulated body fluid (SBF) with ion concentrations nearly equal to those of human blood plasma to form apatite nuclei on their surfaces (first treatment). They then were soaked in modified SBFs, the ion concentrations of which were changed to give a variation in ionic activity product of apatite (IP), in order to make the apatite nuclei grow (second treatment). The Ca/P atomic ratio and the lattice constant c of the formed apatite decreased from 1.54 to 1.40 and from 6.880 to 6.838 A, respectively, with increasing ion concentrations from 0.75 to 2.00 times those of SBF, that is, with increasing IP from 10(-96.6) to 10(-91.9). This was attributed to an increase in the concentration of HPO4(2-) ion substituting for the PO4(3-) ion sites, which gave an increase in the Ca2+ in the apatite. Even the apatite formed in 1.00 SBF showed a Ca/P ratio of 1.51 and lattice constants a of 9.432 A and c of 6.870 A. The Ca/P ratio and lattice constant c were smaller and the lattice constant a was larger than those of the bone apatite; its Ca/P ratio and its lattice constants a and c, were 1.65, 9.419 A, and 6.88 A, respectively. This was attributed to the lower content (2.64 wt%) of the CO3(2-) ion substituting for the PO4(3-) ion sites of the apatite compared to that of the bone apatite (5.80 wt%). The lower content of the CO3(2-) ion in the apatite might be caused by the lower concentration of HCO3- ion in 1.00 SBF compared to that in human blood plasma.

Heterogeneous expression of Toll-like receptor 4 and downregulation of Toll-like receptor 4 expression on human gingival fibroblasts by Porphyromonas gingivalis lipopolysaccharide.

Porphyromonas gingivalis (P. gingivalis) is implicated in the initiation and progression of periodontitis. Human gingival fibroblasts (HGFs) are the major constituent of gingival connective tissue. P. gingivalis or its components such as lipopolysaccharide (LPS) upregulate the production of various inflammatory cytokines including interleukin (IL)-1 and IL-6 in HGFs. Recently, we demonstrated that the binding of P. gingivalis LPS to Toll-like receptor 4 (TLR4) on HGFs activates various second messenger systems (Biochem. Biophys. Res. Commun. 273, 1161-1167, 2000). In the present study, we examined the level of TLR4 expression on HGFs by flow cytometric analysis (FACS), and studied the levels of IL-1 and IL-6 in the culture medium upon LPS stimulation of HGFs by enzyme-linked immunosorbent assay (ELISA). Upon stimulation by P. gingivalis LPS for 24 h, HGFs that expressed a high level of TLR4 secreted significantly higher levels of IL-1 and IL-6 than HGFs that expressed a low level of TLR4. On the other hand, after stimulation with P. gingivalis LPS for 24 h, the level of TLR4 on the surface of HGFs decreased. These results suggest that the level of TLR4 expression on HGFs reflects the extent of inflammation in the gingival tissue, and that P. gingivalis LPS downregulates TLR4 expression on HGFs. These findings may be used to control inflammatory and immune responses in periodontal disease.