Biomedical subjects
Publications and source records attributed to T Kokubo.
A 68-year-old woman who had a history of Recklinghausen neurofibromatosis from 1964 showed nephrotic syndrome in 1989. Renal biopsy revealed membranous nephropathy. Various suspected causes of secondary membranous nephropathy were not found. Coexistence of Recklinghausen neurofibromatosis and membranous nephropathy has rarely been reported.
The most reliable factors predicting a long-term prognosis in individual progressive IgA nephropathy patients were evaluated. Ninety-eight cases who showed moderate to severe histological alterations (total score of 7 or more) and were continuously followed up for 10 years or more from the first biopsy were the subjects of this study. During the follow-up period of 10 years, 52 of the 98 cases went into hemodialysis treatment (HD). All 24 cases who had a higher total score of 17 or more, or severe renal dysfunction of less than 60 ml/min in their initial Ccr values went into HD. The remaining 74 cases who showed both a total score below 16 and initial Ccr values of 60 ml/min or more were divided into two groups in their clinical courses: 28 (group I) of the 74 cases went into HD and the other 46 (group II) did not. The degree of initial proteinuria was significantly different between groups I and II, but the degree of individual proteinuria was in a similar range from 0.5 to 2.5 g/day in most cases of both groups. Therefore, in order to clarify a more precise factor associated with the individual prognosis, the % duration of massive proteinuria (% DP) was calculated in individual cases. The % DP was more significantly different between groups I and II (82.6 +/- 26.1 vs 19.6 +/- 27.3%, p < 0.01). Moreover, all cases but one in group I showed 30% or more in % DP and 35 of 46 cases in group II showed 30% or less.(ABSTRACT TRUNCATED AT 250 WORDS)
A TATA box-binding initiation factor, TFIID, plays a central role in the transcriptional regulation by activators. Using anti-TFIID tau (a TATA box-binding component of native TFIID) immunoaffinity chromatography, nine polypeptides (230, 110, 85, 62, 58, 42, 28, 22, and 21 kDa) were identified as native Drosophila TFIID components that are tightly associated with TFIID tau. To verify the functional activity of the purified TFIID complex, template DNA and other transcription factors were reconstituted with purified TFIID bound to the antibody-Sepharose matrix. Immobilized TFIID mediated not only basal transcription but transcriptional activation by upstream stimulatory factor (USF). On the other hand, recombinant TFIID tau immobilized on the same antibody-Sepharose matrix could not mediate activation by USF. These results suggest that one or more of these additional polypeptides are required as functional TFIID subunits for activator-dependent transcription in conjunction with TFIID tau. As further evidence of the relevance of the Drosophila TFIID components identified in this analysis, including the previously unrecognized p230 (Dynlacht, B. D., Hoey, T., and Tjian, R. (1991) Cell 66, 563-576), protein blot analysis showed that TFIID tau interacts specifically and exclusively with p230. This suggests that p230 is an integral subunit of TFIID and that it may play a major role in tethering other subunits to TFIID tau.
Transcription initiation factor TFIIB recruits RNA polymerase II to the promoter subsequent to interaction with a preformed TFIID-promoter complex. The domains of TFIIB required for binding to the TFIID-promoter complex and for transcription initiation have been determined. The carboxyl-terminal two-thirds of TFIIB, which contains two direct repeats and two basic residue repeats, is sufficient for interaction with the TFIID-promoter complex. An extra 84-residue amino-terminal region, with no obvious known structural motifs, is required for basal transcription activity. Basic residues within the second basic repeat of TFIIB are necessary for stable interaction with the TFIID-promoter complex, whereas the basic character of the first basic repeat is not. Functional roles of other potential structural motifs are discussed in light of the present study.
Transcription initiation factor TFIID is a multimeric protein complex that plays a central role in transcriptional regulation by facilitating promoter responses to various activators. cDNAs encoding the 110-kDa subunit of Drosophila TFIID (p110) were isolated with a degenerate oligodeoxynucleotide probe based on an amino acid sequence of the purified protein. The entire cDNA sequence contains an open reading frame encoding a 921-amino acid polypeptide with a calculated molecular mass of 99,337 Da. The recombinant protein expressed in Sf9 cells via a baculovirus vector interacts directly with the 230-kDa subunit of TFIID (p230). Together with the previous observation that the TATA box-binding subunit of TFIID (TFIID tau or TBP) interacts directly with only p230 among the TFIID subunits, this result suggests that p110 forms a complex with TFIID tau via p230. A binding study using various p230 mutants indicated that both p110 and TFIID tau interact with the N-terminal 352-amino acid portion of p230, suggesting a functional communication between p110 and TFIID tau via p230 interactions.
Magnetic resonance (MR) images and conventional radiographs were compared in 142 hips with avascular necrosis, and a staging system for the disease based on MR imaging was developed. MR images were classified into three patterns: class I: a band of low signal intensity, class II: an area of low signal intensity with internal spot (s) of high signal, and class III: an area of low signal intensity without internal spots of high signal. Most MR class I lesions were in radiographic stage I (normal) or II (sclerotic or cystic changes without collapse). Most MR class II lesions were in radiographic stage III (segmental collapse), and most MR class III lesions were in stage III or IV (secondary degenerative changes). The MR image classification was closely correlated with radiographic staging (p < 0.01, using chi square test). We considered that this classification closely reflected the different stages of the disease according to the histopathology of the bone marrow.
The interface between bone and a bioactive glass cement--a mixture of bioactive glass powder and ammonium phosphate solution, previously reported on by the authors--was evaluated quantitatively and histologically. The materials tested were (1) the original bioactive glass cement (BCI cement); (2) an improved type of bioactive glass cement (BCII cement); (3) polymethylmethacrylate (PMMA) bone cement; and (4) a bioactive, apatite-wollastonite-containing, glass ceramic (A-WGC). Hardened cylindrical specimens of each cement were inserted loosely into canine femora and the interfacial shear strengths were measured using a push-out test. The interfacial strength values of the bioactive glass cements increased with prolonged implantation time. At each postimplantation time studied (8, 12, and 24 weeks), the interfacial strength value of BCI cement did not differ significantly from that of A-WGC. BCII cement interfacial strength was greater than that of BCI cement, whereas the interfacial strength of PMMA bone cement remained at a very low level throughout the study. Histological examinations revealed that direct bonding of both bioactive glass cements to bone had occurred without pathologic degradation. After 24 weeks, the defects between the bone and the bioactive glass cements had been filled with mature lamellar bone. Because the bioactive glass cement system developed by the authors, especially BCII cement, shows excellent osteoconductivity and bonds to bone tightly, we consider it to be a promising material for fixing prostheses into bone.
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We have developed a bioactive bone cement consisting of silane-treated CaO-SiO 2-P 2O 5-CaF 2 glass powder as the filling particles and bisphenol-a-glycidyl methacrylate (BIS-GMA) diluted with triethylene-glycol dimethacrylate (TEGDMA) as the organic matrix. Histological examination demonstrated direct bonding between the cement and bone along the circumference of the cement at 4 weeks after implantation in rat tibia. The compressive strength and toughness of the cement were two and four times greater than those of polymethylmethacrylate (PMMA) cement, respectively. The inflammatory reaction of the skin caused by the new cement was not as intense as that for PMMA 3 days after subcutaneous implantation. This new cement may be applicable as a bioactive bone cement with high mechanical strength.
Hydroxyapatite induction by a synthesized pure silica hydrogel was examined in various simulated body fluids (SBFs) having different magnesium, calcium, and phosphate ion concentrations as well as pH values. The silica hydrogel generated biologically active apatite on its surface by taking up calcium and phosphorous ionic groups from a surrounding SBF that was prepared to emulate the human plasma in inorganic composition. The induction period for apatite nucleation on the surface of the silica was largely decreased with the addition of a small amount of the calcium or phosphate ions to the SBF and with an increase in pH, but increased with the addition of magnesium ion. Bioactivity of bioactive materials like Bioglass and glass-ceramic A-W was well interpreted in terms of the rate of apatite formation reflected in these results. Moreover, the results provide the basic knowledge for designing new bioactive materials.
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Apatite formation on the surface of three kinds of bioactive material at an early stage after implantation in bone was studied using transmission electron microscopy (TEM). The materials were apatite- and wollastonite-containing glass-ceramic (A-W GC) as a surface-active glass-ceramic, dense sintered hydroxyapatite (HA) as a surface-active ceramic, and dense sintered beta-tricalcium phosphate (beta-TCP) as a resorbable ceramic. Particles of these materials, ranging from 100-300 microns in diameter, were implanted into rat tibiae, and specimens were prepared at 3, 7, 10, and 14 days after implantation. For A-W GC, dissolution of the glassy and probably wollastonite phase was observed in the surface region on and after the third day, and a collagen-free thin apatite layer on the surface of the material was evident on and after the seventh day. This apatite layer was observed before the mineralization of the surrounding bone matrix and was sometimes evident even where the material bordered on the bone marrow. On and after the tenth day, the surrounding bone matrix calcified and A-W GC-bone bonding through an apatite layer was completed. For HA, a mineralized collagen-free layer was observed on the surface of the ceramic on and after the tenth day. This layer was always present near calcifying bone and it was difficult to distinguish from immature bone. For beta-TCP, such a surface mineralized layer was rarely evident, even just before bone-ceramic contact, and finally the bone bonded to beta-TCP directly. Cell-mediated degradation of beta-TCP was frequently observed.(ABSTRACT TRUNCATED AT 250 WORDS)
Localized hyperthermic treatment was carried out with use of a metastatic bone tumor model in rabbits. The experimental bone tumor was created by transplantation of pieces of tumor line VX2 into the medullary canal of rabbit tibiae. Two weeks after the transplantation, a ferromagnetic ceramic pin was inserted in the medullary canal. Then, hyperthermia (HT) of the tumor was accomplished with use of an alternating magnetic field for 50 min. All the rabbits were killed 5 weeks after tumor transplantation, and the therapeutic effect was evaluated histologically and roentgenographically. Almost all the tumor cells within the bone marrow were killed by this procedure. The area of tumor necrosis in the HT group was significantly larger than in the control group. The pathological fracture rate and displacement rate were reduced significantly by this treatment (38.5 and 0%) compared with the controls (92.3 and 92.3%). Therefore, HT with the use of ferromagnetic ceramics was effective for local control of malignant bone tumors and seems to be a promising new method of treatment.
It has been reported that bioactive glass-ceramics containing crystalline oxy- and fluoroapatite [Ca10(PO4)6(O,F2) and wollastonite (CaSiO3), chemical composition: MgO 4.6, CaO 44.9, SiO2 34.2, P2O5 16.3, CaF2 0.5 in weight ratio] bond to bone tissue through the formation of an apatite (a calcium and phosphorus-rich layer) on the ceramic surface. In this study, the influence of disodium (1-hydroxythylidene) diphosphonate (DHTD) on the bonding between bone and glass-ceramics containing apatite and wollastonite was investigated. Rectangular ceramic plates (15 mm x 10 mm x 2 mm, abraded with #2000 alumina powder) were implanted into the tibial bone of mature male rabbits. DHTD was administered daily by subcutaneous injection to groups 1-5: group 1-4 at doses of 20, 5.0, 1.0, and 0.1 mg/kg body wt/day for 8 weeks; and group 5 at a dose of 5 mg/kg body wt/day for 4 weeks. Group 6 was given injections of saline as a control. At 8 weeks after implantation, the rabbits were killed. The tibiae containing the ceramics were dissected out and used for a detachment test. The failure load, when an implant became detached from the bone, or when the bone itself broke, was measured. The failure loads for groups 1-6 were 0 kg, 0 kg, 8.08 +/- 2.43 kg, 7.28 +/- 2.07 kg, 5.56 +/- 1.63 kg, and 6.38 +/- 1.30 kg, respectively. Ceramic bonding to bone tissue was inhibited by a higher dose of DHTD (groups 1 and 2).(ABSTRACT TRUNCATED AT 250 WORDS)
Calcium phosphate ceramics, beta-calcium pyrophosphate (Ca2P2O7), beta-tricalcium phosphate (Ca3(PO4)2), hydroxyapatite (Ca10(PO4)6(OH)2) and tetracalcium phosphate (Ca4(PO4)2O), were prepared. The calcium:phosphorus ratios and microporosities were 1 (31.6%), 1.5 (1.6%), 1.66 (1%) and 2 (34.6%) respectively. Samples (15 mm x 10 mm x 2 mm), abraded with No. 2000 alumina powder, were implanted into the tibial metaphysis of mature male rabbits. Failure load, when an implant detached from the bone or the bone itself broke, was measured. At 10 wk after implantation, the failure loads in beta-calcium pyrophosphate, beta-tricalcium phosphate, hydroxyapatite and tetracalcium phosphate were 31.65 +/- 9.90 N, 72.81 +/- 19.01 N, 49.49 +/- 17.25 N and 43.22 +/- 14.99 N respectively. At 25 wk after implantation, the values were 47.04 +/- 14.90 N, 71.34 +/- 19.50 N, 69.09 +/- 16.17 N and 62.03 +/- 18.62 N respectively. Histologically, bone bonding behaviour of calcium phosphate ceramics did not vary with the calcium:phosphorus ratio, as observed by contact microradiogram, Giemsa surface staining and scanning electron micrograph-electron probe micro analysis. There was no intervening soft tissue at the interface of bone and ceramics. Hydroxyapatite or tricalcium phosphate are used as bone substitutes. However, their mechanical strength is insufficient for weight-bearing and they are used as bone filler. This study showed that the apparent insignificance of strict calcium:phosphorus ratio with respect to the biological results greatly simplifies processing of calcium phosphate ceramics for clinical application. In clinical application, calcium phosphate ceramics with different Ca:P can be used as bone fillers for bone defects or bone cavities under non-weight-bearing conditions.
Hydroxyapatite crystallization is induced at 37 degrees C by sol-gel prepared silica from metastable calcium phosphate solutions. The morphology of the apatite forming on the silica surface depends on the nature of the solutions. For example, apatite grew in a flake-like form at pH 7.4. The morphology changed to plate-like when the pH was adjusted to pH 7.2. At this lower pH, the apatite plate even exhibited a hexagonal feature, reflecting the unique hexagonal structure of hydroxyapatite. An increase in either Mg or P ion concentration of the fluid can cause apatite to grow in a rod-like shape while addition of F ions to the fluid leads to a perfect needle pattern. The flake geometry of apatite was not altered by increasing Ca concentration from 2.5 to 3.8 mM in the solution. From this we conclude that sol-gel prepared silica is an efficient apatite inducer and the morphology of the hydroxyapatite deposit is determined by factors of the fluid such as pH, Ca/P molar ratio, Mg and F concentrations.
A strain of Aeromonas hydrophila was found to show prolyl endopeptidase activity. The enzyme gene was cloned and expressed in Escherichia coli JM83. A 12 kbp EcoRI fragment containing the enzyme gene was subcloned at the HincII site of pUC19 to construct plasmid pAPEP-3 with a 3.5 kbp insert. E. coli JM83 transformed with this plasmid showed about 100-fold higher activity than the parent Aeromonas. Analysis of the nucleotide sequence of the insert revealed that the mature enzyme-encoding sequence starts just after the ATG initiation codon of the open reading frame. The enzyme was a single polypeptide composed of 689 amino acid residues with a molecular weight of 76,383. It showed properties very similar to those of Flavobacterium prolyl endopeptidase, except that the isoelectric point was 5.5. The amino acid sequence was 56 and 41% homologous to those of Flavobacterium and porcine brain prolyl endopeptidases, respectively. From a survey of sequence homology with other members of the prolyl endopeptidase family, the amino acid residues involved in the catalytic triad were deduced to be Ser-537, His-656, and Asp-512 (or Asp-621).
A Drosophila cDNA encoding the largest TFIID subunit (p230) was isolated using a degenerate oligodeoxynucleotide probe based on an amino acid sequence of the purified protein. The entire cDNA sequence contains an open reading frame encoding a polypeptide of 2068 amino acids, corresponding to a calculated molecular mass of 232 kD. The deduced amino acid sequence showed a strong sequence similarity with the protein encoded by a human gene (CCG1) implicated in cell cycle progression through G1, suggesting that p230 may be a target for cell cycle regulatory factors. The recombinant protein expressed in Sf9 cells via a baculovirus vector interacts directly with the TATA box-binding subunit of TFIID (TFIID tau or TBP) from Drosophila, human, and yeast. Surprisingly, recombinant p230 inhibits the TATA box-binding activity and function of TFIID tau, suggesting that p230 interactions with TFIID tau, and possible modulations thereof by other factors may play an important role in TFIID function.