The effect of calcium ion concentration on osteoblast viability, proliferation and differentiation in monolayer and 3D culture

Our research group aims to develop an osteochondral composite using type II collagen gel with hydroxyapatite (HAp) deposited on one side. Soaking gels in Ca2+ and phosphate solution is indispensable to HAp deposition, so relationships between cell behavior and Ca2+ concentration were examined in two- and three-dimensional cultures. The present results indicate that 2-4 mM Ca2+ is suitable for proliferation and survival of osteoblasts, whereas slightly higher concentrations (6-8 mM) favor osteoblast differentiation and matrix mineralization in both 2- and 3-dimensional cultures. Higher concentrations (>10 mM) are cytotoxic. Purely from the perspective of calcium deposition, higher concentrations lead to increased accumulation of Ca2+. Culturing cells in phosphate-containing gel in media with Ca2+ also leads to time-dependent formation of HAp in the gel. Considering the viability of embedded cells, culturing scaffolds in media with Ca2+ concentrations around 5mM is useful for both HAp deposition and osteoblast behavior.     

Cloning of a human immunoglobulin gene fragment containing both VH-D and D-JH rearrangements: Implication for VH-D as an intermediate to VH-D-JH formation

In an Epstein-Barr virus-transformed human B cell line we found an unusual immunoglobulin heavy chain gene rearrangement. Restriction mapping and sequencing analysis led us to conclude that V_H-D and D-J_H recombination took place in a single allele. Both V_H-D and D-J_H complexes still had their recombination signal sequences adjacent and the DNA sandwiched by these two complexes retained a germ line configuration, suggesting the potential for a secondary rearrangement resulting in a V_H-D(-D)-J_H formation. With this finding, we propose a novel pathway, in which the V_H-D complex is an intermediate in the formation of a functional V_H exon.     

NECROTIZING MYOPATHY AS A REMOTE EFFECT OF GASTRIC CANCER ACCOMPANIED WITH HASHIMOTO'S THYROIDITIS

An autopsy case of a 74-year-old male who had shown clinically hypothyroidism due to chronic atrophic thyroiditis (Hashimoto's thyroiditis), and pathologically necrotizing myopathy as a remote effect of gastric cancer was reported.
Morphological features of this necrotizing myopathy was those of carcinomatous myopathy rather than those of hypothyroid or diabetic myopathy.
As for the pathogenesis of the necrotizing myopathy (as a Group IV of polymyositis of Walton and Adams), the malignancy might have played an important role as a trigger of the secondary immunological abnormality upon a pre-exizting longstanding immune disorder of Hashimoto's thyroiditis. Pseudomembranous colitis, which was thought to be related to antibiotics (Lincomycin), was also briefly discussed.     

Vinyl polymerization. 294. Decomposition of tetramethyltetrazene in the presence of p-substituted benzyl chlorides

A study was made of the polymerization of acrylonitrile in dimethylformamide (DMF) initiated by the binary systems of tetramethyltetrazene (TMT) and p-substituted benzyl chlorides. The polymerization rate increased linearly with the σ-constants of substituents as electron-releasing groups were introduced to the phenyl ring of benzyl chloride. In order to elucidate the initiation mechanism of the polymerization, a kinetic investigation was also undertaken of the decomposition of TMT in the presence of p-substituted benzyl chlorides in DMF. The decomposition rate was first-order in TMT and first-order in p-substituents benzyl chloride. The decomposition rate also increased with the σ constants of substituents as electron-releasing groups were introduced. On the basis of the results, the initiation mechanism for the polymerization was discussed.     

HIGH-GRADE SURFACE OSTEOSARCOMA OF THE LEFT ILIUM A Case Report and Review of the Literature

A rare case of high-grade surface osteosarcoma of the left ilum is reported. Trephine biopsy performed on a 31-year-old woman suffering from a huge tumor of the left buttock revealed high-grade osteosarcoma, and hemipel-vectomy was undertaken. The surgical specimen showed that the tumor was present on the surface of the left ilium. Ten months after the operation, the patient died of brain metastasis. From these results, we considered this case to be one of high-grade surface osteosarcoma. ACTA PATHOL JPN 38: 235 -240, 1988.     

Synthesis and side-chain crystallization of comb-like polymers obtained from isopropenyl-1,3,5-triazines carrying two,three, and four long alkyl groups

Monomers containing several octadecyl groups, e.g., 2-isopropenyl-4,6-bis(octadecylamino)-1,3,5-triazine ( 2 ), 2-dioctadecylamino-4-isopropenyl-6-octadecylamino-1,3,5-triazine ( 3 ) and 2,4-bis(dioctadecylamino)-6-isopropenly-1,3,5-triazine ( 4 ) were prepared by the alkylation reaction of 2,4-diamino-6-isopropenyl-1,3,5-triazine ( 1 ) with 1-bromooctadecane in the presence of sodium hydride. In the free-radical homopolymerization of these monomers, the polymer yield of 3 was lower than that of 2 due to a decrease in the ceiling temperature, and the polymerization of 4 did not proceed. Copolymerizations of these monomers with styrene or methyl methacrylate were carried out and the monomer reactivity ratios (r₁ and r₂) were determined. The monomer reactivity decreased with increasing the number of octadecyl groups in the monomers. Crystallinity of the octadecyl side chains in the resulting comb-like polymers was evaluated by differential scanning calorimetry.     

Suppressive effect of leflunomide metabolite (A77 1726) on metalloproteinase production in IL-1beta stimulated rheumatoid synovial fibroblasts

Leflunomide, an isoxazol derivative structurally unrelated to other immunomodulatory drugs, has proven to be efficacious in the treatment of rheumatoid arthritis (RA). This study was conducted to elucidate the mechanism by which leflunomide mediated antirheumatic effects. We investigated the effects of A77 1726, leflunomide's active metabolite, on mitogen-activated protein kinase (MAPK) activation in IL-1beta-stimulated rheumatoid synovial fibroblasts. The effects of A77 1726 on the secretion of matrix metalloproteinases (MMPs) from rheumatoid synovial fibroblasts were also examined. A77 1726 partially suppressed IL-1beta-induced ERK1/2 and p38 kinase activation. In contrast, A77 1726 efficiently suppressed IL-1beta-stimulated JNK1/2 kinase activation. Although no suppressive effect was demonstrated on MMP-2, A77 1726 markedly inhibited MMP-1, 3, and 13 secretions from IL-1beta-stimulated rheumatoid synovial fibroblasts. Tissue inhibitor of metalloproteinases-1 (TIMP-1) was constitutively produced from rheumatoid synovial fibroblasts and the suppressive effects of A77 1726 on TIMP-1 production were minimal. Our results suggest that the suppression of the MAPK signalling pathway and MMP synthesis in rheumatoid synovial fibroblasts is a possible mechanism for the inhibitory activity of leflunomide against rheumatoid arthritis.     

A possible role of two hydrophobic amino acids in antigen recognition by synovial T cells in rheumatoid arthritis

Synovial T cells play a crucial role in the pathogenesis of rheumatoid arthritis (RA) synovitis. We have quantitatively analyzed the T cell receptor (TcR) variable (V) region gene repertoire of freshly isolated synovial fluid (SF) T cells, comparing it with that of peripheral blood (PB) T cells in RA. The TcR V gene repertoire of PB and SF T cells in RA and osteoarthritis was heterogeneous. In contrast, Vail in SF was expressed to a greater degree in three of five RA patients, and increased levels of Vp6, 1-3 were found in the SF of four of six RA, compared with paired PB. Of note, Vβ6, 1–3 was universally used in four RA patients with a disease duration of less than 10 years, irrespective of their HLA-DR types. This was in contrast to two other RA patients, suffering for more than 20 years, who showed different Vα and Vβ usages. β-chain sequence analysis in RA patients with a preference for Vβ6, 1–3 has shown that a few clones dominated in SF, whereas polyclonality was observed in PB. These findings suggest oligoclonal expansion of T cells in response to specific antigen(s) in the SF of these patients with RA of relatively short duration. Concomitant use of two hydrophobic amino acids, leucine and valine, in the Dβ region was noticeable among the predominant SF clones. These two amino acids might directly contact a peptide specific for the induction of synovitis in RA patients. TcR-directed therapy may, therefore, be useful for the treatment of early RA synovitis.     

Determination of the depth of 50% of maximum ionization, I50, for electron beams by the divided difference method

A new characterization of depth-ionization parameters for electron beams is empirically deduced from our data analysis based on the divided difference method (the DD method), which employs the numerical differential of an ionization curve. The important feature of the present method is that it does not necessarily require normalized percent depth-ionization (NPDI) data. The depth of 50% of maximum ionization, I50, which is an important parameter for electron beam dosimetry, can be deduced from the analysis of an unnormalized (or partial) depth-ionization (UDI) curve obtained over a short interval of depth. The values of I50 determined by the DD method are in agreement to within 0.1 mm for energies of 4, 6, and 9 MeV, compared with the ones determined by the TG-51 protocol method (or the conventional method), and the difference was 0.9 mm for 12 and 15 MeV. The dose at the reference depth, dref, calculated from I50 by the DD method, is found to be in agreement with TG-51 to within 0.1%. The field size dependence of the DD method using UDI data was studied for three field sizes: 6 x 6, 10 x 10, and 20 x 20 cm2. For all energies, the discrepancies of I50 as determined by both methods were 0.9 mm on average for the 6 x 6 cm2 fields and 0.6 mm for the other two field sizes. This dependence was remarkable for 6 x 6 cm2 fields for 12 and 15 MeV, and the discrepancies shown by the DD method were 1.2 mm for 12 MeV and 1.8 mm for 15 MeV, respectively. Since the reference field size in clinical dosimetry is usually 10 x 10 cm2, this dependence will not affect clinical dosimetry. The DD method could be an alternative option for checking beam quality in dose calibration.