Characterization of the glycosphingolipids of pig cortical bone and cartilage.

Neutral glycosphingolipids and gangliosides were extracted from pig cortical bone and cartilage. To ensure the completeness of extraction, the cortical bone was demineralized and reextracted. Globotriaosylceramide and globoside were noted to be present at high content in the cortical bone. It contained glucosylceramide, lactosylceramide, globotriaosylceramide and globoside as neutral glycosphingolipids at a ratio of 1:0.7:3.1:2.7. In articular cartilage, the ratio was 1:0.7:0.4:0.8. GM3 and GD3 were the major gangliosides in both these tissues. GM3, GM1, GD3, GD1 and GT1 were present at ratios of 1:0.9:0.9:0.1:0.1 in the cortical bone and 1:0:1.2:0.06:0.02 in the cartilage. Neutral glycosphingolipids could be extracted from the cortical bone without the need for demineralization, while most of the gangliosides were extracted after this treatment, implying the occurrence of interactions between gangliosides and minerals in the bone.

Partial Amino Acid Sequences of Peptidyl-Prolyl Isomerases ofFusarium sporotrichioides.

Peptidyl-proprylyl cis-trans isomerase (PPIase) activity was observed from crude extract of Fusarium sporotrichioides. Proteins from this fungi were separated by two-dimensional polyacrylamide gel electrophoresis and more than one thousand protein spots were separated. Two cytosolic PPIases were found by the N-terminal sequencing from the two separated spots. The N-terminal 41 residues of the major protein spot showed high sequence identity (63.4%) with PPIase from Neurospora crassa. This protein was designated as PPIase a, having an apparent molecular mass of 20 kD and pI 7.0. The minor other protein spot, having a similar molecular mass but distinguishable pI 6.4, was also sequenced and the N-terminal twenty residues were almost identical to PPIase a and was designated as PPIase b. Copyright 1995 S. Karger AG, Basel

Resolution of Fusarium sporotrichioides Proteins by Two-Dimensional Polyacrylamide Gel Electrophoresis and Identification by Sequence Homology Comparison in Protein Data Base.

Proteins from Fusarium sporotrichioides M-1-1, a T2-toxin-producing strain, were separated by two-dimensional polyacrylamide gel electrophoresis. One thousand two hundred and forty-four protein spots were resolved and 103 protein spots were subjected to N-terminal sequencing. Fifty-eight protein spots were sequenced and 48 proteins were observed to have blocked N termini. Forty out of 58 sequenced proteins were identified by homology search against the PIR protein sequence data base and protein superfamily data base, while the residual 18 sequences were not identified. Twenty-seven of the N-terminal-blocked proteins were subjected to mild anhydrous hydrazine vapor deblocking. Twenty-four spots were not deblocked indicating the presence of acyl groups at the N termini, while 3 proteins were deblocked showing the blocked group to be pyrroglutamyl carboxylic acid residues. The results can provide a more global view of cellular genetic expression than any other technique. The created data may offer a unique opportunity to link information with DNA sequence data. Copyright 1995 S. Karger AG, Basel

Dysphasia accompanied by periodic lateralized epileptiform discharges.

We encountered a 67-year-old man who presented with repetitious dysphasia accompanied by periodic lateralized epileptiform discharges (PLEDs) on the electroencephalogram. A good correlation was established between the dysphasia and the PLEDs. A persistent partial seizure accompanied by PLEDs originating in the left hemisphere presented with dysphasia clinically. None of the previously reported patients with epileptic dysphasia had accompanying PLEDs. The administration of carbamazepine was successful to terminate the seizure, however valproate was not. This case indicates that PLEDs may produce significant, however reversible, functional damage, and may advance our understanding of the pathophysiology of epileptic dysphasia.

Germinoma arising in the basal ganglia in early stage: CT and MR findings.

We report a patient with a germinoma originating in the left basal ganglia. Unenhanced computed tomography (CT) revealed a relatively well-defined high-density lesion, which was slightly enhanced with contrast medium. No tumor stain was noted on angiography. The mass showed hypointensity on T1-weighted magnetic resonance imaging (MRI), and heterogeneous hyperintensity on T2-weighted imaging. The lesion was homogeneously enhanced after the administration of Gd-DTPA. To our knowledge, the MR findings of basal ganglia germinoma have been retrospectively reviewed in just four reported cases, only two of which were examined with Gd-DTPA enhancement. The two cases revealed relatively large masses with cysts and peritumoral edema, and were slightly enhanced after the intravenous injection of Gd-DTPA. Our case was thought to be in the early stage, and its CT and MR findings were slightly different from those of previously reported cases.

[The importance of dose intensity in preoperative chemotherapy for osteosarcoma--retrospective analysis of a short intensive chemotherapy regimen preoperatively using high-dose methotrexate and cisplatinum].

A retrospective investigation was performed on the significance of dose intensity in preoperative chemotherapy for osteosarcoma. In this paper, we studied 29 patients, who had classical stage II-B osteosarcoma and received preoperative treatment consisting of high dose methotrexate (HD-MTX) and/or cisplatinum (CDDP) between 1985 and 1991. Preoperative dose intensity was calculated in the following two ways. 1. Total dose of MTX and CDDP administered during initial five weeks: MTX (g)/10/m2 + CDDP(mg)/100/m2 2. Dose intensity of MTX and CDDP from initiation of treatment (I) to resumption of postoperative chemotherapy (R): MTX (g)/10/m2 + CDDP (mg)/100/m2 / days from (I) to (R)/ideal period of new regimen (63 days) Continuous disease free survival rate (CDFR) was significantly higher in the group receiving preoperative chemotherapy at higher dose intensity (> 3.0). Supplementary administration of adriamycin (ADR) to CDDP/MTX was not favorable in the preoperative stage, because it put off the following therapy and diminished the dose intensity. Since July 1989, we have tried a short intensive preoperative chemotherapy regimen: weekly two cycles of CDDP 100 mg/m2 intravenously, followed by weekly two cycles of HD-MTX 10g/m2. Surgery was scheduled six weeks after beginning treatment, and postoperative chemotherapy was started three weeks after surgery. Twenty patients have been treated by this regimen to date. All patients remain disease free, and six are CDF beyond 24 months.

Preservation of bone morphogenetic protein in heat-treated bone.

In operations of bone tumors, reimplantation of resected bone after boiling or autoclaving is a simple means of obtaining both tumor necrosis and skeletal reconstruction. However, such reimplants lose their osteogenesity. We investigated whether bone inductive ability could be maintained in heat-treated bone. Bone morphogenetic protein (BMP) extracted from rabbit bone after heating for various periods at different temperatures was implanted into the muscles of mice to evaluate osteogenetic activity. The maximum new bone formation was observed in specimens treated at 70 degrees C for 10 minutes, followed by those treated at 70 degrees C for 15 minutes. We then measured the temperature in the center of a cortical bone heated in 0.15 N NaCl solution at 50 degrees, 60 degrees, 70 degrees, 80 degrees, and 90 degrees C. Cortical bone center temperature reached that of the surrounding solution within 2.5 minutes. These results indicated that heating at 70 degrees for 10 to 15 minutes was suitable for heat treated-bone to maintain bone inductive ability.

Two-dimensional mapping by the high-performance liquid chromatography of oligosaccharides released from glycosphingolipids by endoglycoceramidase.

A two-dimensional sugar mapping method has been developed by which sensitive, reproducible, and simple analysis can be carried out on the structures and compositions of oligosaccharides released from glycosphingolipids by endoglycoceramidase. The oligosaccharides were labeled quantitatively with an ultraviolet-absorbing compound, p-aminobenzoic acid ethyl ester (ABEE). The ABEE-oligosaccharides were separated first on an amide-silica column and then on a C4-silica column by high-performance liquid chromatography. The acidic ABEE-oligosaccharides were eluted as a group at the start of the chromatography while the neutral ABEE-oligosaccharides were separated according to size and structure on an amide-silica column using an eluent without salt. The acidic oligosaccharides were separated according to size and structure when rechromatographed on the same column using an eluent containing KH2PO4. NeuAc-containing ABEE-oligosaccharides were extensively separated from the corresponding NeuGc derivatives. The ABEE-oligosaccharides separated on an amide-silica column were then chromatographed on a column of C4-silica on which lactotriose and neolacto-series oligosaccharides were clearly shown to be separated from the others. On the basis of the retention times of the individual ABEE-oligosaccharides on two separate columns, 9 neutral and 15 acidic oligosaccharides derived from glycosphingolipid standards were two-dimensionally mapped without overlapping. The gangliosides of a human chondrosarcoma tissue and glycosphingolipids of tumor tissue of FBJ virus-transformed murine osteosarcoma cells were analyzed by this method in conjunction with exoglycosidase treatment. At least 11 species of glycosphingolipids were identified in both cases.