An experimental study on in vivo dynamic mechanical properties of the knees of cats in valgus loading

In these experiments an attempt was made to investigate in vivo dynamic mechanical properties of the knee in valgus loading. Fifteen adult cats were used. Sinusoidal bending oscillation of 0.3, 1.0, 1.6, 3.3, 5.0, 8.6 and 10.0 Hz was transmitted to the knee-leg complex under each pre-bending strain in the valgus direction, and the bending moment was measured. Dynamic mechanical properties of the knee-leg complex were given as a complex bending elastic coefficient. Bending elastic and bending damping coefficient of the knee could be calculated by the least square method. As a first approximation, the knee was represented as a non-linear Kelvin model. Both bending elastic and bending damping coefficient of the knee increased as pre-bending moment was increased, and decreased as amplitude of bending oscillation was increased. Mechanical loss tangent of the knee-leg complex decreased within the range of pre-bending moment larger than 1.0 X 10(6) dyn cm as bending moment was increased, and decreased as amplitude of bending oscillation was increased.     

Mutagenicity of 1,2-dibromo-3-chloropropane (DBCP) in the mouse spot test

The spot test with 1,2-dibromo-3-chloropropane (DBCP) was carried out using male PW and female C57BL/6 mice. DBCP induced recessive colour spots in offspring with a significantly high frequency of 2.9%, showing that this chemical is mutagenic for somatic cells of mice in vivo.     

Juxtacardiac pressure in closed-chest dogs

We studied pressure (Ppc)-volume (Vpc) relationships of the pericardial sac by inserting air into it at constant end-diastolic heart volume in six dogs. The lungs were inflated by positive alveolar pressure while pleural pressure was monitored using the esophageal balloon technique. Ppc-Vpc relationships were measured at transpulmonary pressures (PL) of 30, 10, and 5 cmH2O in each of three states: closed chest, open chest with lung separation, and open chest with the pericardium dissected free of its mediastinal attachment. Ppc in the closed-chest condition was more positive than Ppc in the open chest with lung separation, with increase of Vpc and PL, which suggests that the lungs compress the pericardium. Ppc in the open-chest condition with lung separation was also more positive than Ppc in the pericardium after it was dissected free, which suggests that mediastinal attachment compresses the pericardium. It is suggested that lungs in the closed-chest condition as well as mediastinal attachment reduce the heart expansion by a similar magnitude.     

Imaging and uptake mechanism of 131I-meta-iodobenzylguanidine in medullary thyroid carcinoma

131I-meta-iodobenzylguanidine (131I-MIBG) was also taken up by medullary thyroid carcinoma (MTC) as well as by pheochromocytoma in two patients with Sipple's syndrome. However, the mechanism of 131I-MIBG uptake by MTC has not been clarified yet. We measured tissue catecholamine levels in three MTC, since MTC can produce several active substances. Catecholamines were detected in various amounts in all MTC, but not in normal thyroid tissues. These findings suggest that MTC can produce catecholamines and therefore, 131I-MIBG is taken up and stored in catecholamine vesicles of MTC, like pheochromocytoma and neuroblastoma. We conclude that 131I-MIBG may be applied not only to diagnosis but also for the treatment of patients with MTC.     

Relationship between potency of blocking type thyrotropin-binding inhibitor immunoglobulin in three women with primary myxedema and thyroid function of their neonates

Three neonates born to three mothers with primary myxedema who have thyrotropin-binding inhibitor immunoglobulin (TBII) were continually examined after birth. One neonate showed a high TSH level in mass-screening for congenital hypothyroidism and developed transient hypothyroidism. Her TBII disappeared at 114 days of age, and she remained euthyroid after discontinuation of thyroxin replacement at 146 days of age. The other two neonates were euthyroid, though they had positive TBII. In three mothers, the doses of IgGs that inhibited 125I-TSH binding to the level of 50% were compared. The potency of IgG from the mother whose neonate developed hypothyroidism was stronger than that of IgG from the other two mothers. And the elevation of cAMP induced by bovine TSH in suspension culture with porcine thyroid follicles was significantly reduced in the presence of IgG from the three mothers when compared with normal IgG. The thyroid-stimulation blocking activity was more potent in the mother whose neonate developed hypothyroidism than in the other two mothers. This study suggests that the thyroid function of neonates born to primary myxedema with blocking type TBII is influenced by the potency of TSH-binding inhibitor and thyroid-stimulation blocking activity of the mother.     

Identification of the plasminogen-binding site of human alpha 2-plasmin inhibitor

A plasminogen-binding site of human alpha 2-plasmin inhibitor was studied. The chromatogram of digest from the amidinated alpha 2-plasmin inhibitor (67K-daltons, plasminogen-binding form) with trypsin was almost identical with that obtained from the 65K-daltons derivative (non-plasminogen-binding form) treated with the same procedure, except for the three tryptic peptides. One of the three peptides, the deamidinated peptide T-11, was found to have a strong ability to inhibit the interaction of alpha 2-plasmin inhibitor with human plasmin. Moreover, the dissociation constant Kd for interaction between the peptide T-11 and plasmin was estimated to be 5.5 microM, indicating that Kd is about 10-fold lower than that of epsilon-aminocaproic acid. The sequence of the peptide T-11 was determined by the Edman method as follows: NH2-G-D-K-L-F-G-P-D-L-K-L-V-P-P-M-E-E-D-Y-P-Q-F-G-S-P-K-COOH. alpha 2-Plasmin inhibitor and its 65K-daltons derivative were found to have the same NH2-terminal sequence of Asn(Asp)-Gln-Glu-Gln-. These results indicated that the plasminogen-binding site(s) of alpha 2-plasmin inhibitor could be located in the COOH-terminal region of its molecule and that some of epsilon-NH2-groups in the deamidinated peptide T-11 may be involved in the lysine-binding site(s) of plasmin(ogen).     

Structural studies on N-acetylmannosaminuronic acid-containing and glucuronic acid-containing teichuronic acids in the cell wall of Bacillus megaterium AHU 1375

Structural studies were carried out on two kinds of teichuronic acid-glycopeptide complexes (designated as TU-GP-I and TU-GP-II) isolated from lysozyme digest of N-acetylated cell walls of Bacillus megaterium AHU 1375 by ion-exchange chromatography and gel chromatography. TU-GP-I, accounting for about 25% of the cell walls, contained N-acetylmannosaminuronic acid, N-acetylglucosamine, glucose, galactose, glycerol, and phosphorus in an approximate molar ratio of 1:1:2:1:0.5:0.5, together with small amounts of glycopeptide components. TU-GP-II, accounting for about 9% of the cell walls, contained glucuronic acid, glucose, and fucose in a molar ratio of about 2:1.5:1, together with small amounts of glycopeptide components. The results of analyses involving Smith degradation, chromium oxidation, methylation, acetolysis, and H-NMR measurement led to the conclusion that the polysaccharide chain of TU-GP-I comprised repeating units,----6) Glc(alpha 1----3)-ManNAcUA(beta 1----4)[Gal(alpha 1----3)][Glc(beta 1----6)]GlcNAc(beta 1----. About half of the repeating units were substituted by glycerophosphoryl residues at C-6 of the beta-glucosyl residues linked to the N-acetylglucosamine residues. By means of a similar procedure, the polysaccharide chain of TU-GP-II was shown to comprise repeating units,----4)GlcUA(alpha 1----3)GlcUA(alpha 1----3)Glc(alpha 1----3)Fuc(alpha 1----, of which about half were substituted by alpha-glucosyl residues at C-3 of the 4-substituted glucuronosyl residues.     

Purification of scrapie agent from infected animal brains and raising of antibodies to the purified fraction

The fraction (P4) containing scrapie infectivity was obtained by treatment of scrapie-infected mouse brains with the detergent sarcosyl, differential centrifugation, and proteolytic enzyme digestion. Scrapie infectivity in the P4 fraction was purified 239-2,390 times with respect to protein. Similar fractions were also prepared from the brain of a sheep naturally infected with scrapie. Morphological observation of the P4 fractions revealed that the main components were unique rods of 3-5 X 60-200 nm, which resembled scrapie-associated fibrils (SAF) or prion rods. The P4 fractions formed three major broad bands of polypeptides with molecular weights (MWs) of about 24.5K, 21K, and 17K dalton (Kd) in sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and some low MW polypeptides were also present in the fraction. Rabbits immunized with this fraction prepared from mouse brains raised antibodies against the three major polypeptides.     

Design of a new automatic chromatography system for efficient enzyme purification equipped with a time-shared multiple activity analyzer

A new system equipped with a computer-controlled multiple activity analyzer has been developed for the efficient purification of multiple enzymes. The system consists of the following units: conventional enzyme fractionation system with a peristaltic pump, liquid chromatographic column, fraction collector, and uv monitor; computer-operated uv-vis spectrophotometer equipped with a thermo-regulated metal block and a flow-through type silica cuvette; personal computer; dot matrix printer; cooling facility; and automatic sampling-mixing system. The whole system is operated by a newly designed time-sharing computer program for periodic and repetitive sampling of the column eluants containing multiple kinds of enzymes and of designated assay mixtures for each enzyme and for measurement of the initial velocity of spectrophotometric signals. For example, a mixture of aspartase (EC 4.3.1.1) and malate dehydrogenase (EC 1.1.1.39) and also a mixture of these two enzymes and glutamate dehydrogenase (EC 1.4.1.3 or EC 1.4.1.4) were analyzed by the above system using gel permeation chromatography, and the two or three enzyme activities were repeatedly monitored within 4 min. Based on the above results further possibilities for the application of the system for a variety of purposes are discussed.     

Unusual metabolism of sulfur-containing amino acids in rats treated with DL-propargylglycine

S-(2-Hydroxy-2-carboxyethyl)homocysteine, S-(3-hydroxy-3-carboxy-n-propyl)-cysteine, N-acylated S-(beta-carboxyethyl)cysteine, and N-acylated S-(3-hydroxy-3-carboxy-n-propyl) cysteine were excreted in the urine after DL-propargylglycine treatment. Cystathionine was also accumulated in several tissues of DL-propargylglycine-treated rats. N-Monoacetylcystathione was found in the liver of rats and was also detected in the kidney and serum. Cystathionine gamma-lyase activity in liver decreased to about 4% of that of control rats 24 h after the DL-propargylglycine injection, and alanine aminotransferase activity decreased to about 35% of that of control rats. On the other hand, aspartate aminotransferase and cystathionine beta-synthese activity did not show significant changes from those of control rats. The ability of normal tissues to synthesize cystathionine utilizing cystathionine beta-synthase was 1.98 +/- 0.40 mumol/min/g in liver, 0.61 +/- 0.13 in kidney, and 0.18 +/- 0.015 in brain. The maximal contents of cystathionine in rat tissues and the administered amounts of DL-propargylglycine agreed well with the ability to synthesize cystathionine in each tissue.