Tojo S

Biosynthesis of Methylguanidine in Isolated Rat Hepatocytes and in vivo

To clarify the organ in which methylguanidine is synthesized, high doses of creatinine, which is known to stimulate the synthesis of methylguanidine, were administered to male Wistar rats intraperitoneally. Various tissues of the rats were frozen by a freeze clamp method before and 1, 2 and 3 h after injection, and methylguanidine was determined by high-pressure liquid chromatography using 9,10-phenanthrenequinone for fluorometric determination. We found evidence that the liver, kidney, lung, muscle, red blood cells and gut flora synthesize methylguanidine. In addition, we measured the synthesis of methylguanidine in isolated hepatocytes prepared from normal rats following the addition of creatinine, arginine and guanidinoacetic acid to the incubation medium. Synthesis of methylguanidine was observed only in those incubations which contained creatinine, and was dependent on the concentration of creatinine in the media and on the incubation period. Isolated rat hepatocytes also synthesized guanidine in the presence of guanidinoacetic acid. These results indicate that the liver is one of the organs which synthesize methylguanidine and also that creatinine is the precursor.

Active oxygen in methylguanidine synthesis.

Methylguanidine (MG), a toxin reported in uremia, is thought to be a product of creatinine oxidation. This study is designed to demonstrate the role of active oxygen in the oxidation of creatinine under conditions compatible with those found in uremia. MG synthesis is moderately stimulated by the superoxide radical derived from 3 mM hypoxanthine and 0.015 units/ml xanthine oxidase and inhibited by the addition of superoxide dismutase. This is increased markedly by the addition of 0.05% hydrogen peroxide and augmented to about 56,000 times the control rate in the presence of hydroxyl radicals derived from the reaction of 10 mM FeSO4 and 0.05% hydrogen peroxide. In addition, MG synthesis is inhibited by the addition of sorbitol, lactulose or ethanol, the scavengers of hydroxyl radicals. These results indicate that creatinine can be oxidized to MG by various species of active oxygen and that one of the mechanisms of MG synthesis is such oxidation. MG, therefore, may be a useful indicator of peroxidation in vivo.

Glomerular lesions associated with the Crow-Fukase syndrome

Three cases of the Crow-Fukase syndrome without radiographic changes of multiple myeloma are reported, with special reference to the glomerular changes seen. Proteinuria was detected in one case, although decreased renal function was observed in all (GFR: 41.0, 62.0, 74.1 ml/min respectively) at the time of renal biopsy. Glomerular changes were similar in all three cases. The main characteristic changes were mesangial proliferation and thickening of the glomerular capillary walls. Pictures by light microscopy were therefore similar to that of MPGN. On electron microscopy, the thickened capillary walls showed circumferential mesangial interposition and the subendothelial zone was electron-lucent and contained small dense granules or flocculent deposits. By immunofluorescent microscopy, no immunoglobulins, complement components or light chain were detected in the glomeruli except in one case.

Ultrastructural Study of Gaps of the Glomerular Basement Membrane in IgA Nephropathy

Renal biopsy specimens from 163 patients with IgA nephropathy were examined by electron microscopy to clarify the significance of gaps of the glomerular basement membrane (GBM) in IgA nephropathy. Gaps of the GBM were observed in 21 cases. In 1 case the capillary lumen was partially filled with fibrin-like materials, basement membrane-like materials, and epithelial cells, and in 6 cases wide gaps of the GBM were observed. In 10 cases the gaps of the GBM were covered by epithelial and/or endothelial cells, and in 4 cases spherical microparticles were seen at the gaps of the GBM. Microscopic hematuria of less than 5 red blood cells per high-power field of vision at the time of biopsy was less frequently observed in patients with gaps of the GBM than in patients without (p less than 0.05). Marked local thinning or splitting of the GBM and electron-dense deposits or spherical microparticles on peripheral capillary walls were more frequently observed in patients with gaps of the GBM than in those without (p less than 0.001, less than 0.005, less than 0.05, and less than 0.001, respectively).

Dipyridamole Therapy in the Nephrotic Syndrome

Dipyridamole was used in 30 cases of nephrotic syndrome, mostly of intractable type. The results indicate that the drug therapy proved to be effective in decreasing urinary protein and controlling nephrotic condition in 40% of the cases after an initial period of treatment. Long-term results of the drug on urinary protein and on nephrotic condition were rated as good in 36.7 and 53.3%, respectively, of the cases treated. The exact mechanism of action of dipyridamole in the nephrotic syndrome is still obscure in many respects. However, the fact that the drug shares its anti-platelet action with the non-steroid anti-inflammatory drugs, e.g. aspirin and indomethacin, and the rapidity with which it produces its urinary protein-decreasing effect, strongly suggests that it inhibits the release of vasoactive amines and other chemical mediators from blood platelets. As far as the present study is concerned, adverse side effects of dipyridamole were few or minimal, even when the drug used in large doses over a prolonged period of time. From these results it is considered that dipyridamole provides a new remedy which is worthy of trying in nephrotic syndrome as a means of reducing the requirement of steroids and immunosuppressive drugs.

Studies on platelet function in patients with prostatic cancer: Preliminary report

Platelet function was evaluated as an index of the thromboembolic tendency in patients with untreated, advanced prostatic cancer. Patients with benign prostatic hypertrophy (BPH) and similar age distribution served as a comparison group. Platelet aggregations were elevated in both groups, but not significantly different from each other. Platelet serotonin level in patients with prostatic cancer was lower than in patients with BPH (p less than 0.01), whereas plasma serotonin level in patients with prostatic cancer (within normal ranges in our series) was lower than in patients with BPH (p less than 0.001). Levels of 2 intraplatelet proteins, beta-thromboglobulin (beta-TG) and platelet factor 4 (PF4) in the two groups of patients were similar. However, levels of beta-TG were elevated significantly in both groups of patients compared with those of healthy individuals. These studies revealed that the platelet serotonin levels in advanced prostatic cancer patients differed significantly from those in patients with BPH. The platelet serotonin level thus may provide an index of platelet activation in patients with prostatic cancer.

Circulating Immune Complexes and Immunosuppressive Acidic Protein in Patients with Renal Cell Carcinoma

Sera from 23 patients with renal cell carcinoma were tested for circulating immune complexes by Clq binding assay and immunosuppressive acidic protein by single radial immunodiffusion. The mean values of circulating immune complexes and immunosuppressive acidic protein in the patients were significantly higher than those of normal controls, and over-all positive rates were 52 and 35 per cent, respectively. There was a significant correlation between immunosuppressive acidic protein levels and the extent of tumor invasion. Immunosuppressive acidic protein may be a more useful marker than circulating immune complexes. However, presently synthetic evaluations of patients with various nonspecific markers, including these 2 factors, will be necessary.

Elastase--does this enzyme play any role in bladder cancer invasion?

Elastase activity in bladder cancer tissue and normal bladder epithelium was assayed with succinyl trialanine paranitroanilide (Suc-Ala3-NA), a synthetic substrate of elastase. Elastase in the tissue extract liberated paranitroanilide (NA) from Suc-Ala3-NA and coexistant endopeptidases produced alanine-NA and dialanine-NA. These products were detected by high performance liquid chromatography. All activities of elastase and endopeptidases in bladder cancer tissues from 20 patients are significantly higher than those in 17 normal bladder epithelia. In 20 bladder cancer cases evaluated, the relation between the activities of elastase and endopeptidases and the background factors of each bladder cancer case was examined. There was a significantly different activity of elastase between the superficial bladder cancer group (less than or equal to T1) and the invasive group (T2 less than or equal to). Elastase is assumed to play an important role in cancer invasion.

Fluorescence study of renal cell carcinoma with antibodies to renal tubular antigens, intermediate filaments, and lectins.

A fluorescence study was performed in 16 renal cell carcinomas using antibodies to renal tubular antigens (RTA), two intermediate filaments, cytokeratin and vimentin, and two lectins, soybean agglutinin (SBA) and peanut agglutinin (PNA). We observed the presence of RTA, cytokeratin, and vimentin in all of our specimens. The expression of vimentin, the cytoskeletal protein of mesenchymal cells, was considered to be very interesting feature of the tumor. Binding sites of SBA, normally present in glomeruli, proximal and distal tubules, were detectable in the neoplastic cells in only 37.5% of our specimens. PNA did not react with the tumor except for the small area of 2 specimens. Lectins may be useful for estimating the characteristics or renal cell carcinoma including its malignant potentials, and antibodies to RTA and intermediate filaments seem to be available for the diagnosis of the tumor in metastatic lesions.

Antibody Avidity for Native DNA in Lupus Nephritis

To study the participation of the antibody avidity in the pathogenesis of lupus nephritis, we measured the antibody avidity to native DNA by the method of Minden and Farr (14) in patients with SLE. The avidity to native DNA was almost less than 40%. The avidity and the histological activity of lupus nephritis were well correlated; the antibody avidity of the active-group sera was higher than that of the inactive-group sera and the group without nephritis. In the group with antibodies of relatively high avidity, the complexes were deposited mainly in the subendothelial side of GBM and in the mesangium. On the other hand, in the group with low-avidity antibodies, the complexes were localized in the subepithelial side of GBM. On investigating the quality of the complexes, we noted that the complexes composed of high-avidity antibodies prepared in vitro were larger than those of low avidity and the former were larger than 19S. If the native-DNA-anti-native DNA system is the mechanism basic to lupus nephritis, the differences in immune response of the host, namely the degree of antibody avidity may greatly affect complex formation and influence the histological activity and nephritogenicity of lupus nephritis.