Radioimmunoassay of chenodeoxycholic acid-3-sulfate in urine

A sensitive and specific radioimmunoassay (RIA) for the measurement of urinary total chenodeoxycholic acid-3-sulfate (SCDCA) was developed and the accuracy was confirmed. SCDCA bound to bovine serum albumin as the antigen and emulsified with Freund's complete adjuvant was injected into rabbits. The antiserum obtained was capable of binding 75% of [11,12-3H]SCDCA at 1:1000 dilution. The percentage of bound radioactivity decreased linearly with logarithmic increases in unlabeled SCDCA, from 8 to 200 pmol/ml. The antiserum showed an extremely high specificity for SCDCA (free and conjugated), and the values determined by RIA indicated a close correlation with those found by gas-liquid chromatography. The daily urinary SCDCA level was determined using SCDCA-RIA in 12 disease-free humans and 74 patients with chronic liver diseases. In the normal subjects the daily urinary SCDCA level was 0.74 +/- 0.83 mg/day and increased levels were evident in all groups with chronic liver diseases. The daily urinary SCDCA level corresponds closely with the extent of hepatic dysfunction.     

Cancer chemotherapy and somatic cell mutation

The occurrence of a second neoplasm is one of the major obstacles in cancer chemotherapy. The elucidation of the genotoxic effects induced by anti-cancer drugs is considered to be helpful in identifying the degree of cancer risk. Numerous investigations on cancer patients after chemotherapy have demonstrated: (i) an increase in the in vivo somatic cell mutant frequency (Mf) at three genetic loci, including hypoxanthine–guanine phosphoribosyl-transferase (hprt), glycophorin A (GPA), and the T-cell receptor (TCR), and (ii) alterations in the mutational spectra of hprt mutants. However, the time required for and the degree of such changes are quite variable among patients even if they have received the same chemotherapy, suggesting the existence of underlying genetic factor(s). Accordingly, some cancer patients prior to chemotherapy as well as patients with cancer-prone syndrome have been found to show an elevated Mf. Based on the information obtained from somatic cell mutation assays, an individualized chemotherapy should be considered in order to minimize the risk of a second neoplasm.     

Comparative studies of phosphoenolpyruvate carboxylase from c(3) and c(4) plants

Phosphoenolpyruvate carboxylase (PEPC) from several C₃ plants was compared to maize PEPC by immunoblotting using an antibody against maize PEPC and by peptide mapping. In C₃ gramineous plants, PEPCs of slightly different monomeric sizes were detected as two bands for wheat and barley leaves, as three bands for etiolated maize leaves and as four bands for rice leaves by SDS-polyacrylamide gel electrophoresis and immunoblotting, whereas only one PEPC band was detected for maize leaves, a C₄ plant, or tobacco leaves, a dicotyledonous C₃ plant. The peptide fragment patterns of the lower molecular weight PEPC (major band in immunoblotting) in wheat leaves was similar to that of maize PEPC in peptide mapping by protein staining or by immunological detection, but the upper one (minor band) had a different pattern from the lower one in peptide mapping by immunological detection and few peptide fragments from this were recognized by the anti-(maize) PEPC antibody. These results suggest that there are multiple forms of PEPC subunits in the gramineous plants tested, and the major PEPC has a primary structure similar to that of maize PEPC. To obtain information about the expression of PEPCs in C₃ plants, changes in the amount of PEPC protein were investigated during the greening of rice and wheat seedlings. Judging from the regulation by light, there were two types of PEPCs in greening rice seedlings, one induced by light and the other reduced by it. Greening wheat seedlings also show a PEPC band induced by light. These findings indicate that some PEPCs in C₃ gramineous plants not only have structures similar to that of maize PEPC, but also are regulated by light in a similar manner.     

Nucleotide sequence of the PR-1 gene of Nicotiana tabacum

A gene encoding one of the pathogenesis-related proteins, PR1a, and two related pseudogenes were isolated from Nicotiana tabacum. The cloned PR1a gene (pPR-gamma) and one of the pseudogenes (pPR-alpha) were sequenced and found to have similar structures. The sequence of pPR-gamma was quite similar to that of the cDNA clone of PR1a. The plasmid pPR-gamma did not contain an intron and had a typical promoter sequence in the 5'-flanking region.     

Mutagenicity of ptaquiloside, the carcinogen in bracken, and its related illudane-type sesquiterpenes II. Chromosomal aberration tests with cultured mammalian cells

The chromosomal aberration test using a Chinese hamster lung cell line (CHL) was carried out on ptaquiloside and its related compounds, hypoloside B, hypoloside C, illudin M and illudin S. Ptaquiloside induced chromosomal aberrations at doses as low as 4.5 μg/ml (0.0113 mM). The clastogenic effect was ph-dependent. The same activity was observed at a 90-fold higher dose at pH 5.3 in the culture medium compared with the activity at pH 74. or pH 8.0. Both hypoloside B and hypoloside C were also clastogenic at almost the same dose levels as that of ptaquiloside. Illudin M and illudin S were also potet clastogens and induced aberrations at much lower doses than ptaquiloside. These results suggest that the clastogenic effect is involved in the mechanism of carcinogenic potency of ptaquiloside in animals.     

Involvement of the acyl chain of ceramide in carbohydrate recognition by an anti-glycolipid monoclonal antibody: the case of an anti-melanoma antibody,M2590, to GM3-ganglioside

The effect of the chain length of the fatty acid residue of the ceramide moiety of ganglioside G_M3 on the binding ability of monoclonal antibody M2590, which is specific for the carbohydrate structure of G_M3-ganglioside, was examined by means of a direct binding assay on thin layer chromatography plates (TLC immunostaining) and a quantitative enzyme-linked immunosorbent assay (ELISA). Derivatives of G_M3 with a long fatty acid chain reacted with the M2590 antibody, but those with a short fatty acid chain showed no reaction in either assay system. These results suggested that the acyl fatty acid moiety of the ganglioside played an important role in the formation or maintenance of the antigenic structure of the carbohydrate moiety of the ganglioside.     

Pharmacological intervention in oxidant-induced calcium pump dysfunction of dog heart

Micromolar concentrations of HOCl, an oxidant produced by activated neutrophils, inhibited Ca2+ uptake and Ca2+ATPase of isolated dog heart sarcoplasmic reticulum (SR). DTT antagonized completely the HOCl effect only when it was given within 5 min after the addition of HOCl. When the pharmacological intervention was delayed, the recovery with DTT was not complete, and administration of DTT 30 min after the start of HOCl's reaction with SR resulted in only a small improvement in SR Ca2+ uptake. Although H2O2 and Fe ion-chelate (a free radical-generating procedure) also inhibited Ca2+ uptake and ATPase, the concentrations required were very large. The response of cardiac sarcolemmal and skeletal muscle SR calcium pumps to oxidants was similar to that of the cardiac SR calcium pump.