Biomedical subjects
Publications and source records attributed to R Goto.
99mTc-labeled L-cysteine, DL-homocysteine, and S-carbamyl-L-cysteine were prepared and tested for effectiveness in gamma camera imaging and for bioradiodistribution in mice bearing Ehrlich solid tumors. Among these labeled compounds, 99mTc-DL-homocysteine seems most promising for the nuclear medical imaging of tumors.
A group of EMT6 tumor bearing male BALB/c mice which had been treated with alpha-difluoromethylornithine (DFMO, a specific, irreversible inhibitor of ornithine decarboxylase, the enzyme which catalyzes the biosynthesis of putrescine), 8 mg/mouse, ip, 20 and 5 hrs before the 14C-putrescine dose, and a group of control animals were administered 14C-putrescine (0.5 muCi, 0.1 mCi/mmol, iv) 60 min prior to sacrifice. Radioactivity uptake data was obtained for the tumor and 13 major normal organs. In the control animals the tumor exhibited one of the highest uptakes of radioactivity. For DFMO-pretreated mice the radioactivity distribution among most of the normal tissues was not very different from that obtained for the control animals. However, the uptake into the tumor was enhanced by a factor of approximately 4. So, high tumor-to-tissue ratios (3.8, lung to 38, brain) were attained as a result of DFMO treatment.
LE cell formation is one feature of systemic lupus erythematosus exhibited by virtually all mice of the NZB/NZW strain and is the result of accumulation of antibodies directed against components of cell nuclei. A hybrid cell line which produces antibodies capable of inducing LE cell formation in vitro has been isolated in a hybridoma fusion using the splenocytes of unimmunized NZB/NZW mice. These monoclonal autoantibodies provide an intense staining of the chromatin in cells of a number of divergent species and tissues. They bind strongly to the histone rich (2 M NaCl) fraction of extracted, isolated nuclei. Further analyses using the antibodies in immune precipitations and in antibody labeling of capillary blots on nitrocellulose sheets of calf thymus histone demonstrate that the antibodies are directed against histones and are capable of reacting with H1, H2a, H2b, H3, and H4 histones individually. In contrast to human autoantibodies with histone specificity, no cross-reactivity of this monoclonal autoantibody with the lymphocyte surface could be detected by either immunofluorescence or immunoelectron microscopy.
The role of exchange transfusion with fresh whole blood in septic shock infants was evaluated. Endotoxin was positive in 8 of 10 infants and completely removed from 6 by exchange transfusion. All 6 infants with negative endotoxins after exchange transfusions survived, but 2 with positive reactions died despite the exchange transfusion. Staphylococcus aureus was cultured in the blood from 2 infants with negative reactions and one of them survived. These findings suggested that the clearance of endotoxin has an important role in exchange transfusion among septic shock neonates.
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In order to ascertain the significance of the hydrophobic region in the carrier protein responsible for the D-leucine transport reported previously, the effects of p-chloromercuribenzene sulfonate (PCMBS) were investigated on the transport of D- and L-leucines in Ehrlich ascites tumor cells. The uptake of D-leucine in the presence of PCMBS was inhibited more strongly than that of L-leucine. Km values for D-leucine uptake increased with increasing concentration of PCMBS, while those for L-leucine showed only a little increase. D-Leucine uptake was restored considerably from the PCMBS inhibition by washing with buffer, whereas L-leucine uptake was only slightly affected. The inhibition of D-leucine exit by PCMBS was also higher than that of L-leucine. These results suggest that the binding site of PCMBS for the carrier protein of Ehrlich cells is involved in its hydrophobic region that would be more significant for the binding of alkyl side chain D-leucine than that of L-leucine.
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As an extension of the previous finding that radioactivity of 14C-labeled D-amino acids after injection is localized preferentially in the tumor and the pancreas of tumor-bearing animals as compared with the corresponding L-amino acids tested, the results of similar uptake experiments using other tumors araa reported here. The present studies show high radioactivity uptake by human colon cancer, human thyroid cancer, and human leiomyosarcoma transplanted into nude mice, and by solid leukemia L1210 and solid sarcoma 180, but not by Morris hepatoma 7316A or 3'-methyl-4-(dimethylamino)azobenzene-induced rat hepatoma. The results suggest the potential utility of 11C-labeled D-amino acids for the detection of some cancers.
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The selective affinity of amino acids for pancreas, due to its high rate of protein synthesis, has prompted us to synthesize new radio-labeled amino acids for pancreatic imaging. Labeling of 19 unnatural amino acids with 99mTc was tried in the usual way, and 10 labeled products were obtained with low yields. The highest distribution of radioactivity from all these labeled amino acids, which had been injected intravenously into mice, was found in the kidney, whereas little activity was found in the pancreas. Thier low distributions in stomach suggest little dissociation of the labeled amino acids into 99mTcO4- in mice. It is presumed that these labeled amino acids behave unlike their original acids in the body.
Nonhistone chromasomal proteins (NHP) of Ehrlich ascites tumor cells were fractionated into four subfractions by successive extractions with 0.35 M, 0.45 M, and 0.60 M NaCl, and finally 2.0 M NaCl + 5.0 M urea. The CD spectra and transcriptional activities of complexes reconstituted from DNA, histone, and each of these four NHP subfractions obtained from Ehrlich tumor cells were measured in order to identify the effect of each NHP subfraction. The positive ellipticity in the CD of the DNA-histone complex was considerably increased when reconstituted with 0.35 M- or 0.45 M-NHP. The 0.35 M- and 0.45 M-NHP also increased the transcriptional activity of DNA-histone complex. The activities of DNA-histone-0.35 M NHP complex increased progressively with increase in the 0.35 M NHP/DNA ratio. These results suggest that there is a correlation between the increase in transcriptional activity of DNA-histone complexes with 0.35 M- or 0.45 M-NHP and the increase in the percentage of DNA in the B conformation in complexes.
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The present investigation was undertaken to examine an evidence for the exchange diffusion of D-leucine as well as L-leucine and further to study the effect of temperature reduction on the exchange diffusion, in Ehrlich ascites tumor cells in vitro. The homoexchange diffusion for D-leucine was observed at 37 degrees C, the extent being nearly equal to that for L-leucine. The heteroexchange diffusion between D- and L-leucines also occurred at 37 degrees C. These findings were compatible with an assumption that there is a common pathway for D- and L-leucine transport in the tumor cells. It was demonstrated that the exchangeability of D-leucine was depressed remarkably at 0 degree C, while L-leucine still had a considerable exchangeability. This difference in exchangeability of D- and L-leucines in temperature reduction seems to reflect that the bulky hydrocarbon side chains of both the isomers are bound to the different site of carrier as described in the preceding paper.
In a previous paper, we reported that various D-amino acids were taken up several times more effectively than the corresponding L-isomers into several tumors tested in vivo. In order to investigate this further, the in vitro uptake of D-[14C]leucine by Ehrlich ascites tumor cells was investigated in comparison with that of L-[14C]leucine. The distribution ratio, the effects of amino acids and pH, and an approximately linear Lineweaver-Burk plot indicated that D-leucine was transported by an active transport system for L-leucine. Vmax and ku, the first-order rate constant for the unsaturable component, for the uptake of D- and L-leucines decreased with a fall in temperature. The activation energies for Vmax and ku were in the range of 5-10 and 18-21 kcal/mol, respectively. The values for L-leucine were greater than those for D-leucine. Km for D-leucine transport increased with decreasing temperature, whereas Km for L-leucine decreased. This difference suggests that the large alkyl chains of D- and L-leucines bind to different portions of a carrier protein.
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