High-resolution HLA-DRB1 and DQB1 genotyping in Japanese patients with testicular germ cell carcinoma.

We report for the first time the frequency distributions of HLA-DRB1 and -DQB1 genes in 55 patients with testicular germ cell carcinoma (TGC) using the modified PCR-RFLP method and compare the results with those for 1216 healthy Japanese control subjects. The modified PCR-RFLP method produced accurate, reproducible cleavage patterns that are easily discriminated. HLA-DRB1*0410 was the susceptibility allele (RR = 3.26, P = 0.006) and DQB1*0602 appears to be a candidate protective allele (RR = 0.26, P = 0.02) for TGC in the Japanese. None of the HLA-DRB1 and -DQB1 alleles showed a specific tendency for histological type or clinical stage of the tumours. Previous studies based on serotyping methods failed to show these allelic associations. High-resolution genotyping is essential because the peptide-binding domain of MHC class II molecules is determined more precisely by their genotypes than by their serotypes. In addition, inherent technical difficulties and typing errors of up to 25% make serotyping inefficient. Our results suggest that high-resolution genotyping is a useful genetic marker to determine risk for TGC.     

Antitumor effect of KT6124, a novel derivative of protein kinase inhibitor K-252a and its mechanism of action

Summary Novel derivatives of K-252a, (8R*,9S*,11S*)-(–)-9-hydroxy-9-methoxycarbonyl-8-methyl-2,3,9,10-tetrahydro-8, 11-epoxy-1H,8H,11H-2,7b,11a-triazadibenzo [a,g]-cycloocta[cde]trinden-1-one, an inhibitor of protein kinases and calmodulin-dependent phosphodiesterase, were synthesized and evaluated for their antitumor activity in vitro and in vivo. Of ten derivatives tested, four were active against the P388 murine leukemia i. p.-i. p. system, although K-252a was inactive. Among these derivatives, KT6124 was selected for further biological evaluation studies because its efficacy was the highest. KT6124 was also active against sarcoma 180 and B16 melanoma. It exerted a relatively broad spectrum of antiproliferative activity against 20 human tumor cell lines in vitro. To determine the mechanism(s) of action underlying the antitumor activity of KT6124, we tested the drug for inhibition of protein kinases, including Ca²⁺-and phospholipid-dependent protein kinase (PKC), in intact A431 human epidermoid carcinoma cells in comparison with the PKC-inhibitory activity of K-252a. KT6124 did not antagonize the action of phorbol 12-myristate 13-acetate (PMA) in A431 cells, whereas K-252a did, suggesting that KT6124 may not act on protein kinases in the cells. The interaction of KT6124 with DNA in living cells was examined by the alkaline elution method. KT6124 apparantly exhibited DNA scission both dose-and time-dependently in the target cells. The DNA breakage was dependent on proteinase K treatment, suggesting its possible interaction with DNA-related enzyme(s). These results indicate that KT6124 exerts antitumor activity by acting on DNA or on DNA-related enzyme(s) in tumor cells rather than via the inhibition of protein kinases.     

Relapse of colon cancer followed by polymyositis: Report of a case and review of the literature

The direct causal relationship between dermatomyositis-polymyositis (PM) and malignancy remains controversial. We describe herein the case of a patient who underwent surgical treatment for colon cancer, which had preceded the onset of PM with tumor relapse. The PM markedly improved following the initiation of steroid therapy, and has remained under control, probably as a result of chemotherapy. The current concepts of variable clinical courses and the possible mechanism for the association of PM with malignancy are discussed following this case report.     

Therapeutic efficacy of miconazole on deep-seated fungal infections in the respiratory tract system

We evaluated the therapeutic efficacy of miconazole (MCZ, Florid-F inj.), a new antifungal agent for parenteral use, in deep-seated fungal infections of respiratory tract system. A daily dose of 400-1,800 mg of MCZ was given intravenously for 12-38 days (mean: 23.4 days) to 7 patients: 2 patients with pulmonary aspergillosis, 1 patient with bronchial aspergillosis, 1 patient with pulmonary candidiasis and 3 patients with candidemia. One additional patient with pulmonary aspergillosis received three instillations of 20 mg of MCZ into the thoracic cavity. The clinical effects were excellent in 1, good in 4 and poor in 3 patients. The efficacy rate was 100% in 5 cases with respiratory fungal infections but 3 cases with candidemia did not respond well to the treatment. Four strains each of Aspergillus sp. and Candida sp. were identified as causative organisms. Seven of the 8 strains were eradicated by administration of MCZ. Side effects observed were irritation and heat in a leg in 1 patient, hyperlipoidemia in 2 patients and eosinophilia in 1 patient. The adverse reactions disappeared after the completion of the therapy. From the above results, we conclude that MCZ is one of the most useful antifungal agents for parenteral use as a first choice on deep-seated fungal infections in the respiratory tract.     

Neurogenic stress ulceration caused by laparotomy under anesthesia plus restraint. The device of a new rat model

We report herein, a new method devised of producing neurogenic stress ulceration in rats. An experimental subarachnoid hemorrhage (SAH) was produced in rats by injecting 0.2 ml of arterial blood from other rats into the cisterna magna. Three days later, the rats were laparotomized for 1 hour under ether anesthesia, followed by restraint for 3 hours in wakefulness. The SAH rats were found to develop stress ulcers (UL-I) in the glandular stomach, which were significantly (p less than 0.001) more marked than those in non-SAH rats. Measurements were performed on gastric acid secretion, an important aggressive factor. It was found that the SAH rats undergoing the laparotomy-restraint stress showed a more marked increase in gastric acid secretion and a more marked reduction in MBF, than the non-SAH rats. The effects of bilateral vagotomy, upper abdominal sympathectomy and bilateral adrenalectomy were examined, and it was revealed that the SAH rats were under the condition of hyperreactivity both in the sympathetic and parasympathetic systems and, on this basis, the laparotomy-restraint stress caused the stress gastric ulceration. In this rat model, the laparotomy stress was applied under anesthesia and any exposure to low temperatures which may have interfered with blood circulation was avoided.     

Application of cytopathology in a sensitivity test for anti-tumor agents. I. An experimental study

Although the human tumor clonogenic assay (HTCA) is extremely reliable in determining clinical correlations, it is a complicated process requiring considerable time in order to obtain results. Thus, an experimental study on cytopathologic observation (cytologic assay) and comparative evaluation between it and HTCA were performed in order to establish a more rapid and accurate drug sensitivity test. Materials included Colon 26, a cell line established in our department, malignant effusion and surgical specimens. In carrying out HTCA according to the Hamburger-Salmon method, the cell suspension samples following exposure to anti-tumor agents (MMC, L-PAM, ADM, CDDP) were cultivated in test tubes for 3-8 hours and stained by the Papanicolaou and Giemsa methods. According to Tokita's criteria, when cellular changes showed as nuclear pyknosis and nuclear destruction were found to have increased significantly in comparison with a control group, the cells were judged to be sensitive. Very similar and parallel results were obtained between HTCA and cytologic assay in this study, with a significant correlation. Cytologic assay was proved to be an easy, rapid and accurate method for testing drug sensitivity and its clinical application can be expected in the future.     

Sequence-dependent Termination of Mammalian DNA Polymerase Reaction by a New Platinum Compound, (–)-(R)-2-Aminomethylpyrrolidine(1,1-cyclobutane-dicarboxylato)-2-platinum(II) Monohydrate

We examined the mechanisms of the inhibition of DNA synthesis by a new platinum compound, (-)-( R )-2-aminomethylpyrrolidine(1,1-cyclobutane-dicarboxylato)-2-platinum(II) monohydrate (DWA-2114R), a derivative of the antitumor drug cis- diamminedichloroplatinum(II) (CDDP), using prokaryotic and eukaryotic DNA polymerases. Preincubating activated DNA with CDDP or DWA-2114R reduced its template activity for prokaryotic and eukaryotic DNA polymerases in a dose-dependent manner. DWA2114R required six times greater drug concentration and two times longer incubation time to show the same decrease of the template activity compared to CDDP. Treatment of primed pUC118 ssDNA templates with the two drugs followed by second-strand synthesis by prokaryotic and eukaryotic DNA polymerases revealed that DWA2114R bound to DNA in a similar manner to CDDP and these adducts blocked DNA elongation by DNA polymerases of eukaryotes as well as of prokaryotes. With these two drugs, the elongations by E. coli DNA polymerase I (Klenow fragment), T7 DNA polymerase and calf thymus DNA polymerase α were strongly arrested at guanine-guanine sequences (GG). Stop bands were also observed at adenine-guanine sequences (AG) guanine-adenine-guanine sequences (GAG) and mono-guanine sequence (G). Calf testis DNA polymerase β was also arrested efficiently at AG, GAG and G, but much more weakly at GG. This pattern was common to DWA2114R and CDDP.     

A study on the metabolism of galactose cataract using a 31P-NMR spectroscopy

Using a 31P-NMR spectroscopy, we monitored the metabolic kinetics of energy organophosphate compound in rat lens during the process of generating galactose cataract. The most remarkable metabolic change in the earlier phase of galactose cataract formation was found in alpha-glycerophosphate. This increased significantly, as compared to controls, since the day 3 of giving feed containing 25% galactose. The high level lasted for up to three weeks, decrease followed by a gradual decrease and subsequently a significant decrease at five weeks. Adenosine triphosphate (ATP) showed a significant decrease in the galactose group compared to the controls from two weeks after beginning of the experiment and the decrease continued. Inorganic orthophosphate increased gradually in the galactose group as compared to the controls, the increase being of significance at one week reading a maximum at two weeks followed by a subsegment decrease. Our basic study suggests that 31P-NMR spectroscopy is a useful technique in lens of the metabolic kinetics, to noninvasively determine the pathophysiology of galactose cataract, which has been studied biochemically and histologically.     

Immunohistochemical and ultrastructural study on effect of fibroblast growth factor on transformation of fibroblasts to regenerated mesothelial cells

To elucidate the effect of fibroblast growth factor on the phenotypical conversion of fibroblasts to mesothelial cells, both immunohistochemical and ultrastructural examinations were carried out on cultured spheroids that were composed of fibroblasts obtained from the parietal pleura of rats with and without addition of antifibroblast growth factor receptor antibody. In the present study, antifibroblast growth factor receptor antibody was employed to block the effect of the autocrine component of fibroblast growth factor in the culture medium. Phenotypical conversion from fibroblast to mesothelial cells was clearly blocked in the experimental group, to which culture medium had been added with antifibroblast growth factor receptor antibody, whereas the control group, cultured without addition of antifibroblast growth factor receptor antibody, showed phenotypical conversion of fibroblasts that was confirmed by the development of macula adherens, microvilli, and positive expression of cytokeratin. These results indicate the possibility that fibroblast growth factor plays a key role in the process of phenotypic conversion of fibroblasts to regenerated mesothelial cells.     

Kindling of the visual cortex in cats: comparison with amygdaloid kindling

Kindling of the primary visual cortex (VC) was compared with that of the amygdala in cats. VC kindling was basically similar to kindling of the amygdala in that daily electrical stimulation can lead to the development of a generalized convulsion in most subjects, a growth of afterdischarges in their configuration and duration, and a reduction of the afterdischarge threshold. The kindling response of the VC differed from that of the amygdala in a number of respects, i.e., a high afterdischarge threshold, a different pattern of behavioral seizure development, an abrupt growth of electroclinical seizures coincident with the onset of a generalized convulsion, an intersubject variability in seizure susceptibility, and a marked seizure instability. In VC kindling the afterdischarge propagation into the amygdala was not observed until the generalized convulsion developed, and the early involvement of afterdischarge was seen in the pulvinar, lateral geniculate body, and superior colliculus. These data suggest that a neural mechanism different from amygdaloid kindling may participate in VC kindling, and that the subcortical structures of the visual system are involved in the preferential pathway for a seizure generalization from the VC.