Suppressor cells in the effector phase of autologous cytotoxic reactions in cancer patients

Summary Cytotoxicity was induced in lymphocytes (CL) from 10 out of 15 patients by autologous mixed lymphocyte tumor cell culture and further cultivation with recombinant interleukin-2. In cells from 3 of the 10 patients, cytotoxicity was suppressed by more than 50% when autologous peripheral blood mononuclear cells (PBMC) from the patients with large tumors were added to the autologous killing system. The cells responsible for suppressing the cytotoxicity in the effector phase were adherent or nonadherent to plastic depending on the patient examined. The T cell fraction from 1 patient significantly suppressed the cytotoxic activity, and this suppression was seen only in the autologous system. On the other hand, plastic adherent cells but not T cells from PBMC of 2 subjects suppressed the cytotoxic activity of CL. The reason why the main cell population suppressing the CL activity differed among the patients is unclear. However, the findings that the suppression was mostly abrogated following resection of the tumor mass suggested that suppressor cells, either of macrophage lineage or T cells, are induced in patients with a large tumor mass. This speculation is supported by the finding that the PBMC from a patient with tumor recurrence regained the suppressive activity.     

Effects of two treatments on semen from different bulls on in vitro fertilization results of bovine oocytes

The semen of six different bulls was used to examine the effects of treatment with caffeine or caffeine plus Ca-ionophre on in vitro fertilization, cleavage and development into morulae of in vitro matured bovine oocytes. In vitro fertilization results (formation of both pronuclei, cleavage and development to >/= four-cell stage were significantly (P<0.01) higher using caffeine plus Ca-ionophre than those using only caffeine. The rates of fertilization and first cleavage were only slightly variable among the bulls. However, the present data showed significant variability in formation of both pronuclei (36 to 75%) of fertilized ova and development to the >/=4cell stage (39 to 71%) by different bulls. Development into morulae of ova recovered from the rabbit oviduct did not show any significant differences in relation to sperm treatments or individual bulls.     

Bromodeoxyuridine-immunohistochemistry on cellular differentiation and migration in the fundic gland of Xenopus laevis during development

Summary Cellular differentiation and migration in the fundic glands of adult and larval Xenopus laevis have been examined using bromodeoxyuridine-immunohistochemistry. In the adult fundic gland, cumulative labeling with bromodeoxyuridine revealed a proliferative cell zone between the surface mucous cells and mucous neck cells, in what is referred to as the neck portion of the gland. The labeling-index of mucous neck cells had rapidly increased by week-5. The labeling-index of oxynticopeptic cells showed a more delayed increase until week-7, coincident with the decrease in the labeling of mucous neck cells. In the immature fundic glands of larvae, the labeled proliferating cells were randomly distributed throughout the developing gastric mucosa. During metamorphosis, the labeling-index of immature epithelial cells was highest at stage 63. Following administration of bromodeoxyurdine at this, stage, there was no significant loss of labeled epithelial cells during the metamorphosing period. Furthermore, there was no significant difference in the labeling-indices among the epithelial cells, such as surface mucous cells/generative cells, mucous neck cells, and oxynticopeptic cells, 7 days after administration. Cellular differentiation and migration pathways of epithelial cells in the fundic gland of adult X. laevis and its larvae are discussed.     

Cloning and sequence analyses of cDNAs encoding vasotocin and isotocin precursors of chum salmon,Oncorhynchus keta: evolutionary relationships of neurohypophysial hormone precursors

Summary The nucleotide sequences of cloned cDNAs were used to determine the primary structures of the precursors of vasotocin (sVT) and isotocin (sIT) from the hypothalamus of the chum salmon,Oncorhynchus keta. Two different cDNAs were obtained for each of sVT and sIT precursors (sVT-I and sVT-II; sIT-I and sIT-II). Both sVT and sIT precursors were found to contain a signal peptide and hormone that is connected to a neurophysin by a Gly-Lys-Arg sequence. Northern and Southern blot analyses showed that the sVT and sIT genes are expressed by the same chum salmon hypothalamus, but not by the liver and kidney. Microheterogeneity was found in the nucleotide and amino acid sequences of sVT precursors between our results and the previously reported data (Heierhorst et al. 1990). The conspicuous difference is the occurrence of a stop codon in the middle of sVT-II cDNA. The carboxyl termini of both sVT and sIT neurophysins are about 30 amino acids longer than neurophysins of toad and mammalian neurohypophysial hormone precursors. Although these extended regions do not contain a glycosylation site, they show striking similarity with the glycopeptide moiety (copeptin) of toad vasotocin and mammalian vasopressin precursors. The central portion of the neurophysins shows highest homology among corresponding regions of sVT and sIT precursors. Moreover, calculation of nucleotide substitution rates suggests that a recent gene conversion may have occurred which encompasses the exon that encodes the central segment of the sVT and sIT precursors. A possible pathway for the evolution of precursor molecules of neurohypophysial hormones is discussed.Abbreviations AVP vasopressin - C carboxyl - h human - IT isotocin - MT mesotocin - N amino - OXT oxytocin - S chum salmon - SDS sodium dodecyl sulfate - t toad - VT vasotocin     

Killer cells induced by stimulation with allogeneic tumor cells and subsequent culture with recombinant interleukin-2

Summary Peripheral blood lymphocytes were cultured for 5 days with allogeneic tumor cells (allogeneic mixed lymphocyte/tumor cell culture), and subsequently cultured with recombinant interleukin-2 for 12 days. These cultured cells were found to be cytotoxic to autologous tumor cells. Results of two-color analysis using monoclonal antibodies to cell markers showed that more than 80% of their cultured cells were CD3⁺ cells, and CD4⁺ cells showed a higher distribution than CD8⁺ cells. However, CD8⁺ cells had a much higher killing activity with autologous tumor than did CD4⁺ cells, when estimated by an elimination study using monoclonal antibodies to T cell phenotypes and complement. The cold-target inhibition test showed that the cytotoxicity of these cells for autologous tumor cells was inhibited by unlabeled autologous tumor cells but not by unlabeled stimulator cells. Furthermore, about 40% of the cytotoxicity was suppressed by blocking of HLA class I antigen with a monoclonal antibody on autologous tumor cells. Thus, cytotoxic activity of lymphocytes to autologous tumor restricted by target cell HLA class I antigen is possibly induced by allogeneic tumor-stimulation.     

Analysis of bacterial populations in a grassland soil according to rates of development on solid media

Abstract The process of colony formation by bacteria from grassland soil sampled in April, July and September was simulated by a colony-forming curve (CFC). The CFC was a super-imposition of several component curves (cCFC) given theoretically by the first order reaction (FOR) model [3,6]. The pattern of FOR model curves was not influenced by the time of sampling and four cCFCs were always recognized during an incubation period of 160 h. It was considered that the CFC describes an inherent property of the bacterial population of the field. Bacterial isolates were obtained from colonies produced in each of four cCFCs on agar plates. Isolates corresponding to one cCFC were classified as one group. The bacterial isolates were characterized by morphological and physiological tests and subsequently clustered. Few oligotrophic bacteria were obtained among bacteria which produced visible colonies within 63 h of incubation time. On the other hand, approx. 50% of bacteria which produced v colonies after 63 h were oligotrophic bacteria. The time required for the appearance of the first colony, t _r of the FOR model, was very similar in the isolates belonging to one group. A close linear relationship was observed between t _r value and doubling time of isolates.     

Cervicovaginal and endometrial cytology in ovarian cancer

The clinical significance of cytologic examination was studied in 114 patients with ovarian cancer who had received preoperative cytologic examinations. The overall positive rate of the cytologic examinations was 26.3% (30 of 114): 22 (19.3%) of the 114 cases had positive cervicovaginal smears while 13 of 31 endometrial aspiration smears (41.9%) were positive. The positive rate was not related to the volume of ascites but rather to its presence or absence. Thus, if ascites was observed, the positive rate was about 2.1 times higher than if it was absent. In two of four cases of ovarian cancer with no endometrial invasion but a positive cytologic examination of ascitic fluid, fallopian tube specimens contained cancer cells; this suggests that ovarian cancer cells may reach the cervix and/or vagina by passing through the fallopian tube, particularly if ascites is present. Since cytologic examination, especially of endometrial aspiration smears, shows a high positive rate if ovarian cancer cells are observed in the abdominal cavity, cytology should be used as an important ancillary method for the assessment of ovarian cancer.     

Analysis of strongly acidic amino acids by the conventional amino acid analyzer: application to determination of protein-bound cysteine and glutathione

A rapid analysis method of strongly acidic amino acids and related compounds by a simple modification of an existing amino acid analyzer is presented. In this method, an anion-exchanger column (2.6 X 150 mm) packed with Hitachi 3013-N resin was developed with 0.2 M citric acid. Complete separation of phosphothreonine, phosphoserine, phosphotyrosine, cysteic acid, homocysteic acid, and glutathionesulfonic acid was achieved within 35 min, with no regeneration of the column being required. Tyrosine-O-sulfate was analyzed by the same column using 2 M sodium acetate buffer, pH 5.5. Performic acid oxidation of a variety of proteins and direct analysis of the products by this system successfully detected cysteine, homocysteine, and/or glutathione bound to proteins through disulfide bonds. This suggest the potential use of the method for analysis of the states of protein thiol groups, especially those of clinically significant mutant proteins where mutation of arginine to cysteine is rather frequently recognized.     

Structures of the sugar chains of recombinant human granulocyte-colony-stimulating factor produced by Chinese hamster ovary cells

Recombinant human granulocyte-colony-stimulating factor (G-CSF) was purified from Chinese hamster ovary cells transfected with human G-CSF cDNA. The recombinant human G-CSF was treated with alkaline borohydride and the oligosaccharide-alditols liberated were fractioned by gel filtration on a Bio-Gel P-4 column, followed by high-performance liquid chromatography by use of a strong anion exchanger. Two oligosaccharide-alditols were obtained and their structures were identified by component analysis and 500-MHz 1H-NMR spectroscopy. The structures of the sugar chains were NeuAc alpha 2-3Gal beta 1-3GalNAcol and NeuAc alpha 2-3Gal beta 1-3(NeuAc alpha 2-6)GalNAcol.