Immunologic characterization and molecular profile of carcinoembryonic antigen detected by monoclonal antibodies

Four distinct monoclonal antibodies, which reacted with CEA preparations but not with nonspecific cross-reacting antigen or with nonspecific cross-reacting antigen 2, were established. Except for monoclonal antibody AS001 , all of these monoclonal antibodies immunoprecipitated molecular forms of 200K and 180K daltons that are not bridged by disulfide bonds. Immunodepletion experiments and sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis revealed that these monoclonal antibodies recognized the same antigenic structure when 125I-CEA preparation was used. Monoclonal antibody AS001 is of particular interest, because this antibody reacted only with a 200K dalton molecule which is a part of the molecules recognized by the other three monoclonal antibodies. The rosette inhibition assay and the immunoprecipitation experiments suggest that each monoclonal antibody recognizes a different antigenic determinant. The antigenic determinants recognized by monoclonal antibodies YK013 and AS001 may be peptides in nature, whereas the determinants recognized by antibodies YK024 or AS005 might be carbohydrate. The radioimmunoassay with monoclonal antibody AS001 was established, and the results clearly indicate that the incidence of positivity for the sera from digestive tract cancer patients and from lung cancer patients obtained by monoclonal antibody AS001 was higher than that obtained by the polyclonal antibody. Monoclonal antibody AS001 was able to detect the corresponding antigen in the sera, which the polyclonal antibody failed to detect. This study therefore suggests that monoclonal antibodies may enhance and improve the diagnostic value in cancer patients with undetectable or lower CEA levels detected by conventional anti-CEA antibodies.     

Control of insecticide resistance in a field population of houseflies,Musca domestica,by releasing susceptible flies

Susceptible houseflies, Musca domestica, were released at a waste disposal site to control insecticide resistance in a field housefly population. In the first experiment, a total of 163,000 pupae of the susceptible Takatsuki strain were released in October–November 1977. LD₅₀ values to fenitrothion and diazinon decreased to about one-sixth in April 1978, five months after the releases, of those before the releases. For the second experiment, a susceptible colony was derived by cross and backcross between a white-eyed substrain of the Takatsuki and a field colony. This susceptible colony consisted of whiteeyed flies with low activity and normal-eyed flies bearing no or one white eye gene. The results of large cage experiments suggested that the normal-eyed males of the susceptible colony had half the mating competitiveness of wild males. Approximately 31,000–46,000 susceptible pupae were used in each of five releases from October to November 1980. The population number of each sex, estimated by a mark-release-recapture method, increased from 12,000 in late September to 35,000–43,000 in middle November and then decreased to 5,000–8,000 in early December. The frequency of field-collected males bearing one white eye gene and those bearing one male determining factor, which were characteristics of the susceptible colony released, increased gradually during the period of releases. The susceptibility of the field population to fenitrothion and diazinon was examined five times in the period from September to December 1980. With time, the dosage-mortality regression gradually shifted towards that of the susceptible colony after starting the releases. LD₅₀ values to fenitrothion and diazinon decreased to about one-sixth and one-fifth, respectively, in June 1981, six months after the second series of susceptible fly releases, of those before the releases. Ratios of the wild flies to the released fiies were estimated to be between 4.7∶1 and 9.8∶1 in males and between 3.0∶1 and 3.9∶1 in females by taking the quality of the released colony and the population parameters of the field houseflies into consideration. Under several assumptions, the manner of resistant phenotype reduction was discussed, based on the dosage-mortality regressions and the ratios of released flies. These results showed that the releases of susceptible flies were successful in suppression of insecticide resistance in the field housefly population.     

Inhibition of pentagastrin-stimulated gastric acid secretion by intraileal administration of bile and elevation of plasma concentrations of gut glucagon-like immunoreactivity in anesthetized dogs

The effects of intraileal administration of bile on gastric acid secretion stimulated by a submaximal dose of intravenous pentagastrin infusion and on plasma concentrations of gut glucagon-like immunoreactivity (gut GLI) were studied in anesthetized dogs. Gastric acid secretion was measured for a 2-h period at 15-min intervals before and after intraluminal instillation of test solutions. 100 ml of canine bladder bile diluted to 10% in saline evoked a significant inhibition (20%) of gastric acid secretion. The inhibition of gastric acid secretion was accompanied by an elevation of plasma concentration of gut GLI, whereas saline instillation (in controls) caused no responses. Although the inhibition of gastric acid secretion and the elevation of plasma gut GLI are parallel phenomena, gut GLI can be reasonably postulated as one of the candidate mediators of bile-induced inhibition of gastric acid secretion, since its structurally related peptides, pancreatic glucagon, glicentin and oxyntomodulin have been reported as inhibitors of gastric acid secretion.     

The rapid polyphosphoinositide metabolism may be a triggering event for thrombin-mediated stimulation of human platelets

The metabolism of polyphosphoinositides was examined in human platelets activated by thrombin. The addition of thrombin to [3H]glycerol-labeled platelets induced an initial loss and a subsequent increase of the radioactivity in phosphatidylinositol-4,5-bisphosphate (TPI) without any significant change in phosphatidylinositol-4-phosphate (DPI). A marked enhancement of [32P]Pi incorporation into TPI occurred in parallel with an increase in this lipid content, which was accompanied with a conccurent decrease in phosphatidylinositol (PI). The rate of this subsequent increase in TPI was smaller than that observed in [3H]arachidonic acid-labeled platelets, suggesting that formed TPI in activated platelets may contain much greater amount of arachidonate than preexisting TPI in resting platelets. These data indicate that thrombin causes a rapid change in TPI metabolism (initial degradation of preexisting TPI and subsequent production of arachidonate-rich TPI), which might be a primary candidate to modulate thrombin-induced function in human platelets.     

Purification and properties of rat brain dipeptidyl aminopeptidase

Dipeptidyl aminopeptidase, which hydrolyzes the 7-(Gly-Pro)-4-methylcoumarinamide, has been purified from the brains of 3 week-old rats. It was purified about 2,600-fold by column chromatography on CM-cellulose, hydroxyapatite and Gly-Pro AH-Sepharose. This enzyme hydrolyzed Lys-Ala-beta-naphthylamide well with an optimum pH of 5.5. It was inhibited by diisopropyl fluorophosphate, phenyl-methanesulfonyl fluoride, some cations, and puromycin, but was not inhibited by p-chloromercuribenzoate, N-ethylmaleimide, dithiothreitol, EDTA, iodoacetic acid, and bacitracin, indicating that rat brain dipeptidyl aminopeptidase is a serine protease. This enzyme showed a molecular weight of 220,000 by gel filtration and of 51,000 by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The properties of purified rat brain dipeptidyl aminopeptidase were similar to those of bovine pituitary dipeptidyl peptidase II, but the molecular weight and substrate specificity of these enzymes were different.     

Proton magnetic resonance studies of 7 Fe ferredoxins: Lower potential of the [4 Fe–4 S] redox center than the [3 Fe–3 S] cluster

Two types of iron-sulfur clusters, [3 Fe–3 S] and [4 Fe–4 S], were identified by ¹H-NMR in ferredoxins from Thermus thermophilus, Mycobacterium smegmatis and Pseudomonas ovalis. The [4 Fe–4 S] clusters always showed the redox couples which had potentials lower than that of the [3 Fe–3 S] clusters.     

Cell cycle-dependent expression of antigen-binding and I-J-bearing molecules on suppressor T cell hybridomas

The I-J and antigen-binding chains with constant region determinant (Ct) that compose an antigen-specific suppressor T cell factor were found on the surface of suppressor T cell hybridomas, serologically and morphologically demonstrated by a fluorescence-activated cell sorter (FACS) and immunoelectron microscopic analyses. Moreover, the surface expression of the I-J and Ct-bearing chains fluctuating with the same kinetics depended entirely upon the cell cycle. The maximum expression of these two chains was observed in the early stage of the M phase, and the minimum in the S phase. Similarly, the magnitude of the suppressor activity was maximal in the late stage of the M phase, and was minimal in the S phase. The results therefore demonstrated that there exists good correlation between the cell surface expression of the I-J and Ct-bearing chains and the magnitude of the suppressor activity produced. The antigen recognition units on suppressor T cell hybridomas have serologically and morphologically been characterized by using radiolabeled antigens or monoclonal antibodies against the I-J or Ct on the antigen-binding molecule. Cell-binding assay and radioautographic analysis demonstrated that the suppressor T cell hybridoma possesses the capacity to bind native antigen in an antigen-specific fashion as does the hybridoma-derived, antigen-specific suppressor factor composed of the I-J and the Ct-bearing chains, indicating that the recognition unit on the cell surface is composed of a structure similar to the factor.     

Evidence that cyclic AMP may regulate Ca2+-mobilization and phospholipases in thrombin-stimulated human platelets

The regulation of human platelet responses by cyclic AMP (cAMP) has been investigated by measuring thrombin-stimulated serotonin release, Ca²⁺ uptake and phospholipase activity. Thrombin-induced 1,2-diacylglycerol (DG) formation as a result of phospholipase C activation was inhibited by pretreatment with dibutyryl cAMP (dbcAMP) in a dose-dependent manner. Subsequent failure to produce phosphatidic acid (PA), which is converted from 1,2-DG by phosphorylation and would serve as intracellular Ca²⁺ ionophore, appeared to parallel the decrease in Ca²⁺ uptake activity. Phospholipase A₂ activity, monitored by the production of [³H]lysophosphatidylcholine and [³H]lysophosphatidylethanolamine, was also suppressed by dbcAMP. These data indicate that the intracellular cAMP level may be closely associated with Ca²⁺ uptake and phospholipases activation. In addition, it is suggested that alteration of intracellular cAMP regulates phospholipase activation and consequently platelet responses, perhaps by controlling available Ca²⁺ content.     

Cytological evidence for holocentric chromosomes of the silkworms,Bombyx mori and B. mandarina, (Bombycidae,Lepidoptera)

The nature of the centromere and the orientation in meiosis of silkworm chromosomes were investigated using the trivalent of the F₁ hybrid between the wild and domestic silkworm and X-ray-induced aberrant chromosomes as well as normal silkworm chromosomes. The results of the experiments were as follows: (1) Pro-metaphase chromosomes showed no distinct primary constriction even after treatment with hypotonic solution, (2) sister chromatids separated in parallel along the entire length of the chromosome at mitotic anaphase, (3) chiasmata underwent complete terminalization during diakinesis and thus chromosome dyads were always connected end-to-end by a terminal chiasma at metaphase I, (4) radiation-induced aberrant chromosomes were stably transmitted throughout a number of cell generations, and (5) although the homomorphic bivalents generally orientated axially at metaphase I and equatorially at metaphase II, this normal sequence tended to be inverted or modified in the X-ray-induced aberrant chromosomes and in the trivalent of the F₁ hybrid silkworms. These observations may be best interpreted by assuming the holocentric nature of silkworm chromosomes.