Effect of 2-methoxy-4-aminoazobenzene (2-MeO-AAB) and 3-methylcholanthrene (MC) on the induction of microsomal cytochrome P-448 isozymes in primary cultured rat hepatocytes was examined by means of immunochemical methods such as protein A-enzyme-linked immonosorbent assay and immuno-blots using anti-rat cytochrome P-448 monoclonal antibodies and by means of bacterial mutation tests. Although 2-MeO-AAB selectively induced cytochrome P-448H and MC induced both cytochrome P-448H and a low spin form of cytochrome P-448 (P-448L) in the liver of rats, addition of these chemicals to primary cultured rat hepatocytes resulted in selective induction of cytochrome P-448L, as determined by the immunological methods. This was substantiated by the bacterial mutation test using Salmonella typhimurium TA 98 bacteria and two aromatic amine substrates with different specificities to the cytochrome P-448 isozymes. These results suggest that the responses of rat hepatocytes to cytochrome P-450 inducers are different in in vivo and in vitro.

Animals↗

PubMed1987

Thickness of uninvolved fibromuscular stroma and extrauterine spread of carcinoma of the uterine cervix.

The minimum thickness of cervical fibromuscular stroma remaining uninvolved with invasive cervical carcinoma was examined in relation to pelvic node metastases and 5-year cancer death rate, using specimens from Stage IB, IIA, and IIB patients who underwent radical hysterectomy and pelvic lymphadenectomy. The nodal metastasis and 5-year cancer death rates were 7% and 8%, respectively, in patients with the uninvolved fibromuscular stroma thickness above 3 mm, and 37% and 26%, respectively, in patients with the thickness below 3 mm. The thickness of cancer-unaffected cervical fibromuscular stroma seemed to be closely related to and to be a more useful parameter of the biological behavior of invasive cervical carcinoma than the depth of the cancer invasion. A threshold value of the minimum thickness of the tissue as a barrier against extrauterine spread of cervical cancer could not be identified in this study.

Adenocarcinoma↗

PubMed1987

In vivo selection of a low spin form of cytochrome P-448 from 3-methylcholanthrene-induced rat cytochrome P-450 isozymes by carbon tetrachloride.

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Animals↗

PubMed1987

Characterization of Lyt-2-, L3T4- class I-specific cytolytic clones in C3H-gld/gld mice. Implications for functions of accessory molecules and programmed development.

We report the first demonstration of Thy-1+, Lyt-2-, L3T4- MHC-specific CTL clones derived from the Lyt-2-, L3T4- subset of lymph node cells of C3H-gld/gld mice. These clones express alpha/beta heterodimeric TCRs on the cell surface and specifically recognize class I molecules on target cells. Lyt-2 and L3T4 molecules are therefore not essential for the induction, recognition, and killing of antigen-specific CTL. In addition, these studies suggest that antigen specificity development for class I structures may occur before Lyt-2 gene activation in the differentiation of T cells.

Animals↗

PubMed1987

Combination tumor-immunotherapy with recombinant tumor necrosis factor and recombinant interleukin 2 in mice.

Recombinant human tumor necrosis factor (r-TNF) inhibits growth of various mouse tumor cell lines both in vitro and in vivo. Treatment of established tumor nodules with intratumoral (i.t.) injection of r-TNF caused hemorrhagic necrosis of tumor and temporary disappearance of tumor mass. However, a small number of tumor cells remained and later formed fresh nodules. In striking contrast, combination therapy with r-TNF and recombinant human interleukin-2 (r-IL-2) resulted in a marked inhibition of regrowth of tumor cells. More than 60% of MBL-2-bearing mice were completely cured of tumor by treatment with r-TNF and r-IL-2. Cured mice could also reject rechallenged MBL-2 lymphoma cells, indicating the generation of anti-tumor effector cells in vivo. However, lymphocytes obtained from mice cured of MBL-2 showed no significant in vitro cytotoxic activity against MBL-2 lymphoma cells. In contrast, in vitro sensitization of spleen cells from cured mice with mitomycin-C-treated MBL-2 lymphoma cells resulted in the generation of cytotoxic cells against MBL-2 lymphoma cells. Moreover, spleen cells from mice cured of MBL-2 by treatment with r-TNF and r-IL-2 revealed a strong anti-tumor activity upon in vivo neutralization tests. These results strongly suggest that tumor-bearing mice can acquire systemic immunological memory after combination therapy with r-TNF and r-IL-2.

Animals↗

PubMed1987

Autophosphorylation of Ca2+/calmodulin-dependent protein kinase II. Effects on total and Ca2+-independent activities and kinetic parameters.

Incubation of purified rat brain Ca2+/calmodulin-dependent protein kinase II for 2 min in the presence of Ca2+, calmodulin (CaM), Mg2+, and ATP converted the kinase from a completely Ca2+-dependent kinase to a substantially Ca2+-independent form with little loss of total activity. Subsequent addition of EGTA to the autophosphorylation reaction enhanced further autophosphorylation of the kinase which was associated with a suppression of total kinase activity to the Ca2+-independent value. Protein phosphatase 1 rapidly increased the suppressed total activity back to the control value and slowly decreased the Ca2+-independent activity. Kinetic analysis showed that the kinase not previously autophosphorylated had a Km for the synthetic peptide syntide-2 of 7 microM and Vmax of 9.8 mumol/min/mg when assayed in the presence of Ca2+ and CaM. The partially Ca2+-independent species, assayed in the presence of EGTA, had a Km of 21 microM and Vmax of 6.0. In the presence of Ca2+ and CaM the Km decreased and the Vmax increased to approximately control nonphosphorylated values. The completely Ca2+-independent form generated by sequential autophosphorylation first in the presence of Ca2+ and then EGTA had similar kinetic parameters to the partially independent species when assayed in the presence of EGTA, but addition of Ca2+ and CaM (up to 1 mg/ml) had little effect. These results suggest that separate autophosphorylation sites in the Ca2+/CaM-dependent protein kinase II are associated with formation of Ca2+-independent activity and suppression of total activity.

Animals↗

PubMed1987

Relationship between age and plasma t-PA, PA-inhibitor, and PA activity.

A positive correlation between age and the occurrence of thrombosis has been suggested. We studied the relationship between age and fibrinolytic activities; namely levels of tissue plasminogen activator (t-PA) antigen, plasminogen activator inhibitor (PA inhibitor) activity, and plasminogen activator activity (PA activity). A dramatic increase in both t-PA antigen and PA inhibitor was shown in persons with increasing age. PA activity decreased with age. Therefore it is suggested that the tendency of decreased PA activity with increasing age may be related to the high incidence of thrombosis in older persons.

Adolescent↗

PubMed1987

Anesthesia for a patient with total lipodystrophy--a clinical report.

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Journal Article↗

PubMed1987

Amino acid analysis by high-performance liquid chromatography of a single stained protein band from a polyacrylamide gel.

A single stained band containing approximately 5 micrograms of protein was cut out of a polyacrylamide gel and subjected to hydrolysis together with the gel. The hydrolysate was subsequently analyzed for its amino acid content by high-performance liquid chromatography and postlabeling with o-phthalaldehyde. Bovine serum albumin, ribonuclease B, ovalbumin, pepsin, and chymotrypsinogen A were analyzed by this method, and their amino acid compositions were found to be in good agreement with the reported values. By this method, it is possible to quantitate 16 amino acids: Asx, Thr, Ser, Glx, Pro, Cys, Gly, Ala, Val, Ile, Leu, Tyr, Phe, His, Lys, and Arg. Thioglycolic acid is effective protection against the decomposition of Tyr, Cys, and Met; however, the recovery of Met is inconsistent. This method might be very helpful for the amino acid analysis of proteins of multicomponent systems, especially, those which can be resolved only by polyacrylamide gel electrophoresis.

Amino Acids↗

PubMed1987

Calcium . calmodulin-dependent protein kinase II and calcium . phospholipid-dependent protein kinase activities in rat tissues assayed with a synthetic peptide.

Rat tissue levels of Ca2+ . calmodulin-dependent protein kinase II (protein kinase II) and Ca2+ . phospholipid-dependent protein kinase (protein kinase C) were selectively assayed using the synthetic peptide syntide-2 as substrate. The sequence of syntide-2 (pro-leu-ala-arg-thr-leu-ser-val-ala-gly-leu-pro-gly-lys-lys) is homologous to phosphorylation site 2 in glycogen synthase. The relative Vmax/Km ratios of the known Ca2+-dependent protein kinases for syntide-2 were determined to be as follows: protein kinase II, 100; protein kinase C, 22; phosphorylase kinase, 2; myosin light chain kinase, 0.005. Levels of protein kinase II were highest in cerebrum (3.36 units/g tissue) and spleen (0.85 units/g) and lowest in testis (0.05 units/g) and kidney (0.04 units/g). Protein kinase II activity was localized predominantly in the 100,000g particulate fraction of cerebrum and testis, in the supernatant fraction of heart, liver, adrenal, and kidney, and about equally distributed between particulate and supernatant in spleen and lung. Likewise, protein kinase C activity was highest in cerebrum (0.56 units/g) and spleen (0.47 units/g), and the majority of activity was present in the cytosolic fraction for all tissues measured except for cerebrum and testis in which the kinase activity was equal in both fractions. Finally, the ratios of protein kinase II to protein kinase C were different in various rat tissues and between particulate and supernatant fractions. These results suggest somewhat different functions for these two Ca2+-regulated, multifunctional protein kinases.

Animals↗

PubMed1987

Inhibition of neuronal Cl-stimulated Mg-ATPase by alpha-human atrial natriuretic polypeptide.

Effects of alpha-human atrial natriuretic polypeptide (alpha-hANP) on neuronal membrane-bound adenosine triphosphatases (ATPases) (Na, K-ATPase, Cl-stimulated Mg-ATPase and anion-insensitive Mg-ATPase) were examined using rat brain microsomes. This peptide inhibited Cl-stimulated Mg-ATPase in a dose-dependent manner. The inhibition was non-competitive with respect to ATP, and was not affected by Cl-, the most potent stimulator of this enzyme. The activities of Na,K-ATPase and anion-insensitive Mg-ATPase were not changed by alpha-hANP at all.

Animals↗

PubMed1987

Destruction of cultured vascular endothelial cells and red blood cells by immune-activated polymorphonuclear leukocytes.

The cytotoxicity of human polymorphonuclear leukocytes (PMNs) against autologous red blood cells (RBC) and cultured vascular endothelial cells (EC) was investigated. PMNs were activated by phorbol myristate acetate (PMA) together with immune stimuli such as immune complexes and aggregated IgG. In the standard 51Cr-release assay, in which PMA concentration was 5 ng/ml and effector versus target ratio was 5, 76.7% and 34.2% specific 51Cr release was observed from RBC and EC, respectively. Significant levels of 51Cr were released, albeit to a lesser degree, when PMNs were stimulated by immune stimuli. Further experiments which employed various scavengers of oxygen radicals suggested that hydrogen peroxide was the most potent mediator of this cytotoxicity; the implications of these in vitro observations with the pathogenesis of immune vasculitis are of clinical interest.

Antigen-Antibody Complex↗

PubMed1987

Kinetics of ajmaline disposition and pharmacologic response in beagle dogs.

Pharmacokinetics and pharmacodynamics of ajmaline were studied in four healthy dogs after intravenous administration of the drug at the infusion rate of 1.0 mg/min for 45 min. Ajmaline exhibited a saturable binding to plasma protein. One kind of binding site was found in the range of observed drug concentrations and its binding capacity showed nearly threefold interindividual difference. The time course of ajmaline concentration in whole blood Cb could be described by the two-compartment open model and the unbound concentration of ajmaline in plasma Pf was estimated from Cb by using the hematocrit value and the parameters of plasma protein binding and erythrocyte partitioning. The pharmacologic responses to ajmaline were assessed by recording ECG, and the changes in PQ and QRS interval were studied in relation to ajmaline disposition. When ECG changes were related to the ajmaline concentration, a significant degree of hysteresis was observed. The relationship between the unbound drug concentration and the pharmacologic effect was analyzed by a combined pharmacokinetic-pharmacodynamic model, where the hypothetical effect compartment is connected to the Pf in the central compartment by a first-order process. This model allows estimation of the changes in PQ and QRS intervals after intravenous administration of ajmaline. By comparing the drug effect on PQ and QRS intervals, it was suggested that ajmaline distributes to the atrial and the ventricular tissue in a similar degree and causes a reduction in the conduction rate in both sites with similar activity.

Ajmaline↗

PubMed1987

Monoclonal antibodies against human scirrhous carcinoma of the stomach.

We established three monoclonal antibodies (MoAbs) S43, S74 and S202 which reacted strongly with scirrhous carcinomas of the stomach. MoAb S43 reacted with culture supernatants of the scirrhous gastric cancer cell line MK-01, and thus may be useful for in vivo detection of markers for this tumor. MoAb S74 reacted with the majority of gastric cancer tissues, but showed no reaction with normal tissues. It may be useful as a drug carrier in targeting chemotherapy against scirrhous gastric cancer, because of its tumor specificity. MoAb S202 reacted strongly with 100% of scirrhous gastric cancer samples and with the culture supernatants of colonic and pancreatic cancers.

Adenocarcinoma, Scirrhous↗