MINAKUCHI, J. and YATA, J. The Lymphocyte Subpopulations Involved inCytotoxicity Generated by Co-Culture with Autologous and Allogeneic Epstein-BarrVirus-Transformed Cell Line. Tohoku J. exp. Med., 1988, 155 (2), 107-115 -When peripheral blood lymphocytes from healthy adults are cultured with autologous (auto) or allogeneic (allo) Epstein-Barr virus-transformed cells (LCL), non-specific killer activity against NK-sensitive K562 and NK-resistant Raji, as well as specific killer activity against LCL is enhanced or generated. We analyzed the cell subsets possessing such cytotoxicity using monoclonal antibodies (MoAb). OKT3, a MoAb to T cell receptor-associated molecule, added in the effector phase suppressed the killer activity against LCL but not against Raji or K562. In contrast, OKT3 added in the induction phase abolished the generation of cytotoxicity against all targets. The addition of OKT8 in either the effector or induction phase inhibited anti-LCL killing induced by stimulation with alloLCL. This suggests that CD8 is required for recognition of alloLCL. The treatment of effector cells with MoAb and complement(C) revealed that killers against LCL were OKT8+ Leu11-, and those against K562 were OKT8- Leu11+ When autoLCL were used as stimulator, removal of OKT4+ cells in the induction phase diminished the cytotoxicity against all targets, indicating that CD4+ T cells recognize autoLCL. Elimination of CD8+ cells from responder did not decrease the generation of killer activity. Further experiments suggested that this was caused by the coexistence of CD4+ killer cells or by the increase of residual CD8+ effector cells.
FUKUSATO, T., MOHAMAD, A., GERBER, M.A, and THUNG, S.N. SynergisticEffect of 5-Azacytidine and γ-Interferon or Dimethyl Sulfoxide on Expression ofHLA Class I Antigens by PLC-PRF-5 Cells. Tohoku J. exp. Med., 1988, 155 (2), 117-128 - In order to elucidate the mechanism by which HLA antigens expression is induced or enhanced on the injuried or transformed hepatocytes, we have made in vitro studies using human hepatic tumor-derived cell lines as a model system. In the present study, PLC-PRF-5 cells that have the integrated form of hepatitis B virus genome in DNA were treated with 5-azacytidine (5-azaC) in combination with γ-interferon (IFN-γ) or dimethyl sulfoxide (DMSO). HLA antigens on the cell surface were quantitated by using a modified cell-ELISA method. As a result, it was demonstrated that DMSO- or IFN-γ-treatment enhanced expression of HLA class I antigens on the cell surface. In addition, enhanced expression of the antigens on PLC-PRF-5 cells treated with 5-azaC in combination with IFN-γ or DMSO represented a synergistic effect of these inducers on HLA class I antigens expression although no changes in HLA antigens expression were induced after 5-azaC-treatment alone in short-term experiments. Furthermore, an indirect immunofluorescent analysis of hepatitis B surface antigen on the cells demonstrated increased expression of the antigen after 5-azaC-treatment alone. HLA class II antigens and hepatitis B core antigen were not induced even after those treatments and also not after a long-term experiment. These results might indicate possible modulation of HLA class I and hepatitis B virus antigens expression on the cultured cells by a DNA hypomethylating agent, 5-azaC.
TORIYABE, S., MIURA, Y., KIMURA, S., MEGURO, Y., SUGAWARA, T., NOSHIRO, T., TAKAHASHI, M., OHASHI, H., SANO, N., WATANABE, H. and YOSHINAGA, K. APlasma Inhibitor of Sodium and Potassium Activated Adenosine Triphosphatase inPatients with Essential Hypertension. Tohoku J. exp. Med., 1988, 155 (2), 129-137 - The purpose of this study is to evaluate the plasma Na, K-ATPase inhibitor (NKI) in patients with essential hypertension and to compare the mode of its biochemical actions on the Na, K-ATPase with that of ouabain. Plasma NKI was extracted through a reversed-phase cartridge column and its inhibitory action on hog brain Na, K-ATPase was measured in vitro. Plasma NKI activity was significantly greater in patients with essential hypertension (44±2.8% (S.E.), n=28, p<0.01) than in normotensive controls (25±2.4%, n=21). No significant correlation was demonstrated between the values of plasma NKI and mean arterial pressure in either group. Both plasma NKI and ouabain showed a dose-dependent inhibition on the Na, K-ATPase reaction. An action of ouabain was competitively antagonized by increased concentration of potassium in the reaction mixture, while plasma NKI showed a constant inhibition on the Na, K-ATPase independently of potassium concentrations. The action of plasma NKI was of rapid onset and linear with time, while ouabain showed a delayed onset of the reaction over 30sec, followed by a progressively increasing inhibition on the enzyme reaction. Finally, the inhibitory action of plasma NKI on Na, K-ATPase was completely abolished in the presence of bovine serum albumin even at the concentration of 500μg/ml in the reaction mixture, which did not have any influence on the actions of ouabain. To sum up, the results showed a markedly different nature of plasma NKI from ouabain in the mode of biochemical actions on the Na, K-ATPase in vitro. This study may also raise a question whether plasma free NKI, supposedly an active from of NKI, is actually working as a physiological regulator in vivo. It seems thus premature to assume it as a pathogenic factor in essential hypertension.
AKITA, H., KOBAYASHI, Y. and KAGAYAMA, M. A Histochemical Study onLectin Binding in the Immature Enamel and Secretory Ameloblasts of RatIncisors. Tohoku J. exp. Med., 1988, 155 (2), 139-149 - Four lectin-fluorescent marker conjugates (Con A-F, MPA-F, PNA-F, WGA-R) were used for visualizing their binding sites on tissue sections by fluorescent microscopy. The immature enamel, Tomes' processes and distal cytoplasm of ameloblasts were stained with MPA-F and WGA-R on paraformaldehyde-fixed and paraffin-embedded tissue sections. The MPA- and WGA-binding glycoconjugates seemed to be main components of the organic enamel matrix because of the intense fluorescence from the tissue sections. The central portion of distal cytoplasm of ameloblasts was stained with PNA-F but the immature enamel and Tomes' processes were not. This suggests the PNA-binding glycoconjugates to be intermediates during the process of modifying their oligosaccharide chains or to be elements of Golgi area. Tomes' processes, the distal cytoplasm and periphery of nuclei of ameloblasts were stained with Con A-F. The Con A-binding glycoconjugates except those localizing on Tomes' processes may also be intermediates and/or intracellular elements like the PNA-binding glycoconjugates. Cells of stratum intermedium were stained with Con A-F, MPA-F and WGA-R. Cells of stellate reticulum and outer enamel epithelium were stained with Con A-F and WGA-R. It was indicated that enamel proteins, major organic components of immature enamel, are highly possible candidates for the MPA- and WGA-binding glycoconjugates.
SATOH, J., SHINTANI, S., NOBUNAGA, T., SUGAWARA, S., MAKINO, S., TOYOTA, T. and GOTO, Y. NOD Mice with High Incidence of Type 1 Diabetes are not TLymphocytopenic. Tohoku J. exp. Med., 1988, 155 (2), 151-158 - An autoimmune pathogenesis has been indicated in insulin-dependent (type 1) diabetes mellitus (IDDM). Previously we reported that non-obese diabetic (NOD) mice as an animal model of spontaneously developing IDDM were immunologically characterized by T lymphocytopenia and impaired cellular immunities. The cumulative incidence of diabetes in the T lymphocytopenic female NOD mice was 10-20% by 24 weeks of age. On the other hand, the incidence of diabetes are 80-90% in the female NOD/Shi-Sendai (S), in whom proportion of lymophocyte subsets has not been known yet. Therefore, we examined the spleen cells of female NOD/ Shi-S and female NOD/Shi with high incidence of diabetes, and of female Jcl: ICR as a control. Cell numbers, populations of T cells (Thy 1.2+, Lyt-1+ and Lyt-2+), B cells (surface-Ig+), NK cells (acialo GM1+) and responsiveness to Concanavalin A were analyzed as immunological parameters. In contrast to the T lymphocytopenic NOD, these immunological parameters were not impaired in the NOD/Shi-S and NOD/Shi in comparison to those of Jcl: ICR. The results indicate that there may be a positive association between the incidence of diabetes and T cell number and functions in female NOD mice.
KOIZUMI, F., KAWAMURA, T., ISHIMORI, A., EBINA, H. and SATOH, M. PlasmaParacetamol Concentrations Measured by Fluorescence Polarization Immunoassayand Gastric Emptying Time. Tohoku J. exp. Med., 1988, 155 (2), 159-164 -Radionuclide labeled gastric emptying was compared with plasma paracetamol (acetaminophen) concentration measured by fluorescence polarization immunoassay. A significant correlation was found between both gastric emptying half-time of radionuclide (t1/2) and % isotope remained in the region of the stomach and paracetamol absorption kinetic parameters. The strongest correlation was found between t1/2 and the time required to reach the peak plasma level of paracetamol (r=0.680, p<0.01). This study indicates that simple paracetamol marker method has enabled an indirect quantitative measurement of gastric emptying by sampling at varying time intervals.
MIYASAKA, K., NAKAMURA, R., FUNAKOSHI, A. and KITANI, K. InhibitoryEffect of CR-1409, a Competitive Inhibitor of Cholecystokinin, on PancreaticExocrine Secretion in the Conscious Rat. Tohoku J. exp. Med., 1988, 155 (2), 165-172 - The inhibitory effect of CR-1409, a new glutaramic acid derivative and a competitive inhibitor for cholecystokinin (CCK), on the basal and CCK-stimulated pancreatic secretion was examined in the conscious rat in vivo. Rats were prepared with cannulae draining pure bile and pancreatic juice separately and with a duodenal cannula and right and left jugular vein cannulae. Plasma CCK level increased to 3.65±0.79 and 19.9±4.47pM (mean±S.E.) by a 2-hr infusion of 100 and 300pmole/kg/hr of CCK-octapeptide (CCK-8), respectively. Simultaneous infusion of 170nmole/kg/hr of CR-1409 completely abolished pancreatic responses to 100pmole/kg/hr of CCK-8. Infusion of CR-1409 at rate of 57nmole/ kg/hr slightly but significantly inhibited CCK-8 (100pmole/kg/hr)-stimulated secretion. Pancreatic responses to 300pmole/kg/hr of CCK-8 were partially inhibited but not completely abolished by the 170nmole/kg/hr of CR-1409. Neither the basal pancreatic secretion nor the bile secretion was affected by CR-1409. We conclude that CR-1409 inhibited CCK-stimulated pancreatic secretion in vivo.
HATA, K., KIKUCHI, K., TADA, K. and TSUIKI, S. Analysis of Glucose-6-Phosphate Translocase and Hexose-6-Phosphate Phosphohydrolase, the Two ObligatoryComponents of Microsomal Glucose-6-Phosphatase System, in Rat Liver. Tohoku J. exp. Med., 1988, 155 (2), 173-181-A membrane filter procedure developed by Igarashi et al. (1984) for the measurement of glucose 6-phosphate uptake by the microsomes has been demonstrated to be a good method for assaying glucose-6-phosphate translocase, an obligatory component of the microsomal glucose-6-phosphatase system. When glucose-6-phosphate translocase was assayed in developing and diabetic rat livers independently of hexose-6-phosphate phosphohydrolase, another obligatory component of the glucose-6-phosphatase system, the two activities were found to undergo alterations, whose profiles, however, were quite distinct from each other. The profile of the microsomal glucose-6-phosphatase activity resembles the profile of the phosphohydrolase activity rather than that of the translocase activity, suggesting that the phosphohydrolase may be rate-limiting at least under these conditions. AH-109A, a strain of transplantable rat ascites hepatoma, was found to lack both glucose-6-phosphate translocase and hexose-6-phosphate phosphohydrolase activities.
LIU, Y.-T., JIN, C., CHEN, Z., CAI, S.-X., YIN, S.-N., LI, G.-L., WATANABE, T., NAKATSUKA, H., SEIJI, K., INOUE, O., KAWAI, T., UKAI, H. and IKEDA, M. Increased Subjective Symptom Prevalence among Workers Exposed to Trichloroethyleneat Sub-OEL Levels. Tohoku J. exp. Med., 1988, 155(2), 183-195 - Over 100 workers exposed to trichloroethylene (TRI) mostly at less than 50ppm during the production or vapor degreasing operation and about an equal number of the non-exposed control workers were examined for subjective symptoms, hematology, serum biochemistry, and sugar, protein and occult blood in urine. Essentially all the clinico-laboratory tests stayed normal, and there was no significant difference in the findings between the exposed and the controls. Thus, no clinically significant effects of TRI exposure were found in the blood and liver functions among the exposed workers as compared with the controls. The prevalence of the subjective symptoms was, however, significantly higher in the exposed group than in the controls, and dose-response relationship could be established in some selected symptoms such as nausea, heavy feeling in the head, forgetfulness, tremor in extremities, cramp in extremities and dry mouth, although the exposure was low. The findings warrant further attention to the effects of TRI especially on the central nervous system at the concentration lower than, e.g., 50ppm.
OHNEDA, A. and OHNEDA, M. Effect of Glicentin-Related Peptides upon theSecretion of Insulin and Glucagon in the Canine Pancreas. Tohoku J. exp. Med., 1988, 155 (2), 197-204 - In order to clarify responses of the endocrine pancreas to glicentin, four glicentin-related peptides were investigated in a local circulation preparation of the canine pancreas. These peptides were administered in a dosage of 200pmole for 10min into the pancreaticoduodenal artery under the continuous infusion of 0.5% arginine solution. In a group of six dogs, the administration of glicentin-related pancreatic peptide (GRPP) and glicentin 1-16 resulted in an increase in plasma insulin (IRI) and a decrease in plasma glucagon (IRG). In the other group of six dogs, the administration of glicentin 62-69 induced are increase in plasma IRI and a decrease in plasma IRG. Following the successive infusion of oxyntomodulin, both plasma IRI and IRG increased slightly. Porcine glucagon administered at the end of each experiment exerted a rise in blood glucose and plasma IRI in addition to an increase in plasma IRG. In comparison of the maximal responses of plasma IRI and IRG to these glicentin-related peptides, the administration of glicentin 1-16, 62-69 or GRPP elicited an increase in plasma IRI and a decrease in plasma IRG. In contrast, oxyntomodulin and glucagon increased both plasma IRI and IRG. The present study indicates that glicentinrelated peptides, both the N- and C-terminal portions, affect the endocrine function of the pancreas and suggests that glicentin released by nutrient ingestion plays an important role in the enteroinsular axis.
OSUMI, S., MORISHITA, S., WADA, K., USUI, H., KANDA, M., MATSUI, H. and KAKEYA, N. Antihypertensive Effect of NKY-722, a New Water-Soluble 1, 4-Dihydropyridine Derivative, on Conscious Spontaneously Hypertensive Rats. Tohoku J. exp. Med., 1988, 155 (2), 205-206 - 3-(4-Allyl-1-piperazinyl)-2, 2- dimethylpropyl methyl 1, 4-dihydro-2, 6-dimethyl-4-(3-nitrophenyl)-3, 5-pyridine dicarboxylate dihydrochloride (NKY- 722) produced a dose-dependent antihypertensive effect in conscious spontaneously hypertensive rats (SHR). In this respect NKY-722 was more potent and longer-acting than nicardipine. In canine isolated mesenteric arteries, exposed to a Ca2+-free medium containing high K+, NKY-722 inhibited Ca2+-induced contraction in concentration-dependent manner, suggesting the calcium antagonism as the mechanism of action.
YAMASHITA., M., INABA, T., KAWASE, Y., HORII, H., WAKITA, K., FUJII, R. and NAKAHASHI, H. Quantitative Measurement of Renal Function Using Ga-68-EDTA. Tohoku J. exp. Med., 1988, 155 (2), 207-208 - We studied renal function using a positron emission tomography with C15O gas and 68Ga-EDTA in 6 normal subjects (male, 21-77 years old). The blood volume (ml/100g (kidney)) was 19.2±1.8 for the one 21 years old subject and 11.8±2.0 for the rest (65-77 years old). The glomerular filtration rate (ml/min/100g (kidney)) was 57.8±1.1 for the 21 years old subject and 30.4±2.8 for the rest.
TOMITA, Y., MAEDA, K. and TAGAMI, H. Stimulatory Effect of Histamine onNormal Human Melanocytes in Vitro. Tohoku J. exp. Med., 1988, 155 (2), 209- 210 - Normal human epidermal melanocytes became swollen and more dendritec and the immunoreactive tyrosinase increased markedly when they were cultured for 2 days with 5μM of histamine. These results suggest that high dermal concentrations of histamine may be responsible for the induction of skin pigmentary changes associated with local proliferation of mast cells such as in urticaria pigmentosa and systemic mastocytosis.
OKUDA, T. and TACHIBANA, T. Involvement of Mac-1 Antigen in Tumor CellKilling by Macrophages. Tohoku J. exp. Med., 1988, 155 (2), 211-212 - Some tumor cells such as YAC-1 and RL _??_1 were killed by mouse peritoneal macrophages elicited with OK-432 one day before the harvest. The killing reactions were inhibited specifically by anti-Mac-1. The results suggest that some macrophages generated early after the injection of OK-432 kill tumor cells with the aid of CR3.