Glucose does not influence the insulin-like growth factor (IGF) binding to carrier proteins (IGFBPs): Analysis of rat and human serum by western ligand blotting

The insulin-like growth factors (IGFs) circulate bound to specific proteins (termed IGFBP-1 through IGFBP-6) that modulate IGF bioactivity in tissues. The aim of this study was to analyse the effects of glucose on IGF binding to IGFBPs in rat and human serum by means of western ligand blotting. Serum samples were incubated with increasing concentrations of glucose (0 to 50 mmol/l), and EDTA (25 mmol/l) was added to inhibit protease activity. To analyse the effect of glucose on protection of IGFBPs from protease activity, serum from pregnant women (reported to be very rich in proteolytic activity against IGFBPs) was added to rat serum previously incubated with glucose. Glucose did not affect the¹²⁵I-IGF binding to rat and human serum IGFBPs. The intensity of IGFBP-3 bands decreased considerably during the incubation. This appeared to be due to endogenous protease activity, since the decrease was blocked by addition of EDTA. The incubationi of rat serum with pregnant human serum produced a marked attenuation of IGFBP-3 and disappearance of IGFBP-4 bands. In conclusion, our study shows that glucose does not influence the IGF binding to IGFBP-3 either in rat or in human serum, confirms the presence of endogenous proteolytic activity in normal non-pregnant serum, and demonstrates that glucose has no protective action against protease activity.     

Superoxide dismutase activity in rabbit reticulocytes

Summary Reticulocytosis was induced in rabbits by bleeding anemia and erythrocyte superoxide dismutase activity was determined. Reticulocytes were found to contain about 1.3 times as much activity as mature erythrocytes.This work was supported in part by a Grant-in-Aid from the Japan Medical Research Foundation and a Grant-in-Aid (Project No. 56570363) from the Ministry of Education, Science and Culture of Japan.     

Antidiuretic activities of substance P and its analogs

Summary Substance P (SP) and analogs, including 5 nucleoside (ARA or HRA)-peptides, were examined for antidiuretic activity in ethanolized rats. The activity was potent in the analogs embodying the C-terminal hexapeptide, weak in the nucleoside-pentapeptide, and negligible in the nucleoside-tetrapeptide. In addition, the activity was increased by acylation of the hexapeptide. The antidiuretic potencies were also compared with the hypotensive potencies.This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture, Japan, and by a grant from the Imanaga Foundation, Nagoya, Japan.     

Inducement of blood vessel formation by ovarian extracts from mice injected with gonadotropins

Summary Ovarian extracts prepared from immature mice injected with human chorionic gonadotropin (hCG) and pregnant mare serum gonadotropin (PMSG) were assayed for angiogenic activity. The assay method consisted of implanting a film coated with ovarian extracts to the lateral wall of the m.rectus abdominus of a mouse for 20 days and examining the site for vascularization. The higher angiogenic activity obtained with PMSG-treated extract may be related to its follicle stimulating activity.Acknowledgment. Part of this work was supported by a grant from The Ministry of Education, Japan (No. 576165).     

Immunochemical identity of dipeptidyl aminopeptidase IV from pig serum,liver, submaxillary gland and kidney

Summary The enzymes which were extracted by autodigestion from the microsomal fractions of the pig kidney, liver and submaxillary gland and from the serum showed an immunochemical identity by a double immunodiffusion test. But the kidney enzyme had a different pI-value from the pI-values of the enzymes of other organs.This investigation was supported by a grant from the Ministry of Education, Japan to K. M. F. We would like to thank Dr Toshiharu Nagatsu (Department of Life Chemistry, Graduate School at Nagatsuta, Tokyo Institute of Technology, Yokohama, Japan) for helpful suggestions, Mr Moritoshi Sato (Matsumoto Meat Inspection Station, Matsumoto, Japan) for their generous supply of pig organs and Ajinomoto Co. Inc., Tokyo, Japan, for the gift of Gly-Pro-p-nitroanilide tosylate. Technical assistance of Miss K. Yanagisawa is gratefully acknowledged.     

Effect of fluoride administration on renal glucose-6-phosphatase activity in rats

Summary Renal glucose-6-phosphatase activity was found to be significantly elevated by fluoride administration (NaF 35 mg/kg, i.p.). The elevation of the enzyme activity was markedly suppressed by adrenalectomy.This work was supported in part by the Scientific Research Foundation (Grant 7014-267337) from the Education and Culture Ministry of Japan.     

Morphogenesis of human colon cancer cells with fetal rat mesenchymes in organ culture

Summary The morphogenesis and cytodifferentiation of human colon cancer cells (LS174T and HT29) were examined by combining cancer cells with fetal rat digestive-tract mesenchyme in organ culture. LS174T cells migrated into the mesenchyme to form glandular structures composed of single columnar cells with their nuclei oriented basally, while HT29 cells formed cell masses with little lumen formation. Immunohistochemical studies with antibodies against carcinoembryonic antigen and secretory components showed that the composition of cell surface glycoproteins was not necessarily reversed to the normal type, even when neoplastic cells exhibited normal glandular structures.This work was supported by grants-in-aid for Cancer Research from the Ministry of Education, Science, and Culture, Japan, and by the Veterans Administration Medical Research Service, USA. Y.S. Kim is the recipient of a Medical Investigator Award of the Veterans Administration.     

Effects of 2-deoxy-D-glucose,glucose and insulin on efferent activity in gastric vagus nerve

Summary Intracarotid injection of 2-deoxy-D-glucose and insulin increased the efferent activity in the gastric vagus nerve of anesthetized rats, while glucose injection transiently decreased vagus activity.This work was supported by research grant No. 212105 from The Ministry of Education, Science and Culture of Japan.     

A Laser-Doppler-Velocimeter using an optical fiber and its application to local velocity measurement in the coronary artery

Summary A Laser-Doppler-Velocimeter with an optical fiber has been developed to measure arterial blood velocity accurately in a small sample volume. After fundamental experiments to evaluate the accuracy of the present method, blood flow velocity was measured in canine coronary arteries.Acknowledgment. We thank Dr J. Koyama, Vice President of the Institute of Electronics and Communication Engineering of Japan, for his keen interest and continuous encouragement throughout the work. Thanks are also due to Dr H. Kita for English revision, and Dr K. Mito and Mr O. Hiramatsu for their excellent technical assistance. This investigation was supported by the Grants from the Ministry of Education of Japan (No. 521125 and No. 337049 and No. 587008).     

Superoxide anion scavenging effect and superoxide dismutase activity of Ginkgo biloba extract

Ginkgo biloba extract is known to be efficient in diseases associated with free radical generation. The purpose of this work was to study, under in vitro conditions, the action of Ginkgo biloba extract (Gbe) against superoxide anion (O2-.), which is directly or indirectly implicated in cell damage. Gbe appears to have both an O2-. scavenging effect and also a superoxide dismutase activity. Its antiradical effect was demonstrated by low temperature electron spin resonance and in a non-enzymatic system (phenazine methosulfate-NADH), and its enzymatic activity was shown by polarographic determination.     

Development of the hypothalamic-hypophysial-gonadotrophic activities in fetal rats

Summary Pituitary responsiveness to LHRH and anti-LHRH serum was investigated in fetal rats aged 18.5–22.5 days. Synthetic LHRH injection in utero into fetuses brought about a remarkable depletion of pituitary-LH with a corresponding increase of serum-LH on day 18.5. On the contrary, anti-LHRH serum administration to day-20.5 fetuses caused a significant augmentation of pituitary-LH 1 day later. These data indicate that LH-gonadotrophs respond to LHRH even on day 18.5, and that endogenous LHRH begins to affect LH-gonadotrophs on day 20.5.This work was supported by a grant from Ministry of Education, Science and Culture, Japan. The exert technical assistance of Mr Yoshihito Okamura is gratefully acknowledged.     

The growth responses of hamster chondrocytes,dermal fibroblasts and embryo cells to whole blood serum and plasma-derived serum

Summary Autoradiographic studies with³H-thymidine demonstrated that the growth responses of hamster chondrocytes, dermal fibroblasts and embryo cells, respectively, differed in media containing whole blood serum (WBS) and plasmaderived serum (PDS). Dermal fibroblasts seemed to require a growth factor from platelets for growth, but chondrocytes did not. Embryo cells showed an intermediate pattern of growth response to this factor.This work was supported by a Grant-in-Aid for Cancer Research from the Ministry of Education, Science and Culture and the Ministry of Health and Welfare of Japan.We thank Miss M. Tanaka and Miss K. Kawana for technical assistance.     

Increased activity of dipeptidyl peptidase IV in serum of hepatoma-bearing rats coincides with the loss of the enzyme from the hepatoma plasma membrane

The specific activity of dipeptidyl peptidase IV (DPP IV E.C. 3.4.14.-) in the plasma membrane of Morris hepatoma 9121 or hepatoma 7777 was 3.5% and 2.9%, respectively, of that in the plasma membrane of rat liver. The enzyme activity in the serum of hepatoma-bearing rats was 141% (hepatoma 91219) and 162% (hepatoma 7777) of the normal value. Cytochemical investigation showed that the DPP IV activity was almost completely absent from the hepatoma cell plasma membrane and was not sequestered within these cells. Indirect immunofluorescence staining with a polyclonal antibody directed against DPP IV indicated that the loss of activity was due to the absence of DPP IV molecules in the plasma membrane. The possibility that the enzyme is transferred from the membrane into the serum as a result of structural alterations is discussed.     

Increase in activity of natural killer cells (NK cells) by serum thymic factor]

The serum thymic factor (FTS) and one of its structural analogues have been administered to adult normal Mice for 2 to 12 weeks. This treatment induced a significant increase in spleen cell natural killer (NK) activity, evaluated by a cytotoxicity assay against YAC cells.     

Increased activity of dipeptidyl peptidase IV in serum of hepatoma-bearing rats coincides with the loss of the enzyme from the hepatoma plasma membrane

Summary The specific activity of dipeptidyl peptidase IV (DPPIV E.C. 3.4.14.-) in the plasma membrane of Morris hepatoma 9121 or hepatoma 7777 was 3.5% and 2.9%, respectively, of that in the plasma membrane of rat liver. The enzyme activity in the serum of hepatoma-bearing rats was 141% (hepatoma 91219) and 162% (hepatoma 7777) of the normal value. cytochemical investigation showed that the DPP IV activity was almost completely absent from the hepatoma cell plasma membrane and was not sequestered within these cells. Indirect immunofluorescence staining with a polyclonal antibody directed against DPP IV indicated that the loss of activity was due to the absence of DPP IV molecules in the plasma membrane. The possibility that the enzyme is transferred from the membrane into the serum as a result of structural alterations is discussed.     

Appearance of sperm activation factors in the ovary of the sea urchinHemicentrotus pulcherrimus with maturation

Summary Ovaries of the sea urchinHemicentrotus pulcherrimus were extracted with 70% ethanol. The extracts were polarographically assayed for respiration-stimulating activity toward homologous spermatozoa. The data of the bioassay and light microscopic observation on thin sections of the ovaries showed detectable activity only in mature ovaries.We thank the Misaki Marine Biological Station for kindly supplying us with specimens. This work was supported in part by a grant from the Ministry of Education, Science and Culture of Japan.     

A basic protein from bovine brain that co-precipitates with tubulin in vitro

A 193 kDa protein consisting of 58 kDa subunits, which has pI values of 8.50 and 8.65, was purified from bovine brain cytosol. It formed heavy precipitates with tubulin, and the molar ratio of tubulin dimer to this protein in the precipitate was 3.2. In contrast to microtubules containing ordinary microtubule-associated proteins, these complexes remained stable against cold and 1 mM CaCl₂.Acknowledgments. This work was supported in part by a Grant-in-Aid for Scientific Research from the Ministry of Education, Science and Culture of Japan.     

A microanalysis of colloid osmotic pressure

Summary A needle type of colloid osmometer has been devised with special attachments which made it useable with 3-l samples. It has been used in oncometry of fresh water teleosts to give the following results: 5.1±1.0 mm Hg for goldfish, 5.2±0.71 mm Hg for carp, and 14.3±0.75 mm Hg for eel.This study was supported in part by Research Grant 387036 from the Ministry of Education of Japan.invited to National Institute of Physiological Sciences, Okazaki, Japan.     

Low-molecular-weight enzyme inhibitors suppress the development of experimental allergic encephalomyelitis

Summary We tested the activity of low-molecular-weight enzyme inhibitors with immunomodifying actions on the suppression of experimental allergic encephalomyelitis (EAE). Of the agents tested the inhibitors of alkaline phosphatase, aminopeptidase B and esterase gave significant protection againts the clinical expression of EAE in guinea pigs.Acknowledgments. This work was partly supported by a grant-in-aid for New Drug Development Research from the Ministry of Health and Welfare, Japan.     

Immunization of female rabbits against testosterone stimulates testosterone accumulation by isolated ovarian follicles

Summary Ovarian follicles isolated from female rabbits after active immunization against testosterone-3-oxime bovine serum albumin produced more testosterone than similar follicles from controls.This work was supported by the Medical Research Council of Canada MT 4192.