Additive stimulation of hepatic putrescine production by glucagon and insulin after partial hepatectomy in rats

When rats received glucagon or insulin every 2 h after partial hepatectomy (Hx), hepatic putrescine content was increased above control levels at 6 and 12 h, respectively. When the two hormones were combined, the increased levels were additive. Hepatic ornithine decarboxylase activity was above control levels at 12 h after insulin treatment. Hepatic spermidine N1-acetyltransferase activity was enhanced at 6 h only when glucagon was dosed. Putrescine administration from 0 to 4 h or from 6 to 10 h increased hepatic DNA synthesis to similar levels 22 h after Hx. These results suggest that glucagon and insulin additively stimulate hepatic putrescine production after Hx. This may explain the cooperative stimulation of liver regeneration by both hormones.     

Quantitative analysis of development of mitochondrial ultrastructure in differentiating mouse hepatocytes during postnatal period

Between birth and 10 days of age, the volume density (volume/unit cytoplasmic volume) of the matrix, and the surface density (area/unit cytoplasmic volume) of the inner membrane and cristae increased in both periportal and perihepatic hepatocytes, and did not differ significantly between the cells of the two zones. After 10 days of age, however, the volume density of the matrix decreased in perihepatic cells and remained unchanged in periportal cells, and, therefore, it became greater in periportal cells than in perihepatic cells in 20-day-old and adult animals. The surface density of the inner membrane and cristae decreased in the cells of both zones. Further, the hepatocyte volume increased markedly, especially in perihepatic zones between 20 days of age and the adult. The results show that, in postnatally differentiating hepatocytes, mitochondria are likely to develop during early postnatal period, then the structural heterogeneity of mitochondria arises, and hepatocyte volume increases markedly during late postnatal period after weaning. Thus, the process of postnatal hepatocyte differentiation includes such several phases of development.     

Microinjection of the monoclonal anti-tubulin antibody YL1/2 inhibits cleavage of sand dollar eggs

Two monoclonal antibodies against alpha-tubulin (YL1/2 and D2D6) were microinjected into the egg of the sand dollar Clypeaster japonicus, and their effects on cleavage of the egg were investigated. They had already been shown by immunoblotting to react specifically with egg tubulin and by immunofluorescence to stain the mitotic apparatus [OKA et al., (1990). Cell Motil. Cytoskel. 16:239-250]. Injection of YL1/2 prevented chromosome movement and cleavage, although the cleavage furrow developed in some cases. In all eggs injected at prometaphase, metaphase, or anaphase, the birefringence of the mitotic apparatus disappeared immediately after injection. Injection of D2D6 had no significant effect on mitosis or cleavage of whole eggs injected after nuclear disappearance, although it prevented the disappearance of the nuclear envelope in 54% of the eggs injected before the disappearance. FITC-conjugated D2D6 did not accumulate in the spindle when injected into the dividing sand dollar egg. These results indicate that YL1/2 disassembled microtubules, whereas D2D6 did not bind to microtubules in the living cell.     

Deletion of C-terminal 12 amino acids of GLUT1 protein does not abolish the transport activity.

We engineered the GLUT1 cDNA to delete C-terminal 12 amino acids of encoded GLUT1 protein. This mutated GLUT1 protein expressed in CHO cells by transfection of its cDNA was demonstrated to reside on the plasma membrane by cell surface labeling technique, and retain the transport activity, similar to that of the wild-type GLUT1. In addition, metabolic labeling of the intact cells with 35S indicated that the half-life of the mutated GLUT1 was not significantly different from that of the wild-type GLUT1. These results suggest that C-terminal 12 amino acids of GLUT1 are not important for the transport activity and the stability of the protein. Taken together with our previous results on the mutant without C-terminal 37 amino acids, the amino acids between the 37th and the 13th from the C-terminus appear to be essential for the transport activity.     

Protein Kinases Are Involved in Prolonged Acetylcholine Release from Rat Hippocampus Induced by Thyrotropin-Releasing Hormone Analogue NS-3

Abstract: The effects of various protein kinase inhibitors on acetylcholine release from the rat hippocampus induced by the local application of NS-3 (montirelin hydrate, CG-3703), a thyrotropin-releasing hormone analogue, into the medial septum-diagonal band were examined using in vivo microdialysis. Perfusion of NS-3 (1 µ M ) into the medial septum-diagonal band for 20 min produced a pronounced and prolonged increase in the hippocampal acetylcholine efflux. Pretreatment of the medial septum-diagonal band with either K-252a, a nonselective protein kinase inhibitor, or selective protein kinase A inhibitor H-89 almost completely blocked the acetylcholine efflux evoked by NS-3, and selective protein kinase C inhibitor calphostin C inhibited the action of NS-3. On the other hand, NS-3 (0.1–10 µ M ) or TRH (1–100 µ M ) increased the cyclic AMP efflux from the medial septum-diagonal band in a concentration-dependent manner, as measured by microdialysis. These findings suggest that protein kinases A and C in the neurons of the medial septum-diagonal band are involved in the mechanism of the prolonged stimulation of acetylcholine release from the hippocampus induced by thyrotropin-releasing hormone and its analogue, NS-3.     

A unique electrophoretic slow-moving glucose 6-phosphate dehydrogenase variant (G6PD Asahikawa) with a markedly acidic pH optimum

Summary A new glucose 6-phosphate dehydrogenase (G6PD) variant associated with chronic nonspherocytic hemolytic anemia was discovered in Japan. The patient showed hemolytic crises after upper respiratory infections. The enzyme activity was about 3.8% of the normal. The partially purified enzyme revealed slow anodal electrophoretic mobility, high Km NADP, marked thermal-instability, and increased affinity for a substrate analogue (deamino-NADP). A particular characteristic of this enzyme was a biphasic pH curve with a greatly increased activity at low pH values. From these results, this variant was clearly different from hitherto observed G6PD variants, and was designated G6PD Asahikawa.     

Isolation of citrate-positive variants of Escherichia coli from domestic pigeons, pigs, cattle, and horses.

Twenty-seven isolates of citrate-positive variants of Escherichia coli were obtained from domestic pigeons, pigs, cattle, and horses. With the exception of citrate utilization, all isolates closely resembled typical E. coli in their biochemical reactions. These isolates were multiply resistant to antibiotics in in vitro susceptibility tests. Transfer experiments of multiple-drug resistance to the E. coli K-12 strain showed that all citrate-positive isolates from domestic pigeons, pigs, and cattle, resistant to three or more drugs, carried R plasmids showing temperature-sensitive transfer.     

Overexpression of glucose transporter modulates insulin biosynthesis in insulin producing cell line

Glucose transporter (GT) has been suggested to be involved in the insulin biosynthesis. However, the functional relationship between GT and insulin biosynthesis is not well understood. In this report, we have generated rat pancreatic B cell lines (RINr) that stably overexpress a cDNA encoding the brain type GT. These cell lines showed 3- to 4-fold increase in insulin mRNA and protein. These results suggest that GT might have some relationship to the insulin biosynthesis in the pancreatic B cells.     

Effect of glucose on protein phosphorylation in rat pancreatic islets

Isolated rat pancreatic islets, incubated in the presence of extracellular ³²Pi to steady state ³²P incorporation into cellular phosphopeptides, were exposed to glucose for 10 min. Glucose (16.7 mM) significantly stimulated the phosphorylation of six phosphoproteins with molecular weights of 15,000, 35,000, 49,000, 64,000, 93,000 and 138,000. Mannoheptulose (16.7 mM) markedly inhibited glucose-stimulated phosphorylation of these six phosphoproteins. This protein phosphorylation might be important in mediating glucose-stimulated insulin release.     

Clinicopathological assessment of cancer/testis antigens NY-ESO-1 and MAGE-A4 in osteosarcoma

The cancer/testis antigens (CTAs), New York esophageal squamous cell carcinoma-1 (NY-ESO-1) and melanoma antigen gene (MAGE)-A4 are normally restricted to male germ cells but are aberrantly expressed in several cancers. Considering the limited information regarding their significance in osteosarcoma (OS), the purpose of this study was to determine the clinical significance of NY-ESO-1 and MAGE-A4 expression in OS. Nine patients with OS treated at Kindai University Hospital were included in the study. The median age was 27 years, and median follow-up period was 40 months. The specimens obtained at the time of biopsy were used to perform immunostaining for NY-ESO, MAGE-A4, p53, and Ki-67. The positive cell rates and positive case rates of NY-ESO, MAGE-A4, p53, and Ki-67 were calculated. The correlation between the positive cell rate of immunohistochemical markers was also calculated. The correlation between the positive cell rate of NY-ESO-1 or MAGE-A4 and tumor size or maximum standardized uptake (SUV-max) was also determined. The positive cell rates of NY-ESO-1 or MAGE-A4 in continuous disease-free (CDF) cases were also compared with those in alive with disease (AWD) or dead of disease (DOD) cases. The average positive cell rates of NY-ESO, MAGEA4, p53, and Ki-67 were 71.7%, 85.1%, 16.2%, and 14.7%, and their positive case rates were 33.3%, 100%, 44.4%, and 100%, respectively. The positivity rates of NY-ESO-1 and p53 were strongly correlated, whereas those of NY-ESO-1 and Ki-67 were moderately correlated. The MAGE-A4 and p53 positivity rates and the MAGE-A4 and Ki-67 positive cell rates were both strongly correlated. The NY-ESO-1 and MAGE-A4 positivity rates were moderately correlated. The positive correlation between the NY-ESO-1 positive cell rate and tumor size was medium, and that between the MAGE-A4 positivity rate and SUV-max was very strong. There was no significant difference in the positive cell rates of NY-ESO-1 or MAGE-A4 between CDF cases and AWD or DOD cases. Overall, our results suggest that NY-ESO-1 and MAGE-A4 may be involved in the aggressiveness of OS.Key words: New York esophageal squamous cell carcinoma-1 (NY-ESO-1), melanoma antigen gene (MAGE)- A4, osteosarcoma, prognosis, cancer/testis antigen (CTA), immunohistochemistry