Synergistic regulation of inducible nitric oxide synthase gene by CCAAT/enhancer-binding protein beta and nuclear factor-kappaB in hepatocytes

Intratumoral (i.t.) injection of a plasmid DNA vector encoding the murine interleukin-2 (IL-2) gene was used to treat established renal cell carcinoma (Renca) tumors in BALB/c mice. Tumor regression was observed in 60-90% of mice that were injected i.t. for 4 days with IL-2 plasmid DNA complexed with the cationic lipid DMRIE/DOPE ((+/-)-N-(2-hydroxyethyl)-N,N-dimethyl-2,3-bis(tetradecyloxy)-1-propa naminium bromide/dioleoylphosphatidylethanolamine). The mice remained tumor-free until the conclusion of the study, which was 4 months after tumor challenge. In a rechallenge experiment, mice that were rendered tumor-free for 6 months by IL-2 plasmid DNA treatment rejected a subsequent challenge of Renca cells but could not reject a challenge with the unrelated, syngeneic CT-26 tumor. Spleen cells from cured mice contained Renca-specific cytotoxic T lymphocytes, and adoptive transfer of mixed lymphocyte cultures into naive mice at 2 days after challenge with Renca cells prevented tumor growth. In vivo depletion of T-cell subsets at the time of i.t. injection with IL-2 plasmid DNA demonstrated that CD8+ T cells, but not CD4+ T cells, were the primary effectors of the antitumor response.     

Quantitative morphology of the rat kidney during diabetes mellitus and insulin treatment

A morphometric study was performed on moderately hyperglycaemic streptozotocin diabetic rats after 10 and 50 days of diabetes, and on groups of rats that, after initial hyperglycaemia for 50 days, were insulin treated for 2 h or for 5, 15 or 38 days. A group of hyperglycaemic diabetic animals were fasted for 18 h. Another group of rats had acute hyperglycaemia induced by intravenous glucose injection. After 10 and 50 days of diabetes, kidney weight was increased by 55 and 93%. Glomerular volume, tubule length, and tubular and interstitial volume increased in diabetic animals compared with controls. After 4 h insulin treatment, the kidney weight was 20% decreased; after 5 days it was 31% decreased. After 38 days the kidney weight was still 26% greater than in controls. In diabetic animals, 18 h fasting induced a 30% decrease in kidney weight. In normal animals, acute hyperglycaemia induced a 22% increase in kidney weight. Volume fractions of most kidney structures remained similar in all groups. However, the glomerular volume fraction was smaller during kidney enlargement, and the tubular volume fraction was larger after induced hyperglycaemia compared with controls. In conclusion, high blood glucose levels in diabetic and normal animals are associated with increased kidney weight. In hyperglycaemic diabetic animals, normalization of blood glucose after insulin treatment or fasting was followed by a decrease in kidney weight.     

Differential localization and expression of alpha and beta isoenzymes of protein kinase C in the rat retina

Expression and cellular localization of three isoenzymes of Ca2+-dependent protein kinase C (PKCalpha, PKCbeta, and PKCgamma) in the adult rat retina were revealed by immunohistochemistry and in situ hybridization histochemistry with isoenzyme-specific antibodies and cRNA probes. Immunoreactivities and mRNA signals for PKCalpha were conspicuous in rod bipolar cells. A subgroup of amacrine cells expressed PKCalpha. The cells in the ganglion cell layer also displayed PKCalpha gene products. Positive immunoreactivities for PKCbeta were localized as stripe patterns in the inner plexiform layer, corresponding to the stratification levels of axon terminals of cone bipolar cells. The somata of cone bipolar cells expressed PKCbeta. Amacrine cells and retinal ganglion cells also displayed PKCbeta gene products. The results obtained by immunohistochemistry were confirmed with colocalization of mRNA signals for PKCalpha and PKCbeta on the somata. The cell membranes showed stronger immunoreactivities than did the cytoplasms for both PKCalpha and PKCbeta. Neither immunoreactivities nor mRNA signals for PKCgamma were detected in all retinal regions. The differential roles of Ca2+-dependent PKC isoenzymes could be revealed in physiological defined retinal neurons.     

Bombesin activation of phospholipase C beta 1 in rat acinar pancreatic cells involves the pertussis toxin-sensitive G alpha i3 protein

Bombesin stimulation of inositol 1,4,5-trisphosphate (Ins P3) formation in rat sonicated pancreatic acinar cells was inhibited by an antibody directed against the pertussis toxin (PTX)-sensitive GTP-binding G alpha i3 protein but not by an anti-G alpha q-11 antibody. After solubilization and gel filtration, [125I-Tyr4]bombesin binding sites were recovered in a peak of protein of 67 approximately 90 kDa with a maximal enrichment corresponding to a molecular mass of 83-kDa. Results obtained from the non-hydrolysable GTP analog guanosine-5'-[gamma-thio]triphosphate (GTP gamma S) binding, PTX-stimulated ADP-ribosylation and immunoblotting showed that the 83-kDa fraction contained the G alpha i3 protein but not the G alpha q-11 protein. Furthermore, GTP gamma S increased the bombesin binding dissociation constant (KD) from 0.32 to 0.60 nM, while the anti-G alpha i3 antibody decreased the maximal binding capacity (Bmax) from 50 to 25 fmol/mg protein without affecting the KD. Mixing solubilized bombesin binding sites with a phospholipase C (PLC) preparation from rat pancreas reconstituted a bombesin-stimulated PLC activity which was markedly inhibited by the anti-G alpha i3 antibody but unaffected by the anti-G alpha q-11 antibody. In addition, this stimulation was inhibited by an anti-PLC beta 1 antibody. This result supports the involvement of the PLC beta 1 isoform in bombesin receptor activation.     

Control of expression of the gene for the arginine transporter Cat-1 in rat liver cells by glucocorticoids and insulin

A general mathematical model for the dynamic behaviour of a single-compartment respiratory system in response to an arbitrary applied inspiratory airway pressure and arbitrary respiratory muscle activity is investigated. The model is used to compute explicit expressions for ventilation and pressure variables of clinical interest for clinician-selected and impedance-determined inputs. The outcome variables include tidal volume, end-expiratory pressure, minute ventilation, mean alveolar pressure, average pleural pressure, as well as the work performed by the ventilator and the respiratory muscles. It is also demonstrated that under suitable conditions, there is a flow reversal that can occur during inspiration.     

Changes in transforming growth factor beta1 gene expression and immunoreactivity levels during development of chronic radiation enteropathy

Previously, we reported that pirarubicin (THP), an anthracycline, was taken up, at least in part, by both human leukemic HL60 cells and mononuclear cells (MNCs) via a carrier-mediated system. In this study, the possibility of a contribution of nucleoside transport systems to the uptake of THP by HL60 cells and MNCs was investigated. The experiments were performed after both types of cells had been pretreated with a metabolic inhibitor, 2,4-dinitrophenol, to deplete cellular ATP. In HL60 cells, THP uptake was increased and decreased significantly by treatment with equilibrative nucleoside transport inhibitors, nitrobenzylthioinosine (NBMPR), nitrobenzylthioguanosine and dilazep, in the presence and absence, respectively, of an inwardly directed Na+-gradient. THP uptake by HL60 cells showed an overshoot in the presence of the gradient, and was decreased by treatment of the cells with monensin, indicating that the uptake partially depended on the Na+-gradient. In HL60 cells in which equilibrative nucleoside transport was inhibited by NBMPR, THP uptake in the presence of the gradient was inhibited by Na+-dependent concentrative nucleoside transport inhibitors, but no inhibition was observed in the absence of the gradient. In MNCs, conversely, there was no effect of any equilibrative nucleoside transport inhibitor or the Na+-gradient on THP uptake. These results suggested that THP was taken up, at least in part, via both equilibrative and concentrative nucleoside transport systems in HL60 cells, but not in MNCs.     

Mapping of murine diabetogenic gene mody on chromosome 7 at D7Mit258 and its involvement in pancreatic islet and beta cell development during the perinatal period

Mutation of the murine maturity-onset diabetes mellitus of the young (Mody) locus induces diabetes, but the effects of its homozygosity on the pancreas remain unknown. F2 mice were obtained by F1 (diabetic C57BL6 x normal Mus musculus castaneus) crosses. About 20% of the F2 progeny developed diabetes by 2 wk of age, 50% of the progeny were normal at 2 wk and developed diabetes between 5 and 8 wk of age, and the remaining 30% did not develop diabetes. Quantitative trait locus analysis using blood glucose levels of 118 F2 mice at 2 wk of age and 5-8 wk of age located Mody within 3 cM of D7Mit258. Histopathological investigation revealed hypoplastic islets (approximately 33% of that of wild-type mice) and a lower density of beta cells (approximately 20% of wild-type) with a reciprocal dominance of alpha cells (four times that of wild-type) in Mody homozygotes. Electron microscopic observations revealed a specific decrease in the number of insulin secretory granules and a lower density of beta cells. Ratios of insulin to glucagon contents confirmed specific decreases in insulin content: 0.01 for homozygotes, 0.54 for heterozygotes, and 1.11 for wild-type mice on day 14. These results suggest that Mody is involved in both islet growth and beta cell function.     

Characterization of expression of phosphofructokinase isoforms in isolated rat pancreatic islets and purified beta cells and cloning and expression of the rat phosphofructokinase-A isoform

We prepared polymers having a phospholipid polar group, poly [omega-methacryloyloxyalkyl phosphorylcholine (MAPC)-co-n-butyl methacrylate(BMA)], as new biomedical materials and evaluated their blood compatibility with attention to protein adsorption and platelet adhesion. The total amount of proteins adsorbed on the polymer surface from human plasma was determined, and the distribution of adsorbed proteins on the plasma-contacting surface was analyzed. The amount of proteins adsorbed on every poly (MAPC-co-BMA) was small compared with that observed on polymers without the phospholipid polar group. However, there was no significant difference in the amount of adsorbed proteins on the poly(MAPC-co-BMA) even when the methylene chain length between the phospholipid polar group and the backbone in the MAPC moiety was altered. Platelet adhesion on the polymer surface from a platelet suspension in a buffered solution was evaluated with and without plasma treatment on the surface. When a rabbit platelet suspension was brought into contact with the poly(BMA) surface after treatment with plasma, many platelets adhered and aggregated. However, a reduced amount of platelet adhered on the poly(BMA) was found in the case of direct contact with the platelet suspension. On the other hand, the poly(MAPC-co-BMA)s could inhibit platelet adhesion under both conditions. Based on these results, it can be concluded that the proteins adsorbed on the surface play an important role in determining the platelet adhesion and suppression of the protein adsorption on the surface, which is one of the most significant ways of inhibiting platelet adhesion.