Galanin-receptor ligand M40 peptide distinguishes between putative galanin-receptor subtypes

The galanin-receptor ligand M40 [galanin-(1-12)-Pro3-(Ala-Leu)2-Ala amide] binds with high affinity to [mono[125I]iodo-Tyr26]galanin-binding sites in hippocampal, hypothalamic, and spinal cord membranes and in membranes from Rin m5F rat insulinoma cells (IC50 = 3-15 nM). Receptor autoradiographic studies show that M40 (1 microM) displaces [mono[125I]iodo-Tyr26]galanin from binding sites in the hippocampus, hypothalamus, and spinal cord. In the brain, M40 acts as a potent galanin-receptor antagonist: M40, in doses comparable to that of galanin, antagonizes the stimulatory effects of galanin on feeding, and it blocks the galaninergic inhibition of the scopolamine-induced acetylcholine release in the ventral hippocampus in vivo. In contrast, M40 completely fails to antagonize both the galanin-mediated inhibition of the glucose-induced insulin release in isolated mouse pancreatic islets and the inhibitory effects of galanin on the forskolin-stimulated accumulation of 3',5'-cAMP in Rin m5F cells; instead M40 is a weak agonist at the galanin receptors in these two systems. M40 acts as a weak antagonist of galanin in the spinal flexor reflex model. These results suggest that at least two subtypes of the galanin receptor may exist. Hypothalamic and hippocampal galanin receptors represent a putative central galanin-receptor subtype (GL-1-receptor) that is blocked by M40. The pancreatic galanin receptor may represent another subtype (GL-2-receptor) that recognizes M40, but as a weak agonist. The galanin receptors in the spinal cord occupy an intermediate position between these two putative subtypes.     

The role of cysteine residues of spinach ferredoxin-NADP+ reductase As assessed by site-directed mutagenesis

To investigate the functional role of the cysteine residues present in the spinach ferredoxin-NADP+ oxidoreductase, we individually replaced each of the five cysteine residues with serine using site-directed mutagenesis. All of the mutant reductases were correctly assembled in Escherichia coli except for the C42S mutant protein. C114S and C137S mutant enzymes apparently showed structural and kinetic properties very similar to those of the wild-type reductase. However, C272S and C132S mutations yielded enzymes with a decreased catalytic activity in the ferredoxin-dependent reaction (14 and 31% of the wild type, respectively). Whereas the C132S was fully competent in the diaphorase reaction, the C272S mutant flavoprotein showed a 35-fold reduction in catalytic efficiency with respect to the wild-type enzyme (0.4 versus 14.28 microM-1 s-1) due to a substantial decrease of kcat. NADP+ binding by the C272S mutant enzyme was apparently quantitatively the same (Kd = 37 microM) but qualitatively different, as shown by the differential spectrum. Stopped-flow experiments showed that the enzyme-FAD reduction rate was considerably decreased in the C272S mutant reductase, along with a much lower yield of the charge-transfer transient species. It is inferred from these data that the charge transfer (FAD-NADPH) between the reductase and NADPH is required for hydride transfer from the pyridine nucleotide to flavin to occur with a rate compatible with catalysis.     

Primary hepatic carcinoid tumor

A case of primary carcinoid tumor arising in the liver of a 69 year old woman with no endocrine symptoms is reported. Histopathologically, the tumor was diagnosed initially as a hepatocellular carcinoma in the biopsy specimen, and was shown subsequently to be a carcinoid tumor, demonstrating diffuse positive staining with Grimelius method. Mucin stained with periodic acid-Schiff (PAS), alcian-blue, and mucicarmine, and was shown partially in the glandular structures. Immunohistochemically, most of the tumor cells stained positively for chromogranin-A, epithelial membrane antigen (EMA) and neuron specific enolase (NSE). Ultrastructural examination revealed electron-dense core granules, measuring 40-120 nm in diameter in some of the tumor cells. Intensive and careful searches pre- and post-operatively revealed no other primary source of tumor other than the liver. The patient was reported well with no symptoms 3 1/2 years after the operation. This case is considered to be a primary hepatic carcinoid tumor. The recent literature is reviewed, and the possible histogenesis of hepatic carcinoid tumor is discussed.     

Cytokines affecting survival and differentiation of an astrocyte progenitor cell line

The effects of various cytokines on survival and differentiation of an astrocyte progenitor cell line (AP-16) were examined. Epidermal growth factor (EGF) deprivation caused death of AP-16 cells by apoptosis. Transforming growth factor-alpha (TGF-alpha) and basic fibroblast growth factor (bFGF) prevented the apoptosis occurring in the absence of EGF. Leukemia inhibitory factor (LIF) and ciliary neurotrophic factor (CNTF) induced glial fibrillary acidic protein (GFAP) and decreased A2B5 antigen in AP-16 cells, indicating that these cytokines induced AP-16 cells to differentiate into astrocytes.     

Neurohormonal activity in heart insufficiency

The maintenance of cardiac pumping ability in the presence of a primary disturbance of myocardial contractility and/or an excessive haemodynamic strain on the heart is dependent on several compensatory mechanisms. Particular attention has formerly been paid to the importance of the Frank-Starling mechanism and cardiac hypertrophy and dilatation in maintaining a blood supply sufficient to cover the metabolic needs of various tissues in heart failure. In recent years, however, it has been found that certain neurohormonal systems (the sympathetic nervous system, the renin-angiotensin-aldosterone system, atrial natriuretic peptide and several locally acting vaso-active substances) undergo considerable changes according to the degree of heart failure. These compensatory mechanisms support the circulation wholly or partially in acute heart failure, however sustained neurohormonal activation may be harmful in chronic heart failure, where several neurohormonal factors may be activated to ill-effect. The most significant neurohormonal systems and their importance in heart failure are reviewed on the basis of the available literature.     

High dietary calcium level decreases colonic phytate degradation in pigs fed a rapeseed diet

The degradation of phytate (inositol hexaphosphate) in rapeseed meal diet not containing phytase activity was studied in 15 growing ileum-fistulated pigs. Stomach and small intestinal degradation and total gastrointestinal degradation were compared. The effect of addition of calcium carbonate to the rapeseed meal diet at two levels (9.2 and 18.5 g/kg diet) was investigated. A commercial barley-wheat-soybean diet with intrinsic phytase activity was used as reference. Phytate and its hydrolysis products in diets, ileal digesta and feces were determined by HPLC ion-pair chromatography. Hydrolysis of phytate in the stomach and small intestine was 35-45% in pigs fed the rapeseed meal diet independent of calcium addition, and 65% in pigs fed the reference diet. Total gastrointestinal degradation of phytate in pigs fed the rapeseed diet was 97, 77 and 42% (P < 0.001) when calcium intakes were 4.5, 9.9 and 15 g/d, respectively; total gastrointestinal degradation was 72% in pigs fed the reference diet. The intestinal phytate degradation pattern, when rapeseed diet was fed, indicated the activity of an unspecific phosphatase, whereas that of the reference diet indicated intrinsic dietary phytase activity. We conclude that dietary supplementation of calcium carbonate decreases the phytate degradation in the colon of pigs, but not in the stomach and small intestine.     

Fully methylated oriC with negative superhelicity forms an oriC-membrane complex before initiation of chromosome replication

In an in vitro assay, the oriC DNA has been shown to bind to the outer membrane fraction only when it is hemimethylated (G.B. Ogden et al., Cell, 54, 127-135,1988). In this report, however, we demonstrated that a significant amount of the oriC DNA was recovered from the cells just before initiation with the oriC DNA being fully methylated. Formation of this preinitiation oriC-membrane complex and following initiation of chromosome replication were strongly inhibited by novobiocin, a DNA gyrase B subunit inhibitor, which reduced the superhelicity of the reporter plasmid in the cells. On the other hand, both reactions proceeded in the presence of nalidixic acid, a DNA gyrase A subunit inhibitor, which did not have the effect of reducing the superhelicity. These results suggest that the negative superhelicity of the DNA is required for preinitiation oriC-membrane complex formation and following initiation event of replication.     

Induced-fit association between DNA and esperamicin. Importance of structural flexibility in host DNA duplex

In this study, a series of synthetic oligonucleotide duplexes are tested as a substrate for esperamicin. The duplexes contain a typical binding sequence of esperamicin, 5'-GGA/TCC, but have different flexibilities in helix structure from each other. When cleavage activities of these oligonucleotides by esperamicin were estimated by using DNA sequencing method, a substantial increase of the cleavage at 3'-NAGG was observed with increasing the helix flexibility. This observation indicates that structural flexibility of host DNA duplex is important in an induced-fit association between esperamicin and DNA.     

Reoperations after operation on the thoracic aorta: etiology, surgical techniques, and prevention

Recurrent aortic aneurysms, persistent or new dissection, new onset of valvular and coronary artery disease, graft infection, and prosthetic endocarditis are not rare after thoracic aortic operations; they can be difficult to diagnose and represent a formidable surgical challenge. Between 1977 and 1991, 876 operations were performed on the thoracic aorta in our institution: 340 in dissections, 299 in true aneurysms, 150 for aortic remodeling and external wall support during aortic valve replacement, and 87 for miscellaneous causes. During the same period, there were 193 additional reoperations. Vascular reoperations on abdominal aorta and peripheral arteries accounted for 73 cases and are not further discussed in this study. The reasons for reoperation (n = 130) in 120 patients were: failure of biologic valves (n = 23); aneurysm recurrence in a proximal or distal aortic segment (n = 21); pseudoaneurysm formation at suture lines (n = 13); new dissection or dilatation involving ascending aorta (n = 11), aortic arch (n = 13), and descending aorta (n = 10); aneurysm after aortic remodeling (n = 13); new onset of valvular disease (n = 5); and new onset of coronary disease (n = 5). Infected aortic graft and prosthetic endocarditis accounted for 10 reoperations, and a planned two-staged procedure was performed in 6 patients. Omitting the failed biologic valves, reoperations were performed on the aortic segment previously operated on in 69.3% of the cases and on other thoracic segments in 30.7%. Overall hospital mortality rate after reoperation was 5.8%. A significant decrease in operative mortality was observed in the most recent period (3.0% between 1989 and 1991). Reoperations are technically demanding, and some of them are preventable; therefore (1) graft inclusion technique should be abandoned in ascending aortic operation due to formation of false aneurysms; (2) in patients with Marfan syndrome, complete repair of the diseased aorta should be attempted during the initial operation; (3) aortic arch dissection should be repaired definitively during the first operation in low-risk patients; (4) biological valves should be avoided in aneurysm operations; and (5) homograft replacement is the treatment of choice in prosthetic endocarditis or in infected composite graft after an aortic valve or ascending aortic operation.     

Cholesteatoma surgery today

Four present-day surgical techniques are reviewed to assess their respective merits in surgery for cholesteatoma. The oldest method with an open cavity in ears with mastoid extension of cholesteatoma if combined with partial obliteration is still suitable for less experienced surgeons. Transcanal atticotympanotomy is suitable for limited epitympanic and tympanic cholesteatomas as long as it provides a direct view of the operative field. In similar ears, canal wall up surgery is employed if, additionally, mastoidectomy is needed because of chronic inflammation. Canal wall down surgery with full cavity obliteration with a musculoperiosteal flap, bone chips and bone pate should be the method of choice for all cholesteatomas extending beyond the facial nerve canal. The canal skin is kept as an intact tube and provides quick healing. Open cavities should be revised using similar obliteration techniques but, because of the lack of an intact canal skin tube, making use of a large modified K?rner skin flap.