Synthesis of human endothelin-1 precursors in Escherichia coli

Endothelin, the most potent vasoconstrictor found in nature, is thought to be important in the regulation of blood pressure and/or local blood distribution. Human placenta cDNA fragment encoding preproendothelin-1 (preproET-1) and its carboxyl terminal mature precursor (C-matured precursor) was expressed in E. coli. These products were characterized by both enzyme immunoassay and Western blot analysis.     

Amino acid sequence and relative biological activity of eel atrial natriuretic peptide

A peptide exhibiting vasodepressor and natriuretic activities in rats was isolated from eel atria, and its primary structure was determined as H-Ser-Lys-Ser-Ser-Ser-Pro-Cys-Phe-Gly-Gly-Lys-Leu-Asp-Arg-Ile-Gly-Ser-Tyr-Ser- Gly-Leu-Gly-Cys-Asn-Ser-Arg-Lys-OH. This peptide, termed eel atrial natriuretic peptide (ANP), has sequence homology of 59% to mammalian (human or rat) ANP, 52% to fowl ANP, and 46% to frog ANP. When the biological activity of synthetic eel ANP was compared with that of human ANP, the eel peptide was 110 times more potent for the vasodepressor activity in eels, nearly equipotent for the vasodepressor activity in quails, and 20 times less potent for the vasodepressor and natriuretic activity in rats.     

Two distinct types of endothelin receptors are present on chick cardiac membranes

Competitive displacement experiments of 125I-endothelin (ET)-1, -2, or -3 binding to chick cardiac membranes were performed with unlabeled ET-1, -2, -3, and sarafotoxin S6b (STX) as competitors. 125I-ET-1 and -2 binding was competitively inhibited by increasing concentrations of these unlabeled peptides in the same order; i.e. ET-2 greater than or equal to ET-1 greater than ET-3 greater than STX. In contrast, the order of potency in displacing 125I-ET-3 binding was ET-3 greater than ET-2 greater than or equal to ET-1 greater than STX. Affinity labeling of the membranes by cross-linking with 125I-ET-1 and -2 via disuccinimidyl tartarate yielded one major specific band with an apparent Mr = 53,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis followed by autoradiography. On the other hand, affinity labeling with 125I-ET-3 showed that two major and one minor bands of Mr = 34,000, 46,000, and 53,000, respectively, were specifically labeled. These results indicate the presence of two distinct types of ET receptors, one of which has higher affinity for ET-1 and -2 than ET-3 and the other is conversely ET-3-preferring.     

Induction of manganese superoxide dismutase by tumor necrosis factor in human breast cancer MCF-7 cell line and its TNF-resistant variant

Tumor necrosis factor (TNF)-resistant variant of human mammary cancer MCF-7 cell line was isolated by stepwise selection. The final TNF-resistant variant Tnf-1000 showed more than 100-fold higher resistance than the parental MCF-7 cell. Saturation kinetics for 125I-TNF binding showed that TNF-1000 cells had similar TNF receptor numbers as MCF-7 cells, but of a lower affinity. Induction of superoxide dismutase (SOD) was compared between MCF-7 and Tnf-1000 cells treated with TNF: SOD scavenges potentially toxic superoxide radicals. TNF induced more mitochondrial manganese SOD (SODm) in MCF-7 than in Tnf-1000 whereas there appeared to be no significant induction of cytosolic copper/zinc SOD (SODc) by TNF in both MCF-7 and Tnf-1000 cell lines. Acquirement of TNF-resistance in MCF-7 cells might be correlated with expression of SODm.     

Mitochondrial DNA polymorphism in native Philippine cattle based on restriction endonuclease cleavage patterns

An analysis of patterns of cleavage of mtDNA by restriction endonucleases was performed for nine individuals from the Philippine population of native cattle. MtDNA polymorphisms were detected in the restriction patterns generated by the following six enzymes,BamHI,BglII,EcoRV,HindIII,PstI, andScaI. The restriction patterns showing polymorphisms were distributed nonrandomly among the nine individuals examined from the Philippine population of native cattle, indicating the existence of two separate types of mtDNA. These two types of mtDNA are very different from each other, at the level of subspecies. Since the native Philippine cattle are considered to represent an admixture of European and Indian cattle, the two types of mtDNA must be derived from the mtDNAs of both varieties. The polymorphic sites in mtDNA have been located on a restriction map, and the nucleotide substitutions at some of the sites have also been estimated.     

Hypoxia-induced fibre type transformation in rat hindlimb muscles Histochemical and electro-mechanical changes

Twelve male Sprague-Dawley rats (21 days old) were randomly assigned into two experimental groups: sea level control (CONT) and hypobaric hypoxia (HYPO). The HYPO rats were kept in an hypobaric chamber maintaining a simulated altitude of 4000 m (61.1 kPa). After 10 weeks of treatment, the rat hindlimb muscles [soleus (SOL) and extensor digitorum longus (EDL)] were subjected to histochemical and electro-mechanical analyses. Results indicated that compared to CONT the HYPO SOL muscle had a significantly greater relative distribution of fast-twitch-oxidative-glycolytic (FOG) fibres (28.9% SEM 2.0 vs 18.3% SEM 1.8, P less than 0.01) with a significant decrease in slow twitch oxidative fibre distribution (69.5% SEM 2.4 vs 82.9% SEM 3.1, P less than 0.01). Compared to CONT the HYPO EDL muscle also manifested a significant increase in FOG fibre distribution (51.6% SEM 0.8 vs 46.6% SEM 1.1, P less than 0.01), but this was accompanied by a significant decrease in fast twitch glucolytic fibres (44.3% SEM 0.9 vs 49.2% SEM 1.7, P less than 0.05). These histochemical fibre type transformations accompanied significant and expected changes in the electro-mechanical parameters tested in situ, e.g. maximal twitch force, maximal rate of force development, contraction time, half relaxation time, force: frequency curve, and fatigability. It was concluded that chronic hypobaric hypoxia could have a potent influence upon the phenotype expression of muscle fibres.     

Appearance of retrogradely labeled neurons in the rat superior cervical ganglion after injection of wheat-germ agglutinin-horseradish peroxidase conjugate into the contralateral ganglion

Summary Injection of wheat-germ agglutinin-horseradish peroxidase conjugate (WGA-HRP) into the superior cervical ganglion (SCG) of the rat results in accumulation of WGA-HRP in sympathetic postganglionic neurons in the contralateral SCG. The sympathetic pathways involved and the mechanism underlying the labeling were investigated. The labeling in neurons in the contralateral SCG was apparent 6 h after injection and increased in intensity with longer survival times. The number of labeled neurons reached 1300 at 72 h after the injection. Transection of the external (ECN) or internal carotid nerves (ICN) resulted in considerable reduction in the number of labeled neurons. Combined transection of both ECN and ICN virtually eliminated labeling in the contralateral SCG. This provides strong evidence that these two nerves are the major pathways for WGA-HRP transport out of the SCG. No labeling was observed in the contralateral SCG following injection of horseradish peroxidase (HRP). Therefore, it seems unlikely that a direct nerve connection exists between the bilateral ganglia. Instead, the labeling of contralateral SCG neurons appears to depend on the transneuronal transport capacity of WGA-HRP, which conveys the marker in an anterograde direction along the postganglionic fibers to terminals in sympathetic target organs, and then delivers it transneuronally to contralateral SCG neurons. We suggest that the sympathetic nerve fibers originating in the bilateral SCGs run intermingled and are in close contact in their peripheral target organs.     

Immunohistochemical colocalization of glucagon,serotonin, and aromatic L-amino acid decarboxylase in islet A cells of chicken pancreas

Summary The identity of monoamine-emitted, formaldehyde-induced fluorescence in some pancreatic islet cells was studied in pancreatic tissue of male chickens by fluorescence and immunohistochemistry either on the same tissue section or on serial tissue sections. Pancreatic islet cells emitting intense formaldehyde-induced fluorescence also react immunohistochemically with antisera directed against glucagon, serotonin and aromatic L-amino acid decarboxylase. These results show that chicken pancreatic islet A cells contain glucagon, serotonin, and aromatic L-amino acid decarboxylase, an enzyme involved in the synthesis of serotonin. The islet B cells identified with anti-insulin immunoreactivity, which displayed a very weak formaldehyde-induced fluorescence, did not react with anti-serotonin serum.