Evidence for FSH-like material in ACTH granules of certain corticotropic cells in the pituitary of the pig

Summary The ultrastructural immunocytochemical peroxidase-antiperoxidase technique used on serial sections reveals that certain corticotropic cells contain both ACTH and FSH, but not LH (ACTH/FSH cells). To determine the specificity of the anti-FSH staining in these cells, immunocytochemical absorption experiments were performed. The results indicate that (1) anti-FSH and -ACTH antisera are bound to different antigens in the corticotropic cell, and (2) anti-FSH staining is specific for a FSH-like antigen. In the ACTH/FSH cells both hormones are stored in the same secretory granules, distributed among other granules that contain only ACTH.     

Characterization of gangliosides from Ehrlich ascites tumour cells and their variants

Differences in the nature of the gangliosides present in two types of Ehrlich ascites tumour (EAT) cells, the adherent and non-adherent EAT cells, were studied. Gangliosides were isolated by DEAE Sephadex column chromatography and analysed by high-performance thin-layer chromatography (HPTLC). The non-adherent EAT (na-EAT) cells which grow in the peritoneal cavity of mice were selected for growth on basement membrane and tissue culture plastic to give the adherent EAT (a-EAT) cells. na-EAT cells contained 1.57 nmol lipid-bound sialic acid per mg protein and at least 12 different gangliosides, including major gangliosides such as GM3, GM2, GM1, GD3, GD1a and GT1b. On the other hand, the ganglioside pattern of a-EAT cells differed significantly from that of na-EAT cells, both quantitatively and qualitatively. The content of lipid-bound sialic acid in a-EAT cells was only 0.24 nmol per mg of protein. The gangliosides in a-EAT cells were characterized as GD1a and trisialogangliosides and, significantly, a-EAT cells did not contain monosialogangliosides. Neutral glycolipids were isolated from both cell lines and their patterns were compared. In contrast to the gangliosides pattern, their neutral glycolipid patterns were similar. Glucosylceramide and lactosylceramide were the major components in both types of cells. In addition to na- and a-EAT cells, a-EAT cells were passaged in mice by intraperitoneal injection, giving rise to a third variant (c/m EAT cells). We analysed the gangliosides in c/m EAT cells to determine whether there was a change in the ganglioside pattern found in na-EAT cells. After repeated passage of c/m EAT cells in mice, the pattern of gangliosides shifted to that of na-EAT cells. Alterations of ganglioside composition may be associated with the growth environment of the murine peritoneal cavity; alternatively, a selection process may have occurred.Abbreviations EAT cells Ehrlich ascites tumour cells - na-EAT cells non-adherent EAT cells - a-EAT cells adherent EAT cells - c/m EAT cells cultured a-EAT cells passaged in mice - HPTLC high-performance thin-layer chromatography - PBS 10 mM phosphate-buffered saline, pH 7.2, containing 0.15 M NaCl - EDTA ethylene-diaminetetraacetic acid - TFA trifluoroacetic acid - TG thioglycollate - Cer ceramide (N-fatty acyl sphingosine) - GM3 NeuAc2-3Gal1-4Glc-Cer - GM2 GalNAc1-4(NeuAc2-3)Gal1-4Glc-Cer - GM1a Gal1-3GalNAc1-4(NeuAc2-3)Gal1-4Glc-Cer - GD3 NeuAc2-8NeuAc2-3Gal1-4Glc-Cer - GD1a NeuAc2-3Gal1-3GalNAc1-4(NeuAc2-3)Gal1-4Glc-Cer - GT1b NeuAc2-3Gal1-3GalNAc1-4(NeuAc2-8NeuAc2-3)Gal1-4Glc-Cer - LacCer Gal1-4Glc-Cer - Gb3 Gal1-4Gal1-4Glc-Cer - Gb4 GalNAc1-4Gal1-4Gal1-4Glc-Cer This paper is dedicated to my esteemed colleague, Sen-itiroh Hakomori on the occasion of his 65th birthday.     

α-Gustducin immunoreactivity in the airways

The G-protein subunit -gustducin is a marker of chemoreceptive cells. In the present study, we examined the immunohistochemical localization of -gustducin in rat airway epithelium both by light and electron microscopy. -Gustducin immunoreactivity was found in solitary cells that presented ultrastructural features of chemoreceptor cells, i.e. flask-shaped or pear-shaped, with an apical process with thin microvilli protruding into the lumen. The immunostaining was mainly concentrated in the apical process and along the basolateral cell surface. To investigate whether -gustducin-immunoreactive cells represented a distinct cell subset in rat airways, we performed double-label immunocytochemistry with antibodies to protein gene groduct (PGP) 9.5, a marker of neuroendocrine cells, and to phospholipase C beta2 (PLC2), a component of the bitter signalling pathway. -Gustducin-immunoreactive cells were present in a subset of PGP-9.5-immunoreactive elements, although not all -gustducin-positive cells expressed PGP 9.5 labelling. In addition, a subset of -gustducin-expressing cells colocalized PLC2. This work thus demonstrates that solitary -gustducin-immunoreactive cells exist throughout the airways and represent a specialized cell type with morphological and immunohistochemical characteristics of chemoreceptor cells.     

Identification of beta subunit of the rhesus monkey chorionic gonadotropin (rmCG)

Chorionic gonadotropin (CG) is a placental derived hormone that plays a crucial role in successful implantation and establishment of early pregnancy in the primates. The rhesus monkey was chosen as a model to understand the feasibility of developing human DNA immuno-contraceptive. The coding region of rhesus monkey CG -subunit (rmCG) was isolated by the TDRT-PCR method. The nucleotide sequence including the leader peptide was 499 nucleotide long and encoded 166 amino acids. In comparing with the previous known primates CG -subunits, the rmCG was the highest degree of homology with baboon CG -subunit at the deduced amino acid sequence (94%), 79.5% homology with human CG -subunit and 70.4% homology with marmoset monkey CG -subunit. The eukaryotic expression vector pCMV4-rmCG inserted full-coding cDNA sequence of rmCG was constructed, and the expression of rmCG -subunit in HeLa cells transient expressing system in vitro and BALB/c mice in vivo was determined. The results demonstrated that the recombinant PCMV4-rmCG eukaryotic expression vector could express rmCG -subunit in vitro and in vivo.     

Evolutionary origin of the class A and class C β-lactamases

Summary The protein sequences of 18 class A -lactamases and 2 class C -lactamases were analyzed to produce a rooted phylogenetic tree using the DD peptidase of Streptomyces R61 as an outgroup. This tree supports the penicillin-binding proteins as the most likely candidate for the ancestoral origin of the class A and class C -lactamases, these proteins diverging from a common evolutionary origin close to the DD peptidase. The actinomycetes are clearly shown as the origin of the class A -lactamases found in other non-actinomycete species. The tree also divides the -lactamases from the Streptomyces into two subgroups. One subgroup is closer to the DD peptidase root. The other Streptomyces subgroup shares a common branch point with the rest of the class A -lactamases, showing this subgroup as the origin of the non-actinomycete class A -lactamases. The non-actinomycete class A -lactamase phylogenetic tree suggests a spread of these -lactamases by horizontal transfer from the Streptomyces into the non-actinomycete gram-positive bacteria and thence into the gram-negative bacteria. The phylogenetic tree of the Streptomyces class A -lactamases supports the possibility that horizontal transfer of class A -lactamases occurred within the Streptomyces.     

Histology and histochemistry of the testis and ovary of the platyfish,Xiphophorus maculatus,from birth to sexual maturity

Summary The gonads of 3-day- to 7-month-old male and female platyfish (Xiphophorus maculatus) were examined for the presence of 5-3-hydroxysteroid dehydrogenase (3-HSD) and glucose-6-phosphate dehydrogenase (G6PD) by histochemical means. In 3-day-old males a positive response for both enzymes is localized in the Leydig cells. With subsequent testicular development, these cells increase in number and display greater activity at the periphery of the testis and around the efferent ducts. In 3-day-old females 3-HSD and G6PD are localized in the stromal cells of the ovary. These cells increase in number and activity as the animals become sexually mature. Sertoli cells, efferent duct epithelium, and ovarian granulosa cells are negative at all stages of development examined. Our findings suggest that the gonads of neonatal fish possess the potential for steroidogenesis. The role played by sexsteroid hormones in the maturation of the brain-pituitary-gonad axis is discussed.     

β-Alanine metabolism and high salinity stress in the sea anemone,Bunodosoma cavernata

Summary During high salinity stress, -alanine accumulates to high levels in the sea anemone,Bunodosoma cavernata. Following a salinity increase from 26 to 40 -alanine increased 28-fold from 1.5 to 41.9 moles/g dry weight. Both whole animal studies and experiments with cell free homogenates indicate that under high salinity conditions an increase in the rate of -alanine synthesis from aspartic acid as well as a decrease in the rate of -alanine oxidation are responsible for the observed accumulation of -alanine. The rate of aspartic acid decarboxylation to -alanine is about 3 times greater in anemones acclimated to 40 than for those in normal salinity water (26). -alanine oxidation to CO₂ and acetyl-CoA proceeds 2.5 to 3 times slower in high salinity adaptedB. cavernata than in those acclimated to normal salinity. There is always a rapid degradation of uracil to -alanine, but this does not change with salinity.Abbreviations CASF cold acid soluble fraction - FAA free amino acids - MES 2(N-morpholino) ethane sulfonic acid - NPS ninhydrin positive substances - PCA perchloric acid - TCA trichloroacetic acid     

Binding of the galactose-specificPseudomonas aeruginosa lectin,PA-I,to glycosphingolipids and other glycoconjugates

The carbohydrate-binding specificity ofPseudomonas aeruginosa lectin I (PA-I) in iodinated or biotinylated form was studied. A large number of glycosphingolipids, as well as some glycoproteins and neoglycoproteins were used as ligands. Also, inhibition by free saccharides of PA-I binding to glycosphingolipids was tested. It was found that the lectin binds most strongly to terminal and nonsubstituted Gal3Gal- or Gal4Gal-structures.Abbreviations PA-I Pseudomonas aeruginosa lectin I - Cer ceramide - lactosylceramide Gal4GlcCer - iso globotriaosylcerami Gal3Gal4GlcCer - globotriaosylceramide Gal4Gal4GlcCer - globoside or globotetraosylceramide GalNAc3Gal4Gal4GlcCer - Forssman glycolipid GalNAc3GalNAc3Gal4Gal4GlcCer - P1 glycolipid Gal4Gal4GlcNAc3Gal4GlcCer - lactoneotetraosylceramide Gal4GlcNAc3Gal4GlcCer - B5 glycolipid Gal3Gal4GlcNAc3Gal4GlcCer - gangliotetraosylceramide Gal3GalNAc4Gal4GlcCer - GM1 Gal3GalNAc4(NeuAc3)Gal4GlcCer - RBC red blood cells - BSA bovine serum albumin - PBS phosphate-buffered saline - SDS sodium dodecyl sulfate - TLC thin-layer chromatography - HPLC high pressure liquid chromatography - MS mass spectrometry - FAB fast-atom bombardment - EI electron impact     

Differential Expression and Association of Calcium Channel Subunits in Development and Disease

Voltage-gated calcium channels (VDCC) are essential to neuronal maturation and differentiation. It is believed that important signaling information is encoded by VDCC-mediated calcium influx that has both spatial and temporal components. VDCC are multimeric complexes comprised of a pore-forming ₁ subunit and auxiliary and ₂/ subunits. Changes in the fractional contribution of distinct calcium conductances to the total calcium current have been noted in developing and differentiating neurons. These changes are anticipated to reflect the differential expression and localization of the pore-forming ₁ subunits. However, as in vitro studies have established that regulates the channel properties and targeting of ₁, attention has been directed toward the developmental expression and assembly of isoforms. Recently, changes in the component of the omega-conotoxin GVIA (CTX)-sensitive N-type VDCC have indicated differential assembly of _1B with in postnatal rat brain. In addition, unique properties of 4 have been noted with respect to its temporal pattern of expression and incorporation into N-type VDCC complexes. Therefore, the expression and assembly of specific ₁/ complexes may reflect an elaborate cellular strategy for regulating VDCC diversity. The importance of these developmental findings is bolstered by a recent study which identified mutations in the 4 as the molecular defect in the mutant epileptic mouse (lethargic; lh/lh). As 4 is normally expressed in both forebrain and cerebellum, one may consider the impact of the loss of 4 upon VDCC assembly and activity. The importance of the lb and 4 isoforms to calcium channel maturation and assembly is discussed.     

Enzyme-catalyzed synthesis of alkyl β-D-glycosides with crude homogenate ofSulfolobus solfataricus

Summary Crude homogenate of thermophilic archaebacteriumSulfolobus solfataricus, possessing a -glycosidase, has been used to synthesize different alkyl -D-glycosides starting from phenyl -D-glucoside, phenyl -D-galactoside and lactose as carbohydrate donors. High product yield (95% with respect to the carbohydrate donor) of octyl -D-glucoside has been obtained in a two-phase system containing 5% of water. The enantioselection for the galactosyl transfer to the secondary hydroxyl group of propane-1,2-diol is higher than that found using -galactosidase fromE. coli.     

Détection par immunofluorescence des cellules corticotropes et mélanotropes dans l'hypophyse de la grenouille,Rana temporaria L., au cours du développément

Summary During the embryonic and larval developmental stages of the frog, Rana temporaria L, anti- 1–24, 17–39 corticotropine, and MSH antibodies were used to define, with immunofluorescence technique, the appearence of corticotropic and melanotropic cells.A very small number of fluorescent corticotropic cells appears for the first time during the embryonic stage (10 mm), just before the differentiation of the pars intermedia. The cells are small, their large nucleus is surrounded by a fine rim of fluorescent cytoplasm.During premetamorphic stage, the anti-ACTH antibodies (anti- 1–24 and anti- 17–39 corticotropine) reveal more fluorescent cells in the whole pars distalis. The pars intermedia cells can also be visualized by both antisera.At the end of prometamorphosis and during climax the corticotropic cells show a more precise localization. As in adult frog pars distalis, they are concentrated in the rostral half of the lobe. With the same technique anti- and MSH antibodies reveal only the cells of the pars intermedia. No other cell type of the pars distalis reacts with these antibodies. This technique has the advantage to show that the ACTH and the MSH cells appear very early during the embryonic life.

     

Genetic heterogeneity of propionic acidemia: Analysis of 15 Japanese patients

Summary Propionic acidemia is an autosomal recessive metabolic disease resulting from a deficiency of propionyl CoA carboxylase (PCC) activity. We have analyzed the molecular heterogeneity of Japanese propionic acidemia patients using anti-human PCC antiserum and cDNA clones coding for the two protein subunits ( and ) of the enzyme. The steady state levels of both and subunits of PCC from 15 Japanese patients were determined by Western blot. Three patients had neither nor subunits, and the amounts of both and subunits were low in 3 other patients. According to our previous data, we classified these 6 patients as having subunit deficiency. In the remaining 8 patients, subunits were normal, but the subunits were aberrant. Two patients had low levels of normal-sized subunits and 6 had subunits smaller than normal in size and greatly reduced in quantity. These 8 patients were assigned to the subunit deficiency category. One patient had apparently normal and subunits. We could not determine this patient's primary defect. These data reveal the genetic heterogeneity of molecular defects causing propionic acidemia in the Japanese. Southern blot analysis did not reveal any gross alteration in gene structure when DNA was digested withHindIII,EcoRI andTaqI. However, DNA from 3 -subunit-deficient patients, when digested withMspI and probed with PCC cDNA, revealed a unique 2.7-kb band not observed in blots of DNA from any other patient or 15 normal controls. We conclude that this alteredMspI restriction map is the result of a mutation in the subunit gene of these patients.     

Synthesis of selectively radiolabeled hexasaccharides for the determination of enzymatic regioselectivity

Poly-N-acetyllactosamines provide backbone structures for functional modifications such as sialyl Lewis X. To understand how the biosynthesis of poly-N-acetyllactosamines is regulated, two branched oligosaccharides of the structure Gal1,4GlcNAc1, 6(Gal1,4GlcNAc1,2)-Man1,6Man-octyl 1 and 2 were synthesized in which one of the terminal galactose units was selectively radiolabeled. Hexasaccharides 1 and 2 were assembled from the chemically synthesized pentasaccharide precursors GlcNAc1,6(Gal1,4GlcNAc1,2)-Man1,6Man-octyl3 and Gal1,4GlcNAc1,6(GlcNAc1, 2) - Man1,6 Man-octyl 4 respectively, through treatment with UDP-1-[³H]-Gal and 1,4 galactosyltransferase. Compounds 1 and 2 were subsequently incubated with UDP-GlcNAc and the UDP-GlcNAc: Gal1-4Glc(NAc)1,3-N-acetylglucosaminyltransferase (i-GlcNAc transferase) resulting in a partial conversion to a mixture of heptasaccharides which were purified by HPLC. The branch selectivity of the addition of N-acetylglucosamine to compounds 1 and 2 was then characterized by endo--galactosidase digestion of the heptasaccharides, followed by isolation of the resultant pentasaccharides on C18 reverse-phase silica cartridges. Comparison of the amount of radiolabel to a control reaction lacking endo--galactosidase indicated the favored site of GlcNAc addition to be the lower 1,2-branch over the 1,6-branch by a 3:1 ratio.     

Feedback regulation of RNA polymerase subunit synthesis after the conditional overproduction of RNA polymerase in Escherichia coli

Summary The and subunit of RNA polymerase are thought to be controlled by a translational feedback mechanism regulated by the concentration of RNA polymerase holoenzyme. To study this regulation in vivo, an inducible RNA polymerase overproduction system was developed. This system utilizes plasmids from two incompatibility groups that carry RNA polymerase subunit genes under lac promoter/operator control. When the structural genes encoding the components of core RNA polymerase (, and ) or holoenzyme (, , and ⁷⁰) are present on the plasmids, induction of the lac promoter results in a two fold increase in the concentration of functional RNA polymerase. The induction of RNA polymerase overproduction is characterized by an initial large burst of synthesis followed by a gradual decrease as the concentration of RNA polymerase increases. Overproduction of RNA polymerase in a strain carrying an electrophoretic mobility mutation in the rpoB gene results in the specific repression of synthesis off the chromosome. These results indicate that RNA polymerase feedback regulation controls synthesis in vivo.     

Steroid Polyols from the Far Eastern Starfish Henricia sanguinolenta and H. leviuscula leviuscula

Two new asterosaponins, (20R)-3-O--D-(2-O-methylxylopyranosyl)-24-propylcholest-4-ene-3,6,8,15,16,29-hexaol (sanguinoside A) and (20R,24S)-3-O--D-(2,3,4-tri-O-methylxylopyranosyl)-5-cholestane-3,4,6,8,15,24-hexaol (sanguinoside B), were isolated from two species of Pacific Far Eastern Starfish Henricia sanguinolenta and H. leviuscula leviuscula, collected in the Sea of Okhotsk. Both glycosides contain aglycones with pentahydroxysteroid nuclei of similar structures, which are substituted at the 3-hydroxy group with differently methylated -D-xylosyl residues. Sanguinoside A has an unusual structure of its aglycone side chain, whereas sanguinoside B has a unique permethylated carbohydrate chain. In addition, laevisculoside G, a known glycoside, was identified in the H. leviuscula starfish. The structures of the isolated glycosides were established by interpreting their spectral data and by comparing their spectral characteristics with those of known compounds.     

The influence of mutations upon the synthesis of RNA polymerase subunits in Escherichia coli cells

Summary The influence of mutations in structural genes of and subunits of RNA polymerase upon the synthesis of these subunits in E. coli cells have been investigated. An amber-mutation ts22 in the subunit gene decreases the intracellular concentration of this subunit and the rate of its synthesis. At the same time the concentration and the rate of subunit synthesis is increased. These suggest the compensatory activation of the RNA polymerase operon that takes place under the conditions of shortage of one of the subunits. Reversions, as well as more effective supression of ts22 amber mutation, achieved by streptomycin addition, substitution of su2 by su1, or by specific mutations, result in a rise of and drop of subunit concentration and synthesis in ts22 mutant. TsX missense-mutation in the subunit gene alters the properties of the enzyme increasing, at the same time, the concentration and the rate of synthesis of both and subunits, particularly at a nonpermissive temperature. This points to an inversely proportional relationship between the rate of synthesis of RNA polymerase subunits and the total intracellular activity of the enzyme. Extra subunits are rapidly degraded in ts22 and tsX mutants.The whole complex of our data and those of others suggest that the regulation of the synthesis of RNA polymerase subunits is accomplished by interaction of a negative and a positive mechanisms of regulation which include not only activators and repressors but the enzyme itself as well.     

Ganglioside-cholera toxin interactions: a binding and lipid monolayer study

Summary On t.l.c. plates ¹²⁵I-cholera toxin binds to a disialoganglioside tentatively identified as GD_lb with about 10 times less capacity than to ganglioside GM₁. Binding of labeled toxin to both gangliosides was abolished in presence of excess amounts of unlabeled B subunit. Ganglioside extracts from human or pig intestinal mucosa showed toxin binding to gangliosides GM₁ and GD_1b. In ganglioside-containing lipid monolayers the penetration of the toxin was independent of the ganglioside binding capacity.Abbreviations GM₂ Gal-NAc14Gal(3-2NeuAc)14G1c1Cer - GM₁ Gal3Ga1-NAc14Gal(32NeuAc)14G1c11Cer - GD_1a NeuAc23Ga113Gal-NAc14Gal(32NeuAc)14G1c11Cer - GD1b Gall3Gal-NAcl4Gal(32NeuAc82NeuAc)14Glc11Cer - GT_1b NeuAc23Ga113Ga1-NAcal4Gal(3-2NeuAc82NeuAc)14G1c11Cer - dpPC 1,2-hexadecanoyl-sn-glycero-3-phosphocholine - dpPE 1,2-hexadecanoyl-sn-glycero-3-phosphoethanolamine     

Immunohistochemical localization of FSH and LH in the pars distalis of vervet (Cercopithecus aethiops) and babbon (Papio hamadryas) pituitaries

Summary Antisera against oLH¹, oLH and hFSH were used to localize gonadotropic cells in the pars distalis of Cercopithecus aethiops and Papio hamadryas. Three separate cell types were observed for FSH and LH: 85% of immunohistochemically identified gonadotropic cells reacted to all the various antisera; 10% reacted with the anti-LH antibody only; and 5% with the anti-hFSH antibody only. Comparisons between adjacent serial sections treated with various antisera, other than anti-gonadotropic hormones, demonstrated that the gonadotropic cells of these monkeys did not respond to these antisera.

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Résumé Des anticorps anti-LH ovine, anti-LH ovine et anti-FSH humaine ont été utilisés pour localiser les cellules gonadotropes dans la pars distalis de l'hypophyse des Singes Cercopithecus aethiops et Papio hamadryas. Trois catégories cellulaires distinctes, réagissant avec des anticorps anti-hormones gonadotropes, ont été observées. 85% des cellules immunoréactives identifiées en tant que cellules gonadotropes réagissent simultanément avec les différents anticorps mentionnés; 10% des cellules gonadotropes réagissent seulement avec l'anticorps anti-oLH et 5% de ces cellules seulement avec l'anticorps anti-hFSH. La comparaison avec des coupes adjacentes traitées par divers anticorps autres que les anticorps anti-gonadotropines prouve que les cellules gonadotropes de ces Singes ne réagissent jamais simultanément avec l'un ou l'autre de ces anticorps.

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Abbreviations used in this Article oLH ovine luteinizing hormone - hFSH human follicle stimulating hormone - ACTH corticotropin - GH growth hormone - LPH lipotropin - TSH thyrotropin     

Comparison of the apoptotic processes induced by the oxysterols 7β-hydroxycholesterol and cholesterol-5β,6β-epoxide

Oxysterols have been shown to induce apoptosis in a variety of cell lines. The mechanism of oxysterol-induced apoptosis is mainly known at the post-mitochondrial level. The aim of the present study was to compare the pathway of apoptosis induced by the oxysterols 7-hydroxycholesterol (7-OH) and cholesterol-5,6-epoxide (-epoxide) in U937 cells. To this end, we employed a range of inhibitors of apoptosis; a broad-spectrum caspase inhibitor, a specific caspase-3 inhibitor and an inhibitor of cytochromec release and the antioxidants; trolox, ebselen and resveratrol. The three inhibitors of apoptosis prevented cell death induced by 7-OH; however, in -epoxide-treated cells, the inhibitor of cytochromec release did not protect against apoptosis. The cellular antioxidant glutathione was depleted in 7-OH-treated cells but not in cells incubated with -epoxide. Trolox, a water-soluble synthetic analogue of -tocopherol, prevented 7-OH-induced apoptosis but did not protect against cell death induced by -epoxide. Ebselen and resveratrol did not protect U937 cells against apoptosis induced by either 7-OH or -epoxide. Our results suggest that differences occur in the pathways of apoptosis induced by 7-OH and -epoxide in U937 cells.     

Detection and partial characterization of two antigenically distinct β-amylases in developing kernels of wheat

A -amylase (EC 3.2.1.2) was identified in the outer pericarp (P) of developing seeds of wheat (Triticum aestivum L.) and compared with the well known -amylase which is synthesized during seed development in the starchy endosperm (E). The enzyme P already exists in the tissues before anthesis and vanishes at the time when E starts to accumulate. The isoelectric-focusing patterns of P and E are very similar. The relative molecular weight (M_r) of P is slightly higher than that of E (66 and 64.5 kDa, respectively). Both P and E exhibit common epitopes in addition to epitopes specific for each of them. The two enzymes were identified in small amounts in the green tissues of the developing seeds (inner pericarp and testa). No antigenic difference was detected between P and the -amylases of roots and leaves.Abbreviations P pericarp -amylase - E endosperm -amylase - IS1 anti--amylase immune serum - IS2 anti- and anti- amylase immune serum - IS3 anti- amylase immune serum - IEF isoelectric focusing - IgG immunoglobulin G The authors thank Dr. P. Ziegler (Universität Bayreuth, FRG) for stimulating discussion and for useful suggestions during the writing of the text. The authors thank Miss C. Mayer for her skillful technical assistance.