[Influence of 20 % sulfur hexafluoride (SF6) on human corneal endothelial cells : An in vitro study]. / Einfluss von 20 % Schwefelhexafluorid (SF6) auf humane korneale Endothelzellen : Eine In-vitro-Studie.

BACKGROUND: In posterior lamellar keratoplasties, such as Descemet membrane endothelial keratoplasty (DMEK) and Descemet's stripping automated endothelial keratoplasty (DSAEK) an air bubble is left inside the anterior chamber to promote graft attachment during the early postoperative period. In the case of insufficient graft adhesion a renewed intracameral air injection is often necessary. The use of sulfur hexafluoride diluted with air (SF6 20 %) as an alternative to pure air may further enhance graft attachment and reduce the rebubbling rate. The effect of SF6 20 % on corneal endothelium is currently unclear and was therefore examined in vitro. MATERIAL AND METHODS: For this study 12 human corneoscleral discs were mounted in artificial anterior chambers, the systems were continuously filled with culture medium and the anterior chambers with air (n = 5) or SF6 20 % (n = 7) as tamponade. After 6 days of storage in the incubator endothelial cell density, toxicity on endothelial cells and corneal thickness were evaluated. RESULTS: There were no significant differences in endothelial cell loss (p = 1.000), endothelial cell count (p = 0.648), toxicity on endothelial cells (p = 0.048) and central corneal thickness (p = 0.905) between the two groups after 1 week. The level of significance was defined as p ≤ 0.05 and adjusted to p ≤ 0.0056 according to the Bonferroni correction for multiple testing. CONCLUSION: The use of SF6 20 % as tamponade in the anterior chamber for posterior lamellar keratoplasty can be proposed as a safe alternative to pure air filling related to endothelial cell loss. Increased toxic effects on the corneal endothelium by SF6 20 % were not detected in this study; however, further prospective clinical trials are needed to examine the long-term effects in humans.

Acute effects of short-chain alkylglycerols on blood-brain barrier properties of cultured brain endothelial cells.

BACKGROUND AND PURPOSE: The blood-brain barrier (BBB) restricts drug penetration to the brain preventing effective treatment of patients suffering from brain tumours. Intra-arterial injection of short-chain alkylglycerols (AGs) opens the BBB and increases delivery of molecules to rodent brain parenchyma in vivo. The mechanism underlying AG-mediated modification of BBB permeability is still unknown. Here, we have tested the effects of AGs on barrier properties of cultured brain microvascular endothelial cells. EXPERIMENTAL APPROACH: The effects of two AGs, 1-O-pentylglycerol and 2-O-hexyldiglycerol were examined using an in vitro BBB model consisting of primary cultures of rat brain endothelial cells, co-cultured with rat cerebral glial cells. Integrity of the paracellular, tight junction-based, permeation route was analysed by functional assays, immunostaining for junctional proteins, freeze-fracture electron microscopy, and analysis of claudin-claudin trans-interactions. KEY RESULTS: AG treatment (5 min) reversibly reduced transendothelial electrical resistance and increased BBB permeability for fluorescein accompanied by changes in cell morphology and immunostaining for claudin-5 and ß-catenin. These short-term changes were not accompanied by alterations of inter-endothelial tight junction strand complexity or the trans-interaction of claudin-5. CONCLUSION AND IMPLICATIONS: AG-mediated increase in brain endothelial paracellular permeability was short, reversible and did not affect tight junction strand complexity. Redistribution of junctional proteins and alterations in the cell shape indicate the involvement of the cytoskeleton in the action of AGs. These data confirm the results from in vivo studies in rodents characterizing AGs as adjuvants that transiently open the BBB.

[Heavy tamponades--background and future perspectives]. / Schwere Tamponaden - Hintergrund und Zukunftsperspektiven.

Proliferative vitreoretinopathy in the inferior retina remains clinically challenging. Heavier-than-water intraocular tamponades have been developed to improve inferior tamponading properties. The parameters of an optimal intraocular tamponade are defined and the influences of the specific gravity, buoyancy, interfacial tension, and viscosity are discussed. Perfluorocarbon liquids and partially fluorinated alkanes were associated with tamponade emulsification, intraocular inflammation, and rises in intraocular pressure that were less prominent in admixtures of these substances with silicone oil (heavy silicone oils). Two recently developed heavy silicone oil tamponades, Oxane HD and Densiron 68, are well tolerated and have entered clinical practice. The side effects are associated with the chemical properties of the tamponading agent and seem comparable to those seen with conventional silicone oil. Heavy silicone oil tamponade improves inferior tamponade and may be considered as a new generation of intraocular tamponades.

Downregulation of Apaf-1 and caspase-3 by RNA interference in human glioma cells: consequences for erucylphosphocholine-induced apoptosis.

Erucylphosphocholine (ErPC) exerts strong anticancer activity in vivo and in vitroand induces apoptosis even in chemoresistant glioma cell lines. We investigated the contribution of Apaf-1 and caspase-3 to the apoptotic response to ErPC using RNA interference (RNAi) in human glioblastoma cells. We could demonstrate that human glioma cell lines are susceptible to RNAi. Apaf-1 and caspase-3 are amenable to specific small interfering RNA (siRNA)-induced degradation resulting in a reduction of protein levels to 8-33% (Apaf-1) and to 30-50% (caspase-3). Transfection of siRNA directed to Apaf-1 and caspase-3 specifically reduced caspase-3 processing induced by ErPC treatment and yielded a reduction in cells that undergo ErPC-induced apoptosis to 17-33% (Apaf-1) and to 38-50% (caspase-3). The caspase-3 siRNA experiments were corroborated in caspase-3-deficient and -reconstituted MCF-7 breast cancer cells. Survival assays and morphological observations revealed that caspase-3 reconstitution significantly sensitized MCF-7 cells to ErPC. Exploring the caspase cascade responsible for ErPC-induced apoptosis MCF-7 cells provided evidence that caspase-3 is required for the activation of caspases-2, -6 and -8 and also participates in a feedback amplification loop. Our results provide evidence that Apaf-1 and caspase-3 are major determinants of ErPC-induced apoptosis and the possible use of ErPC in a clinical setting is discussed.

Induction of differentiation and tetraploidy by long-term treatment of C6 rat glioma cells with erucylphosphocholine.

Induction of differentiation represents a promising concept for chemotherapy of malignant gliomas, which are often refractory even to the combined treatment with surgery, irradiation and chemotherapy. Since anti-neoplastic alkylphosphocholines can induce differentiation of leukemic cell lines, the effects of the intravenously applicable alkylphosphocholine-derivative erucylphosphocholine (ErPC) on proliferation, morphology and differentiation of the rat glioma cell line C6 was examined in vitro. Short-term exposure to ErPC induced accumulation of the cells in the G2/M-phase of the cell cycle and apoptotic cell death. In contrast, continuous exposure of C6 rat glioma cells to sublethal ErPC doses (30 and 50 microM) caused both the formation of a slower growing tetraploid cell population and astrocytic differentiation. No resistance to in vivo obtainable ErPC concentrations was observed during this treatment. We conclude that ErPC-induced differentiation might be beneficial for a long-term adjuvant chemotherapy of low grade glioma.

Post-transcriptional effects of interleukin-3, interferon-gamma, erythropoietin and butyrate on in vitro hemoglobin chain synthesis in congenital hemolytic anemia.

BACKGROUND AND OBJECTIVES: Various agents modulate hemoglobin synthesis. In vitro modulation of translation in hemoglobin chain synthesis was analysed in patients with congenital hemolytic anemia (n=32) and healthy controls (n=17). DESIGN AND METHODS: Enriched reticulocytes were co-incubated with (3)H-leucine and cytokines or butyrate. Reversed-phase chromatography enabled separation of alpha-, beta- and gamma-globin chains. Globin chain synthesis was calculated from measured (3)H-leucine incorporation. Transferrin, erythropoietin, interleukin-3 and interferon-gamma receptors were detected by flow cytometry. Reverse-transcription polymerase chain reaction (RT PCR) was used to demonstrate changes of RNA stability. RESULTS AND DISCUSSION: Interleukin-3, interferon-gamma and butyrate caused a significant 2-fold increase (range 1.8-2.4; p<0.01) of the alpha- and beta-chain synthesis in congenital hemolytic anaemias. Analysis of gamma-globin chain synthesis revealed a lower, i.e. 1.4 fold increase (range 1.32 to 1.41; p<0.03). The absolute amount of globin synthesis was calculated to be 2.9 x 10(-12) g/reticulocyte/24h. After incubation with interleukin-3 the absolute additional synthesis of the alpha-globin chain reached 1.31 x 10(-12) g/reticulocyte/24h, of the beta-globin chain, 1.15 x 10(-12) g/reticulocyte/24h and of the gamma-globin chain, 0.26 x 10(-12) g/reticulocyte/24h. Butyrate and interferon-gamma had no or even an inhibiting effect on reticulocytes from normal controls, while interleukin-3 stimulated alpha- and gamma-chain synthesis (1.4 and 2.4 fold, respectively; p<0.03) suggesting an increase of fetal hemoglobin (HbF). Erythropoietin showed no stimulating influence. Membrane associated interleukin-3 receptors were detected in 0.78+/-0.14%, and interferon-gamma receptors in 0.1+/-0.015% of the red cells. Erythropoietin receptors were extremely rare (0.05+/-0.015%). The expression of transferrin receptors (CD71) correlated with the extent of globin chain stimulation. The alpha-, and beta-globin mRNA content of the reticulocytes after interleukin-3 incubation, as measured by RT-PCR, increased. INTERPRETATION AND CONCLUSIONS: Hemoglobin chain synthesis could be modulated post-transcriptionally by interleukin-3, interferon-gamma and butyrate. Transferrin receptor and globin RNA stability might be involved in this phenomenon.

Molecular basis of recessive congenital methemoglobinemia, types I and II: Exon skipping and three novel missense mutations in the NADH-cytochrome b5 reductase (diaphorase 1) gene.

Hereditary methemoglobinemia due to reduced nicotin amide adenine dinucleotide (NADH)-cytochrome b5 reductase (b5r) deficiency is classified into an erythrocyte type (I) and a generalized type (II). We investigated the b5r gene of three unrelated patients with types I and II and found four novel mutations. The patient with type I was homozygous for a c.535 G-->A exchange in exon 6 (A179T). The patients with type II were found to be homozygous for a c.757 G-->A transition in exon 9 (V253M) and compound heterozygous for two mutations, respectively. One allele presented a c.379 A-->G transition (M127V). The second allele carried a sequence difference at the invariant 3' splice-acceptor dinucleotide of intron 4 (IVS4-2A-->G) resulting in skipping of exon 5. To characterize a possible effect of this mutation on RNA metabolism, poly(A)(+) RNA was analyzed by RT-PCR and sequencing. The results show that RNA is made from the allele harboring the 3'-splice site mutation. Furthermore, western blot analysis revealed a complete absence of immunologically detectable b5r in skin fibroblasts of this patient. The compound heterozygosity for the splice site and the missense mutations apparently caused hereditary methemoglobinemia type II in this patient. Hum Mutat 17:348, 2001.

Erucylphosphocholine-induced apoptosis in chemoresistant glioblastoma cell lines: involvement of caspase activation and mitochondrial alterations.

Intrinsic chemoresistance constitutes a major problem in the therapy of malignant gliomas. In vitro experiments with four astrocytoma/glioblastoma (AC/GBM) cell lines revealed that the chemoresistance of two cell lines, A172 and T98G, to cisplatin and etoposide was due to resistance to drug-induced apoptosis. In contrast, all the AC/GBM cell lines tested were sensitive to treatment with the lipophilic ether lipid erucylphosphocholine, ErPC. ErPC-induced apoptosis was independent of wild-type p53-signaling and triggering of the CD95/CD95 ligand (CD95L) system. Inhibition of protein and RNA synthesis by cycloheximide and actinomycin D did not abrogate ErPC-induced apoptosis. However, expression of members of the bcl-2 protein family was modulated during ErPC treatment. Activation of caspase 3 and mitochondrial alterations were central to ErPC-induced apoptosis. We conclude that ErPC-induced activation of the mitochondrial pathway enables cell death in the chemoresistant AC/GBM cells.

Glucose-6-phosphate isomerase deficiency.

Most of the metabolic needs of erythrocytes are covered by glycolysis, the oxidative pentose phosphate pathway and the glutathione cycle. Hereditary enzyme deficiencies of all these pathways have been identified, among which glucose-6-phosphate isomerase (GPI) deficiency is the second most frequent erythroenzymopathy in glycolysis, being associated with non-spherocytic haemolytic anaemia of variable severity. This autosomal recessive genetic disorder may be associated in some cases with neurological impairment. GPI is a dimeric enzyme that catalyses the reversible interconversion of fructose-6-phosphate and glucose-6-phosphate. Virtually all the mutant gene products reported are characterized by marked instability and normal substrate affinities, but altered catalytic activity and electrophoretic migration rates. At the nucleotide level, 29 mutations have been reported. This chapter reviews (a) the clinical pattern of the condition; (b) biochemical and molecular studies; (c) structure-function relationships; (d) the molecular basis of neurological dysfunctions sometimes associated with GPI deficiency; and (e) the correlation between the severity of the anaemia and the molecular defect.

Endocrine status in elderly men with lower urinary tract symptoms: correlation of age, hormonal status, and lower urinary tract function. The Prostate Study Group of the Austrian Society of Urology.

OBJECTIVES: To correlate endocrine parameters in elderly men with lower urinary tract symptoms (LUTS) to patient age and clinical parameters such as prostate volume, prostate-specific antigen (PSA) levels, and uroflowmetry and to compare the clinical and endocrinologic parameters in men with or without hypogonadism. METHODS: Men (40 years old or older) with untreated LUTS as defined by an International Prostate Symptom Score (IPSS) of 7 or greater due to benign prostatic hyperplasia were included in this study and underwent the following investigations: IPSS, free uroflow study, postvoid residual volume, transrectal ultrasound for assessment of prostate volume, serum PSA determination, and an endocrine study, including testosterone, human luteinizing hormone, human follicle-stimulating hormone, prolactin, dehydroepiandrostendione-sulphate (DHEA-S), and prolactin. RESULTS: Three hundred twelve men (mean age 62.8 +/- 10.6 years, range 40 to 91) were analyzed. The serum levels of estradiol (correlation coefficient [r] = 0.19), human luteinizing hormone (r = 0.32), human follicle-stimulating hormone (r = 0.19), and DHEA-S (r = -0.39) correlated (P <0.05) with age; no such correlation was seen for testosterone (r = 0.04; P0.05) or prolactin (r = 0.09; P0.05). Estradiol (but not testosterone) correlated (r = 0.17, P = 0.01) with prostate volume. The peak flow rate and PSA did not correlate with any endocrinologic parameter. Hypogonadism (serum testosterone less than 3.0 ng/mL) was detected in 22.1% of patients and had no impact on clinical (IPSS, peak flow rate, prostate volume, and PSA level) or endocrine (human luteinizing hormone, human follicle-stimulating hormone, estradiol, prolactin, and DHEA-S) parameters. CONCLUSIONS: A number of age-related endocrine changes are seen in elderly men with LUTS. Hypogonadism is seen in approximately one fifth of elderly men with LUTS, but in our study it had no impact on symptom status, PSA level, prostate volume, uroflowmetry, or endocrine parameters.

Eight novel mutations and consequences on mRNA and protein level in pyruvate kinase-deficient patients with nonspherocytic hemolytic anemia.

Pyruvate kinase (PK) deficiency (PKD) is an autosomal recessive disorder with the typical manifestation of nonspherocytic hemolytic anemia. We analyzed the mutant enzymes of 10 unrelated patients with PKD, whose symptoms ranged from a mild, chronic hemolytic anemia to a severe anemia, by sequence analysis for the presence of alterations in the PKLR gene. In all cases the patients were shown to be compound heterozygous. Eight novel mutations were identified: 458T-->C (Ile153Thr), 656T-->C (Ile219Thr), 877G-->A (Asp293Asn), 991G-->A (Asp331Asn), 1055C-->A (Ala352Asp), 1483G-->A (Ala495Thr), 1649A-->T (Asp550Val), and 183-184ins16bp. This 16 bp duplication produces a frameshift and subsequent stop codon resulting in a drastically reduced mRNA level, and probably in an unstable gene product. Surprisingly, the existence of M2-type PK could be demonstrated in the patient's red blood cells. The study of different polymorphic sites revealed, with one exception, a strict linkage of the 1705C, 1738T, IVS5(+51)T, T(10) polymorphisms and the presence of 14 ATT repeats in intron 11. Our analyses show the consequences of a distorted structure on enzyme function and we discuss the correlations between the mutations identified and the parameters indicative for enzyme function.

Identification of a novel promoter mutation in the human pyruvate kinase (PK) LR gene of a patient with severe haemolytic anaemia.

Using direct sequencing we analysed the pyruvate kinase (PK) LR gene of a patient with severe haemolytic anaemia due to PK deficiency. A novel promoter mutation -249delA relative to the translation initiation site and the common 1529A mutation in exon 11 of the gene could be identified. Reverse transcription (RT)-PCR analysis combined with restriction digestion revealed that the -249delA mutation leads to a reduction in the amount of mRNA produced from this allele to about 6% of normal. We assume that both mutations would account for the PK deficiency in the compound heterozygous patient.

Erucylphosphocholine, a novel antineoplastic ether lipid, blocks growth and induces apoptosis in brain tumor cell lines in vitro.

A potential benefit of alkylphosphocholines in brain tumor therapy was evaluated. The in vitro effects of the intravenously applicable erucylphosphocholine (ErPC) on proliferation, viability, morphology and cell cycle distribution of a rat glioma, four human astrocytoma/glioblastoma and a human medulloblastoma cell line were analyzed daily after continuous drug-exposure for up to six days. ErPC exerted strong cytostatic and direct cytotoxic effects on all cell lines tested at drug concentrations that are achieved in the rat brain after repeated intravenous injections of nontoxic drug doses. Concentrations of 70 microM (T98G, A172, 85HG66, 86HG39) and 110 microM (C6, D283 Med) led to complete cell death within 48-96 h. Particular characteristics of ErPC action are i) the accumulation of cells with a 4n DNA content corresponding to the G2/M-phase of the cell cycle, ii) the formation of two- and multinucleated cells and iii) the induction of apoptosis.

Molecular basis of neurological dysfunction coupled with haemolytic anaemia in human glucose-6-phosphate isomerase (GPI) deficiency.

Glucose-6-phosphate isomerase (GPI) deficiency, an autosomal recessive genetic disorder with the typical manifestation of nonspherocytic haemolytic anaemia, can be associated in some cases with neurological impairment. GPI has been found to be identical to neuroleukin (NLK), which has neurotrophic and lymphokine properties. To focus on the possible effects of GPI mutations on the central nervous system through an effect on neuroleukin activity, we analysed DNA isolated from two patients with severe GPI deficiency, one of them with additional neurological deficits, and their families. The neurologically affected patient (GPI Homburg) is compound heterozygous for a 59 A-->C (H20P) and a 1016 T-->C (L339P) exchange. Owing to the insertion of proline, the H20P and L339P mutations are likely to affect the folding and activity of the enzyme. In the second family studied, the two affected siblings showed no neurological symptoms. The identified mutations are 1166 A-->G (H389R) and 1549 C-->G (L517V), which are located at the subunit interface. We propose that mutations that lead to incorrect folding destroy both catalytic (GPI) and neurotrophic (NLK) activities, thereby leading to the observed clinical symptoms (GPI Homburg). Those alterations at the active site, however, that allow correct folding retain the neurotrophic properties of the molecule (GPI Calden).

Ankyrin-1 mutations are a major cause of dominant and recessive hereditary spherocytosis.

Hereditary spherocytosis (HS) is the most common inherited haemolytic anaemia in Northern Europeans. The primary molecular defects reside in the red blood cell (RBC) membrane, particularly in proteins that link the membrane skeleton to the overlying lipid bilayer and its integral membrane constituents. Ankyrin-1 is the predominant linker molecule. It attaches spectrin, the major skeletal protein, to the cytoplasmic domain of band 3, the RBC anion exchanger. Two-thirds of patients with HS have combined spectrin and ankyrin-1 deficiency; deficiency of band 3 occurs in about 15 to 20% (ref.1). These data suggest that ankyrin-1 or band 3 defects may be common in HS. To test this we screened all 42 coding exons plus the 5' untranslated/promoter region of ankyrin-1 and the 19 coding exons of band 3 in 46 HS families. Twelve ankyrin-1 mutations and five band 3 mutations were identified. Missense mutations and a mutation in the putative ankyrin-1 promoter were common in recessive HS. In contrast, ankyrin-1 and band 3 frameshift and nonsense null mutations prevailed in dominant HS. Increased accumulation of the normal protein product partially compensated for the ankyrin-1 or band 3 defects in some of these null mutations. Our findings indicate that ankyrin-1 mutations are a major cause of dominant and recessive HS (approximately 35 to 65%), that band 3 mutations are less common (approximately 15 to 25%), and that the severity of HS is modified by factors other than the primary gene defect.

Nuclear degradation of nonsense mutated beta-globin mRNA: a post-transcriptional mechanism to protect heterozygotes from severe clinical manifestations of beta-thalassemia?

Nonsense mutations of the beta-globin gene are a common cause of beta-thalassemia. It is a hallmark of these mutations not only to cause a lack of protein synthesis but also a reduction of mRNA expression. Both the pathophysiologic significance and the underlying mechanisms for this surprising phenomenon have so far remained enigmatic. We report that the reduction of the fully spliced mutant beta-globin mRNA already manifests itself within the nucleus. In contrast, the levels of mutant pre-mRNA are normal. The promoter and the 5'-untranslated region (5'-UTR) of the herpes simplex virus type 1 thymidine kinase (HSV1 Tk) gene can independently circumvent this recognition/response mechanism in cis and restore nonsense mutated beta-globin mRNA expression to normal levels. These two genetic elements can thus exert a dominant influence on the post-transcriptional control of nonsense mutated beta-globin gene expression. While wild-type mRNA levels are restored by fusion of the HSV1 Tk 5'-UTR to the nonsense mutated beta-globin reading frame, translation of a wildtype reading frame in such a hybrid is precluded. In contrast, the HSV1 Tk promoter appears to efficiently deliver the mRNA to the translational apparatus. The 5'-UTR and the promoter sequences therefore control the nuclear fate of nonsense mutated beta-globin mRNA by separable pathways. The nuclear mRNA degradation mechanisms examined here may prevent the synthesis of C-terminally truncated beta-globin chain fragments and may protect heterozygotes from clinically relevant symptoms of beta-thalassemia.

Determination of mRNA fate by different RNA polymerase II promoters.

Translational stop mutations of the human beta-globin gene cause a reduction of cytoplasmic mRNA accumulation in thalassemia patients and in transfection models. The exact mechanism underlying this phenomenon has remained enigmatic but is known to be post-transcriptional. We have used transfected HeLa cells to study the expression of beta-globin mRNAs with nonsense or frameshift mutations within the three exons of this gene. Mutations in exons 1 or 2 reduce cytoplasmic mRNA accumulation whereas a mutation in exon 3 permits essentially normal expression. We report here that the post-transcriptional fate of mutated beta-globin mRNAs is differentially affected by the type of RNA polymerase II promoter driving expression. Replacement of the beta-globin promoter with the herpes simplex virus type 1 thymidine kinase gene promoter but not the cytomegalovirus immediate early promoter rescues the cytoplasmic accumulation of mutated mRNA to wild-type levels. This effect is shown to be independent of the absolute quantity and the kinetics of accumulation of mutated mRNA synthesized, and primer-extension analyses confirm that both viral promoters accurately utilize identical transcription start sites. These data thus reveal an unexpected property of RNA polymerase II promoters: determination of the post-transcriptional fate of the maturing mRNA, presumably by influencing alternative choices between as yet undefined processing and/or transport pathways.

Liver-specific gene expression: A-activator-binding site, a promoter module present in vitellogenin and acute-phase genes.

The A2 vitellogenin gene of Xenopus laevis, which is expressed liver specifically, contains an A-activator-binding site (AABS) that mediates high in vitro transcriptional activity in rat liver nuclear extracts. Footprint experiments with DNase I and gel retardation assays revealed the binding of several proteins to AABS. Using binding sites of known DNA-binding proteins as competitors in the gel retardation assay, we found that the transcription factor C/EBP and/or one of its "iso-binders" as well as LFB1/HNF1 bound AABS. These interactions were confirmed by in vitro transcription experiments using various oligonucleotides as competitors. However, saturating amounts of C/EBP- and LFB1/HNF1-binding sites as competitors only partially blocked AABS-mediated transcriptional activity. This finding implies that at least a third distinct transcription factor interacts with AABS. In vitro transcription experiments revealed that AABS was present not only in the closely related Xenopus A1 vitellogenin gene but also in acute-phase genes as a liver-specific regulatory element known to confer the interleukin-6 response. Both AABS and the interleukin-6 response element are promoter modules interacting with at least three distinct transcription factors, including C/EBP and LFB1/HNF1.

[Continent coronary diversion--Mainz Pouch. 3-year personal experience]. / Kontinente Harnableitung--Mainz-Pouch. Eigene Erfahrungen über 3 Jahre.

Our experiences with the continent urinary diversion and the bladder substitution by means of the Mainz-Pouch technique are reported. We were one of the first Urological Departments in Austria to introduce this technique in late 1987. We present the operative technique, the patient material of 15 patients and their outcome with early and late complications over a follow up period of 16.6 months.

BAP, a rat liver protein that activates transcription through a promoter element with similarity to the USF/MLTF binding site.

The vitellogenin genes of Xenopus are liver-specifically expressed. An in vitro transcription system derived from rat liver nuclei allowed us to define the cis-element BABS (B-activator binding site) in the promoter of the B1 vitellogenin gene. An oligonucleotide encompassing the region from -53 to -44 linked to a TATA box is sufficient for a tenfold increase of the transcriptional activity. Gel retardation assays with nuclear rat liver proteins reveal two DNA-protein complexes: Complex 1 can be competed by the USF/MLTF binding site of the adeno major late promoter whereas complex 2 is a distinct protein we refer to as BAP (B-activator protein). In vitro transcription experiments in the presence of USF/MLTF binding site as competitor show that BAP is an efficient transcription factor. Based on UV cross-linking we estimate that BAP has a molecular weight of 58 kd. Phosphatase treatment reveals that DNA binding of BAP requires phosphorylation. BABS is also present in the hepatitis B virus enhancer suggesting that it might play a role in the tumorigenic potential of the virus.