Biomedical subjects

H Busch

Publications and source records attributed to H Busch.

At least 37 records · Page 2Linked to original sources

PubMed1997

Nucleolar coefficient and cytochemistry of human blood monocytes.

Nucleoli in peripheral blood monocytes represented mostly by micronucleoli were investigated by means of cytochemical procedures for the demonstration of RNA, proteins of silver stained nucleolus organizer region, nucleophosmin, nucleolin and fibrillarin. The values of the nucleolar coefficient (mean number of nucleoli per cell) were influenced by the procedure used for the visualization of nucleoli. The differences in the values of the nucleolar coefficient of monocytes in one and the same person using above mentioned procedures suggested that micronucleoli in blood monocytes represent heterogeneous nuclear structures and only some of them possess both RNA and characteristic nucleolar proteins. Therefore, the procedures used for the visualization of nucleoli must be always taken into consideration when the values of the nucleolar coefficient are evaluated.

Adult↗

PubMed1997

Adducts and tamoxifen.

In a variety of organs and cells, multiple types of DNA-adducts have been identified before and after treatment with carcinogens. Evidence has been presented that tamoxifen DNA-adducts in treated rat liver cells are related to its rat liver carcinogenicity. Specific metabolites of tamoxifen have been reported to be proximal carcinogens in rat liver, particularly the alpha-hydroxy tamoxifen and possibly other oxygenated metabolites. Similar analyses of human liver and human endometrium of patients treated with tamoxifen did not provide evidence for the presence of tamoxifen-related metabolites. Some preliminary data suggest that tamoxifen may reduce the background DNA-adducts in some human tissues. Inasmuch as there was no increase in tamoxifen-related adducts in human liver samples from tamoxifen-treated patients, it was not surprising that there is no consistent clinical evidence for tamoxifen-related human liver cancer. The lack of tamoxifen adducts in human endometrium indicates that if tamoxifen is a human endometrial carcinogen (International Agency for Research on Cancer, unpublished observations), its mechanism of carcinogenicity is different from that for rat liver, in which DNA-adduct formation has been readily demonstrated.

Animals↗

PubMed1996

Changes in voltage activation, Cs+ sensitivity, and ion permeability in H5 mutants of the plant K+ channel KAT1.

KAT1 is a voltage-dependent inward rectifying K+ channel cloned from the higher plant Arabidopsis thaliana [Anderson, J. A., Huprikar, S. S., Kochian, L. V., Lucas, W. J. & Gaber, R. F. (1992) Proc. Natl. Acad. Sci. USA 89, 3736-3740]. It is related to the Shaker superfamily of K+ channels characterized by six transmembrane spanning domains (S1-S6) and a putative pore-forming region between S5 and S6 (H5). The 115 region between Pro-247 and Pro-271 in KAT1 contains 14 additional amino acids when compared with Shaker [Aldrich, R. W. (1993) Nature (London) 362, 107-108]. We studied various point mutations introduced into H5 to determine whether voltage-dependent plant and animal K+ channels share similar pore structures. Through heterologous expression in Xenopus oocytes and voltage-clamp analysis combined with phenotypic analysis involving a potassium transport-defective Saccharomyces cerevisiae strain, we investigated the selectivity filter of the mutants and their susceptibility toward inhibition by cesium and calcium ions. With respect to electrophysiological properties, KAT1 mutants segregated into three groups: (i) wild-type-like channels, (ii) channels modified in selectivity and Cs+ or Ca2+ sensitivity, and (iii) a group that was additionally affected in its voltage dependence. Despite the additional 14 amino acids in H5, this motif in KAT1 is also involved in the formation of the ion-conducting pore because amino acid substitutions at Leu-251, Thr-256, Thr-259, and Thr-260 resulted in functional channels with modified ionic selectivity and inhibition. Creation of Ca2+ sensitivity and an increased susceptibility to Cs+ block through mutations within the narrow pore might indicate that both blockers move deeply into the channel. Furthermore, mutations close to the rim of the pore affecting the half-activation potential (U1/2) indicate that amino acids within the pore either interact with the voltage sensor or ion permeation feeds back on gating.

Amino Acid Sequence↗

PubMed1996

A nucleolar RNA helicase recognized by autoimmune antibodies from a patient with watermelon stomach disease.

Watermelon stomach is characterized by prominent stripes of ectatic vascular tissue in the stomach similar to stripes on a watermelon; in patients with this disorder chronic gastrointestinal bleeding occurs and approximately half of these patients have associated autoimmune disorders. In the serum of one patient, an antinucleolar antibody titer of 1:25 600 was found; the antibodies specifically recognized an approximately 100 kDa nucleolar protein, which we referred to as the 'Gu' protein. Its cDNA was cloned and sequenced. The Gu protein is a member of a new subgroup of RNA helicases, the DEXD box family. Gu protein fused with glutathione S-transferase contains ATP-dependent RNA helicase activity which preferably translocates in the 5'-->3' direction. Its RNA folding activity, RNA-dependent ATPase and dATPase activities, and its translocation direction are similar to those of RNA helicase II [Flores-Rozas, H. and Hurwitz, J. (1993) J. Biol. Chem. 268, 21372-21383]. Sequencing of 209 amino acids of RNA helicase II peptides showed 96.7% identity with the cDNA-derived amino acid sequence of the Gu protein. The precise biological roles of this RNA helicase in the biogenesis of ribosomal RNA and the pathogenesis of watermelon disease and autoimmune disorder require further study.

Adenosine Triphosphatases↗

PubMed1996

C23 interacts with B23, a putative nucleolar-localization-signal-binding protein.

The human protein C23 (nucleolin) is a major nucleolar protein. Its interactions with other proteins were studied with the two-hybrid system which identified nucleolar protein B23 (nucleophosmin) as being associated with C23. Both proteins were co-immunoprecipitated from HeLa cell nuclear extract by either monoclonal anti-C23 or monoclonal anti-B23. Binding studies utilizing deletion mutants indicated that the binding of C23 and B23 involves specific motifs. In addition to an approximately 46-amino-acid-binding domain in B23 (amino acids 194-239), amino acids 540-628 of C23 were required for binding; this region of C23 is required for the nucleolar localization. In addition, nucleolar protein p120 was also found to be co-immunoprecipitated with B23. A fragment of p120 containing a functional nucleolar localization signal bound to the truncated binding domain of B23, as did C23. These results suggest that the interaction of C23 and B23 may represent a nucleolar-targeting mechanism in which B23 acts as a nucleolar-localization signal-binding protein.

Glutathione Transferase↗

PubMed1996

Overexpression of human nucleolar proteins in insect cells: characterization of nucleolar protein p120.

Nucleolar p120 is a proliferation-associated protein, which becomes detectable early in the G1 phase of the cell cycle and peaks early in the S phase. A variety of human malignant tumor cells contain much higher levels of p120 than normal resting cells. The cellular functions of p120 are unknown, and little information is available on the structural characteristics of the human p120 protein. For biochemical characterization, human p120 protein was expressed in a baculovirus system and purified to approximately 95% purity. By indirect immunofluorescence, most of the recombinant human p120 as well as recombinant human B23, C23, or fibrillarin were localized to insect cell nucleoli and to large globular nuclear inclusions. Like endogenous p120 in HeLa cells, recombinant p120 expressed in insect cells was phosphorylated. On sucrose density gradients, p120 from HeLa cells sedimented in the 60-80S region, in which preribosomal particles sedimented using similar extraction and centrifugation procedures. The sedimentation of p120 shifted to the 5-10S region by treatment with 1 M KCl or with RNAse which suggests that p120 is bound to RNA.

Amino Acid Sequence↗

PubMed1996

[Radiological thorax monitoring in ventilated intensive-care patients].

PURPOSE: At many institutions chest x-rays (CXR) are routinely performed on mechanically ventilated patients on a daily basis. Among the aims of this study was the assessment of a) frequency of clinically relevant findings from routine CXR in surgical patients under mechanical ventilation, b) comparison of expectations of clinically important CXR findings with estimation of clinical relevance of findings once films were obtained, c) indications and clinical consequences of findings from CXR requested in addition to routine films. METHODS: Prospective study, 40 patients with 741 CXR. Standardised film reporting. Daily recording of clinical data by means of an evaluation score. Interview of intensive-care physicians for assessment of: what information was expected from any routine CXR? did CXR offer helpful information for further treatment planning? RESULTS: 26% of routine CXR had a considerable therapeutical impact. 43% of all routine CXR were expected to have an influence on patient management, 57% were not. 16% of those CXR that had been expected not to offer essential information, disclosed relevant findings. Of the additionally obtained CXR, 53% were performed to check position of endotracheal tube and catheters and 47% related to other indications. CONCLUSION: Critically ill surgical patients under mechanical ventilation should have daily CXR follow-up, as one-fourth of these films has an impact on the patient's management.

APACHE↗

PubMed1996

[Pharmacopsychiatric guidelines for treatment of alcohol withdrawal syndrome].

The therapy of the alcohol withdrawal syndrome is very heterogeneous with more than 100 drug combinations. This results from differently qualified physicians and from insufficient pharmacostudies concerning evaluation and psychopathological methodology of the alcohol withdrawal syndrome. The following advices can be given: Physical withdrawal should take place under the conditions of an appropriate hospital with a qualified team. The application of electrolytes and vitamines (B1) may be necessary in alcohol addicts. Less severe alcohol withdrawal syndromes are treated sufficiently with carbamazepine, sometimes it can be necessary to combine it with neuroleptics or benzodiazepines. A secure prevention from delirium tremens and its treatment is only guaranteed by clomethiazole or benzodiazepines. To reduce the doses of those drugs, combinations with neuroleptics are possible. In general voluntary physical withdrawal proves only to be successful if longtime abstinence is planned and a motivation program is setup the for withdrawal treatment.

Alcohol Withdrawal Delirium↗

PubMed1996

Combinatorial effects of monoclonal anti-p120 antibody (MAbp120), liposomes and hyperthermia on MCF-7 and LOX tumor cell lines.

The human nucleolar protein p120 is highly expressed in human cancers. Its high expression in breast cancer correlates with a poor prognosis, and its overexpression in 3T3 mouse fibroblasts causes malignant transformation. This study reports that a combination of monoclonal anti-p 120 antibody (MAbp120), liposomes (Lipo), and hyperthermia (HT) resulted in enhanced antitumor effects in cultured human breast adenocarcinoma (MCF-7) and human amelanotic melanoma (LOX) cells. Monoclonal antibody uptake and intracellular localization of the protein p120 were monitored by double labeling indirect immunofluorescence. Cell growth inhibition by the combination of MAbp120 + Lipo + HT was 65% for MCF-7 cells and 96% for LOX cells. When tested on LOX cells, monoclonal antibodies (MAbB23, MAbC23) to other nucleolar proteins (B23, C23) produced only slight cytotoxicity with similar treatment protocols.

Antibodies, Monoclonal↗

PubMed1996

[Surgery in the elderly--extent and status in surgery].

The results of selected elective and emergency operations of patients treated in the surgical department of Bonn University in 1994 and 1995 are presented and compared to the ASA-score. It was possible to show that old age itself is not a contraindication for major surgical interventions but that lethality mainly depends on concurrent diseases, i.e. the biological age. The numeric age is not an important factor. Physiological ageing only leads to diminished functional capacity, which can be partly compensated by more intensive care combined with early medical treatment. An age-specific factor which cannot be measured is the psychosocial age, the mental health. Mental health has an important influence on surgery in the aged patient.

Aged↗

PubMed1995

Phosphorothioate oligonucleotides bind in a non sequence-specific manner to the nucleolar protein C23/nucleolin.

To design optimal strategies for intracellular delivery of antisense phosphorothioate oligonucleotides, it may be useful to understand their interaction with cellular macromolecules. Nuclear extracts from LOX amelanotic myeloma cells were studied for protein binding to phosphorothioate oligonucleotides using a Southwestern protocol. Multiple nuclear proteins bound to the phosphorothioate oligonucleotides but no detectable protein binding was found to phosphodiester oligonucleotides. The protein with the strongest binding signals was shown by immunoprecipitation to be nucleolar C23/nucleolin, a 110 kDa protein. With glutathione S-transferase/nucleolin fusion protein constructs, the region of nucleolin containing the RNA recognition motifs had binding activity to phosphorothioate oligonucleotides.

Base Sequence↗

PubMed1995

Cloning and electrophysiological analysis of KST1, an inward rectifying K+ channel expressed in potato guard cells.

Potassium uptake by guard cells represents part of the osmotic motor which drives stomatal opening. Patch-clamp measurements have identified inward rectifying K+ channels capable of mediating K+ uptake in guard cells and various other plant cell types. Here we report the molecular cloning and characterization of a voltage-dependent K+ channel (KST1) from potato (Solanum tuberosum L.) guard cells. In situ hybridization shows expression of kst1 in guard cells. Two-electrode voltage-clamp and patch-clamp studies of the gene product after cRNA injection into Xenopus oocytes identified KST1 as a slowly activating, voltage-dependent, inward rectifying K+ channel. The single channel current voltage curve was linear in the range -160 to +20 mV, with a deduced single channel conductance of 7 pS in symmetrical 100 mM K+. This channel type, modulated by pH changes within the physiological range, required ATP for activation. In line with the properties of a K(+)-selective channel, KST1 was permeable to K+, Rb+ and NH4+ and excluded Na+ and Li+. Cs+ at submillimolar concentrations blocked the channel in a voltage-dependent manner. Related studies on potato guard cell protoplasts confirmed the biophysical characteristics of the kst1 gene product (KST1) in the heterologous expression system. Therefore, KST1 represents a major K+ uptake channel in potato guard cells.

Amino Acid Sequence↗

PubMed1995

Specific aspartic acid-rich sequences are responsible for silver staining of nucleolar proteins.

Ag-NOR proteins are silver-stainable proteins in the nucleolar organizer regions and are used to distinguish benign from malignant tumors. B23 and C23 are the two major Ag-NOR proteins. This study shows that only one of the two acidic clusters of B23 is responsible for its silver staining property. Fusion of this region of B23 (amino acids 161-188) to glutathione S-transferase produced an Ag-NOR positive fusion protein. The same result was obtained when amino acids 233-277 of C23 was fused with glutathione S-transferase. The aspartate residues, but not the glutamate residues, were found to be primarily responsible for the silver staining of the acidic clusters.

Amino Acid Sequence↗

PubMed1995

Inward rectifier potassium channels in plants differ from their animal counterparts in response to voltage and channel modulators.

We have investigated the electrophysiological basis of potassium inward rectification of the KAT1 gene product from Arabidopsis thaliana expressed in Xenopus oocytes and of functionally related K+ channels in the plasma membrane of guard and root cells from Vicia faba and Zea mays. The whole-cell currents passed by these channels activate, following steps to membrane potentials more negative than -100 mV, with half activation times of tens of milliseconds. This voltage dependence was unaffected by the removal of cytoplasmic magnesium. Consequently, unlike inward rectifier channels of animals, inward rectification of plant potassium channels is an intrinsic property of the channel protein itself. We also found that the activation kinetics of KAT1 were modulated by external pH. Decreasing the pH in the range 8.5 to 4.5 hastened activation and shifted the steady state activation curve by 19 mV per pH unit. This indicates that the activity of these K+ channels and the activity of the plasma membrane H(+)-ATPase may not only be coordinated by membrane potential but also by pH. The instantaneous current-voltage relationship, on the other hand, did not depend on pH, indicating that H+ do not block the channel. In addition to sensitivity towards protons, the channels showed a high affinity voltage dependent block in the presence of cesium, but were less sensitive to barium. Recordings from membrane patches of KAT1 injected oocytes in symmetric, Mg(2+)-free, 100 mM-K+, solutions allowed measurements of the current-voltage relation of single open KAT1 channels with a unitary conductance of 5 pS. We conclude that the inward rectification of the currents mediated by the KAT1 gene product, or the related endogenous channels of plant cells, results from voltage-modulated structural changes within the channel proteins. The voltage-sensing or the gating-structures appear to interact with a titratable acidic residue exposed to the extracellular medium.

Animals↗

PubMed1995

Immunodominant RNA recognition motifs of human nucleolin/C23.

Nucleolin/C23 is a nucleolar phosphoprotein implicated in the synthesis, processing and transport of ribosomal RNA and gene transcription. Auto-antibodies to human nucleolin/C23 have been reported in patients with systemic lupus erythematosus and other systemic autoimmune disorders. To identify immunodominant regions in nucleolin/C23, deletion fragments of nucleolin/C23 were fused in frame with the glutathione S-transferase gene. Seven monoclonal anti-nucleolin/C23 antibodies were used to determine the immunoreactivity of the bacterially expressed fusion proteins. Two sets of immunogenic regions at amino acids 314-389 and 387-461 were identified; each contained overlapping discontinuous epitopes and a centrally located RNA recognition motif. An auto-immune serum from a patient with systemic lupus erythematosus patient was found to contain antibodies against human nucleolin/C23 which recognized amino acids 387-461 of nucleolin/C23.

Amino Acid Sequence↗

PubMed1995

Elevated levels of activities of beta-hexosaminidase and alpha-mannosidase in human immunodeficiency virus-infected patients.

To determine their value as markers of the clinical stage of human immunodeficiency virus (HIV) disease, plasma activities of lysosomal glycosidases were determined in the plasma of 97 HIV-infected patients: molecular forms of cathepsin D were characterized by Western blot to examine the mode of enzyme release. In patients with Centers for Disease Control and Prevention stage II and III of HIV disease, plasma activity of beta-hexosaminidase was significantly increased. In patients with stage III infection, alpha-mannosidase activity was also significantly increased and cathepsin D was elevated and present only in its premature form. Thus, determination of plasma activities of beta-hexosaminidase and alpha-mannosidase in HIV-positive persons may be useful for distinguishing the clinical stage of disease. The elevation of precursors of cathepsin D in patients with stage III HIV disease indicates that secretion of lysosomal enzymes rather than leakage of enzymes from damaged cells is markedly elevated.

Cathepsin D↗

PubMed1995

Functional domains of nucleolar phosphoprotein p120.

Nucleolar phosphoprotein p120 is a low abundance, proliferation-associated protein. Several functional domains have been characterized and are discussed here such as the antigenic domain recognized by a monoclonal antibody, the nuclear/nucleolar localization domain, phosphorylation domains of casein kinase II (CKII) and protein kinase C, a putative methylation domain and an RNA binding region. By sucrose gradient sedimentation analyses, protein p120 was shown to rapidly sediment with 60-80 S pre-rRNP particles but sedimented more slowly when treated with RNAse or salt suggesting binding to RNA. Nucleolar protein p120 differed from other nucleolar proteins such as C23 (nucleolin) and B23 (nucleophosmin) which sedimented more slowly near the top of the gradient.

Amino Acid Sequence↗

PubMed1994

Apoptosis in human tumor cells following treatment with p120 antisense oligodeoxynucleotide ISIS 3466.

Previously, we reported that treatment of LOX cells in vitro with phosphorothioate oligonucleotide ISIS 3466 (antisense to the human nucleolar protein p120-FB2) produced a 70% cell kill and morphological changes including nucleolar unravelling, chromatin condensation and fragmentation, and a reduction in mitotic figures consistent with apoptosis. This report shows that HeLa cells treated with ISIS 3466 also developed apoptosis: nucleosomal ladders were found when the DNA from the treated HeLa cells was extracted and run on agarose gels. The morphological changes consistent with apoptosis were found more frequently in the floating cells than in the attached cells. The percentages of floating cells and attached cells were indicators of the toxicity of the different oligonucleotides studied. Of these, oligonucleotide ISIS 3466 produced the highest percent of floating cells (78.4%). Treatment of HeLa cells with other oligonucleotides produced fewer floating cells, and the characteristic nucleosomal ladder was not found following DNA extraction.

Apoptosis↗