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Biomedical subjects

B Koch

Publications and source records attributed to B Koch.

At least 73 records · Page 4Linked to original sources
PubMed1997

Detection of p53 in inflammatory tissue and lymphocytes using immunohistology and flow cytometry: a critical comment.

AIMS: To analyse the expression of p53 in lymphatic cells found in inflammatory tissues and the peripheral blood by immunological methods. METHODS: Immunohistological analysis of synovial tissues from patients with rheumatoid arthritis and flow cytometric analysis of peripheral blood lymphocytes were performed with anti-p53 antibodies from different sources. RESULTS: The anti-p53 antibodies PAb240, PAb421, and PAb1801 from one supplier bound to the cytoplasm of lymphocytes, fibroblasts, and endothelial cells in rheumatoid synovial tissue, while the same anti-p53 antibodies from other sources and the p53 specific antibodies PAb1620 and DO1 were negative. Using flow cytometry, the antibodies that labelled cells in inflammatory tissues were shown to bind also to peripheral lymphocytes, while the antibodies that were negative in immunohistology did not react with peripheral blood lymphocytes. p53 expression could be confirmed by western blot in rheumatoid synovial tissue, but not in peripheral blood lymphocytes using PAb421 and PAb240 antibodies from our own laboratory, which had been negative in immunohistology. CONCLUSIONS: Demonstration of p53 by western blot is more sensitive and reliable than immunohistology and flow cytometry. Western blot is the gold standard for the demonstration of p53 expression and should be used, whenever possible, to confirm p53 expression in normal tissue shown by immunohistology or flow cytometry. All other reports on p53 expression, especially those obtained using antibodies with an unusual staining pattern must be interpreted with caution.

Arthritis, Rheumatoid↗
PubMed1996

Pituitary adenylate cyclase-activating polypeptide induces expression of corticosteroid-binding globulin in cultured fetal hepatocytes: synergy with tri-iodothyronine.

The purpose of the present study was to determine whether functional receptors for pituitary adenylate cyclase-activating polypeptide (PACAP) are expressed in cultured rat fetal hepatocytes and eventually play a role in regulating gene expression of corticosteroid-binding globulin (CBG). We found PACAP38 and PACAP27 to elevate cAMP levels in hepatocytes in a dose-dependent manner, with a plateau being achieved at 10 nM and EC50 values of about 0.5-1 nM. PACAP failed to alter the turnover of inositol phosphates, whereas PACAP and VIP stimulated cAMP accumulation in an equipotent manner, suggesting the presence in these cells of type II receptor isoforms. As revealed by measurements of both CBG mRNA levels and concentrations of binding sites, long-term treatment of fetal cells with 10 nM PACAP, although resulting in partial desensitization of peptide-induced cAMP accumulation, caused a significant 3-fold elevation in CBG synthesis. This stimulatory influence of PACAP was mimicked by the cell permeant N6,2'-O-dibutyryladenosine 3',5'-phosphate (dbcAMP). Treatment of hepatocytes with tri-iodothyronine (T3) enhanced CBG expression and, most interestingly, appeared to synergize with PACAP to elicit a 2-3-fold amplification of CBG synthesis. This study thus provides first evidence for the up-regulation by PACAP and cAMP of CBG expression in fetal hepatocytes and for T3's playing a synergistic role in enhancing PACAP-induced synthesis of the binder.

Animals↗
PubMed1996

Demonstration of interleukin-1beta and interleukin-6 in cells of synovial fluids by flow cytometry.

Cytokine levels are increased in the synovial fluid of affected joints from patients with inflammatory joint diseases. The aim of our study was therefore to determine if and to what extent immmunologically defined subpopulations of mononuclear cells (MNC) in the synovial fluid are responsible for the increased levels of interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) in affected joints. Lipopolysaccharide (LPS) stimulated peripheral MNC were used as positive controls. While soluble IL-1beta (median 167 pg/ml) and IL-6 (median 508 pg/ml) levels were significantly elevated in the synovial fluids tested, IL-1beta and IL-6 were demonstrated by flow cytometry in only a small subpopulation (<=11%) of mononuclear synovial fluid cells in 7/13 patients. Our results suggest that elevated IL-1beta and IL-6 levels in the synovial fluid of inflammatory joints are derived mainly from cells in the synovial membrane and only to a minor extent from cells in the synovial fluid itself.

Arthritis, Reactive↗
PubMed1996

Analysis of bcl-2+ lymphocyte subpopulations in inflammatory synovial infiltrates by a double-immunostaining technique.

We used a double-immunostaining technique to analyze the distribution of bcl-2+ B and T lymphocytes within the synovial membranes (SM) of 13 patients with rheumatic diseases: 11 with rheumatoid arthritis (RA), 1 with ankylosing spondylitis (AS), and 1 with osteoarthritis (OA). A high proportion (up to 50%) of the lymphocytes belonged to the B cell subset. Most of both T and B lymphocytes were positive for the bcl-2 protein. In germinal centers B lymphocytes were also negative for bcl-2 protein expression, comparable to the situation in germinal centers of secondary lymphatic organs. We conclude that bcl-2- B lymphocytes are submitted to antigen selection in the inflamed SMs while bcl-2 protein expression provides survival signals for their persistence in the infiltrates. The expression of bcl-2 may be an important factor in protecting lymphocytes in SM from apoptosis by glucocorticoids, cytostatic drugs, and irradiation.

Adult↗
PubMed1996

[Must the recurrent nerve in thyroid gland resection always be exposed? A prospective randomized study].

It was the purpose of this study to establish whether it is necessary to identify the recurrent laryngeal nerve intraoperatively when resecting the endemic nodular goiter. We prospectively formed two groups from 800 subtotal unilateral resections ("nerves at risk"). In group I (382 subtotal resections, 48.4%) the nerve was not identified intraoperatively. In group II (413 subtotal resections, 51.6%) the recurrent laryngeal nerve was routinely identified in all cases. The operative technique was standardized, giving special attention to the "anterior lamella". For all patients, preoperative and postoperative evaluation of the vocal cords was performed routinely. Altogether we saw 4 (0.5%) transient vocal cord palsies: 2 in group I and 2 in group II. All 4 laryngeal nerve palsies recovered within 4 months. These data demonstrated that no benefit is gained from routine dissection of the laryngeal nerve during resection of endemic nodular goiter. Therefore the demand for obligatory intraoperative identification of the recurrent nerve is not tenable.

Adult↗
PubMed1996

[Rare intracranial plasmacytoma manifestations. Case reports and review of the literature in diffuse plasmocytoma, in primary solitary extramedullary plasmacytoma in in primary solitary osseous plasmacytoma].

Plasmacytomas can be divided into multiple, solitary osseous and solitary extraosseous/extramedullary plasmacytomas. Intracranial plasmacytomas of the dura, leptomeninx and cerebrum are well known from the literature. They are manifestations of multiple myeloma, intracranial extramedullary plasmacytoma or metastatic disease of extramedullary plasmacytoma in distant locations. We describe a cerebellar manifestation of a solitary plasmacytoma of the bone, and a leptomeningeal carcinomatosis of a multiple plasmacytoma. A summary of the literature concerning intracranial plasmacytomas is given. Dural manifestations of plasmacytoma have the same features as meningiomas in CT or MRI. Cerebral or cerebellar manifestations cannot be differentiated from brain tumors by means of CT or MRI. In CT, plasmacytomas show high-density lesions. T2w-MRI reveals a low-intensity lesion. In T1w-MRI, intense homogeneous contrast enhancement can be demonstrated.

Bone Neoplasms↗
PubMed1996

Regulation of corticosteroid-binding globulin synthesis by 1alpha,25-dihyroxy-vitamin D3 (calcitriol), 9-cis-retinoic acid and triiodothyronine in cultured rat fetal hepatocytes.

Evidence regarding the nature of the regulatory factors which directly act upon liver cells and extra-hepatic tissues to alter CBG synthesis is scarce. The present study used cultured rat fetal hepatocytes to investigate the involvement and possible interplay in this process of several members of the nuclear receptors superfamily: vitamin D (VDR), retinoic acids (RAR/RXR) and thyroid hormones (TR). Treatment of cells with 1alpha,25-(OH)2D3 (1,25-D) elicited a dose-dependent inhibition of basal CBG concentration in culture medium. Maximum inhibition to about 15% of control level was achieved with 0.1-1.0 nM, with an IC50 of 3.8 x 10(-12) M and with no significant change in binding affinity. Differential activation of RAR and RXR with either 9-cis-retinoic acid (9-cis-RA) or the RAR-selective synthetic retinoid TTNPB revealed that high doses of both drugs diminished CBG expression, though the former proved about 10-times more potent than the latter in this regard. Amplification by triiodothyronine (T3) of CBG synthesis failed to block the inhibitory effects of either 1,25-D or retinoids, as revealed by both binding capacity and mRNA measurements. Relative to CBG, 1,25-D similarly depressed the synthesis of alpha-fetoprotein (AFP), while on the contrary, retinoids and T3 were shown to cause opposite effects, as 9-cis-RA and TTNPB elevated and T3 decreased AFP expression. The present findings identify for the first time ligands of VDR and RAR/RXR as powerful negative regulators of both basal and T3-stimulated CBG biosynthesis in fetal hepatocytes and suggest lack of a functional interplay between TR and VR or RAR/RXR in these processes.

Animals↗
PubMed1996

Sucrase-isomaltase gene expression in suckling rat intestine: hormonal, dietary, and growth factor control.

The effect of hydrocortisone, sucrose, and epidermal growth factor (EGF), alone or in combination, was investigated on the precocious expression of sucrase activity and sucrase-isomaltase (SI) mRNA in normal and adrenalectomized (ADX) suckling rats. In normal rats, each regulatory factor administered separately from 12 to 13 days of age induced a parallel expression of sucrase activity and SI mRNA. A potentiation of both sucrase activity and SI mRNA was observed after administration of hydrocortisone plus sucrose. Only SI mRNA was potentiated after EGF-sucrose administration or EGF-hydrocortisone administration. However, the potentiation of the activity was obtained after a prolonged treatment period of 4 days. In adrenalectomized rats, sucrose and EGF were still able to induce precocious induction of SI mRNA and sucrase activity, but sucrase activity was lowered by 75% by sucrose and by 30% by EGF. The combined treatments gave results similar to those obtained in normal rats. The results obtained in ADX rats indicate that the effects of sucrose and EGF on the precocious expression of sucrase and the potentiation of SI mRNA are independent of endogenous glucocorticoids. The rate of [3H]thymidine incorporation into mucosal DNA of normal sucklings was similar after single or combined treatments. From this study, we conclude that the precocious induction of intestinal sucrase by various regulatory factors, alone or in combination, is primarily regulated at the level of mRNA and is independent of increases in cellular proliferation and circulating glucocorticoids.

Adrenalectomy↗
PubMed1996

Moexipril versus captopril in patients with mild to moderate hypertension.

Moexipril is a new, long-acting angiotensin-converting enzyme (ACE) inhibitor. In contrast to captopril, it is a prodrug of the pharmacologically active agent moexiprilat and will be administered once daily. The objective of this study was to compare the efficacy, safety, and tolerability of moexipril with that of captopril during a 12-week treatment of patients with mild to moderate hypertension. Patients with a sitting diastolic blood pressure (SDBP) of 95-114 mm Hg, inclusive, were randomized in a 2:1 ratio to receive moexipril, 7.5 mg, once daily or captopril, 25 mg, twice daily. After 6 weeks of treatment, the dose of moexipril and captopril was increased to 15 mg once daily and 50 mg twice daily, respectively, if the patient's SDBP remained > or = 90 mm Hg. Blood pressure was measured at biweekly visits. At study endpoint, adjusted mean reductions in SDBP were comparable between the moexipril and captopril groups (-9.8 vs. -8.7 mm Hg), and moexipril was more effective than captopril in reducing sitting systolic blood pressure. Adverse experiences (headache, dizziness, and upper respiratory infection) occurred at similar frequencies in the moexipril and in the captopril groups. The data indicate that moexipril at dosages of 7.5 and 15 mg once daily is as efficacious as twice daily captopril in reducing blood pressure in patients with mild to moderate hypertension.

Adult↗
PubMed1996

In vitro testing of artificial heart valves: comparison between Newtonian and non-Newtonian fluids.

The in vitro testing of artificial heart valves is often performed with simple fluids like glycerol solutions. Blood, however, is a non-Newtonian fluid with a complex viscoelastic behavior, and different flow fields in comparable geometries may result. Therefore, we used different polymer solutions (Polyacrylamid, Xanthan gum) with blood-like rheological properties as well as various Newtonian fluids (water, glycerol solutions) in our heart valve test device. Hydrodynamic parameters of Björk-Shiley heart valves with a tissue annulus diameter (TAD) of 21-29 mm were investigated under aortic flow conditions. Major results can be summarized as follows. The mean systolic pressure differences depend on the model fluids tested. Closing time and closing volume are not influenced by the rheological behavior of fluids. These parameters depend on TAD and the pressure differences across the valve. In contrast, rheological behavior has a pronounced influence upon leakage flow and leakage volume, respectively. Results show furthermore that the apparent viscosity data as a function of shear rate are not sufficient to characterize the rheological fluid behavior relevant to hydrodynamic parameters of the heart valves investigated. Therefore, similarity in the yield curves of non-Newtonian test fluids mimicing blood is only a pre-requisite for a suitable test fluid. More information about the viscous and elastic component of the fluid viscosity is required, especially in geometries where a complex flow field exists as in the case of leakage flow.

Acrylic Resins↗
PubMed1996

Evidence for the presence of receptors for pituitary adenylate cyclase-activating polypeptide in the neurohypophysis that are positively coupled to cyclic AMP formation and neurohypophyseal hormone secretion.

This study examines the neural lobe of the pituitary gland for the presence of receptors for pituitary adenylate cyclase-activating polypeptide (PACAP) and their possible involvement in the regulation of neurosecretion. The presence of PACAP receptors of type I was revealed in the neural lobe, as well as in anterior and intermediate lobes, by means of RT-PCR amplification using selective oligonucleotide pairs of primers. They appeared to be expressed in the tissues as a short form together with an isoform of heavier molecular weight. Activation of receptors in the presence of PACAP stimulated both formation of cyclic AMP (cAMP) and secretion of arginine vasopressin (AVP) in neural lobes, in a dose-related fashion, with half-maximum (EC50) values of 1.0 +/- 0.2 x 10(-9) M and 1.4 +/- 0.3 x 10(-8) M, respectively. Parallel with AVP, PACAP also stimulated oxytocin (OXT) output, with an EC50 value of 0.6 +/- 0.1 x 10(-8) M. In an attempt to localize receptors on cells (mainly astrocyte-like glials or pituicytes) and/or on nerve fibers of the gland, we used cultures of neural lobe cells and explants (in which nerve fibers undergo degeneration), as well as isolated nerve endings. In both cells and nerve terminals, PACAP enhanced accumulation of cAMP, while it triggered AVP secretion from the latter. The stimulatory effect of PACAP on both AVP and OXT release was mimicked by dbcAMP and blocked by H89, an inhibitor of cAMP-dependent protein kinase. We conclude that in the neural lobe, PACAP receptors are localized on both nerve terminals and pituicytes, which participate in the modulation of secretion of neurohypophyseal hormones in an interactive way and mainly through the cAMP signalling route.

Animals↗
PubMed1996

Glucose monitoring as a guide to diabetes management. Critical subject review.

PURPOSE: To encourage a balanced approach to blood glucose monitoring in diabetes by a critical review of the history, power and cost of glucose testing. DATA SOURCES: The Cambridge Data Base was searched and was supplemented by a random review of other relevant sources, including textbooks, company pamphlets, and laboratory manuals. STUDY SELECTION: Keywords used were "glucosuria diagnosis," "blood glucose self-monitoring," "glycosylated hemoglobin," and "fructosamine" for the 10-year period ending 1992, restricted to English language and human. DATA EXTRACTION: About 200 titles were retrieved and reviewed according to the author's judgment of relevance. FINDINGS: "Snapshot tests" (venous and capillary blood glucose) and "memory tests" (urine glucose, glycated hemoglobin fractions and fructosamine) must be employed according to individual patients treatment goals. Day-to-day metabolic guidance is facilitated by capillary blood glucose testing for patients receiving insulin and by urine glucose testing for others. Capillary blood glucose testing is mandatory in cases of hypoglycemia unawareness (inability to sense hypoglycemia because of neuropathy) but is not a substitute for a knowledge of clinical hypoglycemia self-care. Criteria by reason (clinical judgement and cost effectiveness) must be separated from criteria by emotion (preoccupation with technology and marketing). No randomized studies show that any of these tests consistently improve clinical outcome. Optimal metabolic control and cost savings can be expected from a rational selection of tests.

Blood Glucose↗
PubMed1995

Purification and characterization of recombinant cytochrome P450TYR expressed at high levels in Escherichia coli.

The multifunctional tyrosine N-hydroxylase, cytochrome P450TYR (CYP79), from Sorghum bicolor catalyzing the conversion of tyrosine to p-hydroxyphenyl-acetaldoxime in the biosynthesis of the cyanogenic glucoside dhurrin, has been expressed in Escherichia coli using the isopropyl-beta-D-thiogalactopyranoside-inducible vector pSP19g10L, containing the cDNA encoding CYP79. The expression construct was optimized by reducing the length of the N-terminal hydrophobic core of the signal sequence of cytochrome P450TYR and by exchanging the first eight codons with the first eight codons of bovine P45017 alpha. The highest yielding construct provided 200-500 nmol P450TYR/liter cell culture. The recombinant P450TYR was gently and efficiently extracted from E. coli spheroblasts by temperature-induced phase partitioning of Triton X-114 in the presence of 30% glycerol and isolated by DEAE and reactive red chromatography. In reconstitution experiments using saturating amounts of sorghum NADPH-cytochrome P450 reductase, the Km and turnover rate for isolated recombinant P450TYR was 0.22 +/- 0.06 mM and 49.2 +/- 3.8 min-1, respectively, whereas a turnover rate as high as 350 min-1, was obtained using E. coli membranes. Addition of 3 mM glutathione stimulated the activity of reconstituted P450TYR and of sorghum microsomes although the effect was highly variable. Phenylalanine, the precursor of several cyanogenic glucosides, gave a type I binding spectrum, but was not metabolized by P450TYR, demonstrating the high substrate specificity of this P450. Administration of radioactively labeled p-hydroxyphenylacetaldoxime to E. coli cells, showed E. coli metabolized p-hydroxyphenylacetaldoxime independent of the expression of P450TYR.

Amino Acid Sequence↗
PubMed1995

Cytochrome P-450TYR is a multifunctional heme-thiolate enzyme catalyzing the conversion of L-tyrosine to p-hydroxyphenylacetaldehyde oxime in the biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor (L.) Moench.

Cytochrome P-450TYR, which catalyzes the N-hydroxylation of L-tyrosine in the biosynthesis of the cyanogenic glucoside dhurrin in Sorghum bicolor (L.) Moench has recently been isolated (Sibbesen, O., Koch, B., Halkier, B. A., and Møller, B. L. (1994) Proc. Natl. Acad. Sci. U.S.A. 92, 9740-9744). Reconstitution of the enzyme activity in lipid micelles containing cytochrome P-450TYR and NADPH-cytochrome P-450 oxidoreductase demonstrates that cytochrome P-450TYR catalyzes the conversion of L-tyrosine into p-hydroxyphenylacetaldehyde oxime. Earlier studies with microsomes have demonstrated that this conversion involves two N-hydroxylation reactions of which the first produces N-hydroxytyrosine. We propose that the product of the second N-hydroxylation reaction is N,N-dihydroxytyrosine. N,N-dihydroxytyrosine is dehydrated to 2-nitroso-3-(p-hydroxyphenyl) propionic acid which decarboxylates to p-hydroxyphenylacetaldehyde oxime. The dehydration and decarboxylation reactions may proceed non-enzymatically. The E/Z ratio of the p-hydroxyphenylacetaldehyde oxime produced by reconstituted cytochrome P-450TYR is 69:31. Lipid micelles made from L-alpha-dilauroyl phosphatidylcholine are more than twice as effective in reconstituting cytochrome P-450TYR activity as compared to other lipids. The Km and turnover number of the enzyme is 0.14 mM and 200 min-1, respectively, when assayed in the presence of 15 mM NaCl whereas the values are 0.21 mM and 230 min-1 when assayed in the absence of added salt. The multifunctional nature cytochrome P-450TYR is confirmed by demonstrating that binding of L-tyrosine or N-hydroxytyrosine mutually excludes binding of the other substrate. These results explain why the conversion of tyrosine to p-hydroxyphenylacetaldehyde oxime as earlier reported (Møller, B. L., and Conn, E. E. (1980) J. Biol. Chem. 255, 3049-3056) shows the phenomenon of catalytic facilitation ("channeling"). Cytochrome P-450TYR is the first isolated multifunctional heme-thiolate enzyme from plants. N-Hydroxylases of the cytochrome P-450 type with high substrate specificity have not previously been reported.

Catalysis↗
PubMed1995

The B lymphocyte in rheumatoid arthritis: analysis of rearranged V kappa genes from B cells infiltrating the synovial membrane.

The participation of the humoral immune system in rheumatoid arthritis (RA) is characterized by the production of rheumatoid factors (RF). RF are autoantibodies against the Fc part of IgG which are encoded by diverse germ-line genes. Most of the RF-encoding genes are unmutated, but in RA, a substantial quantity is encoded by somatically mutated genes. In addition, the synovial membranes (SM) of the diseased joints of RA patients are infiltrated by B lymphocytes which form germinal center-like aggregates. To analyze the local immune response, B cell foci from two RA SM were isolated by micromanipulation. From DNA of these foci, the rearranged kappa light chain variable region (V kappa) genes were amplified by polymerase chain reaction (PCR), cloned and sequenced. The amplification of different V kappa-J kappa combinations of different foci suggested oligoclonal expansion of B lymphocytes, which was confirmed by sequence analysis: each PCR product contained members of a single B cell clone. The sequence analysis of 29 different clones revealed rearrangements of diverse V kappa genes. Both frequent representatives of the V kappa 3 and the V kappa 1 family, as well as rarely used genes such as the L10 and B2 genes of the V kappa 2 and V kappa 5 families were found. Of the eleven potentially functional gene rearrangements, eight were significantly mutated, indicating their derivation from antigen-selected B cells. Intraclonal diversity in one of these clones may suggest ongoing mutation in the diseased synovial membrane of patients with RA.

Adult↗
PubMed1995

[Apoptosis and cell proliferation in HHV-6 infections. Regulatory mechanisms of p53/bcl-2/ras interactions].

HHV-6 infected immature T (HSB2) and Hodgkin (HDLM2) cells and biopsy tissues from lymph nodes of patients with Hodgkin's disease (HD) and Kikuchi lymphadenitis (KL) were studied immunohistologically for virus antigen expression and for the oncogene/anti-oncogene products ras, bcl-2 and p53. Cell proliferation and cell death were tentatively monitored in tissue culture by PCNA staining, by viability testing and in situ end labeling of fragmented DNA. PCNA was also used in biopsy samples. KL is characterized by high incidences of focal cell death (i.e. histiocytic necrotizing lymphadenitis), while HD is apparently more a proliferative disease. The techniques used revealed no significant differences in the cellular expression of viral DNA or antigens among cell lines, HD or KL. The HDLM2 cell line with the superior survival after HHV-6 infection showed a significantly lower expression of p53 and PCNA than HSB2 cells. Biopsy samples from patients with KL did not express p53, and ras and PCNA were observed in fewer cells than in HD. Bcl-2, however, was significantly more frequently seen than in HD. The interpretation of the data is difficult; they suggest that there are additional regulatory influences in control of cell proliferation and cell death, such as cytokines and growth factors, which are altered after viral infection. Also, virus-induced cell death probably includes other mechanisms besides apoptosis, such as cell damage caused by oxygen radicals.

Apoptosis↗