Biomedical subjects

A Gebhardt

Publications and source records attributed to A Gebhardt.

At least 19 recordsLinked to original sources

PubMed2005

High efficiency, high quality x-ray optic based on ellipsoidally bent highly oriented pyrolytic graphite crystal for ultrafast x-ray diffraction experiments.

By the use of a thin highly oriented pyrolytic graphite crystal (HOPG) bent to a high-performance ellipsoidal shape it was possible to focus monochromatic x-rays of 4.5 keV photon energy with an efficiency of 0.0033, which is 30 times larger than for previously used bent crystals. Isotropic Ti K alpha radiation of a 150 microm source was focused onto a 450 microm spot. The size of the focal spot can be explained by broadening due to the mosaic crystal rocking curve. The rocking curve width (FWHM) of the thin graphite foil was determined to 0.11 degrees. The estimated temporal broadening of an ultrashort K alpha pulse by the crystal is not larger than 300 fs. These properties make the x-ray optic very attractive for ultrafast time-resolved x-ray measurements.

Journal Article↗

PubMed2005

Establishment and development of the National Tuberculosis Control Programme in Vietnam.

OBJECTIVE: To describe the establishment and development of the National Tuberculosis Control Programme (NTP) of Vietnam. METHODS: Data were obtained from the surveillance system established by the new NTP in 1986 and based on the principles now described as the WHO DOTS strategy. RESULTS: The proportion of districts covered by the NTP increased from 40% in 1986 to almost 100% in 2000. The proportion of communes applying NTP guidelines increased from 18% in 1986 to 99.8% in 2000. The total number of tuberculosis cases notified increased from 8737 in 1986 to 89 792 in 2000. Most of these are new smear-positive cases. Based on WHO estimations of the incidence rate, the proportion of new smear-positive cases detected and put on short-course treatment has been over 70% since 1996. Reported cure rates with short-course chemotherapy are consistently over 85%. CONCLUSIONS: DOTS is feasible in a low-income, high-burden country. The main reasons for success were political commitment, a well-functioning health network, integration of tuberculosis control into the general health service at district level, a continuous supply of drugs and effective external support. Major challenges are long-term financial support, expansion to remote areas and vulnerable groups, definition of the role of the private sector, and future developments of the HIV epidemic and multidrug resistance.

Humans↗

PubMed2000

Quadruple tacrolimus-based induction therapy including azathioprine and ALG does not significantly improve outcome after liver transplantation when compared with standard induction with tacrolimus and steroids: results of a prospective, randomized trial.

BACKGROUND: Tacrolimus in combination with prednisolone has been proven to be a safe and effective immunosuppressive induction therapy in solid organ transplantation. However, it remains unclear whether a tacrolimus-based quadruple induction regimen with azathioprine and an antilymphocytic preparation could further improve the results after orthotopic liver transplantation. Therefore, we designed a prospective, randomized study to compare the immunosuppressive efficacy of dual (tacrolimus and prednisolone) and quadruple (tacrolimus, azathioprine, ALG Merieux and prednisolone) induction after liver transplantation. METHODS: After randomization, 120 consecutive patients of primary liver transplants were divided into the dual group (n=59) and the quadruple group (n=61) and followed for a minimum of 3 years. RESULTS: Patient survival at 3 years was 88.2% in the dual versus 94.9% in the quadruple group. Overall 25 patients in each group (41 and 42%, respectively) developed acute rejection. There was no difference in the number and severity of rejections. In each group only four patients required OKT3-therapy, however, although three of four patients in the quadruple group responded to OKT3 and cleared rejection, none of the four patients in the dual group were treated successfully with OKT3 (P<0.02). Rejection in these patients resolved only after additional treatment with mycophenolate mofetil. Adverse events and infections were equally distributed in both groups. Asymptomatic Cytomegalovirus infections were more common in the quadruple group (P<0.02). As of today, only one patient developed posttransplant lymphoproliferative disease (dual group). CONCLUSIONS: The data from our single-center study indicate that both tacrolimus-based dual and quadruple immunosuppressive induction regimens yield similar safety and effectiveness after liver transplantation.

Adult↗

PubMed1998

Effects of antihypertensive drugs on cholesterol metabolism of human mononuclear leukocytes and hepatoma cells.

OBJECTIVES: Primary prevention trials of antihypertensive therapy have shown conflicting results on coronary events. Potential interference of antihypertensive agents with cellular lipid metabolism may alter the atherosclerotic risk of individuals. DESIGN AND METHODS: The effects of the calcium antagonist's verapamil, diltiazem, and nifedipine and of the beta-blockers propranolol and metoprolol on low density lipoprotein (LDL) receptor activity, cholesterol esterification rate, oleate incorporation in triglycerides and sterol synthesis were studied in freshly isolated human leukocytes and HEP G2 cells. RESULTS: Up to a concentration of 3-10 mumol/L, verapamil, propranolol, and metoprolol led to an increased cellular content of 125I-LDL by an inhibition of degradation. In mononuclear cells verapamil stimulated accumulation and degradation. No effect on binding was observed. Diltiazem was only stimulatory on 125I-LDL processing in leukocytes. Beta blockers and verapamil significantly reduced the LDL mediated 14C-oleate incorporation in cholesterol esters. In the presence of 25-hydroxycholesterol the esterification was not diminished, which suggests that cholesterolacyltransferase (ACAT) was not affected per se. Whereas all the agents induced the synthesis of lanosterol, metoprolol inhibited cholesterol synthesis. None of the agents had a significant influence on 14C-oleate incorporation in triglycerides, suggesting a specific influence on cholesterol metabolism. CONCLUSIONS: Antihypertensive drugs affect the cholesterol metabolism on a cellular level. Mechanisms are an interference with degradation of LDL and consequent alterations of cholesterol esterification. Using leukocytes as peripheral cells and HEP G2 as a model of human liver, these results may have importance when antihypertensive long-term therapy is conducted for primary or secondary prevention of atherosclerotic complications.

Antihypertensive Agents↗

PubMed1998

Custom-made cast titanium implants produced with CAD/CAM for the reconstruction of cranium defects.

Titanium implants for the reconstruction of bony skull defects, using data from three-dimensional spiral computer tomography, have been described by other authors. Instead of milling the implants from a titanium block, an advanced method of rapid prototyping for a fine casting process is presented. Casting vs milling offers several advantages. It is possible to form very thinly tapered structures and to obtain more complex geometrical structures with smaller diameters. Many geometrical forms, which cannot be milled for technical reasons, can be produced using this technique.

Adult↗

PubMed1998

Effects of pravastatin on cholesterol metabolism of cholesterol-fed heterozygous WHHL rabbits.

1. We administered the 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor pravastatin at a daily dose of 1 mg kg(-1) body weight to cholesterol-fed (0.03%) heterozygous Watanabe heritable hyperlipidaemic rabbits, an animal model for heterozygous familial hypercholesterolaemia. 2. After 12 months of cholesterol treatment, immunohistochemistry with the monoclonal antibody 9D9 was used to detect hepatic low density lipoprotein (LDL) receptors, which were quantified by densitometry. In addition we determined LDL receptor mRNA by competitive reverse transcriptase polymerase chain reaction. The cholesterol precursor lathosterol and the plant sterol campesterol were analysed by gas-liquid chromatography. 3. The drug reduced total plasma cholesterol levels by 51% (P=0.04), when compared to the control group. Unexpectedly, hepatic LDL receptor density and mRNA showed no significant differences between the groups. Total plasma levels of lathosterol and campesterol also revealed no significant differences between the groups, if expressed relative to plasma cholesterol. 4. The findings suggest that mechanisms other than induced hepatic LDL receptors are responsible for the cholesterol-lowering effect of pravastatin in this animal model. We propose a reduced cholesterol absorption efficiency compatible with similar campesterol levels between both groups observed in our study.

Animals↗

PubMed1997

Vitronectin receptor (alpha(v)beta3) mediates platelet adhesion to the luminal aspect of endothelial cells: implications for reperfusion in acute myocardial infarction.

BACKGROUND: Platelet interaction with endothelium plays an important role in the pathophysiology of coronary microcirculation. We assessed the role of the vitronectin receptor (integrin alpha(v)beta3) in platelet/endothelium adhesion. METHODS AND RESULTS: We investigated the effect on platelet/endothelium adhesion of plasma obtained from patients with acute myocardial infarction during reperfusion (before and 8, 24, 48, and 72 hours and 5 to 7 days after direct angioplasty) and with pretreatment with alpha-thrombin (2 U/mL) and recombinant human interleukin-1beta. Platelet/endothelium adhesion was significantly enhanced by approximately 20% after pretreatment of endothelium with patient plasma for 4 hours (P<.05) compared with endothelium treated with pooled control plasma. Plasma-induced platelet/endothelium adhesion was, in part, RGD peptide dependent. Pretreatment of endothelial cells with alpha-thrombin or recombinant human interleukin-1beta enhanced platelet/endothelium adhesion and surface expression of alpha(v)beta3 on the luminal aspect of endothelium (P<.05). The adhesion of platelets, isolated platelet microparticles, and Chinese hamster ovary cells bearing human recombinant alpha(IIb)beta3 (platelet glycoprotein IIb-IIIa) to activated endothelial cells was inhibited by antiadhesive peptides GRGDSP and c(RGDfV) and monoclonal antibodies 4F10, LM609, and 7E3. CONCLUSIONS: The expression of vitronectin receptor exposed on the luminal aspect of activated endothelium is enhanced and mediates platelet/endothelium adhesion. Vitronectin receptor-mediated platelet attachment to activated endothelium during reperfusion may contribute to reperfusion injury and could be a target for antiadhesive therapy.

Adult↗

PubMed1997

LDL receptor activity in human leukocyte subtypes: regulation by insulin.

OBJECTIVES: LDL receptors of leukocytes play a key role in lipoprotein uptake, immunoregulation and the pathogenesis of atherosclerosis. Numerous studies with different methods of low reliability yielded conflicting results of its regulation in leukocyte subtypes. DESIGN AND METHODS: LDL receptors of human leukocytes were measured with use of the monoclonal antibody C-7. Specific C-7 binding was detected by FACS analysis using phycoerythrin-anti-mouse-IgG. Parallel incubations with FITC-labelled anti-LEU 4 (CD 3), anti-LEU 12 (CD 19) and anti-MY 4 (CD 14) antibodies were used to distinguish C-7 binding of specific cell types (T-, B-lymphocytes and monocytes). RESULTS: In contrast to monocytes, T and B-lymphocytes freshly isolated from healthy blood donors had no detectable binding capacity for C-7. After 24 and 48 h incubation of cells in a lipid-free medium, lymphocytes acquired some C-7 binding, albeit still much less than monocytes. Incubation with insulin for 24 h in a concentration of 0.5 microgram/mL led to an increase in C-7 binding for monocytes (up to 180%). Saturation experiments with the ligand suggests an increase in the number of receptors. In contrast the same insulin concentration inhibited C-7 binding of B- and T-lymphocytes by 35%. CONCLUSIONS: FACS analysis using monoclonal antibodies seems to be a feasible method for the investigation of lipid metabolism in leukocytes. The LDL receptor expression and its regulation by insulin differs in circulating monocytes and lymphocytes.

Blood Donors↗

PubMed1996

Organ- and tissue-specific expression of the low density lipoprotein receptor. Rapid quantitation by competitive reverse transcriptasepolymerase chain reaction.

A rapid and sensitive method of quantifying very low-abundant mRNA species by competitive reverse transcriptase-polymerase chain reaction (cRT-PCR) is presented. For each analysis, a defined amount of total cellular RNA is co-reverse transcribed and co-amplified with a titration series of in vitro synthesized RNA from a clone of the mRNA which carries an internal deletion. The equivalence point, which defines the number of specific mRNA molecules in the sample, can be determined using ethidium bromide stained gels of the reaction products. We present here three examples of the application of this new technique to the quantification of the low abundant mRNA for the low density lipoprotein (LDL) receptor. First, the regulation of LDL receptor mRNA expression by the HMG-CoA reductase inhibitor, pravastatin, has been analysed in vitro, in a human gastric tumour cell line. Second, LDL receptor mRNA from various bovine tissues has been quantified. Third, the expression of LDL receptor mRNA in human tissues originating from colorectal carcinoma and corresponding tumour-tree margins of surgical specimens has been measured.

Animals↗

PubMed1995

Increased LDL receptor mRNA expression in colon cancer is correlated with a rise in plasma cholesterol levels after curative surgery.

It is currently under debate whether the low serum cholesterol levels that are frequently observed in cancer patients represent a risk factor for/or, rather, are a consequence of the tumour. We postulate that malignant tumours are directly involved in an increased catabolism of cholesterol-rich low-density lipoprotein (LDL) particles. In a prospective study of 25 patients with colorectal carcinoma, we measured intraindividual shifts in serum cholesterol levels after surgery, and the expression of LDL-receptor mRNA in surgically removed specimens. A significant rise in plasma cholesterol levels was observed in patients 3 and 12 months after curative surgery, but not after non-curative surgery. In human colon carcinoma tissues LDL receptor mRNA expression, as determined by competitive reverse-transcriptase-polymerase-chain reaction, was found to be significantly increased when compared to tissues from the tumour-free margin (median values, 1.2 x 10(6) vs. 2.0 x 10(5) molecules/micrograms total cellular RNA, respectively, n = 17). The extent of LDL-receptor mRNA expression positively correlated to the percentage rise of plasma cholesterol levels 3 months (n = 7, r = 0.8763) and 12 months (n = 6, r = 0.9181) after curative surgery. This finding provides in vivo evidence that the tumour tissue itself contributes to decreased plasma cholesterol levels in patients suffering from colorectal carcinomas. It supports the hypothesis that low cholesterol levels in cancer patients are a consequence, and not the cause, of the malignancy.

Adult↗

PubMed1995

Effects of pravastatin, a hydroxymethylglutaryl-CoA reductase inhibitor, on two human tumour cell lines.

Competitive inhibitors of 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase are currently used to treat patients with hypercholesterolaemia. These inhibitors affect not only cholesterol biosynthesis, but also the production of non-steroidal mevalonate derivatives, that are involved in a number of growth-regulatory processes. As a consequence, their potential use as anticancer drugs has been suggested. In order to examine long-term effects of this potential therapeutic approach, we cultivated the gastric carcinoma cell line, EPG85-257, and the breast tumour cell line, MDA-MB231, in the presence of increasing concentrations of the HMG-CoA reductase inhibitor, pravastatin. For both cell lines, this procedure led to the selection of resistant variants able to proliferate in more than 1000 microM inhibitor. By competitive reverse transcriptase/polymerase chain reaction assay (cRT-PCR), the expression of the mRNA for two key proteins of cellular cholesterol metabolism, HMG-CoA reductase and low-density lipoprotein (LDL) receptor, were analysed in sensitive and resistant cells. Despite similar growth rates, MDA-MB231 cells expressed approximately four times more HMG-CoA reductase mRNA than EPG85-257 cells and over 30 times more LDL receptor mRNA. Both mRNA species were coordinately regulated in the parental and in the pravastatin-resistant variant cells. Expression was highly stimulated (3- to 4-fold for the HMG-CoA reductase and 2- to 3-fold for the LDL receptor) in the resistant variants when cultured in lipoprotein-deficient medium in the presence of 1000 microM pravastatin. Immunocytological analysis of the expression of the HMG-CoA reductase and LDL receptor protein were in accordance with the data on specific mRNA expression obtained by cRT-PCR. Southern blot analysis revealed a 1.5-fold amplification of the HMG-CoA reductase gene in resistant MDA-MB231 cells, but not in the resistant EPG85-257 variant. Our data provide evidence for resistance mechanisms to pravastatin that are independent of the amplification of the HMG-CoA reductase gene. By analogy to the cell-culture models employed in this study, it is conceivable that similar mechanisms might occur in human tumour cells in vivo during long-term treatment with HMG-CoA reductase inhibitors. This might limit their application as chemotherapeutic anticancer agents.

Blotting, Southern↗

PubMed1994

[Quantitative analysis of mRNA expression of the LDL receptor and HMG-CoA reductase in bovine tissue with competitive RT-PCR].

The present study exemplifies the method of quantitative reverse transcriptase-polymerase chain reaction (cRT-PCR) by quantifying, in bovine tissues, the specific mRNAs for the two key proteins which regulate cellular cholesterol metabolism, the low density lipoprotein (LDL)-receptor and the 3-hydroxy-3-methylglutaryl-Coenzyme A (HMG-CoA) reductase. Our data reveal a broad range of expression for both mRNAs. The LDL-receptor mRNA expression was highest in the adrenal gland (6.7 x 10(4) molecules/micrograms cellular RNA) followed by the corpus luteum of the ovary, intermediate in the liver (8.6 x 10(2) molecules/micrograms cellular RNA) and not detectable in the large intestine. Expression of the HMG-CoA reductase almost paralleled LDL-receptor values. It was found to be highest in the ovary (6.1 x 10(6) molecules/micrograms cellular RNA) followed by the adrenal gland, intermediate in the liver (3.1 x 10(5) molecules/micrograms cellular RNA) and lowest in the large intestine (9.9 x 10(4) molecules/micrograms cellular RNA). These data are consistent with other reports concerning the quantitative analysis of the expression of both proteins as determined by other experimental approaches. Thus cRT-PCR is a valuable tool for the quantitative analysis of the LDL-receptor and the HMG-CoA reductase.

Adrenal Glands↗

PubMed1994

Rapid quantitation of mRNA species in ethidium bromide-stained gels of competitive RT-PCR products.

A rapid method for quantifying the low-abundant mRNAs of the low density lipoprotein receptor and the 3-hydroxy-3-methylglutaryl coenzyme A reductase by competitive polymerase chain reaction is presented. This approach requires neither special labeling nor blotting procedures. For each analysis, a defined amount of total cellular RNA is co-reverse transcribed and co-amplified with a titration series of in vitro synthesized competitor RNA that carries an internal deletion. The equivalence point, which defines the amount of specific RNA in the sample, can be scored in ethidium bromide-stained agarose or polyacrylamide gels of the reaction products. As an example, responses to pravastatin, a competitive inhibitor of the HMG-CoA reductase, in a human tumor cell line were analyzed with this new technique. As a control, the expression of the unregulated gene, glyceraldehyde-3-phosphate dehydrogenase was measured in parallel using the same methodology. The results obtained were compared with those obtained by conventional Northern blotting.

Base Sequence↗

PubMed1992

Elevated lipoprotein(a) levels in patients with acute myeloblastic leukaemia decrease after successful chemotherapeutic treatment.

Twenty-two patients with acute myeloblastic leukaemia (AML) were studied to investigate disease-associated changes in lipid metabolism. Lipoprotein (a) [Lp(a)] levels were found to be elevated at the time of diagnosis (median 23 mg/dl; 41% of patient group had levels greater than 25 mg/dl) and diminished after successful chemotherapeutic treatment in 9 of 10 cases, with a maximum decrease from 56 to 10 mg/dl. In contrast, reduced levels of total cholesterol, low density lipoprotein (LDL) and high density lipoprotein (HDL) (medians 137, 87 and 20 mg/dl, respectively) were observed at the time of diagnosis. Cholesterol and HDL levels increased in all 10 and LDL in 9 cases in which complete remission was achieved. These data suggest that the catabolism of LDL-cholesterol might be even more enhanced than assumed to date. Furthermore, it indicates that the Lp(a) level in acute myeloblastic leukaemia is influenced either directly or indirectly by the leukaemic blasts.

Adult↗

PubMed1991

Pertussis toxin inhibits autophosphorylation and activation of the insulin receptor kinase.

Pertussis toxin is an ADP-ribosyltransferase which alters the function of some of the GTP-binding proteins and inhibits some actions of insulin. In vivo, pertussis toxin (2 micrograms/ml/2h) inhibited insulin-stimulated tyrosyl autophosphorylation of the insulin receptor by 50% in FaO cells, and nearly completely inhibited phosphorylation of the cellular insulin receptor substrate pp185. Similarly, insulin-stimulated autophosphorylation and kinase activity of the insulin receptor purified on wheat germ agglutinin-agarose from pertussis toxin-treated FaO cells was diminished 50%; however, treatment of cells with the catalytically inactive B-oligomer of the toxin had no effect on receptor tyrosine kinase activity in vitro. Pertussis toxin did not alter insulin binding or the cellular levels of ATP, cAMP, and cGMP. Furthermore, immunoprecipitation of the insulin receptor from intact cells with anti-insulin receptor antibodies showed that pertussis toxin did not increase the phosphorylation of serine or threonine residues in the insulin receptor. These results suggest that pertussis toxin can modulate signal transduction of insulin at the level of the insulin receptor kinase.

Animals↗

PubMed1990

Cytotoxicity of a recombinant ricin-A-chain fusion protein containing a proteolytically-cleavable spacer sequence.

Chimeric proteins composed of ricin toxin A chain (RTA) and staphylococcal protein A (PA) have been produced in E. coli. Constructs consisting of N-terminal RTA and C-terminal PA (RTA-PA) or N-terminal PA and C-terminal (PA-RTA) were capable of binding to immunoglobulin G (via PA) and of specifically depurinating 28 S ribosomal RNA (via RTA). However, neither fusion protein was cytotoxic to antigen-bearing target cells in the presence of an appropriate monoclonal antibody presumably because the RTA could not be released from the PA within the cytosol where the ribosomal substrate of RTA is located. The overcome this, a short amino acid sequence from diphtheria toxin was engineered between the RTA and PA to produce a disulfide-linked loop containing a trypsin sensitive cleavage site. Cleavage of this fusion protein with trypsin converted the RTA-DT-PA to the two chain form consisting of RTA linked by a disulfide bond to PA. The cleaved fusion protein was highly toxic to Daudi cells coated with anti-immunoglobulin antibody suggesting that the RTA could be released from the PA by reduction within the cytosol.

Amino Acid Sequence↗

PubMed1989

Analysis of A-MuLV transformed fibroblast lines for amplification of the c-myc, p53 and c-fos nuclear proto-oncogenes.

Aberrations in nuclear proto-oncogene organisation and/or gene expression have been implicated in cell transformation mediated by the v-abl gene. For example, it has been suggested that amplification of the c-myc proto-oncogene is a co-operative event in v-abl induced fibroblast transformation. We have investigated amplification of the c-myc, p53 and c-fos nuclear proto-oncogenes in several Abelson murine leukaemia virus (A-MuLV) transformed fibroblast lines. None of these proto-oncogenes were detectably rearranged or amplified in v-abl transformed Swiss 3T3 lines. In contrast, NIH3T3 fibroblasts transformed by the v-abl gene consistently showed a 4 to 16-fold amplification of the c-myc gene. These data show that c-myc gene amplification is not an obligatory event associated with A-MuLV transformation, but may be restricted to cell lines derived from NIH3T3. c-myc gene amplification also did not correlate with a reduced latency period for tumour induction in nude mice. In addition, c-myc amplification was not selected during tumourigenesis, indicating that this event is not required for A-MuLV transformed Swiss 3T3 cells to display a full tumourigenic phenotype.

Abelson murine leukemia virus↗

PubMed1986

Abelson transformed fibroblasts lacking the EGF receptor are not tumourigenic in nude mice.

Cells transformed by the v-abl-oncogene produce large amounts of the tumour growth factor alpha TGF. alpha TGF is homologous to the epidermal growth factor (EGF) and stimulates cell growth via the EGF receptor pathway. To separate metabolic events in the v-abl-transformed cells mediated by alpha TGF as opposed to the v-abl-encoded protein-tyrosine kinase, we have employed the Swiss 3T3 variant cell line NR6 which lacks a functional EGF receptor. v-abl was found to transform efficiently NR6 cells in vitro. These transformed NR6 cells displayed a variety of in vitro properties which were indistinguishable from transformed wild-type fibroblast lines. However, in contrast to the wild-type lines, v-abl-transformed NR6 cells failed to form tumours when injected into athymic nude mice. These results imply an important function for alpha TGF and the EGF receptor in the establishment of the v-abl-induced fibrosarcomas.

Abelson murine leukemia virus↗