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Biomedical subjects

G Weigel

Publications and source records attributed to G Weigel.

At least 19 recordsLinked to original sources
PubMed2003

Quality of packed red blood cells and platelet concentrates collected by multicomponent collection using the MCS plus device.

The demand for blood components is constantly increasing, while the exclusion criteria for donors are strengthened in order to reach maximal safety for donors and patients. To counterbalance reduced availability of volunteers, multicomponent collections (MCC) is an attractive approach to produce more than one component during a single apheresis procedure from one donor, such as packed red blood cells (PRBCs) and platelet concentrates (PCs). Further, the exposures of patients to a limited number of donors reduces the possibility of alloimmunization and transfusion-related diseases. We measured the quality of PRBCs and PCs obtained by MCC, using the MCS+ device with the LDPRBC program, Revision B, and compared them with the quality of manually collected PRBCs and PCs collected with the Revision C2 of the MCS+. We found higher pH levels and lower hemolysis assessed by means of fHb and K+ in the supernatant of PRBCs over the whole storage period of 42 days in MCC-derived PRBCs. The functional metabolism assessed by intracellular ATP was higher in PRBCs collected by MCC than in manually collected units. Furthermore, PCs obtained during MCC showed an increase in p-selectin expression on day 5 of storage compared to PCs collected with the Revision C2 of the MCS+. The p-selectin expression on MCC platelets was within the range of p-selectin expression found in PCs obtained by other apheresis devices. These results indicate less storage lesion in MCC-derived PRBCs compared to manually collected units and no compromise in the quality of MCC PCs obtained in the same apheresis procedure.

Adenosine Triphosphate↗
PubMed2003

Early failure of the tissue engineered porcine heart valve SYNERGRAFT in pediatric patients.

OBJECTIVES: The first tissue engineered decellularized porcine heart valve, Synergraft (Cryolife Inc., USA) was introduced in Europe as an alternative to conventional biological valves. This is the first report of the rapid failure of these new grafts in a small series. MATERIALS AND METHODS: In 2001, 2 model 500 and 2 model 700 Synergraft valves were implanted in four male children (age 2.5-11 years) in the right ventricular outflow tract as a root. Two patients had a Ross operation and two had a homograft replacement. RESULTS: The cryopreserved Synergraft valves appeared macroscopically unremarkable at implantation. Recovery from surgery was uneventful and good valve function was demonstrated postoperatively. Three children died, two suddenly with severely degenerated Synergraft valves 6 weeks and 1 year after implantation. The third child died on the 7th day due to Synergraft rupture. Subsequently the fourth graft was explanted prophylactically 2 days after implantation. Macroscopically all four grafts showed severe inflammation starting on the outside (day 2 explant) leading to structural failure (day 7 explant) and severe degeneration of the leaflets and wall (6 weeks and 1 year explant). Histology demonstrated severe foreign body type reaction dominated by neutrophil granulocytes and macrophages in the early explants and a lymphocytic reaction at 1 year. In addition significant calcific deposits were demonstrated at all stages. Surprisingly pre-implant samples of the Synergraft revealed incomplete decellularization and calcific deposits. No cell repopulation of the porcine matrix occurred. CONCLUSION: The xenogenic collagen matrix of the Synergraft valve elicits a strong inflammatory response in humans which is non-specific early on and is followed by a lymphocyte response. Structural failure or rapid degeneration of the graft occurred within 1 year. Calcific deposits before implantation and incomplete decellularization may indicate manufacturing problems. The porcine Synergraft treated heart valves should not be implanted at this stage and has been stopped.

Animals↗
PubMed2003

Comparison of different decellularization procedures of porcine heart valves.

BACKGROUND: Tissue engineering of heart valves should avoid the disadvantages of conventional prostheses. In this study we tested different decellularization procedures for their potential of cell removal and their ability to preserve the matrix. METHODS: Specimens of porcine aortic and pulmonary roots were treated with either trypsin or sodium-dodecyl-sulfate (SDS) or Triton-X 100 and sodium-deoxycholate with a range of concentrations. Tissue samples were then processed for scanning electron microscopy and laser scanning microscopy. RESULTS: Trypsin achieved only incomplete decellularization and caused severe structural alterations of the matrix. In contrast SDS removed cells completely but caused strong structural alterations. Treatment with Triton-X100 and sodium-deoxycholate achieved both complete decellularization and preservation of the matrix structure. CONCLUSION: Techniques of decellularization are highly variable in efficiency and matrix preservation and was best achieved in our study with Triton-X100 and sodium deoxycholate.

Animals↗
PubMed2002

A clinical study of a fibrinogen-based collagen fleece for dural repair in neurosurgery.

BACKGROUND: There is frequently a need for dural grafts to cover defects resulting from retraction, shrinkage, or excision following neurosurgical procedures. Many substances have been tried as possible dural substitution, and different tissues and materials have been evaluated for use in dural repair. METHOD: The authors performed a retrospective review of 288 consecutive neurosurgical procedures using a fibrinogen based collagen fleece (TachoComb), a resorbable mesh of collagen from horse tendons, coated with human fibrinogen, bovine thrombin, bovine aprotinin and riboflavin (for marking the coated side), for dural substitution. The fibrinogen and thrombin imitate the last step of the coagulation cascade. On contact with bleeding wounds or other body fluids the coagulation factors dissolve and a link is formed between the collagen carrier and the wound surface. Thrombin converts fibrinogen into fibrin by splitting off peptides. Aprotinin prevents premature lysis of the fibrin clot by plasmin. FINDINGS: Neither superficial or deep wound infections nor aseptic meningitis were noted. We found good fibrous incorporation of TachoComb into the surrounding normal dura. Postoperative cerebrospinal-fluid (CSF) leaks developed in only five cases, who had to be re-operated, upon as well as one patient with a rebleeding. In another four cases, there was notable subcutaneous cerebrospinal-fluid accumulation without CSF-leak. They required a lumbar cerebrospinal-fluid drainage. INTERPRETATION: We conclude that TachoComb is a valuable alternative to the patients fibrous tissues for dural repair in cases in which autogenous tissues are either unavailable or insufficient for proper reconstruction.

Aprotinin↗
PubMed2002

Depletion of intracellular GTP results in nuclear factor-kappaB activation and intercellular adhesion molecule-1 expression in human endothelial cells.

The expression of the intercellular adhesion molecule 1 (ICAM-1) on the surface of endothelial cells plays an important role in immune-mediated processes. The induction by the proinflammatory cytokine interleukin (IL)-1beta is regulated by nuclear transcription factor kappaB (NF-kappaB). We studied the effect of an inosine-5'-monophosphate dehydrogenase (IMPDH) inhibitor, mycophenolic acid (MPA), on constitutive and IL-1beta-induced expression of ICAM-1 in human umbilical vein endothelial cells (HUVECs). Unexpectedly, pretreatment with MPA enhanced the constitutive expression and potentiated the induction of ICAM-1 by IL-1beta, as detected by flow cytometry. Northern blot analysis revealed an increase in ICAM-1 mRNA levels in cells treated with MPA. This was associated with an increase in phosphorylation of IkappaB-alpha (an inhibitor of NF-kappaB), nuclear translocation of the NF-kappaB subunits p50 and p65 and their binding to DNA as detected by Western blotting, confocal microscopy, and electrophoretic mobility shift assay. The up-regulation of ICAM-1 by MPA was prevented by high doses (100 microM) of guanine or guanosine but not by physiological doses (0.1 microM), indicating that guanylates are involved in endothelial responses to IL-1beta. Cultivation of HUVECs in the absence of guanine enhanced further ICAM-1 expression during IMPDH inhibition. These results demonstrate that cytokine-mediated endothelial ICAM-1 expression can be modulated by IMPDH inhibition. We believe this represents a novel interaction between endothelial guanylate metabolism, NF-kappaB activation, and adhesion molecule expression.

Cells, Cultured↗
PubMed2001

Prostaglandin-J2 induces synthesis of interleukin-8 by endothelial cells in a PPAR-gamma-independent manner.

PPARgamma is a transcription factor of nuclear receptor superfamily, involved in the regulation of inflammation. We investigated the influence of PPARgamma-ligands, 15-deoxy-delta12,14 prostaglandin-J2 (15d-PGJ2), and ciglitazone, on the generation of interleukin-8 (IL-8) by the human microvascular endothelial cell line (HMEC- 1). Expression of PPARgamma in HMEC-1 was confirmed by RT-PCR. Both PPARgamma-ligands tested induced the activation of PPAR, but the potency of ciglitazone was higher, as evidenced by luciferase assay. Resting HMEC-1 released about 150 pg/ml of IL-8 protein. Treatment with LPS increased the IL-8 secretion up to 1 ng/ml. 15d-PGJ2 potently and dose-dependently increased both the steady-state and LPS-induced generation of IL-8 mRNA and IL-8 protein. In contrast, neither basal nor LPS-elicited expression of IL-8 was influenced by ciglitazone. We conclude, that 15d-PGJ2 is a potent inducer of IL-8 production and can be a mediator of inflammatory response, but this effect is independent of PPARgamma activation.

Cell Line↗
PubMed2001

Altered intracellular purine nucleotides in gamma-irradiated red blood cell concentrates.

BACKGROUND AND OBJECTIVES: Gamma irradiation at a dose of 30 Gy induces deterioration of erythrocytes, resulting in storage lesions that significantly shorten the shelf-life of packed red cell concentrates (RCCs). The aim of the present study was to investigate the effect of gamma irradiation on intracellular purine nucleotides of red blood cells during storage. MATERIALS AND METHODS: Three-day-old leucocyte-depleted saline-adenine-glucose-mannitol (SAGM)-preserved RCCs, obtained from the Blood Service of the Austrian Red Cross, were gamma irradiated with 30 Gy. Samples were taken on days 1, 2, 3 and 7 after irradiation and subsequently at weekly intervals up to the end the of shelf-life (day 39 after irradiation) and were investigated for the K+ and Na+ content in the supernatant, for intracellular concentrations of ATP, ADP, ITP, IDP, GTP and GDP of erythrocytes, and for haemolysis. RESULTS: Within the first 24 h after gamma irradiation, no metabolic or biochemical changes were detectable in the RCCs. The K+ concentration in the supernatant increased after 24 h, while the Na+ concentration decreased in irradiated units and this ion disequilibrium persisted until the end of the shelf-life. After an initial increase of intracellular ATP, ADP and GTP during the first week of storage, the intracellular concentrations of ATP, ADP, GTP and ITP decreased, while IDP increased. The decrease of ATP and ADP was found to be more pronounced in irradiated units. At the end of the shelf-life, the ATP, GTP and ITP concentrations of irradiated RCCs had decreased to < 10% of the initial level and the critical threshold of 0.8% haemolysis was reached. CONCLUSION: Gamma irradiation of SAGM-preserved RCCs leads to serious deterioration of the purine nucleotide metabolism of erythrocytes during storage, which can reduce the in vivo recovery of the transfused red cells.

Blood Preservation↗
PubMed2001

Effect of mycophenolate mofetil therapy on inosine monophosphate dehydrogenase induction in red blood cells of heart transplant recipients.

BACKGROUND: Mycophenolic acid is reported to provide effective immunosuppression by inhibiting inosine monophosphate dehydrogenase. In an attempt to monitor the biological effects of long-term therapy with mycophenolate mofetil, we measured levels of guanosine 5' triphosphate and adenosine 5' triphosphate in red blood cells (RBCs) of patients after heart transplantations. METHODS: Fifty-two patients enrolled in the study were randomly assigned to one of two groups. Patients in the control group (n = 27) received cyclosporine A (INN, ciclosporin), azathioprine, and prednisone. Patients in the study group (n = 25) were switched from azathioprine to mycophenolate mofetil 3 months after the heart transplantation. Adenosine 5' triphosphate and guanosine 5' triphosphate levels were determined by means of HPLC. The activities of inosine monophosphate dehydrogenase and hypoxanthine-guanine phosphoribosyltransferase, which are responsible for guanine nucleotide formation, were measured in RBCs by radiochemical methods. RESULTS: Adenosine 5' triphosphate levels were unchanged in patients treated with mycophenolate mofetil, whereas those of the control group who received azathioprine (from 142 +/- 26 pmol/10(6) RBCs to 165 +/- 25 pmol/10(6) RBCs; P <.001) increased. As the length of mycophenolate mofetil therapy increased, patients in the study group showed significantly elevated guanosine 5' triphosphate levels (15.6 +/- 6.1 pmol/10(6) RBCs versus 6.6 +/- 2.1 pmol/10(6) RBCs; P <.001) and a 5-fold increase in inosine monophosphate dehydrogenase activity (108.6 +/- 13.3 pmol/mg of protein per hour versus 22.5 +/- 1.7 pmol/mg of protein per hour; P <.001) compared with the control group. In addition, a slight but significant enhancement of hypoxanthine-guanine phosphoribosyltransferase activity was seen in the mycophenolate mofetil group. CONCLUSIONS: Our studies have shown that long-term administration of mycophenolate mofetil is associated with increasing guanosine 5' triphosphate levels in RBCs as the result of an induction of inosine monophosphate dehydrogenase and hypoxanthine-guanine phosphoribosyltransferase activities in erythrocytes.

Adenosine Triphosphate↗
PubMed2000

Adhesion molecule behavior during rejection and infection episodes after heart transplantation.

In cardiac transplant recipients the release of soluble cellular adhesion molecules intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-Selectin into serum is pronounced during immune activation. It is uncertain whether there is a specific pattern of release during infection or cardiac allograft rejection. In a prospective study, 30 consecutive cardiac allograft recipients were followed for a median period of 11.4 months (range 1-34). Soluble ICAM-1 (sICAM-1), soluble VCAM-1 (sVCAM-1) and soluble E-Selectin (sE-Selectin) were measured in addition to acute phase proteins (C-reactive protein, alpha1-antitrypsin), complement factors (C3, C4) and beta2-microglobulin. The measured serum levels were correlated with the clinical status of the transplant recipient: 1) uneventful clinical status; 2) asymptomatic infection; 3) symptomatic infection and 4) rejection. Forty age-matched healthy subjects served as controls. Six days before biopsy-proven cardiac allograft rejection sICAM-1-release started to increase (p < 0.05) as compared to uneventful clinical status. The peak concentration of sICAM-1 was measured three days before rejection. On the day of rejection, serum concentrations of sICAM-1 (p < 0.001) and sVCAM-1 (p < 0.05) were increased, whereas sE-Selectin was not markedly elevated. In symptomatic infections, the serum concentrations of sICAM-1 (p < 0.001) and sVCAM-1 (p < 0.05) were elevated at the day of diagnosis and both parameters reached peak levels three days after onset of chemotherapy. In multivariate analysis soluble adhesion molecules only weakly discriminated between rejection and infection (sensitivity: 13%, specificity: 95%). Although, in combination with routine blood parameters the discriminatory power could be improved (sensitivity: 85%, specificity: 85%) the clinical utility of these markers in non-invasive monitoring is limited.

Adult↗
PubMed1999

Azathioprine and 6-mercaptopurine alter the nucleotide balance in endothelial cells.

Graft vascular disease after solid organ transplantation is a main complication that limits long-term survival of the graft. An injury of the endothelium and subsequent vascular response is considered to be responsible for smooth muscle cell hyperplasia with resulting luminal narrowing. What is less certain are the precise steps leading to endothelial injury and subsequent vessel disease. Since the immunosuppressive drug azathioprine is in clinical use due to its antiproliferative effect on lymphocytes, we were interested in how far it exerts effects on the vascular endothelium. Azathioprine and its metabolite 6-mercaptopurine, a potent inhibitor of purine salvage pathway enzymes, dose dependently led to decreased endothelial cell proliferation as well as to decreases in intracellular purine nucleotides adenosine-triphosphate and guanosine-triphosphate. By increasing the formation of the pyrimidine nucleotide uridine-triphosphate within 24 hours, azathioprine and its metabolite altered the endothelial nucleotide balance. Since not only the formation of toxic thio- and methylthiopurines (thio-guanosine-monophosphate, methyl-thio-inosine-monophosphate) was measured, the activity of the enzyme thiopurinemethyltransferase was induced (3.21+/-2.04 U per 10(9) cells, mean+/-SD). These findings indicate that the vascular endothelium plays an active role in the metabolization of the established immunosuppressant azathioprine that then exerts specific toxic effects on endothelial cells.

Azathioprine↗
PubMed1999

Evaluation of the EMIT Mycophenolic Acid Assay from Dade Behring.

The Emit Mycophenolic Acid Assay, a new homogeneous enzyme immunoassay for the quantitative analysis of mycophenolic acid (MPA) in human plasma, was evaluated and compared to a high-performance liquid chromatography (HPLC) method. Coefficients of variation (CV) of the within-run imprecision (n = 10) varied from 2.5% to 4.4% and from 1.3% to 4.9% for the Emit and the HPLC, respectively. The CV's of between-day imprecision (n = 10) ranged from 7.9% to 10.8% for the Emit and from 4.7% to 12.1% for the HPLC. Mean recoveries were 95.6% and 100.1% for Emit and HPLC, respectively. Serial dilution of a patient pool demonstrated a linear relationship between expected (x) and measured (y) concentrations: Emit, y = 0.998x + 0.086; HPLC, y = 1.006x - 0.016. The detection limit and the lower limit of quantification were 0.087 mg/L and 0.20 mg/L for Emit. The detection limit for HPLC was 0.08 mg/L using a signal-to-noise ratio of three. Sample stability under various storage conditions was satisfactory, although storage at -20 degrees C is recommended for storage longer than one day. No cross-reactivity from the major metabolite mycophenolic acid glucuronide (MPAG) was found. A correlation study on 261 patient samples yielded the following regression equation (bivariate Deming procedure): Emit = 1.012HPLC + 0.244, r = 0.970.

Adult↗
PubMed1999

Mycophenolic acid influences T helper 2 (Th2) cytokine induced expression of intercellular cell adhesion molecule-1 (ICAM-1) on human endothelial cells.

A possible synergistic effect of mycophenolic acid (MPA) and the immunosuppressive cytokines IL-4, IL-10 and IL-13 was investigated in human umbilical vein endothelial cells (HUVEC). These cytokines, which are produced by Th2-lymphocytes, were shown to induce the expression of vascular cell adhesion molecule-1 (VCAM-1) and production of IL-6 in endothelial cells. In this study we found IL-4 to induce both intercellular cell adhesion molecule-1 (ICAM-1) and VCAM-1 expression, whereas IL-13 induced VCAM-1 only. The surface expression of E-selectin was not influenced by any of the cytokines tested. MPA on its own led to statistically significant ICAM-1 expression on HUVEC. The combination of MPA with IL-4 led to a significant ICAM-1 expression in an additive manner compared to the cytokine alone. In contrast, MPA neither induced VCAM-1 nor did it influence the effects of IL-4 and IL-13 on VCAM-1 expression. A clinically relevant concentration of mycophenolic acid (10 micromol/l) decreased intracellular guanosine-5'-triphosphate (GTP) levels significantly. Since intracellular nucleotides are responsible for the glycosylation of proteins, a disturbance of the endothelial nucleotide balance could be responsible for the effects of MPA on ICAM-1. Guanine and guanosine prevented and partially reversed the actions of MPA, on both intracellular GTP and ICAM-1 expression, which strongly implies that MPA by interfering with nucleotide metabolism, affects the adhesive properties of endothelial cells, and by acting synergistically with IL-4 probably influences Th2 cytokine effects.

Cell Adhesion Molecules↗
PubMed1999

Alterations of endothelial nucleotide levels by mycophenolic acid result in changes of membrane glycosylation and E-selectin expression.

The effect of the inhibitor of inosine-5'-monophosphate dehydrogenase (IMPDH), mycophenolic acid, on intracellular nucleotides and the synthesis of cellular glycoproteins was evaluated in human umbilical vein endothelial cells. A clinically attainable concentration (10 micromol/l) of mycophenolic acid decreased guanosine-5'-triphosphate (GTP) levels significantly and led to a strong elevation of uridine-5'-triphosphate (UTP), whereas intracellular adenosine-5'-triphosphate (ATP) pools remained unaffected. The staining of the endothelial cell membranes with lectins specific for fucose and mannose (Ulex europaeus- and Galanthus nivalis agglutinin, respectively) was reduced, reflecting an inhibition of fucose and mannose incorporation into endothelial glycoproteins. The surface expression of E-selectin, an important determinant for leuko-endothelial interactions decreased significantly. Guanine and guanosine prevented the actions of mycophenolic acid and reversed the drug-induced decrease in GTP and its associated effects. The findings that mycophenolic acid produces alterations in the formation of glycoproteins and in the membrane architecture are indicative of metabolic lesions induced by an agent that depresses guanine nucleotide synthesis through inhibition of IMPDH. The pronounced reduction of E-selectin surface expression on endothelial cells accompanied by changes of endothelial cell fucosylation, a prerequisite for the contact with lymphocytic L-selectin, indicates an inhibitory effect of mycophenolic acid in the rolling phase of leukocyte recruitment and strongly implies a new and additional immunosuppressive mechanism of this agent.

Adenosine Triphosphate↗
PubMed1999

A simple HPLC method for monitoring mycophenolic acid and its glucuronidated metabolite in transplant recipients.

Mycophenolic acid (MPA) is nowadays in broad clinical use as a substitute for azathioprine. An immunoassay for MPA recently received approval for clinical applications. The high performance liquid chromatography (HPLC) assay for measuring MPA and its glucuronide conjugate (MPAG) we describe here is not only rapid and simple but also extremely sensitive at plasma levels obtained during standard immunosuppressive regimens. The determination of MPAG is possible without any change of the chromatographic conditions (detection wavelength of 214 nm, mobile phase: acetonitrile and 50 mmol/l o-phosphoric acid (50:50, V/V), run time: 15 min). The required equipment is a standard HPLC system including a simple UV-detector. Sample volume of 400 microl is required for both determinations. Detection limit is 0.25 micromol/l for MPA and 5 micromol/l for MPAG. Linearity is excellent for serial dilutions (0.5-25 micromol/l for MPA, 25-500 micromol/l for MPAG) and high accuracies favour the method described. More than 2000 plasma samples tested for MPA in patients after heart transplantation within one year and more than 500 samples for MPAG underline the clinical applicability of this assay.

Blood Chemical Analysis↗